Patent Application
20240417393
The technical field relates to certain 2,5-diazabicyclo[4.2.0]octanes, to their use in the treatment of cardiovascular disease and metabolic conditions, for example type 2 diabetes, and to pharmaceutical compositions containing them.
Obesity and type 2 diabetes (T2D) are major and growing health problems worldwide (Lancet, 2014, 9922, 1068-1083). The two diseases are strongly associated with each other, with obesity proceeding development of insulin resistance and T2D. T2D is associated with several comorbidities including cardiovascular disease, renal disease, hypertension, stroke, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) (Lancet, 2005, 9468, 1415-1428).
Incretin hormones including GLP-1 (glucagon-like peptide-1) and GIP (glucose-dependent insulinotropic polypeptide) are gut peptides that are secreted after nutrient intake and stimulate insulin secretion (Diabetes ObesMetab., 2018, 20(Suppl.1), 5-21). GLP-1 secretion from the gut is impaired in obese subjects which may indicate a role in the pathophysiology of obesity (Regulatory Peptides, 2004, 122, 209-217).
GLP-1 is secreted from the L-cells in the lower gut in response to food intake. GLP-1 stimulates insulin secretion from the pancreatic P-cells, in a glucose dependent manner (Diabetologia, 1993, 36, 741-744). GLP-1 also inhibits glucagon secretion, reduces appetite and slows down gastric emptying. The GLP-1 receptor is also present in the heart, kidneys and immune system and activation has been shown to reduce blood pressure, increase natriuresis and decrease inflammation.
GLP-1 is a 37-amino acid peptide, post-translationally processed from pro-glucagon, a 158 amino acid precursor polypeptide (www.uniprot.org, pro-glucagon entry P01275). Several other peptides are also derived from proglucagon and processed in a tissue specific manor, including glucagon and oxyntomodulin. GLP-1 has very short half-life in vivo as it is rapidly degraded by dipeptidyl peptidase-4 (DPP-IV) (Front. Endocrinol. 2019, 10, Article 260, 1-10).
Incretin-based glucose- and body weight-lowering medications include GLP-1 receptor agonists, DPP-IV inhibitors and more recently also combinations of GLP-1 agonists and glucose-dependent insulinotropic polypeptide (GIP) agonists (Peptides, 2020, 125, Article 170202). Traditionally GLP-1 analogues are peptide hormones which have been modified to minimize DPP-IV cleavage and are administered as injectables. The first oral GLP-1 peptide was recently approved but bioavailability is low and the drug needs to be administered in the fasting state, 30 min before nutrient intake which may limit patient compliance (JAMA, 2017, 318(15), 1460-1470). The injectable peptides show increased efficacy over the oral peptides but are limited by the route of administration. Small molecule GLP-1 receptor agonists are in development from several companies and are expected to provide a therapeutic benefit versus peptide based therapies due to early use in the treatment paradigm.
Pharmacological stimulation of GLP-1 receptors has been shown to significantly reduce HbAlc levels, provide long term weight loss and reduce blood pressure. GLP-1 receptor agonists have also been shown to reduce cardiovascular events and prolong life in high-risk patients with T2D and are therefore recommended by the European Association for the Study of Diabetes (EASD) and American Diabetes Association (ADA) in patients with multiple risk factors of cardiovascular disease (CVD) independent of the patients glycemic control (Diabetes Care, 2020, 43, 487-493).
There remains a need for an easily-administered prevention and/or treatment for cardiometabolic and associated diseases.
WO2018/109607 discloses 6-carboxylic acids of benzimidazoles and 4-aza-, 5-aza-, 7-aza- and 4,7-diazabenzimidazoles as GLP-1 receptor agonists, processes to make said compounds, and methods comprising administering said compounds to a mammal in need thereof.
WO2019/239319 and WO2019/239371 disclose 6-carboxylic acids of benzimidazoles and 4-aza-, 5-aza- and 7-aza-benzimidazoles as GLP-1 receptor agonists, processes to make said compounds, and methods comprising administering said compounds to a mammal in need thereof.
WO2020/103815 disclose GLP-1 receptor agonist compounds and pharmaceutical compositions thereof, for use in e.g. treating type 2 diabetes mellitus, pre-diabetes, obesity, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis and cardiovascular disease.
WO2020/207474 disclose GLP-1 receptor agonist compounds and pharmaceutical compositions thereof, for use in e.g. treating type 2 diabetes mellitus, pre-diabetes, obesity, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis and cardiovascular disease.
WO2020/234726 disclose combinations of GLP-1 receptor agonist compounds and pharmaceutical compositions thereof and an acetyl-CoA carboxylase (ACC) inhibitor or a diacylglycerol acyltransferase (DGAT2) inhibitor, or a ketohexokinase (KHK) inhibitor or farnesoid X receptor (FXR) agonist, for use in e.g. treating type 2 diabetes mellitus, pre-diabetes, obesity, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis and related diseases.
WO2020/263695 discloses glucagon-like peptide-1 receptor agonists and therapeutic uses of the compounds to treat type II diabetes mellitus.
WO2021/081207 discloses compounds that bind to and act as agonists or modulators of the glucagon-like peptide-1 receptor (GLP-1R) and act as agonists or modulators of GLP-1R. The disclosure further relates to the use of the compounds for the treatment and/or prevention of diseases and/or conditions by said compounds.
WO2021/018023 discloses compounds for modulating a Glucagon-like peptide-1 (GLP-1) receptor, and a pharmaceutical use thereof.
WO2021/096284 and WO2021/096304 discloses compounds that act as GLP-1 receptor agonists, for use as therapeutic agents for metabolic diseases.
WO2021/112538 discloses compounds which serves as a GLP-1 receptor agonist and may be useful in the prevention or treatment of a disease associated with GLP-1 activity.
WO2021/154796 discloses GLP-1R agonists, and compositions, methods, and kits thereof. Such compounds are generally useful for treating a GLP-1R mediated disease or condition.
WO2021/160127 discloses GLP-1 agonists, pharmaceutical compositions, and methods of use thereof.
WO2021116874 discloses of solid forms of 2-[[4-[(S)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl]piperidin-1-yl]methyl]-1-[[(S)-oxetan-2-yl]methyl]-1H-benzo[d]imidazole-6-carboxylic acid, 1,3-dihydroxy-2-(hydroxymethyl)propan-2-amine salt for pharmaceutical use.
CN113493447A discloses a compound that can be used as a GLP-1 receptor agonist. WO2021197464 discloses fused imidazole derivatives, preparation methods and medical use as a therapeutic agent, especially as GLP-1 receptor agonists.
CN113480534A discloses benzimidazole or azabenzimidazole-6-carboxylate compound that can activate GLP-1R downstream signaling pathway.
WO2021154796 discloses compounds as GLP-1R agonists, and compositions, methods, and kits thereof.
WO2021219019 discloses GLP-1 agonists of formula I, including pharmaceutically acceptable salts and solvates thereof, pharmaceutical compositions, and methods of using the same.
WO2021244645 discloses five-membered heteroaromatic imidazole compounds I and their medical use.
WO2021249492 discloses methyl-substituted benzobisoxazole compound and the use thereof in the preparation of drugs for treating related diseases.
CN113816948A discloses fused imidazole derivatives as GLP-1 receptor agonist in the treatment of diabetes.
WO2021254470 discloses preparation of 6-oxo-3,6-dihydropyridine derivative and a pharmaceutical composition containing the derivative, are used as therapeutic agents, in particular as GLP-1 receptors agonist and in the preparation of drugs for the treatment and/or prevention of diabetes.
WO2022007979 discloses a fused imidazole derivative, a preparation method therefor, a pharmaceutical composition containing the derivative, and the use of same as a therapeutic agent, in particular the use thereof as a GLP-1 receptor agonist.
CN113831337A discloses heterocyclic nitrogen compounds as GLP-1 receptor agonist.
WO2022068772 discloses a kind of benzimidazole derivative, its preparation method and application as GLP-1R agonists.
WO2022042691 discloses GLP-1 agonists, including pharmaceutically acceptable salts and solvates thereof, and pharmaceutical compositions including the same.
WO2022040600 discloses compounds that may be used as a glucagon-like peptide-1 receptors (GLP-1R) agonist.
WO2022028572 discloses GLP-1 agonists, including pharmaceutically acceptable salts and solvates thereof, and pharmaceutical compositions including the same.
WO2022031994 discloses compounds and pharmaceutical compositions thereof, for use in, e.g. treating type 2 diabetes mellitus, pre-diabetes, obesity, non-alc. fatty liver disease, non-alc. steatohepatitis, and cardiovascular disease.
CN114591308A discloses piperazine-imidazole containing GLP-1R receptor agonist compounds and application thereof.
WO2022111624 discloses benzimidazole derivatives that are agonists of a glucagon-like peptide-1 receptor (GLP-1R).
WO2022109182 discloses polyheterocyclic benzimidazole compounds and their preparation and use in the treatment of GLP-1R mediated diseases.
CN114478497A discloses a kind of aryl alkyl acid GLP-1 receptor agonist, its preparation method and application in treatment or prevention of GLP-1-mediated diseases and related diseases.
WO2022078380 discloses compounds that are GLP-1 agonists.
WO2022078407 discloses compounds that are GLP-1 agonists.
WO2022078152 discloses a kind of benzimidazolone compounds, their preparation method and application as GLP-1 receptor agonist.
CN114716423A discloses 5,6-dihydro-1,2,4-triazine compounds as GLP-1 receptor agonist.
CN114634510A discloses imidazolopyridine derivatives, which can be used to prepare drugs for treating GLP-1 receptor agonist mediated diseases.
CN114591296A discloses aromatic heterocyclic derivatives as GLP-1R agonists.
WO2022192430 discloses GLP-1R agonists and compositions, methods, and kits thereof.
WO2022192428 discloses GLP-1R agonists and compositions, methods, and kits thereof.
WO2022184849 discloses GLP-1R agonists, uses and pharmaceutical compositions thereof.
CN114907351A discloses tricyclic GLP-1 receptor agonists.
WO2022165076 discloses substituted benzimidazolecarboxylic acids which are GLP-1 receptor modulator compounds.
CN114805336A discloses fused imidazole compounds that are GLP-1 receptor agonists.
CN114763352A discloses benzimidazole derivatives and its application as GLP-1 receptor agonist.
J. Med. Chem. 2022, 65, 12, 8208-8226 discloses A Small-Molecule Oral Agonist of the Human Glucagon-like Peptide-1 Receptor.
Cell Research 2020, (39), 1140-1142 discloses structural insights into the activation of GLP-1R by a small molecule agonist.
An object is to provide novel GLP-1 receptor modulators useful in therapy. A further object is to provide novel compounds having improved safety profile, e.g with regards to selectivity for the GLP-1 receptor over e.g. phosphodiesterase 3 (PDE3) and/or having is improved metabolic stability in the body.
There is provided compounds that are modulators of the glucagon-like peptide-1 (GLP-1) receptor, their use as medicaments, pharmaceutical compositions containing them and synthetic routes to their production.
In one embodiment, there is provided a compound of Formula (I),
The compounds of Formula (I) are modulators of the GLP-1 receptor. Thus, the compounds of Formula (I) can be used as a medicament, in particular for disorders, disease or conditions responsive to modulation of the GLP-1 receptor, and more specifically cardiovascular disease and metabolic conditions.
In another embodiment there is provided a compound of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula (I), wherein the stereochemistry is undefined, e.g. a racemate or a mixture of diastereomers.
In another embodiment there is provided a compound of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula (I), wherein the stereochemistry is defined.
In another embodiment there is provided a pharmaceutical formulation comprising a therapeutically effective amount of a compound of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula (I), and a pharmaceutically acceptable diluent, excipient and/or inert carrier.
In a further embodiment there is provided a pharmaceutical formulation comprising a compound of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula is (I), for use in the treatment of a condition where modulation of the GLP-1 receptor would be beneficial.
In a further embodiment there is provided a compound of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula (I), for use in therapy, especially in the treatment of cancer in a mammal, particularly a human.
In a further embodiment there is provided the use of a compound of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula (I), for the manufacture of a medicament for the treatment of cardiovascular disease and metabolic conditions.
According to another aspect there is provided a process for the preparation of compounds of Formula (I), or pharmaceutically acceptable salts of compounds of Formula (I), and the intermediates used in the preparation thereof.
The compounds of Formula (I) described herein have the advantage that they may be more efficacious, be less toxic, be more selective, be more potent, produce fewer side effects, be more easily absorbed, and/or have a better pharmacokinetic profile (e.g. higher oral bioavailability and/or lower clearance), than compounds known in the prior art.
This detailed description and its specific examples, while indicating embodiments, are intended for purposes of illustration only. Therefore, there is no limitation to the illustrative embodiments described in this specification. In addition, it is to be appreciated that various features that are, for clarity reasons, described in the context of separate embodiments, also may be combined to form a single embodiment. Conversely, various features that are, for brevity reasons, described in the context of a single embodiment, also may be combined to form sub-combinations thereof. is Listed below are definitions of various terms used in the specification and claims.
It is to be understood that where in this specification a group is qualified by “defined above” the said group encompasses the first occurring and broadest definition as well as each and all of the other definitions for that group.
In this specification, the term “modulator” is used to describe a compound that exhibit varying receptor agonism, either full agonism, or partial agonism.
It is to be understood that in this specification “C1-4” means a carbon group having 1, 2, 3 or 4 carbon atoms.
It is to be understood that in this specification “C1-2” means a carbon group having 1 or 2 carbon atoms.
In this specification, unless stated otherwise, the term “alkyl” includes both straight and branched chain alkyl groups and may be, but is not limited to, methyl, ethyl, n-propyl, i-propyl, n-butyl, sec-butyl, iso-butyl or tert-butyl.
It is to be understood that in this specification “(5- to 6-membered)heteroaryl” means an aromatic ring with 5 to 6 atoms and containing one or more heteroatoms independently selected from nitrogen, oxygen or sulphur.
It is to be understood that in this specification “(6-membered)heteroaryl” means an aromatic ring with 6 atoms and containing one or more heteroatoms independently selected from nitrogen, oxygen or sulphur.
It is to be understood that in this specification “(6-membered)heteroaryl” means for example pyridine.
It is to be understood that in this specification “(5-membered)heteroaryl” means an aromatic ring with 5 atoms and containing one or more heteroatoms independently selected from nitrogen, oxygen or sulphur.
It is to be understood that in this specification “(4- to 6-membered)heterocycloalkyl” means a partially or completely saturated ring system with 4 to 6 atoms and wherein at least one of the ring carbon atoms is replaced with a heteroatom independently selected from nitrogen, oxygen or sulphur.
It is to be understood that in this specification a “heterocycloalkyl” substituent may be attached via a nitrogen atom having the appropriate valences, or via any is ring carbon atom.
It is to be understood that in this specification a “heterocycloalkyl” or “heteroaryl” substituent may be further substituted, for example by a substituent selected from C1-2alkyl.
In this specification, unless stated otherwise, the term “pharmaceutically acceptable” is used to characterize a moiety (e.g. a salt, dosage form, or excipient) as being appropriate for use in accordance with sound medical judgment. In general, a pharmaceutically acceptable moiety has one or more benefits that outweigh any deleterious effect that the moiety may have. Deleterious effects may include, for example, excessive toxicity, irritation, allergic response, and other problems and complications.
There is provided compounds of Formula (I) wherein X1, X2, Z1, Z2, Z3, R1-R7, m, n, p and q are as defined in Formula (I).
In one embodiment X1 is N or C1-2.
In a further embodiment X1 is N.
In still a further embodiment X1 is C1-2.
R1 is 0, 1, 2 or 3 substituents independently selected from F, Cl, Br, CN, OCH3, OCFH2, OCF2H, OCF3, CH3, CFH2, CF2H and CF3.
In one embodiment X2 is independently N or C, provided that no more than two atoms in the aromatic ring A are N.
In a further embodiment X2 is C.
In one embodiment Z1 is N or CR3.
In a further embodiment Z1 is N.
In still a further embodiment Z1 is CR3.
R3 is selected from H, F, Cl, N(CH3)2, C1-2alkyl and OC1-2alkyl, wherein said C1-2alkyl is substituted by 0, 1, 2 or 3 F.
In one embodiment Z2 and Z3 are each independently N or CR4, provided that when Z1 or Z3 is N, Z2 is CR4.
In a further embodiment Z1 and Z2 are N.
In still a further embodiment Z1 and Z3 are N.
In still a further embodiment Z2 and Z3 are N.
In still a further embodiment Z1 is N, Z2 and Z3 are CR4.
In still a further embodiment Z2 is N, Z1 and Z3 are CR4.
In still a further embodiment Z3 is N, Z1 and Z2 are CR4.
In still a further embodiment Z1, Z2 and Z3 are CR4.
R4 is independently selected from H, F, Cl, OH, CH3, CFH2, CF2H, CF3, OCH3, OCFH2, OCF2H and OCF3.
In one embodiment R1 is 0, 1, 2 or 3 substituents independently selected from F, Cl, Br, CN, OCH3, OCFH2, OCF2H, OCF3, CH3, CFH2, CF2H and CF3.
In a further embodiment R1 is 0, 1 or 2 substituents independently selected from F, Cl, Br, CN, OCH3, OCFH2, OCF2H, OCF3, CH3, CFH2, CF2H and CF3.
In still a further embodiment R1 is 0, 1, 2 or 3 substituents independently selected from F, Cl, Br, CN, OCH3
In still a further embodiment R1 is 0, 1 or 2 substituents independently selected from F, Cl, Br, CN, OCH3.
In still a further embodiment R1 is 0, 1 or 2 substituents independently selected from F, Cl and CN.
In still a further embodiment R1 is 0 or 1 substituents selected from F, Cl and CN.
In one embodiment R2 is selected from 0 or 1 F, Cl or CN.
In one embodiment R3 is selected from H, F, Cl, N(CH3)2, C1-2alkyl and OC1-2alkyl, wherein said C1-2alkyl is substituted by 0, 1, 2 or 3 F.
In a further embodiment R3 is selected from H, F, Cl, C1-2alkyl and OC1-2alkyl, wherein said C1-2alkyl is substituted by 0, 1, 2 or 3 F.
In still a further embodiment R3 is selected from H, F, Cl, CH3, CFH2, CF2H, CF3, OCH3, OCFH2, OCF2H and OCF3.
In still a further embodiment R3 is selected from H, F, Cl, CH3 and OCH3.
In one embodiment R4 is independently selected from H, F, Cl, OH, CH3, CFH2, CF2H, is CF3, OCH3, OCFH2, OCF2H and OCF3.
In a further embodiment R4 is independently selected from H, F, Cl, OH, CH3 and OCH3-In still a further embodiment R4 is independently selected from H, F, Cl, CH3 and OCH3.
In still a further embodiment R4 is independently selected from H, F and Cl.
In one embodiment R5 is selected from H, CH3, CFH2, CF2H and CF3.
In a further embodiment R5 is selected from H, and CH3.
In still a further embodiment R5 is CH3.
In one embodiment R6 is selected from (4- to 6-membered)heterocycloalkyl, (5- to 6-membered)heteroaryl, CN, C1-4alkyl, O(C1-4alkyl), S(C1-4alkyl), cyclopropyl, cyclobutyl, O(cyclopropyl) or S(cyclopropyl), wherein said (4- to 6-membered)heterocycloalkyl and (5 to 6-membered)heteroaryl is substituted by 0 or 1 substituent selected from C1-2alkyl and wherein said C1-4alkyl is substituted by 0 or 1 substituent selected from CN or OCH3, and 0, 1, 2 or 3 F and wherein said cyclopropyl and cyclobutyl is substituted by 0 or 1 substituent selected from CN, OCH3, OCFH2, OCF2H, OCF3 and CH2CN and 0, 1, 2 or 3 F.
In a further embodiment R6 is selected from C1-4alkyl, O(C1-4alkyl) and S(C1-4alkyl), wherein said C1-4alkyl is substituted by 0 or 1 substituent selected from CN or OCH3, and 0, 1,2 or 3F.
In still a further embodiment R6 is selected from cyclopropyl, cyclobutyl, O(cyclopropyl) or S(cyclopropyl), said cyclopropyl and cyclobutyl is substituted by 0 or 1 substituent selected from CN, OCH3, OCFH2, OCF2H, OCF3 and CH2CN and 0, 1, 2 or 3 F.
In still a further embodiment R6 is selected from (4- to 6-membered)heterocycloalkyl and (5 to 6-membered)heteroaryl, wherein said (4- to 6-membered)heterocycloalkyl and (5- to 6-membered)heteroaryl is substituted by 0 or 1 substituent selected from C1-2alkyl.
In still a further embodiment R6 is selected from (5- to 6-membered)heteroaryl, wherein said (5- to 6-membered)heteroaryl is substituted by 0 or 1 substituent selected from C1-2alkyl.
In still a further embodiment R6 is selected from (4- to 6-membered)heterocycloalkyl, is wherein said (4- to 6-membered)heterocycloalkyl is substituted by 0 or 1 substituent selected from C1-2alkyl.
In still a further embodiment R6 is oxetan-2-yl.
In one embodiment R7 is independently selected from F, C1-2alkyl and OC1-2alkyl, wherein said C1-2alkyl is substituted by 0, 1, 2 or 3 substituents independently selected from F.
In a further embodiment R7 is independently selected from F, C1-2alkyl and OC1-2alkyl.
In still a further embodiment R7 is independently selected from F, CH3 and OCH3.
In one embodiment m is 0, 1, 2 or 3.
In a further embodiment m is 0, 1, or 2.
In still a further embodiment m is 1 or 2
In still a further embodiment m is 0 or 1.
In still a further embodiment m is 1.
In still a further embodiment m is 0.
In one embodiment n is 0 or 1.
In a further embodiment n is 1.
In still a further embodiment n is 0.
In one embodiment p is 1, 2 or 3.
In a further embodiment p is 1 or 2.
In still a further embodiment p is 1.
In one embodiment q is 0, 1 or 2.
In a further embodiment q is 0 or 1.
In still a further embodiment q is 0.
In one embodiment, there is provided a compound of Formula (Ia),
In a further embodiment, there is provided a compound of Formula (Ia), wherein
In still a further embodiment, there is provided a compound of Formula (Ia),
In one embodiment, there is provided a compound of Formula (Ib),
In a further embodiment, there is provided a compound of Formula (Ib), is wherein
In still a further embodiment, there is provided a compound of Formula (Ib),
In one embodiment, there is provided a compound of Formula (Ic),
In a further embodiment, there is provided a compound of Formula (Ic), wherein
In still a further embodiment, there is provided a compound of Formula (Ic), wherein
In one embodiment, there is provided a compound of Formula (Id),
In a further embodiment, there is provided a compound of Formula (Id), wherein
In still a further embodiment, there is provided a compound of Formula (Id), wherein
In one embodiment, there is provided a compound of Formula (Ie),
In a further embodiment, there is provided a compound of Formula (Ie), wherein
In still a further embodiment, there is provided a compound of Formula (Ie), wherein
In one embodiment, there is provided a compound of Formula (If),
In a further embodiment, there is provided a compound of Formula (If), wherein
In still a further embodiment, there is provided a compound of Formula (If), wherein
In one embodiment the compounds of Formula (I) are selected from:
It shall be noted that any one of these specific compounds may be disclaimed from any of the herein mentioned embodiments.
In one embodiment there is provided a process for the preparation of compounds of Formula is (I), or pharmaceutically acceptable salts of compounds of Formula (I), and the intermediates used in the preparation thereof.
Another embodiment is a product obtainable by any of the processes or examples disclosed herein.
The compounds of Formula (I) and their pharmaceutically acceptable salts are believed to be useful in the prevention or treatment of cardiovascular disease and metabolic conditions, including but not limited to type 2 diabetes (T2D), obesity, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH), in a mammal, particularly a human.
For the avoidance of doubt, as used herein, the term “treatment” includes therapeutic and/or prophylactic treatment.
When a compound or salt described herein is administered as therapy for treating a disorder, a “therapeutically effective amount” is an amount sufficient to reduce or completely alleviate symptoms or other detrimental effects of the disorder, cure the disorder, reverse, completely stop, or slow the progress of the disorder or reduce the risk of the disorder getting worse.
The compounds described herein are thus indicated both in the therapeutic and/or prophylactic treatment of these conditions.
The compounds described herein have the advantage that they may be more efficacious, be less toxic, be more selective, be more potent, produce fewer side effects, be more easily absorbed, and/or have a better pharmacokinetic profile (e.g. higher oral bioavailability and/or lower clearance), than compounds known in the prior art.
For the above-mentioned therapeutic indications, the dosage administered will vary with the compound employed, the mode of administration and the treatment desired. However, in general, satisfactory results are obtained when the compounds are administered at a dosage of the solid form of between 1 mg and 2000 mg per day.
The compounds of Formula (I), and pharmaceutically acceptable derivatives thereof, may be used on their own, or in the form of appropriate pharmaceutical compositions in which the compound or derivative is in admixture with a pharmaceutically acceptable is adjuvant, diluent or carrier. Thus, another aspect concerns a pharmaceutical composition comprising a novel compound of Formula (I), or a pharmaceutically acceptable salt thereof, in admixture with a pharmaceutically acceptable adjuvant, diluent or carrier. Administration may be by, but is not limited to, enteral (including oral, sublingual or rectal), intranasal, inhalation, intravenous, topical or other parenteral routes. Conventional procedures for the selection and preparation of suitable pharmaceutical Formulations are described in, for example, Pharmaceuticals—The Science of Dosage Form Designs, M. E. Aulton, Churchill Livingstone, 2nd Ed. 2002. In one embodiment the pharmaceutical composition comprises less than 80% and in another embodiment less than 50% of a compound of Formula (I), or a pharmaceutically acceptable salt thereof.
In one embodiment there is provided a compound selected from any one of the compounds of Formula (I), or a pharmaceutically acceptable salt of a compound of Formula (I), for use in therapy, especially in the prevention or treatment of cardiovascular disease and metabolic conditions, including but not limited to type 2 diabetes (T2D), obesity, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH).
These and other embodiments are described in greater detail herein below, where further aspects will be apparent to one skilled in the art from reading this specification.
The compounds of Formula (I), or pharmaceutically acceptable salts thereof, may also be administered in conjunction with other compounds used for the treatment of the above conditions.
In another embodiment, there is a combination therapy wherein a compound selected from any one of the compounds of Formula (I), or a pharmaceutically acceptable salt thereof, and a second active ingredient are administered concurrently, sequentially or in admixture, for the treatment of one or more of the conditions listed above. Such a combination may be used in combination with one or more further active ingredients.
When used in a combination therapy, it is contemplated that the a compound selected from any one of the compounds of Formula (I), or pharmaceutically acceptable salts thereof, is and the other active ingredients may be administered in a single composition, completely separate compositions, or a combination thereof. It also is contemplated that the active ingredients may be administered concurrently, simultaneously, sequentially, or separately. The particular composition(s) and dosing frequency(ies) of the combination therapy will depend on a variety of factors, including, for example, the route of administration, the condition being treated, the species of the patient, any potential interactions between the active ingredients when combined into a single composition, any interactions between the active ingredients when they are administered to the animal patient, and various other factors known to physicians (in the context of human patients), veterinarians (in the context of non-human patients), and others skilled in the art.
There is provided a method of treatment of a condition where modulation of GLP-1 receptor is required, which method comprises administration of a therapeutically effective amount of a compound selected from any one of the compounds of Formula (I) to a person suffering from, or susceptible to, such a condition.
The compounds of Formula (I) will normally be administered via the oral, topical, parenteral, intravenous, intramuscular, subcutaneous or in other injectable ways, buccal, rectal, vaginal, transdermal and/or nasal route and/or via inhalation, in the form of pharmaceutical preparations comprising the active ingredient or a pharmaceutically acceptable salt thereof, in a pharmaceutically acceptable dosage form. Depending upon the disorder and patient to be treated and the route of administration, the compositions may be administered at varying doses. Conventional procedures for the selection and preparation of suitable pharmaceutical Formulations are described in, for example, Pharmaceuticals—The Science of Dosage Form Designs, M. E. Aulton, Churchill Livingstone, 2nd Ed. 2002.
In one embodiment suitable daily doses of the compounds of Formula (I) in therapeutical treatment of humans are about 0.0001-100 mg/kg body weight, in another embodiment about 0.01-10 mg/kg body weight.
The optimum dosage and frequency of administration will depend on the particular condition being treated and its severity; the species of the patient; the age, sex, size and weight, diet, and general physical condition of the particular patient; brain/body weight ratio; other medication the patient may be taking; the route of administration; the Formulation; and is various other factors known to physicians and others skilled in the art.
According to a further aspect there is thus provided a pharmaceutical Formulation comprising a compound selected from any one of the compounds of Formula (I), or pharmaceutically acceptable derivatives thereof, in admixture with a pharmaceutically acceptable adjuvant, diluent and/or carrier.
The compound of Formula (I) may be present in the pharmaceutical Formulation in a concentration from 0.1 to 99.5%, such as from 0.5 to 95%, by weight of the total Formulation.
The protection and deprotection of functional groups is described in Protective Groups in Organic Synthesis, 4th Ed, T. W. Greene and P. G. M. Wuts, Wiley-Interscience (2006) and Protecting Groups, 3rd Ed, P. J. Kocienski, Georg Thieme Verlag (2005).
A further embodiment encompasses pharmaceutically acceptable salts of the compounds of Formula (I).
A salt of a compound selected from any one of Formula (I) may be advantageous due to one or more of its chemical or physical properties, such as stability in differing temperatures and humidities, or a desirable solubility in H2O, oil, or other solvent. In some instances, a salt may be used to aid in the isolation or purification of the compound. In some embodiments (particularly where the salt is intended for administration to an animal, e.g. a human, or is a reagent for use in making a compound or salt intended for administration to an animal), the salt is pharmaceutically acceptable.
The term “pharmaceutically acceptable” is used to characterize a moiety (e.g. a salt, dosage form, or excipient) as being appropriate for use in accordance with sound medical judgment. In general, a pharmaceutically acceptable moiety has one or more benefits that outweigh any deleterious effect that the moiety may have. Deleterious effects may include, for example, excessive toxicity, irritation, allergic response, and other problems and complications.
Where the compound is sufficiently basic, pharmaceutically acceptable salts include, but are not limited to, inorganic or organic acid addition salts.
For reviews on suitable salts, see Berge et al., J. Pharm. Sci., 1977, 66, 1-19 or Handbook of Pharmaceutical Salts: Properties, selection and use, P. H. Stahl, P. G. Vermuth, is IUPAC, Wiley-VCH, 2002.
Where an acid co-former is a solid at r.t. and there is no or only partial proton transfer between the compound of Formula (I) and such an acid co-former, a co-crystal of the co-former and compound of Formula (I) may result rather than a salt. All such co-crystal forms of the compound of Formula (I) are encompassed herein.
It is also to be understood that certain compounds of Formula (I) may exist in solvated form, e.g. hydrates, including solvates of a pharmaceutically acceptable salt of a compound of Formula (I).
In a further embodiment, certain compounds of Formula (I) may exist as racemates and racemic mixtures, single enantiomers, individual diastereomers and diastereomeric mixtures. Certain compounds of Formula (I) may also contain linkages (e.g. carbon-carbon bonds, carbon-nitrogen bonds such as amide bonds) wherein bond rotation is restricted about that particular linkage, e.g. restriction resulting from the presence of a ring bond or double bond. Stereoisomers may be separated using conventional techniques, e.g. chromatography or fractional crystallization, or the stereoisomers may be made by stereoselective synthesis.
In a further embodiment, the compounds of Formula (I) encompass any isotopically-labelled (or “radio-labelled”) derivatives of a compound of Formula (I). Such a derivative is a derivative of a compound of Formula (I) wherein one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number typically found in nature. Examples of isotopes that may be incorporated include 2H (also written as “D” for deuterium).
In a further embodiment, the compounds of Formula (I) may be administered in the form of a prodrug which is broken down in the human or animal body to give a compound of the Formula (I).
Various forms of prodrugs are known in the art. For examples of prodrug derivatives, see: Nature Reviews Drug Discovery 2008, 7, 255 and references cited therein. Intermediate compounds may also exist in enantiomeric forms and may be used as purified enantiomers, diastereomers, racemates or mixtures.
The following examples are non-limiting examples.
PrepMethod A: The compound was purified by preparative HPLC on a Kromasil C8 column (10 μm, 250×20 mm ID) using a gradient of MeCN in H2O/MeCN/FA (95/5/0.2) as mobile phase.
PrepMethod B: The compound was purified by preparative HPLC on a Waters™ Sunfire™ C18 OBD column (5 m, 150×30 mm ID) using a gradient of MeCN in H2O/FA (0.1 M) as mobile phase.
PrepMethod C: The compound was purified by preparative HPLC on a XSelect CSH OBD column (5 μm, 150×30 mm ID) using a gradient of MeCN in H2O/TFA (0.05%) as mobile phase.
PrepMethod D: The compound was purified by preparative SFC on a Waters™ BEH, (5 m, 250×30 mm ID) using MeOH/H2O/NH3 (20 mM) in CO2 as mobile phase.
PrepMethod E: The compound was purified by preparative HPLC on a Waters™ Sunfire™ column (5 μm, 100×19 mm ID) using a gradient of MeCN in H2O as mobile phase.
PrepMethod F: The compound was purified by preparative HPLC on a Kromasil C8 column (10 μm, 250×50 mm ID) using a gradient of MeCN in H2O/MeCN/FA (95/5/0.2) as mobile phase.
PrepMethod G: The compound was purified by preparative HPLC on a Waters™ Sunfire™ column (5 m, 100×19 mm ID) using a gradient of MeCN in H2O as mobile phase.
PrepMethod H: The compound was purified by preparative HPLC on a Waters™ Sunfire™ C18 OBD column (5 m, 150×30 mm ID) using a gradient of MeCN in H2O/FA (0.1%) as mobile phase.
PrepMethod I: The compound was purified by preparative HPLC on a XSelect CSH C18 OBD column (5 μm, 150×30 mm ID) using a gradient of MeCN in H2O/TFA (0.1%) as mobile phase.
PrepMethod J: The compound was purified by preparative HPLC on an Xbridge Prep OBD C18 column (5 μm, 150×30 mm ID) using a gradient of MeCN in a H2O/NH4HCO3 (10 mM)/NH3 (0.1%, aq) buffer system as mobile phase.
PrepMethod K: The compound was purified by preparative HPLC on an Xbridge Shield RP18 OBD column (5 μm, 150×30 mm ID) using a gradient of MeCN in a H2O/NH4HCO3 (10 mM)/NH3 (0.1%, aq) buffer system as mobile phase.
PrepMethod L: The compound was purified by preparative HPLC on a YMC-Actus Triart C18 (5 m, 150×30 mm ID) using a gradient of MeCN in H2O/NH4HCO3 (10 mM)/NH3 (0.1% aq) buffer system as mobile phase.
PrepMethod M: The compound was purified by preparative HPLC on a XBridge™ C18 column (10 m, 250×50 mm ID) using a gradient of MeCN in H2O/MeCN/NH3 (95/5/0.2) as mobile phase.
Relevant fractions were collected, combined and freeze-dried to give the purified compound or relevant fractions were collected, combined and concentrated at reduced pressure, extracted with DCM or EtOAc, and the organic phase was dried either over Na2SO4 or by using a phase-separator, and then concentrated at reduced pressure to give the purified compound.
ChemDraw is optionally using labels in the graphical representation of stereocenters is such as ‘&’ and ‘or’ to describe the configuration of the stereochemical centers present in the structure.
In general chemical structures of Examples and Intermediates containing the label ‘&’ at a stereocenter, means the configuration of such Example or Intermediate at that stereocenter is a mixture of both (R) and (S); and a label ‘or’ means the configuration of such Example or Intermediate at that stereocenter is either (S) or (R). Absolute, unspecified, ‘&’, and ‘or’ stereocenters can all be present in a single structure.
In general for structures of Examples and Intermediates where all of the stereocenters are designated as ‘&’, the structure is named with a “rac-” prefix. For structures of Examples and Intermediates where all of the stereocenters are designated as ‘or’, the structure is named with a “rel-” prefix.
In general Examples and Intermediate compounds are named using the descriptors (RS) and (SR) to denote general ‘&’ centers for chemical structures with multiple chiral centers where only some are designated as ‘&’. The descriptors (R*) and (S*) are used to denote the general ‘or’ centers for chemical structures with multiple chiral centers where only some are designated as ‘or’.
In general Examples and Intermediate compounds containing stereocenters having a relationship that is either cis or trans, are named using the descriptors (RS, SR) or (RS, RS), to denote chemical structures with multiple chiral centers where only some are designated as ‘&’.
In general, for structures of Examples and Intermediates where all stereocenters present are racemic, no flag is designated to the stereocenter(s) and the structure is drawn with straight bond(s) at each stereocenter.
In general, for structures of Examples and Intermediates where two or more stereocenters are present in a ring and fixed to each other and do not vary independently of each other, e.g. are cis or trans to each other, said stereocenters are drawn with stereobonds representing their internal relationship. Said stereocenters are labelled with an “&1” flag representing a mixture of cis-configuration or a mixture of trans-configuration, or an “orn” flag representing a single cis-isomer or a single trans-isomer with unknown absolute stereochemistry. In general, should the structure of said Example or Intermediate further contain one or more stereocenters that are racemic and not fixed in relation to the former stereocenters, said stereocenter(s) is drawn with straight bond(s) at said stereocenters.
In general the descriptors (r) and (s) are used to describe the absolute configuration of any pseudoasymmetric centers in the structures of Examples and Intermediates.
In general the label “Isomer 1” corresponds to the first eluted isomer, and “Isomer 2” corresponds to the second eluted isomer, on a given chiral TIPLC column and eluent, and are used to distinguish two isomers containing one or more stereocenters with absolute unknown configuration;
(xvii) Unless otherwise specified, all crystallographic measurements were performed at 175K on a Bruker Smart Apex II diffractometer operating in the ω scans mode. The intensity data were collected within the θmax≤26.0° using Mo-Kα radiation (λ=0.71078 Å).
The structure was solved by direct methods and refined by the full-matrix least-squares technique in the anisotropic approximation for non-hydrogen atoms using the Bruker SHELXTL program package.
(xviii) in addition to the ones mentioned above, the following abbreviations and units have been used:
DMAP (16 g, 130.97 mmol) was added to a solution of 4,7-diazaspiro[2.5]octane-5,8-dione (185 g, 1.32 mol) in DCM (2.5 L). Di-tert-butyl dicarbonate (576 g, 2.64 mol) was added in several batches, and the resulting solution was stirred at rt for 2 h. The reaction was quenched by the addition of sat NH4Cl (2.0 mL). The organic layer was combined and washed with brine (1.0 mL). The organic layer was dried over anhydrous Na2SO4, and concentrated under vacuum. The residue was purified by silica gel column chromatography (EtOAc:petroleum ether, 1:6) to give the title compound (380 g, 85%) as a white solid.
Di-tert-butyl 5.8-dihydroxy-4.7-diazaspiro[2.5]octane-4.7-dicarboxylate
A solution of diisobutylaluminium hydride in toluene (1 N, 1.47 L, 103.36 mmol) was added dropwise with stirring at −78° C. to a solution of 4,7-di-tert-butyl 5,8-dioxo-4,7-diazaspiro[2.5]octane-4,7-dicarboxylate Intermediate 1 (125 g, 367.25 mmol) in THE (1.5 L), and the reaction mixture was stirred at −78° C. for 2 h. The reaction was quenched by the addition of Rochelle salt (2 N, 1.5 L). The mixture was diluted with of DCM (2 L), and the solids were filtered out. The filtrate was dried over anhydrous Na2SO4 and concentrated under vacuum. The residue was purified by silica gel column chromatography (DCM:MeOH, 50:1) to give the title compound (340 g, 90%) as a light yellow semi-solid; MS (ESI) m/z [M+H]+ 345.
is BF3·OEt2 (124 g, 873.68 mmol) and triethylsilane (101 g, 868.62 mmol) was added in several batches, at −78° C., to a solution of 4,7-di-tert-butyl 5,8-dihydroxy-4,7-diazaspiro[2.5]octane-4,7-dicarboxylate Intermediate 2 (75 g, 217.77 mmol) in DCM (750 mL). The resulting solution was stirred at −78° C. for 2 h. Another batch was prepared as described above. The reaction was then quenched by the addition of water/ice (500 mL). The resulting solution was extracted with DCM (2×500 mL) and the combined organic layer was dried over anhydrous Na2SO4 and concentrated under vacuum. The residue was purified by silica gel column chromatography (EtOAc/hexane, 1:20-1:10), to give the title compound (75 g, 55%) as an off-white solid; 1H NMR (CDCl3) δ 4.3(s, 1H), 3.4 (m, 2H), 2.1 (s, 2H), 1.47 (s, 9H).
pTsOH (22 g, 127.76 mmol) was added to a solution of di-tert-butyl (1R,6S)-2,5-diazabicyclo[4.2.0]octane-2,5-dicarboxylate Intermediate 3 (40 g, 128.04 mmol) in THE (400 mL). The resulting solution was stirred at 60° C. for 3 h. Three additional batches were prepared as described above. The reaction mixture was cooled to rt, and the solids were filtered out. K2CO3 (320 g) was added with stirring to the resulting solution and stirred at rt for 1 h. The solids were filtered out. The resulting mixture was concentrated under vacuum. The residue was purified by silica gel column chromatography (DCM:MeOH, 1:0-10:1) to give the title compound (45 g, 41%) as yellow oil; MS (ESI) m/z [M+H]+ 213.
is DIPEA (8.45 mL, 48.36 mmol) was added to a solution of methyl 3,5-difluoro-4-nitrobenzoate (3.50 g, 16.12 mmol) and (S)-oxetan-2-ylmethanamine (1.40 g, 16.12 mmol) in THF/DMF (125 mL, 5:2), and the reaction mixture was stirred at 20° C. for 4 h. The solvent was removed under reduced pressure, and the residue was suspended in water (300 mL). The aqueous layer was extracted with EtOAc (3×500 mL), and the combined organic layer was dried over Na2SO4, filtered and evaporated. The crude product was purified by straight phase flash chromatography on silica (gradient: 10-20% EtOAc in petroleum ether) to give the title compound (4.50 g, 98%) as a yellow solid; MS (ESI) m/z [M+H]+ 285.0.
A suspension of methyl (S)-3-fluoro-4-nitro-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 5 (4.2 g, 14.78 mmol) and 10% Pd-C(1.57 g, 1.48 mmol) in THE (150 mL) was stirred under an atmosphere of H2 (g) at 3 atm and 25° C. for 2 h. The reaction mixture was filtered through celite and the filter cake was washed with MeOH (3×100 mL). The filtrate was concentrated under reduced pressure. The crude product was purified by straight phase flash chromatography on silica (gradient: 70-80% EtOAc in petroleum ether), to give the title compound (3.20 g, 85%) as a light red solid; MS (ESI) m/z [M+H]+ 254.95.
pTsOH (0.108 g, 0.57 mmol) was added to a solution of methyl (S)-4-amino-3-fluoro-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 6 (1.45 g, 5.70 mmol) and 2-chloro-1,1,1-trimethoxyethane (1.06 g, 6.84 mmol) in MeCN (10 mL) and the reaction mixture was stirred at rt for 18 h. The solvent was evaporated at reduced pressure and the crude compound was purified by straight phase flash chromatography on silica (gradient: 50-100% EtOAc in heptane) to give the title compound (1.54 g, 86%); MS (ESI) m/z [M+H]+ 313.26.
K2CO3 (5.43 g, 39.27 mmol) was added to a solution of methyl 3-fluoro-5-methoxy-4-nitrobenzoate (3 g, 13.09 mmol) and (S)-oxetan-2-ylmethanamine (1.14 g, 13.09 mmol) in THF/DMF (5:2, 110 mL) and the reaction mixture was stirred at 90° C. for 16 h. The solvent was removed under reduced pressure, and the residue was suspended in water (250 mL). The aqueous layer was extracted with EtOAc (3×250 mL), and the combined organic layer was dried over Na2SO4, filtered and evaporated at reduced pressure. The crude product was purified by straight phase flash column chromatography on silica (10-20% EtOAc in petroleum ether) to give the title compound (1.8 g, 46%) as a yellow solid; MS (ESI) m z [M+H]+ 297.1.
A suspension of Pd-C (0.144 g, 1.35 mmol) and methyl (S)-3-methoxy-4-nitro-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 8 (4 g, 13.50 mmol) in THE (100 mL) was stirred under an atmosphere of H2(g) at 2 atm and 15° C. for 3 h. The reaction mixture was filtered through celite and the filtercake was washed with MeOH (3×300 mL). The filtrate was concentrated under reduced pressure. The crude product was purified by straight phase flash column chromatography on silica (50-70% EtOAc in petroleum ether) to give the title compound (3.00 g, 83%) as a light yellow solid; MS (ESI) m/z [M+H]+ 267.3.
pTsOH (0.119 g, 0.63 mmol) was added to a solution of methyl (S)-4-amino-3-methoxy-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 9 (0.333 g, 1.25 mmol) and 2-chloro-1,1,1-trimethoxyethane (0.387 g, 2.50 mmol) in MeCN (10 mL) and the reaction mixture was stirred at 45° C. for 30 min. The reaction mixture was concentrated under reduced pressure and the residue was purified by straight phase flash column chromatography on silica (50-100% EtOAc in heptane) to give the title compound (0.155 g, 38%); MS (ESI) m/z [M+H]+ 325.0.
Pd2(dba)3 (0.366 g, 0.40 mmol) was added to a solution of 2-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-5-chloropyridine (WO2020234726) (1.306 g, 4 mmol), rac-is tert-butyl (1R,6S)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 4 (0.849 g, 4.00 mmol), RuPhos (0.747 g, 1.60 mmol) and sodium 2-methylpropan-2-olate (1.538 g, 16.00 mmol) in degassed toluene (10 mL). The reaction mixture was evacuated and backfilled with N2 (g) (×3) and then stirred at 100° C. for 10 min. The reaction mixture was cooled to rt and filtered through a pad of celite, and the solid cake was rinsed with toluene. The filtrate was concentrated under reduced pressure and the crude compound was purified by straight phase flash column chromatography on silica (gradient: 0-20% EtOAc:heptane) to give the title compound (0.640 g, 35%); MS (ESI) m/z [M+H]+ 458.1.
TFA (547 μl, 7.10 mmol) was added to a solution of rac-tert-butyl (1R,6S)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 11 (650 mg, 1.42 mmol) in MeCN (30 mL) and the reaction mixture was stirred at rt for 1 h. The reaction mixture was concentrated in vacuo, then co-evaporated with toluene (×1) and MeCN (×2) to give the crude step a) product; MS (ESI) m z [M+H]+ 359.9.
The crude product from Step a) was dissolved in MeCN (30 mL), and K2CO3 (588 mg, 4.26 mmol) and methyl (S)-2-(chloromethyl)-4-fluoro-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 7 (533 mg, 1.70 mmol) was added and the reaction mixture was heated at 30° C. for 18 h. The reaction mixture was cooled to rt and filtered through celite. The filtrate was concentrated in vacuo and the residue was purified by straight phase flash column chromatography on silica (gradient: 0-100% EtOAc in heptane). The product containing fractions was pooled and the solvent was removed by evaporation to give a crude residue. Another batch was prepared as described above (130 mg), and the crude products were combined and purified by preparative HPLC, PrepMethod A, (gradient 40-70%), to give the title compound (0.730 g, 81%); MS (ESI) m/z [M+H]+ 634.47.
The diastereomers of methyl 2-(((1RS,6SR)-5-((SR)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 12 (730 mg, 1.13 mmol) were separated by chiral chromatography on a LUX C4 column (250×30 mm, 5 μm), eluted with 35% EtOH/DEA (100/20 mM) in CO2, 130 bar, at a flow rate of 130 mL/min and detected at 220 nm;
The stereoisomers of the first eluted compound mixture (417 mg) were separated by chiral chromatography on a LUX C3 (OJ) column (150×4.6 mm, 5 μm), eluted with 8% MeOH/DEA (100/20 mM) in CO2, 130 bar, at a flow rate of 130 mL/min and detected at 220 nm;
TFA (169 μl, 2.20 mmol) was added to a solution of rac-tert-butyl (1R,6S)-5-((S)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 11 (202 mg, 0.44 mmol) in DCM (3.0 mL) and the reaction mixture was stirred at rt for 1 h. The mixture was evaporated at reduced pressure to give the sub-title compound; MS (ESI) m/z [M+H]+ 359.9. The crude product from Step a) was dissolved in MeCN (3 mL) and K2CO3 (182 mg, 1.32 mmol) and methyl (S)-2-(chloromethyl)-4-methoxy-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 10 (144 mg, 0.44 mmol) was added, and the reaction mixture was heated at 60° C. for 18 h. The reaction mixture was cooled to rt and filtered through delite. The filtrate was concentrated at reduced pressure and the crude product is was purified by straight phase flash column chromatography on silica (30-100% EtOAc in hexane) to give the title compound (235 mg, 83%); MS (ESI) m/z [M]+/[M+2H]+646.5/648.3.
The diastereomers of methyl 2-(((1RS,6SR)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 16 (235 mg, 0.36 mmol) were separated by chiral chromatography on a CHIRALPAK® IH column (5 μm, 250×30 mm ID), eluted with 25% EtOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 130 mL/min and detected at 230 nm;
The stereoisomers of the first eluted compound mixture (74 mg) were separated by chiral chromatography on a YMC SA (IA) column (5 μm, 250×30 mm ID), eluted with 15% EtOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 130 mL/min and detected at 220 nm;
The stereoisomers of the second eluted compound mixture (88 mg) were separated by chiral chromatography on a LUX C4 column (5 μm, 250×30 mm ID), eluted with 30% EtOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 130 mL/min and detected at 230 nm; The first eluted compound was collected and evaporated to give the title compound Isomer 3 Intermediate 19 (58 mg); MS (ESI) m/z [M+H]+ 646.54.
The second eluted compound was collected and evaporated to give the title compound Isomer 4 Intermediate 20 (47 mg); MS (ESI) m/z [M+H]+ 646.2.
DIPEA (15.44 mL, 88.43 mmol) and 5-bromo-1-chloro-3-fluoro-2-nitrobenzene (7.5 g, 29.48 mmol) were added dropwise to a solution of (S)-oxetan-2-ylmethanamine (2.57 g, 29.48 mmol) in THE (200 mL), and the reaction mixture was stirred at 60° C. for 3 h. The reaction mixture was diluted with EtOAc (300 mL) and washed with sat brine (4×300 mL). The organic layer was dried over Na2SO4, filtered and evaporated. The crude product was purified by flash chromatography on silica (0-50% EtOAc in petroleum ether) to give the title compound (9.00 g, 95%) as a yellow oil; MS (ESI) m/z [M+H]+ 321/323.
Fe(s) (24.66 g, 441.60 mmol) was added to a mixture of (S)-5-bromo-3-chloro-2-nitro-N-(oxetan-2-ylmethyl)aniline Intermediate 21 (14.2 g, 44.16 mmol) and NH4Cl (23.62 g, 441.60 mmol) in MeOH (400 mL) and water (100 mL) at 20° C., and the reaction mixture was stirred at 60° C. for 6 h. The reaction mixture was filtered and the precipitates were washed with MeOH (4×100 mL). The filtrate was concentrated under reduced pressure and the crude product was diluted with EtOAc (500 mL). The organic layer was washed sequentially with water (500 mL) and sat brine (500 mL), dried over Na2SO4, filtered and evaporated. The is crude product was purified by flash chromatography on silica (30-50% EtOAc in petroleum ether) to give the title compound (12.00 g, 93%) as a white solid; MS (ESI) m/z [M+H]+ 292/291.
A mixture of (S)-5-bromo-3-chloro-N1-(oxetan-2-ylmethyl)benzene-1,2-diamine Intermediate 22 (1.5 g, 5.14 mmol), Pd(dppf)Cl2·DCM (0.38 g, 0.51 mmol) and DIPEA (8.99 mL, 51.45 mmol) in MeOH (300 mL) was stirred under an atmosphere of CO(g) at 60 atm and 120° C. for 30 h. The solvent was removed under reduced pressure and the crude product was purified by flash chromatography on silica (20-25% EtOAc in petroleum ether) to give the title compound (1.0 g, 72%) as a white solid; MS (ESI) m/z [M+H]+ 271.
pTsOH (0.357 g, 1.88 mmol) was added to a solution of methyl (S)-4-amino-3-chloro-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 23 (5.08 g, 18.77 mmol) and 2-chloro-1,1,1-trimethoxyethane (3.77 g, 24.40 mmol) in MeCN (20 mL) and the reaction mixture was stirred at 50° C. for 30 min. 2-Chloro-1,1,1-trimethoxyethane (1.16 g, 7.51 mmol) was added and the reaction mixture was stirred at 50° C. for 20 min. The reaction mixture was diluted with EtOAc (10 mL) and extracted with NaHCO3 (aq, 2×3 mL). The organic layer was dried over MgSO4, filtered and concentrated at reduced pressure. The crude compound was purified is by flash chromatography on silica (50-100% EtOAc in heptane) to give the title compound (5.30 g, 86%); MS (ESI) m/z [M+H]+ 329.1.
A mixture of 2-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-5-chloropyridine (WO2020234726) (2.85 g, 8.73 mmol), rac-tert-butyl (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate (2.78 g, 13.1 mmol), RuPhos (815 mg, 1.75 mmol), Pd2(dba)3 (800 mg, 873 mol) and sodium tert-butoxide (2.52 g, 26.2 mmol) in toluene (50 mL) was stirred at 100° C. under an atmosphere of N2(g) overnight, and then cooled to rt. The reaction mixture was poured onto water (300 mL) and extracted with EtOAc (2×300 mL). The combined organic layer was washed with brine, dried over Na2SO4, and the solvent was removed under vacuum.
The crude product was purified by flash chromatography on silica (0-100% MTBE in hexane) to give the title compound (1.2 g 30%) as a yellow sticky oil; MS (ESI) m/z [M+H]+ 458.2.
A solution of HCl in Et2O (2M, 439 mg, 0.6 ml) was added to a solution of rac-tert-butyl (1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 25 (1.2 g, 2.2 mmol) in DCM (20 mL). The solution was stirred at rt for 8 h, and then concentrated under reduced pressure, and is the solids were dried in vacuo to give the hydrochloride of the title compound (1.03 g, 98%) as a beige solid; MS (ESI) m/z [M+H]+ 358.1.
K2CO3 (579 mg, 4.19 mmol) and methyl (S)-4-chloro-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate_Intermediate 24 (598 mg, 1.82 mmol) were added to a solution of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 26 (500 mg, 1.40 mmol) in MeCN (5 mL) and the reaction mixture was heated at 50° C. for 18 h. The reaction mixture was cooled to rt and filtered. The filtrate was purified by flash chromatography on silica (20-100% EtOAc in heptane) to give the title compound (0.555 g, 61%); MS (ESI) m/z [M+H]+ 652.1.
The diastereoisomers of methyl 4-chloro-2-(((1RS,6RS)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 27 (555 mg) were separated by chiral chromatography on a LUX C4 column (250×30 mm, 5 μm), eluted with 30% MeCN/EtOH/DEA (70/30/20 mM) in CO2, 125 bar, at a flow rate of 115 mL/min, and detected at 225 nm;
A mixture of 4-bromo-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxole (WO2020234726) (2.89 g, 8.42 mmol), rac-tert-butyl (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate (2.68 g, 12.63 mmol), RuPhos (786 mg, 1.68 mmol), Pd2(dba)3 (771 mg, 842 μmol) and sodium tert-butoxide (2.43 g, 25.27 mmol) in toluene (40 mL) was stirred under an atmosphere of N2(g) at 100° C. overnight and then cooled to rt. The reaction mixture was poured into water (100 mL) and extracted with EtOAc (3×100 mL). The combined organic layer was washed with brine, dried over Na2SO4, and the solvent was removed under vacuum.
The crude product was purified by flash chromatography on silica (0-100% MTBE in hexane) to give the title compound (1.1 g, 35%) as a yellow oil; MS (ESI) m/z [M+H]+ 475.0.
A solution of HCl in Et2O (2M, 439 mg, 0.6 mL) was added to a solution of rac-tert-butyl (1R,6R)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 31 (1.1 g, 2.3 mmol) in DCM (15 mL). The reaction mixture was stirred at ambient temperature for 8 h and then concentrated under reduced pressure to give the hydrochloride of the title compound (950 mg, 98%) as a beige solid; MS (ESI) m/z [M+H]+ 375.0.
DIPEA (824 mg, 6.38 mmol) was added to a suspension of rac-(1R,6R)-2-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane hydrochloride Intermediate 32 (598 mg, 1.59 mmol), methyl (S)-2-(chloromethyl)-4-methoxy-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 10 (518 25 mg, 1.59 mmol) and NaI (1.2 g, 7.97 mmol) in MeCN (5 mL) at 20° C., and the reaction mixture was heated at 60° C. overnight. The reaction mixture was concentrated under reduced pressure, and then diluted with EtOAc (10 mL). The organic layer was washed with sat NaHCO3 (aq, 20 mL) and brine (20 mL), dried over Na2SO4, and concentrated under reduced pressure. The crude product was purified by preparative HPLC, PrepMethod E, (gradient: 40-65%) to give the title compound (102 mg, 35%); MS (ESI) m/z [M+H]+ 663.2.
The diastereomers of Intermediate 33 (218 mg) were separated by chiral chromatography on a Whelk-01 column (250×50 mm, 5 μm), eluted with 35% MeCN/MeOH/DEA (50/50/20 mM) in CO2, 120 bar, at a flow rate of 400 mL/min and detected at 220 nm; the first eluted compound mixture was collected and evaporated to yield a mixture of isomers (148 mg); and the second eluted compound was collected and evaporated to yield the title compound Isomer 3, Intermediate 36 (28 mg); MS (ESI) m/z [M+H]+ 663.4.
The stereoisomers of the first eluted compound mixture (148 mg) were separated by chiral chromatography on a Chiralpak IH column (250×30 mm, 5 μm), eluted with 10% MeCN/MeOH/DEA (50/50/20 mM) in CO2, 120 bar, at a flow rate of 130 mL/min and detected at 230 nm;
TFA (354 μl, 4.60 mmol) was added to a solution of rac-tert-butyl (1R,6S)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 11 (421 mg, 0.92 mmol) in DCM (3.0 mL) and the reaction mixture was stirred at rt for 20 min. The reaction mixture was concentrated at reduced pressure to give the crude Step a) subtitle compound; MS (ESI) m/z [M+H]+ 358.3.
The crude product from Step a) was dissolved in MeCN (3 mL) and K2CO3 (381 mg, 2.76 mmol) and methyl (S)-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate (270 mg, 0.92 mmol) were added, and the reaction mixture was heated at 75° C. for 1 h. The mixture was cooled to rt and filtered through celite. The filtrate was collected and purified by flash chromatography on silica (30-100% EtOAc in heptane, then 5% EtOH in is EtOAc) to give the title compound (0.450 g, 79%); MS (ESI) m/z [M+H]+ 616.3.
The diastereomers of methyl 2-(((1RS,6SR)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 37 (480 mg) were separated by chiral chromatography on a Chiralpak IH column (250×30 mm, 5 μm), eluted with 25% EtOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 140 mL/min and detected at 230 nm and;
A mixture of methyl 2-(((1RS,6SR)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 and Isomer 2 Intermediate 38 (200 mg, 0.32 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (90 mg, 0.65 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title is compound Isomer 1 and Isomer 2 (0.120 g, 61.4%); MS (ESI) m/z [M+H]+ 602.4.
K2CO3 (463 mg, 3.35 mmol) and methyl (S)-2-(chloromethyl)-4-methoxy-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 10 (399 mg, 1.23 mmol) was added to a solution of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 26 (400 mg, 1.12 mmol) in MeCN (3 mL) and the reaction mixture was heated at 60° C. for 40 h. The reaction mixture was cooled to rt and filtered over celite. The filtrate was collected and purified by flash chromatography on silica (0-100% EtOAc in heptane) to give the title compound (0.47 g, 65%); MS (ESI) m z [M+H]+ 646.4.
The diastereoisomers of methyl 2-(((1RS,6RS)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 42 (470 mg) were separated by chiral chromatography on a DCPakA column (250×20 mm, 5 μm), eluted with 18% MeOH/DEA (100/20 mM) in CO2, 130 bar, at a flow rate of 70 mL/min and detected at 240 nm;
The stereoisomers of the first eluted compound mixture were separated by chiral chromatography on a Whelk-01 column (250×50 mm, 5 μm), eluted with 35% MeOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 400 mL/min and detected at 220 nm;
The stereoisomers of the first eluted compound mixture (107 mg) were separated by chiral chromatography on a Lux C3 (OJ) column (250×30 mm, 5 um), eluted with 15%
MeOH/MeCN/DEA (85/15/20 mM) in CO2, 130 bar, at a flow rate of 130 mL/min, and detected at 230 nm;
The stereoisomers of the second eluted compound mixture (120 mg) were separated by chiral chromatography on a Kromasil XT column (250×20 mm, 5 μm), eluted with 10% MeCN/MeOH/NH3 (50/50/20 mM) in CO2, 130 bar, at a flow rate of 70 mL/min, and detected at 254 nm;
The first eluted compound was collected and evaporated to give the title compound Isomer 3, Intermediate 45, (30 mg); MS (ESI) m/z [M+H]+ 646.5; and the second eluted compound was collected and evaporated to give the title compound Isomer 4, Intermediate 46, (54 mg); MS (ESI) m/z [M+H]+ 646.7.
DIPEA (3.73 ml, 21.35 mmol) was added to a solution of (S)-(tetrahydrofuran-2-yl)methanamine (1.08 g, 10.68 mmol) and 5-bromo-1-fluoro-3-methoxy-2-nitrobenzene (2.67 g, 10.68 mmol) in MeCN (20 mL) and the reaction mixture was stirred at 57° C. for 3 h. The reaction mixture was concentrated at reduced pressure and the residue was purified by flash chromatography on silica (0-20% EtOAc in heptane) to give the title compound (2.39 g, 68%); MS (ESI) m/z [M+H]+ 333.1.
DIPEA (6.07 ml, 34.73 mmol) and HSiCl3 (2.453 ml, 24.31 mmol) was added dropwise at 0° C. to a solution of (S)-5-bromo-3-methoxy-2-nitro-N-((tetrahydrofuran-2-yl)methyl)aniline Intermediate 47 (2.3 g, 6.95 mmol) in MeCN (20 mL). The reaction mixture was stirred at 0° C. for 2 min and then at rt for 30 min. NaHCO3(aq, 10 mL) was added dropwise and the biphasic mixture was stirred at rt for 30 min and then extracted with EtOAc (2×5 mL). The combined organic layer was dried over MgSO4, filtered and evaporated at reduced pressure to give the title compound (2.0 g, 96%); MS (ESI) m/z [M+H]+ 303.1.
DIPEA (8.70 ml, 49.80 mmol) was added to a suspension of (S)-5-bromo-3-methoxy-N1-((tetrahydrofuran-2-yl)methyl)benzene-1,2-diamine Intermediate 48 (1.5 g, 4.98 mmol) and Pd(dppf)Cl2 (292 mg, 0.40 mmol) in MeOH (20 mL) and the reaction mixture was stirred under an atmosphere of CO(g) at 9 atm and 85° C. for 16 h. The reaction mixture was filtered through a pad of celite, and the filter cake was rinsed with MeOH (10 mL). The filtrate was concentrated at reduced pressure and the residue was purified by flash chromatography on silica (20-60% EtOAc in heptane) to give the title compound (444 mg, 32%); MS (ESI) m z [M+H]+ 281.2.
pTsOH (149 mg, 0.78 mmol) was added to a solution of methyl (S)-4-amino-3-methoxy-5-(((tetrahydrofuran-2-yl)methyl)amino)benzoate Intermediate 49 (440 mg, 1.57 mmol) and 2-chloro-1,1,1-trimethoxyethane (485 mg, 3.14 mmol) in MeCN (10 mL) and the reaction mixture was stirred at 50° C. for 30 min. The reaction mixture was diluted with EtOAc (10 mL) and washed with NaHCO3 (aq, 2×3 mL). The organic layer was dried over MgSO4, filtered and evaporated at reduced pressure to give the title compound (450 mg, 85%); MS (ESI) m/z [M+H]+ 399.0.
is K2CO3 (406 mg, 2.93 mmol) and methyl (S)-2-(chloromethyl)-4-methoxy-1-((tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 50 (340 mg, 1.00 mmol) were added to a solution of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 26 (350 mg, 0.98 mmol) in MeCN (5 mL) and the reaction mixture was heated at 50° C. for 18 h. The reaction mixture was cooled to rt and EtOAc (10 mL) was added. The organic layer was washed with NaHCO3 (aq, 10 mL), dried over MgSO4 and the solvent was evaporated at reduced pressure. The crude product was purified by flash chromatography on silica (20-100% EtOAc in heptane, then 3% EtOH in EtOAc) to give the title compound (0.377 g, 58%); MS (ESI) m/z [M+H]+ 660.64.
The diastereoisomers of methyl 2-(((1RS,6RS)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 51 (377 mg) were separated by chiral chromatography on a Whelk-01 column (250×50, 10 μm), eluted with 40% MeOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 400 mL/min, and detected at 220 nm;
The stereoisomers of the first eluted compound mixture (191 mg) were separated by chiral chromatography on a Kromasil XT column (250×20, 5 μm), eluted with 10% MeOH/NH3 (100/20 mM) in CO2, 120 bar, at a flow rate of 100 mL/min, and detected at 254 nm; the first eluted compound was collected and evaporated to give the title compound Isomer 1, Intermediate 52 (29 mg); MS (ESI) m/z [M+H]+ 660.4; and the second eluted compound was collected and evaporated to give the title compound Isomer is 2, Intermediate 53 (52 mg); MS (ESI) m/z [M+H]+ 660.5.
The stereoisomers of the second eluted compound mixture (200 mg) were separated by chiral chromatography on a Kromasil XT (150×4.6, 5 μm), eluted with 6% MeOH/NH3 (100/20 mM) in CO2, 140 bar, at a flow rate of 70 mL/min, and detected at 240 nm; the first eluted compound was collected and evaporated to give the title compound Isomer 3, Intermediate 54 (31 mg); MS (ESI) m/z [M+H]+ 660.7.
K2CO3 (406 mg, 2.93 mmol) and methyl (S)-2-(chloromethyl)-4-methoxy-1-((tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 50 (340 mg, 1.00 mmol) was added to a solution of rac-(1R,6S)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 12 Step a) (350 mg, 0.98 mmol) in MeCN (5 mL) and the reaction mixture was heated at 50° C. for 18 h. The reaction mixture was cooled to rt and filtered through celite, the filtrate was collected and purified by flash chromatography on silica (20-100% EtOAc in heptane) to give the title compound (0.382 g, 59%); MS (ESI) m/z [M+H]+ 660.4.
The diastereoisomers of methyl 2-(((1RS,6SR)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 55 (382 mg) were separated by chiral chromatography on a Whelk-01 column (250×50, 10 m), eluted with 45% MeOH/DEA (100/20 mM) in CO2, 120 bar, at a flow rate of 400 mL/min, and detected at 220 nm;
The stereoisomers of the first eluted compound mixture were separated by chiral chromatography on a LUX C3 (OJ) column (250×30, 5 μm), eluted with 17% MeCN/MeOH/DEA (85/15/20 mM) in CO2, 130 bar, at a flow rate of 120 mL/min and detected at 240 nm;
The stereoisomers of the second eluted compound mixture (162 mg) were separated by chiral chromatography on a LUX C3 (OJ) column (250×30, 5 μm), eluted with 18% MeCN/MeOH/DEA (85/15/20 mM) in CO2, 130 bar, at a flow rate of 120 mL/min and detected at 240 nm;
Pd2(dba)3 (366 mg, 0.40 mmol) was added to a mixture of 4-bromo-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxole (WO2020234726) (1.37 g, 4.0 mmol), rac-tert-butyl (1R,6S)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 4 (849 mg, 4.00 mmol), RuPhos (747 mg, 1.60 mmol) and sodium 2-methylpropan-2-olate (1.54 g, 16.00 mmol) in degassed toluene (10 mL). The reaction mixture was evacuated and backfilled with N2(g) (×3) then stirred at 100° C. for 10 min. The mixture was cooled to rt and filtered through a pad of celite, and the pad was rinsed with toluene. The filtrate was collected and the solvent was evaporated. The crude compound was purified by flash chromatography on silica (0-20% EtOAc in heptane) to give the title compound (950 mg, 50%); MS (ESI) m/z [M+H]+ 475.3.
TFA (770 μl, 10.0 mmol) was added to a solution of rac-tert-butyl (1R,6S)-5-(2-(4-Chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 60 (950 mg, 2.00 mmol) in MeCN (30 mL) and the reaction mixture was stirred at rt for 1 h. The reaction mixture was concentrated under reduced pressure and co-evaporated with toluene (×1) and then MeCN (×2) to give the crude sub-title compound; MS (ESI) m/z [M+H]+ 375.2.
K2CO3 (829 mg, 6.00 mmol) and methyl (S)-2-(chloromethyl)-4-fluoro-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 7 (751 mg, 2.40 mmol) were added to a solution of the Step a) product in MeCN (30 mL) and the reaction mixture was heated at 30° C. for 18 h. The reaction mixture was cooled to rt and filtered. The filtrate was collected and the crude product was purified by flash chromatography on silica (0-100% EtOAc in heptane). The product containing fractions were collected and concentrated, and the crude product was purified by preparative HPLC, PrepMethod F, (gradient: 40-70%) to give the title compound (810 mg, 62%); MS (ESI) m/z [M+H]+ 651.2.
The stereoisomers of methyl 2-(((1RS,6SR)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 61 (810 mg) were separated by chiral chromatography on a Lux C4 column (250×50 mm, 5 μm), eluted with 18% EtOH/DEA (100/20 mM) in CO2, 135 bar, at a flow rate of 350 mL/min, and detected at 220 nm;
The stereoisomers of the first eluted compound mixture (318 mg) were separated by chiral chromatography on a Lux C3 (OJ) column (250×30 mm, 5 μm), eluted with 8% MeOH/DEA (100/20 mM) in CO2, 130 bar, at a flow rate of 130 mL/min, and detected at 220 nm; the first eluted compound was collected and evaporated to give the title compound Isomer 1, Intermediate 62 (114 mg, 14%); MS (ESI) m/z [M+H]+ 651.3; and
Methyl 3-chloro-5-fluoro-4-nitrobenzoate (7.93 g, 33.95 mmol), (1-ethyl-1H-imidazol-5-yl)methanamine (4.25 g, 33.95 mmol) and DIPEA (10.97 g, 84.87 mmol) were mixed in DMF (80 mL), and the reaction mixture was heated at 50° C. for 16 h. The mixture were poured into water (100 mL) and filtered. The precipitate was collected and dried to give the title compound (11.5 g) as dark yellow solid; MS (ESI) m/z [M+H]+ 325.6.
Wet Pt/C (1%, 1.99 g, 10.2 mmol) was added to a suspension of methyl 3-chloro-5-(((1-ethyl-1H-imidazol-5-yl)methyl)amino)-4-nitrobenzoate Intermediate 66 (11.52 g, 34.01 mmol) in MeOH (100 mL) and the reaction mixture was stirred under an atmosphere of H2 (g) at 1 atm and at 20° C. for 16 h. The reaction mixture was filtered, and the solids were washed with is MeOH (50 mL). The filtrate was concentrated under reduced pressure. The crude product was purified by flash chromatography on silica (0-40% MeOH in CHCl3) to give the title compound (4 g, 38%); 1H NMR (400 MHz, CDCl3): δ 7.587 (s, 1H), 7.583 (s, 1H), 7.28 (s, 1H), 6.68 (s, 1H), 4.5 (brs, 1H), 4.24 (s, 2H), 3.99-3.97 (q, 2H), 3.85 (s, 3H), 1.43 (t, 3H)
2,2,2-Triethoxyethan-1-ol (3.11 g, 17.46 mmol) and pTsOH (100 mg, 582 μmol) were added to a stirred solution of methyl 4-amino-3-chloro-5-(((1-ethyl-1H-imidazol-5-yl)methyl)amino)benzoate Intermediate 67 (1.8 g, 5.82 mmol) in MeCN (100 mL) and the reaction mixture was heated at 60° C. 18 h. The reaction mixture concentrated under reduced pressure, and the crude residue was diluted with EtOAc. The organic layer was washed with water and NaHCO3 (aq), dried and evaporated at reduced pressure to give the crude subtitle compound (2.04 g, 60% purity); MS (ESI) m/z [M+H]+ 349.2.
Methyl 4-chloro-1-((1-ethyl-1H-imidazol-5-yl)methyl)-2-(hydroxymethyl)-1H-benzo[d]imidazole-6-carboxylate Step a) (2.04 g, 5.85 mmol) was added in portions to a is vigorously stirred mixture of SOCl2 (6.97 g, 58.55 mmol) and DMF (one drop). After complete addition the reaction mixture was stirred at rt for 1 h, and then concentrated at reduced pressure to give the hydrochloride of the title compound (1.8 g, 71%) as white solid; MS (ESI) m/z [M+H]+ 367.2.
Methyl 4-chloro-2-(chloromethyl)-1-((1-ethyl-1H-imidazol-5-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 68 (281 mg, 766 μmol), rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane dihydrochloride Intermediate 26 (330 mg, 766 μmol), DIPEA (594 mg, 4.6 mmol) and NaI (459 mg, 3.06 mmol) were mixed in DMF (20 mL), and the reaction mixture was heated at 50° C. for 16 h. The mixture was cooled, diluted with water and extracted several times with EtOAc. The combined organic layer was washed with brine, dried over Na2SO4, and filtered.
The filtrate was concentrated under reduced pressure and the crude product was purified by preparative HPLC, PrepMethod E, (gradient: 40-65%) to give the title compound (152 mg, 29%); MS (ESI) m/z [M+H]+ 690.2.
The stereoisomers of rac-methyl 4-chloro-2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-ethyl-1H-imidazol-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 69 were separated by chiral chromatography on o Chiralpak IC III column (250×20 mm, 5 μm), eluted with hexane-IPA-MeOH (50:25:25), at a flow rate of 12 mL/min;
H2θ2(35% v/v) (148 mL, 3.51 mol) was added to a solution of 4-amino-5-fluoro-2-methoxybenzoic acid (25.0 g, 135.02 mmol) in TFA (375 mL, 4.86 mol). The reaction mixture was carefully heated at 50° C. for 16 h and the solution went from a dark orange clear solution to a pale yellow clear solution. The reaction mixture was concentrated in vacuo to give the title compound (30.0 g, 62%); MS (ESI) m/z [M−H]− 214.0. Intermediate 74
SOCl2 (20.74 g, 174.31 mmol) was added dropwise to a solution of 5-fluoro-2-methoxy-4-nitrobenzoic acid Intermediate 73 (25.0 g, 116.21 mmol) in dry MeOH (150 mL), and the reaction mixture was heated to reflux for 16 h. The reaction mixture was concentrated under reduced pressure, and the crude product was dissolved in hot hexane and filtered. The filtrate was concentrated in vacuo to give the title compound (7.0 g, 18%); GC/MS [229.0] retention time 8.439 min.
Methyl 5-fluoro-2-methoxy-4-nitrobenzoate Intermediate 74 (2.0 g, 8.73 mmol), 1-[(2S)-oxetan-2-yl]methanamine (836 mg, 9.6 mmol) and DIPEA (4.56 mL, 26.19 mmol) were mixed in DMSO (10 mL), and the reaction mixture was stirred at 90° C. for 16 h. The reaction mixture was cooled to rt, diluted with water and extracted with MTBE. The combined organic layer was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated to give the title compound (2.1 g, 65%) as a yellow oil; MS (ESI) m/z [M+H]+ 297.0 Intermediate 76
Pd/C (10%, 50 mg) was added to a solution of methyl (S)-2-methoxy-4-nitro-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 75 (2.1 g, 5.67 mmol) in THE (25 mL) and the is reaction mixture was stirred under an atmosphere of H2(g) at ambient pressure and temperature until the reaction was complete. The reaction mixture was filtered and the filtrate was concentrated. The residue was purified by chromatography to give the title compound (260 mg, 13%); MS (ESI) m/z [M+H]+ 267.2.
2-Chloro-1,1,1-trimethoxyethane (184 mg, 1.19 mmol) and pTsOH·H2O (19 mg, 99 μmol) was added to a solution of methyl (S)-4-amino-2-methoxy-5-((oxetan-2-ylmethyl)amino)benzoate Intermediate 76 (265 mg, 995 μmol) in THE (50 mL), and the reaction mixture was stirred at 50° C. overnight. The mixture was poured into water and extracted with EtOAc. The combined organic layer was washed with brine and concentrated under vacuum to give the title compound (270 mg, 75%); MS (ESI) m/z [M+H]+ 325.2.
Methyl 3-fluoro-5-methoxy-4-nitrobenzoate (1.5 g, 6.55 mmol), 2-(2,2-difluorocyclopropoxy)ethanamine (1.36 g, 7.85 mmol) and DIPEA (3.42 mL, 19.63 mmol) were mixed in DMSO (5 mL), and the reaction mixture was stirred at 80° C. for 16 h. The is reaction mixture was cooled to rt, diluted with water and extracted with EtOAc. The combined organic layer was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated to give the title compound (2.2 g, 75%); MS (ESI) m/z [M+H]+ 347.1.
Pt/C (10%, 25 mg) was added to a solution of methyl 3-((2-(2,2-difluorocyclopropoxy)ethyl)amino)-5-methoxy-4-nitrobenzoate Intermediate 78 (2.2 g, 6.35 mmol) in MeOH (15 mL), and the reaction mixture was stirred under an atmosphere of H2(g) at ambient pressure and temperature until the reaction was complete. The reaction mixture was filtered and the filtrate was concentrated. The residue was purified by flash chromatography on silica (hexane:EtOAc, 1:1) to give the title compound (1.15 g, 66%); MS (ESI) m/z [M+H]+ 317.2.
2-Chloro-1,1,1-trimethoxyethane (293 mg, 1.8 mmol) and pTsOH·H2O (30 mg) were added to a solution of methyl 4-amino-3-((2-(2,2-difluorocyclopropoxy)ethyl)amino)-5-methoxybenzoate Intermediate 79 (0.5 g, 1.5 mmol) in THE (50 mL), and the reaction mixture was stirred at 50° C. overnight. The mixture was poured into water and extracted with EtOAc. The combined organic layer was washed with brine and concentrated under vacuum to give the title compound (0.50 g, 87%); MS (ESI) m/z [M+H]+ 375.0.
Methyl 3,5-difluoro-4-nitrobenzoate (4.0 g, 18.42 mmol), 2-(2,2-difluorocyclopropoxy)ethan-1-amine (2.53 g, 18.42 mmol) and DIPEA (6.42 mL, 36.85 mmol) were mixed in THE (15 mL), and the reaction mixture was stirred at 45° C. for 16 h. The reaction mixture was cooled to rt, diluted with water and extracted with EtOAc. The combined organic layer was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated to give the title compound (6.0 g, 97%); MS (ESI) m/z [M+H]+ 335.2.
Pt/C (1%, 40.8 mg) was added to a solution of methyl 3-((2-(2,2-difluorocyclopropoxy)ethyl)amino)-5-fluoro-4-nitrobenzoate Intermediate 81 (6.0 g, 17.9 mol) in MeOH (30 mL) and the reaction mixture was stirred under an atmosphere of H2(g) at ambient pressure and temperature until the reaction was complete. The reaction mixture was filtered and the filtrate was concentrated. The residue was purified by flash chromatography on silica (hexane/EtOAc, 1/1) to give the title compound (4.5 g, 71%); MS (ESI) m/z [M+H]+ 305.0.
2-Chloro-1,1,1-trimethoxyethane (2.74 g, 17.75 mmol) and pTsOH·H2O (281 mg, 1.48 mmol) were added to a solution of methyl 4-amino-3-((2-(2,2-difluorocyclopropoxy)ethyl)amino)-5-fluorobenzoate Intermediate 82 (4.5 g, 14.79 mmol) in THE (50 mL), and the reaction mixture was stirred at 50° C. overnight. The reaction mixture was poured into water and extracted with EtOAc, and the combined organic layer was washed with brine and concentrated under vacuum to give the title compound (3.0 g, 56%); MS (ESI) m/z [M+H]+ 335.2.
The title compound was prepared from rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane hydrochloride Intermediate 26 (222 mg, 0.66 mmol) and methyl (S)-2-(chloromethyl)-5-methoxy-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 77 (201 mg, 0.62 mmol) in analogy with the description for Intermediate 33 to give the title compound (26 mg, 6%); MS (ESI) m z [M+H]+ 646.2.
Methyl 2-(chloromethyl)-1-(2-(2,2-difluorocyclopropoxy)ethyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Intermediate 80 (440 mg, 1.1 mmol) and DIPEA (455 mg, 4.2 mmol)) were added to a solution of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane hydrochloride Intermediate 26 (504 mg, 1.4 mmol) in MeCN (5 mL) and the reaction mixture was stirred at 45° C. overnight. The reaction mixture was diluted with water and extracted with EtOAc. The combined organic layer was washed with sat NaCl (aq), dried over anhydrous Na2SO4, filtered and concentrated, and the crude product was purified by preparative HPLC, PrepMethod G, (gradient: 0-60%) to give the title compound (197 mg, 25%) as a yellow oil; MS (ESI) m/z [M+H]+ 696.2.
The title compound was prepared from rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane hydrochloride Intermediate 26 (403 mg, 1.12 mmol) and methyl 2-(chloromethyl)-1-(2-(2,2-difluorocyclopropoxy)ethyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Intermediate 83 (371 mg, 1.0 mmol) as described for Intermediate 85 to give the title compound (74 mg, 11%); MS (ESI) m z [M+H]+ 684.2.
The enantiomers of 4-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile WO2020207474 (4.5 g, 13.47 mmol) were separated by chiral chromatography on a Chiral Art Amylose-C NEO column (250×50 mm, 10 μm), eluted with 25% IPA in hexane in CO2, 100 bar, at a flow rate of 200 mL/min, and detected at 220 nm;
Cs2CO3 (921 mg, 2.83 mmol) was added to a mixture of rel-(R)-4-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile, Isomer 1 Intermediate 87 (472 mg, 1.41 mmol), rac-tert-butyl (1R,6S)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 4 (300 mg, 1.41 mmol) and Pd-PEPPSI-IHeptCl (119 mg, 0.14 mmol) in 1,4-dioxane (20 mL) and the reaction mixture was stirred at 80° C. for 16 h. The reaction mixture was filtered through silica and then concentrated. The residue was diluted with EtOAc (100 mL), and the organic layer was washed with sat brine (2×50 mL), dried over Na2SO4, filtered and evaporated. The crude product was purified by flash chromatography on silica (gradient: 20-25% EtOAc in petroleum ether) to give the title compound Isomer mixture 1 (480 mg, 73%) as a yellow solid; MS (ESI) m/z [M+H]+ 466.0. pTsOH (533 mg, 3.09 mmol) was added to a solution of the product from Step a) (480 mg) in is DCM (20 mL) and the reaction mixture was stirred at 35° C. for 12 h. The solvent was removed under reduced pressure to give the title compound Isomer mixture 1 as the para-toluenesulfonic acid salt (900 mg, 100%) as a blue solid; MS (ESI) m/z [M+H]+ 365.9.
The title compound was prepared in two steps, as described for Intermediate 89, from rel-(R)-4-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile, Isomer 2 Intermediate 88 (472 mg, 1.41 mmol) and rac-tert-butyl (1R,6S)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 4 (300 mg, 1.41 mmol) to give the title compound Isomer mixture 2 as the para-toluenesulfonic acid salt (522 mg, 79%); MS (ESI) m/z [M+H]+ 366.0.
pTsOH (1.396 g, 8.11 mmol) was added to a solution of rac-tert-butyl (1R,6S)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 60 (0.77 g, 1.62 mmol) in DCM (15 mL) at 20° C. under an atmosphere of N2(g), and the reaction mixture was stirred at 35° C. for 3 h. The solvent was removed under reduced pressure to give the title compound as the para-toluensulfonic acid salt (1.8 g) as a crude yellow solid; MS (ESI) m/z [M+H]+ 374.9.
Methyl 2-(((1RS,6SR)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 1
K2CO3 (1.254 g, 9.07 mmol) was added to a solution of 4-((R*)-4-((1SR,6RS)-2,5-diazabicyclo[4.2.0]octan-2-yl)-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile para toluenesulfonate, Isomer mixture 1, Intermediate 89 (0.8 g, 0.91 mmol) and methyl (S)-2-(chloromethyl)-4-methoxy-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 10 (0.295 g, 0.91 mmol) in MeCN (15 mL) and the reaction mixture was stirred at 60° C. for 4 h. The reaction mixture was filtered through celite, and then concentrated. The residue was diluted with EtOAc (125 mL), and washed with sat brine (50 mL). The organic layer was dried over Na2SO4, filtered and evaporated. The residue was purified by preparative TLC (EtOAc:petroleum ether, 1:5), to give the title compound Isomer mixture 1 (0.540 g, 91%) as a yellow solid; MS (ESI) m/z [M+H]+ 654.3.
The stereoisomers of methyl 2-(((1RS,6SR)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 1
Intermediate 92 (0.5 g) were separated by chiral chromatography on a Chiralpak IF column (250×50 mm, 5 μm), eluted with 50% EtOH in hexane (0.1% 2M NH3 in MeOH), at a flow rate of 15 mL/min, and detected at 220 nm and 254 nm;
The title compound was prepared as described for Intermediate 92 from 4-((R*)-4-((1SR,6RS)-2,5-diazabicyclo[4.2.0]octan-2-yl)-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile para toluenesulfonate Isomer mixture 1, Intermediate 89 (800 mg, 0.91 mmol) and methyl (S)-4-chloro-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 24 (299 mg, 0.91 mmol). The crude product was purified by reversed phase flash chromatography on a C18 column (0-90% MeCN in water) to give the title compound Isomer mixture 1 (370 mg, 62%) as a white solid; MS (ESI) m/z [M+H]+ 658.2.
The title compound was prepared as described for Intermediate 92 from 4-((R*)-4-((1SR,6RS)-2,5-diazabicyclo[4.2.0]octan-2-yl)-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile para toluenesulfonate Isomer mixture 2, Intermediate 90 (680 mg, 0.77 mmol) and methyl (S)-4-chloro-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 24 (254 mg, 0.77 mmol). The crude product was purified by reversed phase flash chromatography on a C18 column (0-90% MeCN in water) to give the title compound Isomer mixture 2 (400 mg, 79%) as a white solid; MS (ESI) m/z [M+H]+ 658.1.
The stereoisomers of methyl 4-chloro-2-(((1RS,6SR)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 1 Intermediate 95 (0.37 g) were separated by chiral chromatography on a Chiralpak IF column (250×50 mm, 5 μm), eluted with 50% EtOH in hexane (0.5% 2M NH3 in MeOH), at a flow rate of 15 mL/min, and detected at 220 nm and 254 nm;
The stereoisomers of methyl 4-chloro-2-(((1RS,6SR)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 2 Intermediate 96 (0.40 g) were separated by chiral chromatography on a Chiralpak IF column (250×50 mm, 5 μm), eluted with 50% EtOH in hexane (0.5% 2M NH3 in MeOH), at a flow rate of 15 mL/min, and detected at 220 nm and 254 nm;
The title compound was prepared as described for Intermediate 92 from 4-((R*)-4-((1SR,6RS)-2,5-diazabicyclo[4.2.0]octan-2-yl)-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile para toluenesulfonate Isomer mixture 1, Intermediate 89 (800 mg, 0.91 mmol) and methyl (S)-2-(chloromethyl)-4-fluoro-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 7 (284 mg, 0.91 mmol). The crude product was purified by preparative TLC (MeOH:DCM, 1:20), to give the title compound Isomer is mixture 1 (450 mg, 77%) as a yellow solid; MS (ESI) m/z [M+H]+ 642.2.
The title compound was prepared as described for Intermediate 92 from 4-((R*)-4-((1SR,6RS)-2,5-diazabicyclo[4.2.0]octan-2-yl)-2-methylbenzo[d][1,3]dioxol-2-yl)-3-fluorobenzonitrile para toluenesulfonate Isomer mixture 2, Intermediate 90 (700 mg, 0.79 mmol) and methyl (S)-2-(chloromethyl)-4-fluoro-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 7 (248 mg, 0.79 mmol). The crude product was purified by preparative TLC (MeOH:DCM, 1:20), to give the title compound Isomer mixture 2 (500 mg, 98%) as a yellow solid; MS (ESI) m/z [M+H]+ 642.3.
The stereoisomers of methyl 2-(((1RS,6SR)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 1 Intermediate 100 (0.45 g) were separated by chiral chromatography on a Chiralpak ID column (250×50 mm, 5 μm), eluted with 30% EtOH in hexane (0.5% 2M NH3 in MeOH), at a flow rate of 20 mL/min, and detected at 220 nm and 254 nm;
The stereoisomers of methyl 2-(((1RS,6SR)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 2 Intermediate 101 (0.495 g) were separated by chiral chromatography on a Chiralpak ID column (250×50 is mm, 5 μm), eluted with 20% EtOH in hexane (0.5% 2M NH3 in MeOH), at a flow rate of 20 mL/min, and detected at 220 nm and 254 nm;
Methyl (S)-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate (132 mg, 0.45 mmol) was added to a mixture of rac-(1R,6S)-2-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane para toluensulfonate Intermediate 91 (500 mg, 0.45 mmol) and K2CO3 (558 mg, 4.03 mmol) in MeCN (15 mL) and the reaction mixture was stirred at 60° C. for 3 h. The reaction mixture was concentrated and then diluted with EtOAc (100 mL). The organic layer was washed with sat brine (3×50 mL), dried over Na2SO4, filtered and evaporated. The residue was purified by preparative TLC (petroleum ether: EtOAc, 5: 1), to give the title compound (200 mg, 70%) as a yellow solid; MS (ESI) m/z [M+H]+ 633.2.
The diastereomers of methyl 2-(((1RS,6SR)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 105 (657 mg, 1.04 mmol) were separated by chiral chromatography on a Chiralpak IE column (250×50 mm ID, 5 μm), eluted with 5% IPA in hexane/DCM (3:1, 0.5% 2M NH3 in MeOH), at a flow rate of 20 mL/min and detected at 220 nm and 254 nm;
The stereoisomers of the first eluted compound mixture (258 mg) were separated by chiral chromatography on a Lux Sum Amylose-1 column (250×50 mm ID, 10 μm), eluted with 20% IPA in hexane (0.5% 2M NH3 in MeOH), at a flow rate of 20 mL/min and detected at 220 and 254 nm;
The stereoisomers of the second eluted compound mixture (256 mg) were separated by chiral chromatography on a CHIRALPAK IE column (250×20 mm ID, 5 μm), eluted with 10% IPA in hexane/DCM (3:1, 0.5% NH3 in MeOH), at a flow rate of 20 mL/min and detected at 220 and 254 nm;
The stereoisomers of methyl 2-(((1RS,6SR)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 109 (800 mg, 1.21 mmol) were separated by chiral chromatography on a Chiralpak IH column (250×30 mm, 5 μm), eluted with 30% IPA/MeCN (1:1, 0.1% 2M NH3 in MeOH) in CO2, 100 bar, at a flow rate of 60 mL/min and detected at 220 nm;
The stereoisomers of the first eluted compound mixture (400 mg) were separated by chiral chromatography on a (R, R)-Whelk-01 Kromasil column (250×21.1 mm, 5 μm), eluted with 10% IPA in MTBE (0.5% 2M NH3 in MeOH), at a flow rate of 20 mL/min and detected at 220 nm and 254 nm;
The stereoisomers of 2-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-5-chloropyridine WO2020234726 (2 g, 6.12 mmol) were separated by chiral chromatography on an UniChiral OD-5H column (250×30 mm, 5 μm), eluted with 10% IPA/hexane (1:4, 0.5% 2M NH3 in MeOH) in CO2, 100 bar, at a flow rate of 100 mL/min and detected at 220 nm; The first eluted compound was collected and evaporated to give the title compound Isomer 1, Intermediate 114 (400 mg, 20%); MS (ESI) m/z [M+H]+ 326/328; and
Cs2CO3 (1.247 g, 3.83 mmol) was added to a mixture of rel-(R)-2-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-5-chloropyridine Isomer 1, Intermediate 114 (500 mg, 1.53 mmol), rac-tert-butyl (1R,6S)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 4 (271 mg, 1.28 mmol), Palladacycle Gen 4 (73 mg, 0.060 mmol) and 2′-(bis(3,5-bis(trifluoromethyl)phenyl)phosphino)-3′,6′-dimethoxy-N2.N2.N6.N6-tetramethyl-[1,1′-biphenyl]-2,6-diamine (48 mg, 0.060 mmol) in 1,4-dioxane (10 mL) and the reaction mixture was stirred at 90° C. for 16 h. The reaction mixture was filtered through celite and the filter cake was washed with EtOAc (3×50 mL). The combined filtrate was collected and concentrated under reduced pressure and the residue was purified by preparative TLC (petroleum ether:EtOAc, 5:1), to give the sub-title compound Isomer mixture 1 (500 mg, 86%) as a white solid; MS (ESI) m/z [M+H]+ 458.2.
pTsOH (0.846 g, 4.91 mmol) was added to a solution of the product from Step a) (450 mg, 0.98 mmol) in DCM (10 mL) and the reaction mixture was stirred at 35° C. for 3 h. The reaction mixture was concentrated under reduced pressure to give the title compound Isomer mixture 1 as the para-toluensulfonic acid salt (1.0 g, 83%); MS (ESI) m/z [M+H]+ 358.0.
The title compound was prepared in two steps in a similar way as as described for Intermediate 116 from Intermediate 115 (400 mg, 1.22 mmol) and Intermediate 4 (200 mg, 0.94 mmol) to give the title compound Isomer mixture 2 as the para-toluensulfonic acid salt (400 mg, 99%) as a yellow solid; MS (ESI) m/z [M+H]+ 358.
K2CO3 (0.635 g, 4.59 mmol) was added to a solution of (1RS,6SR)-2-((S*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane para-toluensulfonate Isomer mixture 1 Intermediate 116 (0.8 g, 0.66 mmol) and methyl (S)-4-chloro-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 24 (0.216 g, 0.66 mmol) in MeCN (15 mL) and the reaction mixture was stirred at 60° C. for 8 h. The reaction mixture was quenched with water (100 mL), extracted with EtOAc (3×50 mL), and the organic layer was dried over Na2SO4, filtered and evaporated. The crude product was purified by reversed phase flash chromatography on a C18-column (0-80% MeCN in water) to give the title compound (0.400 g, 94%) as a white solid; MS (ESI) m/z [M+H]+ 650.2.
Cs2CO3 (1.17 g, 3.58 mmol) was added to a solution of (1RS,6SR)-2-((S*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane para-toluenesulfonate, Isomer mixture 2 Intermediate 117 (380 mg, 0.72 mmol) and methyl (S)-4-chloro-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 24 (236 mg, 0.72 mmol) in MeCN (20 mL) and the reaction mixture was stirred at 60° C. for 8 h. The reaction mixture was filtered through celite and washed with MeCN (3×25 mL. The solvent was removed under reduced pressure and the crude product was purified by reversed phase flash chromatography on a C18 column (0-100% MeCN in water) to give the title compound Isomer mixture 2 (380 mg, 81%) as a yellow solid; MS (ESI) m/z [M+H]+ 650.
The stereoisomers of methyl 4-chloro-2-(((1RS,6SR)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 1 Intermediate 118 (380 mg) were separated by chiral chromatography on a Chiralpak ID column (250×20 mm ID, μm), eluted with 25% EtOH in hexane (0.5% 2 M NH3 in MeOH), at a flow rate of 20 mL/min and detected at 220 nm and 254 nm;
The first eluted compound was collected and evaporated to give the title compound Isomer 1, Intermediate 120 (150 mg, 39%) as a yellow solid; MS (ESI) m/z [M+H]+ 650.2; and The second eluted compound was collected and evaporated to give the title compound Isomer 2 Intermediate 121 (150 mg, 39%) as a white solid; MS (ESI) m/z [M+H]+ 650.1.
Methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4
The stereoisomers of methyl 4-chloro-2-(((1RS,6SR)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer mixture 2 Intermediate 119 (380 mg) were separated by chiral chromatography on a Chiralpak ID column (250×20 mm ID, 5 μm), eluted with 30% EtOH in hexane (0.5% 2 M NH3 in MeOH), at a flow rate of 20 mL/min and detected at 220 nm and 254 nm;
The first eluted compound was collected and evaporated to give the title compound Isomer 3, Intermediate 122 (180 mg, 47%) as a yellow solid; MS (ESI) m/z [M+H]+ 650; and
Methyl (S)-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate (370 mg, 1.26 mmol) was added to a suspension of rac-(1R,6R)-2-(2-(4-Chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane hydrochloride Intermediate 32 (562 mg, 1.26 mmol), DIPEA (974 mg, 7.54 mmol) and NaI (753 mg, 5.0 mmol) in MeCN (150 mL) and the reaction mixture was stirred at 45° C. for 18 h. The reaction mixture was concentrated in vacuo, the residue was diluted with water and extracted with DCM (2×90 mL). The combined organic layer was dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by preparative HPLC, PrepMethod E, is (gradient: 50-70%) to give the title compound (246 mg, 33%); MS (ESI) m/z [M+H]+ 632.2.
The stereoisomers of methyl 2-(((1RS,6RS)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 124 were separated by chiral chromatography on a Chiralcel OZ-H column (250×50 mm, 5p m), eluted with hexane:MeOH:IPA (3:1:1), at a flow rate of 12 mL/min; The first eluted compound was collected and evaporated to give the title compound Isomer 1, Intermediate 125 (78 mg); MS (ESI) m/z [M+H]+ 632.2; and
Methyl 2-(chloromethyl)-1-[(2S)-oxetan-2-yl]methyl-1H-1,3-benzodiazole-6-carboxylate (254 mg, 862 μmol) was added to a suspension of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 26 (370 mg, 1.03 mmol), DIPEA (557 mg, 4.31 mmol) and NaI (517 mg, 3.45 mmol) in MeCN (10 mL), and the reaction mixture was stirred at 60° C. for 12 h. The reaction mixture was concentrated in vacuo, then diluted with water (20 mL) and extracted with DCM (3×20 mL). The combined organic layer was dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC, PrepMethod E, (gradient: 30-70%) to give the title compound (250 mg, 49%); MS (ESI) m/z [M+H]+ 616.2.
The diastereoisomers of methyl 2-(((1RS,6RS)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 127 were separated by chiral chromatography on a CHIRACEL OZ-H column (250×20 mm ID, 5 μm), eluted with hexane:IPA:MeOH (80:10:10), at a flow rate of 18 mL/min;
Methyl 3,5-difluoro-4-nitrobenzoate (5.0 g, 23.03 mmol), 2-(1-(aminomethyl)cyclopropyl)acetonitrile hydrochloride (3.38 g, 23.03 mmol) and DIPEA (12 mL, 69.08 mmol) were mixed in THE (20 mL) and the reaction mixture was stirred at 60° C. for 16 h. The reaction mixture was cooled to rt and diluted with water. The aqueous phase was extracted with MTBE (3×50 mL), and the combined organic layer was washed with brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated in vacuo to give the title compound (5.1 g, 54%); MS (ESI) m/z [M+H]+ 308.0.
Pd/C (10%, 0.5 g) was added to a solution of methyl 3-(((1-(cyanomethyl)cyclopropyl)methyl)amino)-5-fluoro-4-nitrobenzoate Intermediate 130 (5.1 g, 12.45 mmol) in dry MeOH (20 mL) and the reaction mixture was stirred under an atmosphere of H2(g) (1 atm) at ambient temperature until the reaction was complete. The reaction mixture was filtered and concentrated in vacuo. The residue was purified by flash chromatography on silica (0-99% MTBE in hexane) to give the title compound (2.8 g, 77%); MS (ESI) m z [M+H]+ 278.2.
2-Chloro-1,1,1-trimethoxyethane (1.34 mL, 9.92 mmol) and pTsOH (155 mg, 902 μmol) were added to a solution of methyl 4-amino-3-(((1-(cyanomethyl)cyclopropyl)methyl)amino)-5-fluorobenzoate Intermediate 131 (2.5 g, 9.02 mmol) in THE (50 mL) and the reaction mixture was stirred at 50° C. overnight. The reaction mixture was poured into water and extracted with EtOAc (3×10 mL). The combined organic layer was washed with brine and concentrated under vacuum to give the title compound (2.25 g, 67%); MS (ESI) m/z [M+H]+ 336.2.
rac-(1R,6R)-2-(2-(5-Chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane dihydrochloride Intermediate 26 (500 mg, 1.4 mmol), methyl 2-(chloromethyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Intermediate 132 (389 mg, 1.16 mmol), DIPEA (749 mg, 5.8 mmol, 1.01 mL) and NaI (17 mg, 116 μmol) were mixed in MeCN (3 mL) and the reaction mixture was stirred at 40° C. for 16 h. The reaction mixture was diluted with water (5 mL) and extracted with EtOAc (3×5 mL). The combined organic layer was dried over anhydrous Na2SO4, filtered and evaporated. The residue was purified by preparative HPLC, PrepMethod E, (gradient: 30-50%) to give the title compound (330 mg, 43%); MS (ESI) m/z [M+H]+ 657.2.
The stereoisomers of rac-methyl 2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Intermediate 133 were separated by chiral chromatography on a CHIRALPAK IC column (250×21 mm, 5 μm), eluted with IPA:MeOH:CHCl3 (47.5:47.5:5), at a flow rate of 12 mL/min; the first eluted compound mixture was collected and evaporated to yield a mixture of isomers and the second eluted compound mixture was collected and evaporated to yield a mixture of isomers. The stereoisomers of the first eluted compound mixture were separated by chiral chromatography on a CHIRALPAK IF column (250×21 mm, 5 μm) eluted with hexane:IPA:MeOH (80:10:10) at a flow rate of 14 mL/min;
The stereoisomers of the second eluted compound mixture were separated by chiral chromatography on a Chiralpak IA column (250×21 mm, 5 μm) eluted with 20% MeOH in CO2 at a flow rate of 50 mL/min;
The title compound was prepared in three steps as described for Intermediate 132 from methyl 3,5-difluoro-4-nitrobenzoate (3.05 g, 23.03 mmol) and 1-(aminomethyl)cyclopropane-1-carbonitrile hydrochloride (5.0 g, 23.0 mmol) to give the title compound (2.35 g, 69%); MS (ESI) m/z [M+H]+ 322.0.
rac-(1R,6R)-2-(2-(5-Chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 26 (500 mg, 1.4 mmol), methyl 2-(chloromethyl)-1-((1-cyanocyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Intermediate 138 (373 mg, 1.16 mmol), DIPEA (1.01 mL, 5.8 mmol) and NaI (17 mg, 116 μmol) were mixed in MeCN (5 mL) and the reaction mixture was stirred at 40° C. for 16 h. The reaction mixture was diluted with water (5 mL) and extracted with EtOAc (3×5 mL). The combined organic layer was dried over anhydrous Na2SO4, filtered and evaporated. The residue was purified by preparative HPLC, PrepMethod E (gradient: 30-50%) to give the title compound (370 mg, 49%); MS (ESI) m/z [M+H]+ 643.2.
The stereoisomers of rac-methyl 2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Intermediate 139 were separated by chiral chromatography on a CHIRALCEL OZ-H colum (250×21 mm, 5 μm) eluted with hexane:IPA:MeOH (60:20:20) at a flow rate of 14 mL/min; the second eluted compound mixture was collected and evaporated to give a mixture of stereoisomers; and
The stereoisomers of the second eluted compound mixture were separated by chiral chromatography on a Chiralpak IA column (250×30 mm, 5 μm) eluted with hexane:MeOH:IPA (60:20:20) at a flow rate of 12 mL/min;
Methyl 3-fluoro-5-methoxy-4-nitrobenzoate (901 g, 3.93 mmol) and DIPEA (1.5 g, 11.8 mmol) were slowly added to a solution of 1-(aminomethyl)cyclopropyl)-1-carbonitrile hydrochloride (522 mg, 3.93 mmol) in THF (20 mL) and the reaction mixture was stirred at 60° C. for 16 h. The reaction mixture was cooled to rt and diluted with water. The aqueous phase was extracted with DCM (3×30 mL), and the combined organic layer was dried over is anhydrous Na2SO4, filtered and concentrated. The residue was purified by flash chromatography on silica (DCM:MeOH, 4:1). The product containing fractions were collected and evaporated, and the obtained yellow solid was triturated with petroleum ether:EtOAc (30:1) to give the title compound (1.1 g, 94%) as a yellow solid; MS (ESI) m z [M+H]+ 306.1.
Wet Pt/C (10%, 0.3 g) was added to a suspension of methyl 3-(((1-cyanocyclopropyl)methyl)amino)-5-methoxy-4-nitrobenzoate Intermediate 142 (1.104 g, 3.50 mmol) in MeOH (100 mL) and the reaction mixture was stirred under an atmosphere of H2(g) (1 atm) at 20° C. for 36 h. The reaction mixture was filtered and the filter cake was washed with MeOH (50 mL). The filtrate was concentrated under reduced pressure and the crude product was purified by flash chromatography on silica (0-99% EtOAc in hexane) to give the title compound (515 mg, 50%); 1H NMR (500 MHz, CDCl3) δ 7.22 (d, 1H), 7.10 (d, 1H), 3.93-3.82 (m, 6H), 3.24 (s, 2H), 1.36-1.29 (m, 2H), 1.02-0.92 (m, 2H).
2-Chloro-1,1,1-trimethoxyethane (318 mg, 2.06 mmol) and pTsOH (32 mg, 0.19 mmol) were added to a solution of methyl 4-amino-3-(((1-cyanocyclopropyl)methyl)amino)-5-methoxybenzoate Intermediate 143 (515 mg, 1.87 mmol) in MeCN (50 mL) and the reaction is mixture was stirred at 80° C. for 18 h. The reaction mixture was evaporated at reduced pressure and the residue was diluted with EtOAc. The organic layer was washed with NaHCO3 (aq) and water, dried over Na2SO4, filtered and evaporated. The crude product was purified by preparative HPLC, PrepMethod E, (gradient: 0-55%) to give the title compound (211 mg, 47%); MS (ESI) m/z [M+H]+ 334.0.
Methyl 2-(chloromethyl)-1-((1-cyanocyclopropyl)methyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Intermediate 144 (212 mg, 634 μmol) was added to a suspension of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane dihydrochloride Intermediate 26 (273 mg, 634 μmol), DIPEA (491 mg, 3.8 mmol) and NaI (380 mg, 2.54 mmol) in MeCN (100 mL) and the reaction mixture was stirred at 60° C. for 18 h. The reaction mixture was concentrated in vacuo, diluted with water (40 mL) and extracted with DCM (2×70 mL). The combined organic layer was dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC, PrepMethod E, (gradient: 30-70%) to give the title compound (144 mg, 35%); MS (ESI) m/z [M+H]+ 655.0.
The stereoisomers of rac-methyl 2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Intermediate 145 were separated by chiral chromatography on a CHIRALPAK IA column (250×20 mm, μm), eluted with hexane:IPA:MeOH (70:15:15), at a flow rate of 12 mL/min; the first eluted is compound mixture was collected and evaporated to yield a mixture of isomers; and the second eluted compound mixture was collected and evaporated to yield a mixture of isomers.
The stereoisomers of the first eluted compound mixture were separated by chiral chromatography on a CHIRALPAK IB column (250×20 mm, 5 μm) eluted with hexane:IPA:MeOH (85:7.5:7.5) at a flow rate of 18 mL/min;
The stereoisomers of the second eluted compound mixture were separated by chiral chromatography on a Chiralpak IB column (250×20 mm, 5 μm) eluted with hexane:IPA:MeOH (85:7.5:7.5) at a flow rate of 18 mL/min;
Methyl 3-chloro-5-fluoro-4-nitrobenzoate (5.05 g, 21.62 mmol), 2-(1-(aminomethyl)cyclopropyl)acetonitrile hydrochloride (3.17 g, 21.62 mmol) and DIPEA (8.38 g, 64.87 mmol) were mixed in THE (200 mL), and the reaction mixture was heated at 50° C. for 14 h. The reaction mixture was evaporated under reduced pressure and the residue was diluted with EtOAc (150 mL). The organic layer was washed with water (2×50 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give the crude title compound (5.73 g); MS (ESI) m/z [M+H]+ 324.0.
Wet Pt/C (10%, 0.2 g) was added to a suspension of methyl 3-chloro-5-(((1-(cyanomethyl)cyclopropyl)methyl)amino)-4-nitrobenzoate Intermediate 150 (5.73 g, 17.7 mmol) in MeOH (200 mL) and the reaction mixture was stirred under an atmosphere of H2(g) (1 atm) at 20° C. for 64 h. The reaction mixture was filtered, and the catalyst carefully washed with MeOH (100 mL). The filtrate was concentrated under reduced pressure. The crude compound was purified by flash chromatography on silica (0-99% MTBE in hexane) to give the title compound (3.7 g, 71%); 1H NMR (500 MHz, CDCl3) δ 7.66 (s, 1H), 7.34 (s, 1H), 3.87 (s, 3H), 3.18 (s, 2H), 2.61 (s, 2H), 1.58 (s, 3H), 0.76 (s, 4H).
2-Chloro-1,1,1-trimethoxyethane (435 mg, 2.81 mmol) and pTsOH (44 mg, 256 μmol) were is added to a stirred solution of methyl 4-amino-3-chloro-5-(((1-(cyanomethyl)cyclopropyl)methyl)amino)benzoate Intermediate 151 (751 mg, 2.56 mmol) in MeCN (100 mL) and the reaction mixture was heated at 60° C. for 2 h. The reaction mixture was cooled to rt and then evaporated under reduced pressure. The residue was diluted with EtOAc (70 mL) and the mixture was washed with NaHCO3 (30 mL) and water (30 mL). The organic layer was dried over Na2SO4 and evaporated to give the crude title compound (0.72 g); MS (ESI) m/z [M+H]+ 352.0.
Methyl 4-chloro-2-(chloromethyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 152 (1.08 g, 3.08 mmol) was added to a suspension of rac-(1R,6R)-2-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane dihydrochloride Intermediate 26 (1.33 g, 3.08 mmol), DIPEA (2.39 g, 18.47 mmol) and NaI (1.85 g, 12.32 mmol) in MeCN (150 mL) and the reaction mixture was stirred at 60° C. for 18 h. The reaction mixture was concentrated in vacuo, and the residue was diluted with water (60 mL) and extracted with DCM (2×90 mL). The combined organic layer was dried over anhydrous Na2SO4, filtered and evaporated. The residue was purified by preparative HPLC, PrepMethod E, (gradient: 30-50%) to give the title compound (150 mg, 49%); MS (ESI) m/z [M+H]+ 673.
The stereoisomers of rac-methyl 4-chloro-2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 153 is were separated by chiral chromatography on a CHIRALPAK IC column (250×20 mm, 5 μm) eluted with hexane:IPA:MeOH (50:25:25) at a flow rate of 13 mL/min;
The stereoisomers of the first eluted compound mixture were separated by chiral chromatography on a Chiralcel OZ-H column (250×20 mm, 5 μm), eluted with hexane:IPA:MeOH (40:30:30) at a flow rate of 12 mL/min;
The stereoisomers of the second eluted compound mixture were separated by chiral chromatography on a Chiralpak AS-H column (250×20 mm, 5 μm), eluted with hexane:IPA:MeOH (90:5:5) at a flow rate of 12 mL/min;
Methyl 3-fluoro-5-methoxy-4-nitrobensoat (3.0 g, 13.09 mmol), 2-cyclopropoxyethan-1-amine hydrochloride (1.8 g, 13.09 mmol) and DIPEA (5.02 mL, 28.8 mmol) were mixed in DMSO (15 mL) and the reaction mixture was stirred at 80° C. for 16 h. The reaction mixture was cooled to rt, diluted with water (20 mL), and the mixture was extracted with MTBE (3×20 mL). The combined organic layer was washed with brine (20 mL), dried over anhydrous Na2SO4, filtered and concentrated to give the title compound (3.5 g, 69%); MS (ESI) m/z [M+H]+ 311.0.
Pd/C (10%, 0.35 g) was added to a solution of methyl 3-((2-cyclopropoxyethyl)amino)-5-methoxy-4-nitrobenzoate Intermediate 158 (3.5 g, 9.02 mmol) in dry MeOH (15 mL) and the reaction mixture was stirred under an atmosphere of H2(g) (1 atm) at ambient temperature until the reaction was complete. The reaction mixture was concentrated in vacuo, and the residue was purified by flash chromatography on silica (0-99% MTBE in hexane) to give the title compound (1.1 g, 41%); MS (ESI) m/z [M+H]+ 281.2.
2-Chloro-1,1,1-trimethoxyethane (667 mg, 4.32 mmol) and pTsOH (68 mg, 0.39 mmol) were added to a solution of methyl 4-amino-3-((2-cyclopropoxyethyl)amino)-5-methoxybenzoate Intermediate 159 (1.1 g, 3.92 mmol) in THE (50 mL) and the reaction mixture was stirred at 50° C. overnight. The mixture was poured into water (15 mL) and extracted with EtOAc (3×15 mL). The combined organic layer was washed with brine (10 mL), dried over anhydrous Na2SO4, filtered and evaporated to give the title compound (1.2 g, 68%); MS (ESI) m z [M+H]+ 339.0.
rac-(1R,6R)-2-(2-(5-Chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 26 (500 mg, 1.4 mmol), methyl 2-(chloromethyl)-1-(2-cyclopropoxyethyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Intermediate 160 (393 mg, 1.16 mmol), DIPEA (749 mg, 5.8 mmol) and NaI (17 mg, 116 μmol) were mixed in dry MeCN (7 mL), and the reaction mixture was stirred at 40° C. 16 h. The reaction mixture was diluted with water (10 mL) and extracted with EtOAc (3×10 mL). The combined organic layer was separated, dried over Na2SO4 and evaporated. The crude residue was purified by preparative HPLC PrepMethod E (gradient: 30-50%) to give the title compound (180 mg, 24%); MS (ESI) m/z [M+H]+ 660.2.
The stereoisomers of rac-methyl 2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(2-cyclopropoxyethyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Intermediate 161 were separated by chiral chromatography on a CHIRALPAK AD-H column (250×20 mm, 5 μm) eluted with 20% MeOH in CO2, at a flow rate of 50 mL/min;
The stereoisomers of the second eluted compound mixture were separated by chiral chromatography on a Chiralcel OZ-H column (250×20 mm, 5 μm) eluted with hexane:IPA:MeOH (80:10:10) at a flow rate of 12 mL/min;
The stereoisomers of 2-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-5-chloropyridine WO2020234726 (6 g, 18.37 mmol) were separated by chiral chromatography on an LUX Al (AD) column (250×30 mm, 5 μm), eluted with 3% (IPA, 20 mM DEA) in CO2, 120 bar, at a flow rate of 150 mL/min and detected at 220 nm; The first eluted compound was collected and evaporated to give the title compound, Intermediate 163 (2 g, 33%); [α]D20+152 (c 1.00, MeCN); 1H NMR (400 MHz, DMSO-d6) δ 2.07 (3H, d), 6.83 (1H, td), 6.97 (1H, d), 7.06 (1H, d), 7.64 (1H, dd), 8.04 (1H, dd), 8.74 (1H, d).
The absolute configuration of Intermediate 163 was determined by vibrational circular dichroism (VCD) spectroscopy. The experimental spectrum recorded in CDCl3 was compared to a simulated spectrum of the (S) enantiomer calculated using density functional theory at the B3PW91/cc-pVTZ level of theory. Based on the large number of points of agreement between the experimental and simulated spectra, the title compound was assigned as the (S) enantiomer.
TEA (17.51 g, 0.1731 mol), followed by a solution of benzyl (2,5-dioxopyrrolidin-1-yl) carbonate (28.4 g, 0.1154 mol) in dry DCM (100 mL) was added to a solution of rac-tert-butyl (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate (24.5 g, 0.1154 mol) in dry DCM (100 mL) and the reaction mixture was stirred at rt overnight. The reaction mixture was diluted with DCM (50 mL), washed with 10% citric acid (50 mL), sat NaHCO3, (50 mL) and brine (100 mL). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude compound was purified by column chromatography (hexane:EtOAc, 1:1) to give the title compound (39 g, 98%); MS (ESI) m z [[(M −Boc)+H]+247.2; 1H NMR (500 MHz, CDCl3) δ 7.44-7.23 (m, 5H), 5.20-4.98 (m, 2H), 3.83-3.52 (m, 4H), 3.42 (dtt, 2H), 2.20 (d, 2H), 1.78 (s, 2H), 1.45 (d, 9H).
The stereoisomers of rac-2-benzyl 5-(tert-butyl) (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2,5-dicarboxylate Intermediate 164 were separated by chiral chromatography on a CHIRALPAK IA column (250×30 mm, 5 μm), eluted with hexane:IPA:MeOH (80:10:10), at a flow rate of 40 mL/min;
The first eluted compound was collected and evaporated to give the title compound Isomer 1, Intermediate 165 (15.7 g); MS (ESI) m/z [[(M −Boc)+H]+247.4; and
A solution of rel-2-benzyl 5-(tert-butyl) (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2,5-dicarboxylate Isomer 2 Intermediate 166_(14.9 g 0.043 mol, retrospectively) and 10% Pd/C (1.4 g) in MeOH (100 mL) was evacuated and then purged with H2(g) (×3). The reaction mixture was stirred at rt under pressure of H2(g) (2 atm) until complete reaction (monitoring is by NMR). The catalyst was carefully removed by filtration and washed with MeOH (2×20 mL). The filtrate was concentrated in vacuo to give the title compound Intermediate 167 (8.8 g, 96%); [α]D20+6.18 (c 0.5, MeOH); 1H NMR (400 MHz, CDCl3) δ 3.66 (ddd, 1H), 3.08-2.93 (m, 1H), 2.87 (pt, 2H), 2.77-2.48 (m, 3H), 2.15 (q, 1H), 2.01-1.78 (m, 2H), 1.55 (ddd, 1H), 1.39-1.26 (m, 9H).
The absolute configuration of the title compound was determined by converting the title compound in two steps to the 4-chlorobenzoyl derivative Intermediate 169 as described below in the experimental descriptions for Intermediate 168 and Intermediate 169. Based on the X-ray data of Intermediate 169 the title compound Intermediate 167 was assigned as the (1R,6R) enantiomer.
TEA (214 mg, 2.1 mmol), followed by a solution of 4-chlorobenzyl carbonochloridate (241 mg, 1.4 mmol) in dry DCM (15 mL), was added to a solution of tert-butyl (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 167 (250 mg, 1.4 mmol) in dry DCM (5 mL) and the reaction mixture was stirred at rt overnight. The reaction mixture was diluted with DCM (5 mL) and washed with 10% citric acid (10 mL), sat NaHCO3 (20 mL) and brine (10 mL). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated at reduced pressure. The crude product was purified by column (hexane:MTBE, 1:1) to give the title compound (335 mg, 82%); MS (ESI) m/z [(M-Boc)+H]+281.2
2 M HCl in Et2O (2 mL) was added to a solution rel-2-(tert-butyl) 5-(4-chlorobenzyl) (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2,5-dicarboxylate Isomer 2 Intermediate 168 (95 mg, 0.25 mmol) in DCM (10 mL) and the reaction mixture was stirred at ambient temperature for 8 h. The reaction mixture was concentrated under reduced pressure to give the hydrochloride of the title compound (80 mg, quantitative yield) as a white solid; MS (ESI) m/z 281.0.
Crystals for X-ray diffraction studies was grown from acetonitrile. The molecular structure of Intermediate 169 is shown in
Crystallographic data: C14H18ClN2O2, Cl, 2×(H2O), M=380.87, monoclinic, space group P21, Cell Lengths: a=16.437(3), b=7.0509(10), c=14.574(2) A, cell angles: a 90 R 99.923(8) γ 90, cell volume: V=1663.8, crystal size ca. 0.11×0.25×0.49 mm, R-factor (%) 7.62.
A mixture of (S)-2-(4-bromo-2-methylbenzo[d][1,3]dioxol-2-yl)-5-chloropyridine Intermediate 163 (12.36 g, 37.86 mmol), tert-butyl (1R,6R)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 167 (8.461 g, 37.86 mmol), Palladacycle Gen 4 (2.59 g, 2.27 mmol), and Cs2CO3 (24.67 g, 75.73 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). 1,4-Dioxane (60 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at 90° C. for 20 h. The reaction mixture was cooled to rt, diluted with EtOAc (100 mL) and washed with brine (2×50 mL). The organic layer was dried over MgSO4, filtered, and concentrated at reduced pressure. The crude product was purified by flash chromatography on silica (0-12% EtOAc in heptane). The product was dissolved in EtOAc (100 mL), SiliaMetS Thiol (3 g, 40-63 μm) was added and the mixture was stirred at rt for 2 h, then filtered. The is filtrate was collected and concentrated at reduced pressure to give the title compound (10.80 g, 62.3%); [α]D20+79 (c 1.0, MeCN); MS (ESI) m/z [M+H]+ 458.3.
pTsOH hydrate (9.87 g, 51.88 mmol) was added to a solution of tert-butyl (1R,6R)-5-((S)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane-2-carboxylate Intermediate 170 (10.8 g, 23.58 mmol) in EtOAc (80 mL) and the reaction mixture was stirred at 42° C. for 18 h. The reaction mixture was cooled to rt, diluted with EtOAc (50 mL) and sat K2CO3 (aq, 10 mL) was added drop wise. The organic layer was washed with sat K2CO3 (aq, 3×25 mL), dried over MgSO4, filtered and concentrated at reduced pressure to give the title compound (8.30 g, 98%); MS (ESI) m/z [M+H]+ 358.2.
K2CO3 (9.27 g, 67.07 mmol) and methyl (S)-4-chloro-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 24 (7.73 g, 23.47 mmol) were added to a solution of (1R,6R)-2-((S)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octane Intermediate 171 (8 g, 22.36 mmol) in MeCN (50 mL) and the reaction mixture was stirred at 53° C. for 24 h. The reaction mixture was cooled to rt and EtOAc (100 mL) was added. The organic layer was washed with NaHCO3 (aq, 2×50 mL), is dried over MgSO4, filtered and concentrated at reduced pressure. The crude compound was purified by preparative HPLC, PrepMethod M, (gradient: 50-100%). Relevant fractions were pooled and most of the MeCN was evaporated and the residue was extracted with EtOAc (2×30 mL). The combined organic layer was dried over MgSO4, filtered, and the filtrate was stirred with SiliaMetS Thiol (6 g, 40-63 μm) at rt for 2 h. The mixture was filtered and the filtrate was concentrated at reduced pressure. The residue was purified by flash chromatography (50-100% EtOAc in heptane) and the product containing fractions were pooled and concentrated at reduced pressure. MeOH (300 mL) was added to the residue and the mixture was stirred at rt for 30 min whereupon a solid formed. The mixture was cooled to 0° C., and the solid was isolated by filtration, rinsed with several portions of cooled MeOH (10 mL) and finally dried in vacuo to give the title compound (9.59 g, 65.9%) as a white solid; [α]D20+49 (c 1.0, MeCN); MS (ESI) m/z [M+H]+ 652.35.
A mixture of methyl 2-(((1R*,6S*)-5-((S*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 13 (106 mg, 0.17 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (61 mg, 0.44 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight.
The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, is filtered and purified by preparative HPLC, PrepMethod B, (gradient: 5-95%), to give the title compound (0.070 g, 67%); HRMS (ESI) m/z [M+H]+ calcd for C32H32ClFN5O5: 620.2070, found: 620.2098; 1H NMR (600 MHz, DMSO-d6) 1.51-1.57 (1H, m), 1.63-1.74 (1H, m), 1.85-1.91 (1H, m), 1.96 (3H, s), 2.14-2.2 (1H, m), 2.31-2.41 (2H, m), 2.61-2.73 (2H, m), 3.15-3.27 (3H, m), 3.28-3.34 (1H, m), 3.73 (1H, d), 4.15 (1H, d), 4.19-4.26 (2H, m), 4.42-4.48 (1H, m), 4.75 (1H, dd), 4.81-4.87 (1H, m), 5.13-5.2 (1H, m), 6.31 (1H, dd), 6.45 (1H, dd), 6.71 (1H, t), 7.49-7.55 (1H, m), 7.60 (1H, dd), 7.94-7.98 (1H, m), 8.14-8.17 (1H, m), 8.69 (1H, d).
A mixture of methyl 2-(((1R*,6S*)-5-((S*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 14 (138 mg, 0.22 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (61 mg, 0.44 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod B, (gradient: 5-95%), to give the title compound (0.074 g, 54%); HRMS (ESI) m/z [M+H]+ calcd for C32H32ClFN5O5: 620.2070, found: 620.2086 1H NMR (600 MHz, DMSO-d6) 1.52-1.64 (3H, m), 1.99 (4H, s), 2.3-2.41 (2H, m), 2.61-2.69 (1H, m), 2.7-2.76 (1H, m), 3.1-3.16 (1H, m), 3.17 (1H, s), 3.24-3.3 (2H, m), 3.77 (1H, d), 4.11 (1H, d), 4.21-4.28 (2H, m), 4.41-4.48 (1H, m), 4.71-4.84 (2H, m), is 5.11-5.18 (1H, m), 6.25-6.3 (1H, m), 6.44-6.48 (1H, m), 6.71 (1H, t), 7.5-7.55 (1H, m), 7.58 (1H, d), 7.95-8 (1H, m), 8.14-8.18 (1H, m), 8.67-8.71 (1H, m).
2-((1R*,6S*)-5-(R*)-2-(5-Chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid, Isomer 4
A mixture of methyl 2-(((1R*,6S*)-5-((S*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 15 (125 mg, 0.20 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (55 mg, 0.39 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt for 90 min. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO. Acetic acid (0.5 mL) was added and the mixture was purified by preparative HPLC, PrepMethod A, (gradient: 30-60%), to give the title compound (0.059 g, 49%); HRMS (ESI) m/z [M+H]+ calcd for C32H32ClFN5O5: 620.2070, found: 620.2100; 1H NMR (500 MHz, DMSO-d6) 1.69-1.83 (2H, m), 1.93-2.04 (4H, m), 2.19-2.29 (1H, m), 2.32-2.41 (2H, m), 2.43-2.48 (1H, m), 2.6-2.74 (3H, m), 3.14-3.22 (1H, m), 3.61 (1H, d), 4.21 (1H, d), 4.24-4.31 (1H, m), 4.38-4.47 (1H, m), 4.48-4.57 (1H, m), 4.59-4.72 (1H, m), 4.89-5 (1H, m), 5.05-5.16 (1H, m), 6.32-6.37 (1H, m), 6.44-6.49 (1H, m), 6.73 (1H, t), 7.48-7.54 (1H, m), 7.62 (1H, d), 8.00 (1H, dd), 8.16 (1H, s), 8.71 (1H, d).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 17 (33 mg, 0.05 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (14 mg, 0.10 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The mixture was concentrated at reduced pressure, and the residue was purified by preparative HPLC, PrepMethod A, (gradient: 20-90%), to give the title compound (13 mg, 41%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2292; 1H NMR (500 MHz, CD3OD) 1.6-1.68 (1H, m), 1.69-1.76 (2H, m), 2.02 (3H, s), 2.09 (1H, p), 2.39-2.49 (2H, m), 2.69-2.82 (2H, m), 3.13-3.19 (1H, m), 3.37-3.45 (1H, m), 3.82 (1H, d), 4.04 (3H, s), 4.20 (1H, d), 4.26-4.35 (2H, m), 4.55-4.63 (1H, m), 4.72 (1H, dd), 4.87-4.93 (1H, m), 5.21-5.33 (1H, m), 6.31 (1H, dd), 6.44 (1H, dd), 6.72 (1H, t), 7.45 (1H, d), 7.64 (1H, dd), 7.86 (1H, dd), 7.95 (1H, d), 8.11 (1H, s), 8.59 (1H, dd).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 18 (29 mg, 0.04 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (11 mg, 0.08 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight.
The mixture was concentrated at reduced pressure, and the residue was purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (13 mg, 21%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2280; 1H NMR (500 MHz, CD3OD) 1.69-1.81 (2H, m), 1.87-1.95 (1H, m), 2.00 (3H, s), 2.16-2.27 (1H, m), 2.4-2.51 (2H, m), 2.7-2.81 (2H, m), 3.24-3.3 (2H, m), 3.34-3.42 (1H, m), 3.81 (1H, d), 4.05 (3H, s), 4.23 (1H, d), 4.27-4.37 (2H, m), 4.54-4.63 (1H, m), 4.73 (1H, dd), 4.93 (1H, dd), 5.24-5.33 (1H, m), 6.31-6.37 (1H, m), 6.42 (1H, dd), 6.72 (1H, t), 7.45 (1H, d), 7.65 (1H, dd), 7.87 (1H, dd), 7.96 (1H, d), 8.10 (1H, s), 8.57-8.61 (1H, m).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 19 (58 mg, 0.09 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (25 mg, 0.18 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The mixture was concentrated at reduced pressure, and the residue was purified by preparative HPLC, PrepMethod A, (gradient: 20-90%), to give the title compound (22 mg, is 39%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2278; 1H NMR (500 MHz, CD3OD) 1.65-1.72 (1H, m), 1.76-1.91 (2H, m), 2.02 (3H, s), 2.1-2.2 (1H, m), 2.43-2.59 (2H, m), 2.72-2.83 (2H, m), 3.14-3.22 (1H, m), 3.25-3.3 (1H, m), 3.35-3.43 (1H, m), 3.75 (1H, d), 4.03 (3H, s), 4.21 (1H, d), 4.34 (1H, q), 4.45-4.53 (1H, m), 4.57-4.69 (2H, m), 5.00 (1H, dd), 5.2-5.29 (1H, m), 6.32 (1H, dd), 6.44 (1H, dd), 6.72 (1H, t), 7.44 (1H, d), 7.65 (1H, dd), 7.86 (1H, dd), 7.96 (1H, d), 8.09 (1H, s), 8.60 (1H, dd).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 20 (47 mg, 0.07 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (20 mg, 0.15 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The mixture was concentrated at reduced pressure, and the residue was purified by preparative HPLC, PrepMethod A, (gradient: 20-90%), to give the title compound (16 mg, 35%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2280; 1H NMR (500 MHz, CD3OD) 1.81-1.89 (2H, m), 1.91-1.99 (1H, m), 2.00 (3H, s), 2.2-2.33 (1H, m), 2.41-2.5 (1H, m), 2.5-2.61 (1H, m), 2.7-2.83 (2H, m), 3.25-3.3 (2H, m), 3.32-3.4 (1H, m), 3.73 (1H, d), 4.03 (3H, s), 4.23 (1H, d), 4.35 (1H, q), 4.46-4.54 (1H, m), 4.57-4.7 (2H, is m), 5.02 (1H, dd), 5.2-5.3 (1H, m), 6.34 (1H, dd), 6.42 (1H, dd), 6.72 (1H, t), 7.44 (1H, d), 7.61-7.67 (1H, m), 7.84 (1H, dd), 7.96 (1H, d), 8.10 (1H, s), 8.58 (1H, dd).
A mixture of methyl 4-chloro-2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 28 (49 mg, 0.08 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (21 mg, 0.15 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound Isomer 1 (0.023 g, 48%); IRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1818; 1H NMR (500 MHz, CD3OD) 1.44-1.54 (1H, m), 1.74-1.87 (2H, m), 2.01 (3H, s), 2.17-2.25 (1H, m), 2.43-2.64 (3H, m), 2.7-2.9 (3H, m), 2.96-3.03 (1H, m), 3.84-4.08 (3H, m), 4.41-4.49 (1H, m), 4.6-4.68 (1H, m), 4.87-4.92 (overlapping with solvent peak, m), 5.21-5.3 (1H, m), 6.39 (1H, dd), 6.49-6.54 (1H, m), 6.74 (1H, t), 7.66-7.71 (1H, is m), 7.88 (1H, dd), 7.94 (1H, d), 8.27 (1H, d), 8.6-8.63 (1H, m).
A mixture of methyl 4-chloro-2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 29 (150 mg, 0.23 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (64 mg, 0.46 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (0.108 g, 74%); HRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1788; 1H NMR (500 MHz, CD3OD) 1.44-1.54 (1H, m), 1.78-1.88 (2H, m), 1.99 (3H, s), 2.19-2.28 (1H, m), 2.42-2.54 (2H, m), 2.54-2.67 (2H, m), 2.72-2.85 (2H, m), 2.91-2.97 (1H, m), 3.81-3.88 (2H, m), 4.04 (1H, d), 4.43 (1H, dt), 4.56-4.64 (1H, m), 4.78 (1H, dd), 4.82-4.92 (overlapping with solvent peak, m), 5.17-5.25 (1H, m), 6.32-6.38 (1H, m), 6.47-6.53 (1H, m), 6.72 (1H, t), 7.60 (1H, d), 7.80 (1H, dd), 7.92 (1H, d), 8.24 (1H, d), 8.54 (1H, d).
A solution of LiOH (0.953 g, 39.80 mmol) in water (40 mL) was added dropwise to a solution of methyl 4-chloro-2-(((1R,6R)-5-((S)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 172 (9.59 g, 14.74 mmol) in THE (80 mL) and the reaction mixture was stirred at rt for 4 h. The reaction mixture was concentrated at reduced pressure and the solid was suspended in water (200 mL). EtOAc (100 mL) was added and the pH was adjusted to pH 4.5 by addition of 1 M citric acid. The phases were separated and the aqueous layer was extracted with EtOAc (2×100 mL). The combined organic layer was washed with water (2×20 mL), dried over MgSO4, filtered and concentrated under reduced pressure. The white solid was lyophilized for 72 h to give the title compound (8.80 g, 94%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1786; 1H NMR (500 MHz, DMSO) δ 1.42-1.52 (1H, m), 1.7-1.86 (2H, m), 2.21-2.29 (1H, m), 2.34-2.49 (4H, m), 2.52-2.6 (1H, m), 2.68-2.78 (2H, m), 2.82-2.88 (1H, m), 3.26-3.33 (2H, m), 3.66 (1H, d), 3.77-3.83 (1H, m), 3.98-4.04 (1H, m), 4.35-4.43 (1H, m), 4.46-4.53 (1H, m), 4.7-4.77 (1H, m), 4.84-4.92 (1H, m), 5.04-5.13 (1H, m), 6.31-6.36 (1H, m), 6.5-6.56 (1H, m), 6.72 (1H, t), 7.59 (1H, d), 7.79 (1H, d), 7.92-7.98 (1H, m), 8.24-8.27 (1H, m), 8.69 (1H, d).
Single clear colorless blocked-shaped crystal of Example 3b was recrystallized from methyl tertiary-butyl ether. A suitable single crystal was selected and mounted on a mitigen sample holder (Mitegen, USA) in perflouroether oil. X-ray diffraction data was collected at 100 K using Cryostream 800 (Oxford Cryosystem, UK) in co-scan mode with XtaLab Synergy-S (Rigaku, Japan) equipped with Cu Kα micro focus source (50 kV, 0.01 mA) and Hypix-Arc 100 detector. The diffraction pattern was initially indexed and the total number of runs and images was based on the strategy calculation from the program CrysAlisPro 1.171.42.46a (Rigaku, Japan). Data reduction, scaling and absorption corrections were performed using CrysAlisPro 1.171.42.46a (Rigaku, Japan). The integrated and scaled data were corrected is using numerical absorption correction based on gaussian integration over a multifaceted crystal model and empirical absorption correction using spherical harmonics, implemented in SCALE3 ABSPACK scaling algorithm.
The structure was solved by the ShelXT (Acta Cryst. C71 (2015) 3-8) structure solution program using using dual methods and refined by full matrix least squares minimisation on F2 using version 2018/3 of ShelXL 2018/3 (Acta Cryst. C71 (2015) 3-8) within Olex2 (J. Appl. Cryst. 42 (2009) 339-431). All non-hydrogen atoms were refined anisotropically. All hydrogen atoms attached to oxygen atoms were located from the differential Fourier map and were refined isotropically. All other hydrogen atoms were determined geometrically and refined isotropically.
Crystallographic data is listed below. Asymmetric unit contains two molecules of Example 3b and four molecules of methyl tertiary-butyl ether, therefore this crystal structure is Example 3b methyl tertiary-butyl ether disolvate. Flack (Acta Cryst. B69 (2013) 249-259) and Hooft (J. Appl. Cryst. 43 (2010) 665-668) parameters were found to be 0.002(3) and −0.006(3), respectively. Thermal ellipsoid drawing of Example 3b is shown in
By comparison of retention times on chiral HPLC and biological assay data, the title compound 4-chloro-2-(((1R,6R)-5-((S)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid was concluded to be identical to 4-Chloro-2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid, Isomer 2.
A mixture of methyl 4-chloro-2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 30 (68 mg, 0.10 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (29 mg, 0.21 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3) three times. A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (0.051 g, 77%); HRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1816; 1H NMR (500 MHz, CD3OD) 1.24-1.35 (1H, m), 1.62-1.71 (1H, m), 1.7-1.82 (1H, m), 2.02 (3H, s), 2.12-2.2 (1H, m), 2.44-2.56 (2H, m), 2.63-2.72 (1H, m), 2.75-2.89 (3H, m), 3.11-3.18 (1H, m), 3.90 (1H, d), 3.93-4 (1H, m), 4.04 (1H, d), 4.46 (1H, dt), 4.6-4.67 (1H, m), 4.70 (1H, dd), 4.98 (1H, dd), 5.27-5.35 (1H, m), 6.35-6.41 (1H, m), 6.49-6.55 (1H, m), 6.74 (1H, t), 7.66-7.71 (1H, m), 7.88 (1H, dd), 7.94 (1H, d), 8.29 (1H, d), 8.58-8.63 (1H, m).
A solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 34 (85 mg, 0.13 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (43 mg, 0.31 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated and the residue dissolved in DMSO, and purified by preparative HPLC, PrepMethod A, (gradient: 40-80%) to give the title compound (0.054 g, 65) as a white powder; HRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2246; 1H NMR (500 MHz, CD3OD) 1.41-1.58 (1H, m), 1.81-1.93 (2H, m), 2.03 (3H, s), 2.27 (1H, q), 2.42-2.63 (3H, m), 2.63-2.73 (2H, m), 2.77-2.9 (2H, m), 2.96 (1H, dt), 3.83 (1H, d), 3.88 (1H, dt), 4.01 (1H, d), 4.05 (3H, s), 4.45 (1H, dt), 4.64 (1H, q), 4.77-4.86 (2H, m), 5.2-5.33 (1H, m), 6.39 (1H, d), 6.54 (1H, d), 6.74 (1H, t), 7.16-7.24 (1H, m), 7.24-7.32 (1H, m), 7.46 (1H, d), 7.57 (1H, td), 7.95 (1H, d).
A solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 35 (30 mg, 0.05 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (19 mg, 0.14 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated and the residue was dissolved in DMSO and purified by preparative HPLC, PrepMethod A, to give the title compound (0.023 g, 78%) as a white powder; HRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2254; 1H NMR (500 MHz, CD3OD) 1.36-1.53 (1H, m), 1.73-1.89 (2H, m), 2.01 (3H, s), 2.13-2.25 (1H, m), 2.4-2.61 (3H, m), 2.65 (1H, s), 2.7-2.88 (3H, m), 2.98 (1H, dt), 3.83 (1H, d), 3.90 is (1H, dt), 3.98 (1H, d), 4.03 (3H, s), 4.43 (1H, dt), 4.63 (1H, td), 4.78-4.84 (2H, m), 5.18-5.33 (1H, m), 6.37 (1H, dd), 6.50 (1H, dd), 6.72 (1H, t), 7.20 (1H, dd), 7.27 (1H, dd), 7.44 (1H, d), 7.61 (1H, t), 7.94 (1H, d).
A solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 36 (28 mg, 0.04 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (18 mg, 0.13 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated and the residue was dissolved in DMSO, and purified by preparative HPLC, PrepMethod A, to give the title compound (9.0 mg, 33%) as a white powder; HRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2212; 1H NMR (500 MHz, CD3OD) 1.21-1.37 (1H, m), 1.67 (1H, dt), 1.79 (1H, qd), 2.02 (3H, s), 2.15 (1H, dt), 2.43 (1H, td), 2.47-2.56 (1H, m), 2.57-2.65 (1H, m), 2.66 (1H, s), 2.75-2.92 (3H, m), 3.09 (1H, dt), 3.83 (1H, d), 3.91-4.00 (2H, m), 4.03 (3H, s), 4.46 (1H, dt), 4.60-4.73 (2H, m), 4.95 (1H, dd), 5.30 (1H, qd), 6.37 (1H, dd), 6.50 (1H, dd), 6.72 (1H, t), 7.21 (1H, dd), 7.28 (1H, dd), 7.44 (1H, d), 7.62 (1H, t), 7.96 (1H, d).
The stereoisomers of 2-(((1RS,6SR)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid, Isomer 1 and Isomer 2 Intermediate 41 (115 mg) were separated by chiral chromatography on a Chiralcel OD column (250×20 mm, 5 μm), eluted with heptane/IPA (80/20), at a flow rate of 20 mL/min, and detected at 264 nm; the first eluted compound was collected and evaporated to give the title compound, Isomer 1, Example 5a (25 mg); HRMS (ESI) m/z [M+H]+ calcd for C32H33ClN5O5: 602.2164, found: 602.2180; 1H NMR (500 MHz, CD3OD) 1.61-1.68 (1H, m), 1.69-1.76 (2H, m), 2.02 (3H, s), 2.04-2.14 (1H, m), 2.41-2.52 (2H, m), 2.7-2.84 (2H, m), 3.13-3.19 (1H, m), 3.39-3.47 (1H, m), 3.85 (1H, d), 4.20 (1H, d), 4.26-4.38 (2H, m), 4.57-4.63 (1H, m), 4.77 (1H, dd), 4.94-4.87 (partly overlapping with solvent, m), 5.24-5.31 (1H, m), 6.32 (1H, dd), 6.44 (1H, dd), 6.72 (1H, t), 7.64 (1H, dd), 7.69 (1H, d), 7.86 (1H, dd), 7.98 (1H, dd), 8.32-8.35 (1H, m), is 8.59 (1H, dd).
the second eluted compound was collected and evaporated to give the title compound, Isomer 2, Example 5b (36 mg); MS (ESI) m/z [M+H]+ 602.4; 1H NMR (500 MHz, CD3OD) 1.68-1.81 (2H, m), 1.85-1.96 (1H, m), 1.99 (3H, s), 2.16-2.27 (1H, m), 2.41-2.51 (2H, m), 2.7-2.83 (2H, m), 3.23-3.3 (partly overlapping with solvent, m), 3.36-3.44 (1H, m), 3.84 (1H, d), 4.22 (1H, d), 4.26-4.38 (2H, m), 4.56-4.64 (1H, m), 4.77 (1H, dd), 4.93 (1H, dd), 5.25-5.33 (1H, m), 6.33 (1H, dd), 6.42 (1H, dd), 6.68-6.75 (1H, m), 7.6-7.67 (1H, m), 7.69 (1H, d), 7.85 (1H, dd), 7.99 (1H, dd), 8.32-8.36 (1H, m), 8.58 (1H, dd).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 39 (70 mg, 0.11 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (32 mg, 0.23 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and the crude compound was purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (0.050 g, 73%); HRMS (ESI) m/z [M+H]+ is calcd for C32H32ClN5O5: 602.2164, found: 602.2184; 1H NMR (500 MHz, DMSO-d6) 1.62-1.75 (3H, m), 2.01 (3H, s), 2.03-2.11 (1H, m), 2.31-2.4 (1H, m), 2.6-2.75 (2H, m), 3.13-3.2 (1H, m), 3.23 (2H, d), 3.29 (1H, s), 3.60 (1H, d), 4.17 (1H, d), 4.31 (1H, q), 4.38-4.46 (1H, m), 4.48-4.55 (1H, m), 4.57-4.64 (1H, m), 4.84-4.93 (1H, m), 5.05-5.13 (1H, m), 6.29-6.34 (1H, m), 6.45-6.5 (1H, m), 6.73 (1H, t), 7.58-7.67 (2H, m), 7.78-7.83 (1H, m), 8.02 (1H, dd), 8.24-8.28 (1H, m), 8.72 (1H, d).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 40 (70 mg, 0.11 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (32 mg, 0.23 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and the crude compound was purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (0.042 g, 62%); HRMS (ESI) m/z [M+H]+ calcd for C32H32ClN5O5: 602.2164, found: 602.2166; 1H NMR (500 MHz, CD3OD) 1.83-1.9 (2H, m), 1.93-2 (1H, m), 2.01 (3H, s), 2.25-2.36 (1H, m), 2.42-2.51 (1H, m), 2.52-2.63 (1H, m), 2.71-2.77 (1H, m), 2.78-2.85 (1H, m), 3.27-3.33 (partly overlapping with solvent, m), is 3.34-3.42 (1H, m), 3.73 (1H, d), 4.26 (1H, d), 4.33-4.4 (1H, m), 4.53 (1H, dt), 4.63-4.72 (2H, m), 5.07 (1H, dd), 5.24-5.33 (1H, m), 6.36 (1H, dd), 6.4-6.45 (1H, m), 6.7-6.77 (1H, m), 7.67 (2H, td), 7.87 (1H, dd), 7.98 (1H, dd), 8.32-8.37 (1H, m), 8.60 (1H, dd).
A mixture of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 43 (53 mg, 0.08 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (23 mg, 0.16 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (15 mg, 29%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2302; 1H NMR (500 MHz, CD3OD) 1.47 (1H, dd), 1.79-1.88 (2H, m), 2.01 (3H, s), 2.21-2.29 (1H, m), 2.39-2.59 (3H, m), 2.62 (1H, dd), 2.73-2.86 (2H, m), 2.91-2.97 (1H, m), 3.81 (1H, d), 3.83-3.88 (1H, m), 3.95-4.01 (1H, m), 4.03 (3H, s), 4.39-4.46 (1H, m), 4.58-4.65 (1H, m), 4.77-4.83 (2H, m), 5.19-5.28 (1H, m), 6.35-6.41 (1H, m), 6.52 is (1H, dd), 6.74 (1H, t), 7.43 (1H, d), 7.63 (1H, dd), 7.85 (1H, dd), 7.93 (1H, d), 8.57 (1H, dd).
A mixture of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 44 (43 mg, 0.07 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (19 mg, 0.14 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (22 mg, 51%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2282; 1H NMR (500 MHz, CD3OD) 1.47-1.55 (1H, m), 1.74-1.88 (2H, m), 2.01 (3H, s), 2.21 (1H, dd), 2.47-2.58 (2H, m), 2.6-2.69 (1H, m), 2.71-2.84 (2H, m), 2.83-2.92 (1H, m), 3.05 (1H, d), 3.87-3.96 (2H, m), 4.03 (4H, s), 4.44 (1H, dt), 4.6-4.68 (1H, m), 4.82-4.84 (2H, m), 5.21-5.29 (1H, m), 6.39 (1H, dd), 6.52 (1H, dd), 6.75 (1H, t), 7.44 (1H, d), 7.69 (1H, dd), 7.87-7.91 (1H, m), 7.95 (1H, d), 8.61 (1H, dd).
A mixture of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 45 (30 mg, 0.05 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (13 mg, 0.09 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (23 mg, 80%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2302; 1H NMR (500 MHz, CD3OD) 1.3-1.41 (1H, m), 1.67-1.81 (2H, m), 2.02 (3H, s), 2.13-2.22 (1H, m), 2.45-2.56 (2H, m), 2.66-2.74 (1H, m), 2.75-2.91 (3H, m), 3.14-3.2 (1H, m), 3.92 (1H, d), 3.94-3.99 (1H, m), 4.02 (4H, d), 4.41-4.49 (1H, m), 4.6-4.7 (2H, m), 4.93 (1H, dd), 5.25-5.34 (1H, m), 6.39 (1H, dd), 6.52 (1H, dd), 6.74 (1H, t), 7.44 (1H, d), 7.69 (1H, dd), 7.89 (1H, dd), 7.97 (1H, d), 8.59-8.63 (1H, m).
A mixture of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 46 (54 mg, 0.08 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (23 mg, 0.17 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 is mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min and added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated at reduced pressure, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod A, (gradient: 20-90%) to give the title compound (18 mg, 35%); HRMS (ESI) m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2306; 1H NMR (500 MHz, CD3OD) 1.26-1.37 (1H, m), 1.66-1.74 (1H, m), 1.75-1.85 (1H, m), 2.01 (3H, s), 2.16-2.24 (1H, m), 2.42-2.53 (2H, m), 2.58-2.72 (2H, m), 2.72-2.85 (2H, m), 3.05-3.11 (1H, m), 3.84-3.91 (2H, m), 3.96 (1H, d), 4.02 (3H, s), 4.39-4.47 (1H, m), 4.57-4.67 (2H, m), 4.91 (1H, dd), 5.23-5.32 (1H, m), 6.34-6.39 (1H, m), 6.52 (1H, dd), 6.73 (1H, t), 7.43 (1H, d), 7.61-7.66 (1H, m), 7.85 (1H, dd), 7.95 (1H, d), 8.56-8.59 (1H, m).
A solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 52 (29 mg, 0.04 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (21 mg, 0.15 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL), and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (30 mg, 106%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2480; is 1H NMR (500 MHz, CD3OD) 1.73 (1H, dq), 1.82-1.93 (2H, m), 1.93-2.01 (3H, m), 2.03 (3H, s), 2.23 (1H, h), 2.29-2.4 (1H, m), 3.14 (1H, td), 3.33-3.46 (2H, m), 3.60 (1H, td), 3.73-3.81 (1H, m), 3.92-4.01 (2H, m), 4.07 (3H, s), 4.11 (1H, ddd), 4.23 (1H, tdd), 4.51 (1H, dd), 4.68 (1H, dd), 4.74 (2H, s), 6.47 (1H, dd), 6.62 (1H, dd), 6.81 (1H, t), 7.51 (1H, d), 7.66 (1H, dd), 7.89 (1H, dd), 7.98 (1H, d), 8.61 (1H, d).
A solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 53 (52 mg, 0.08 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (25 mg, 0.18 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL), and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (40 mg, 79%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2462; 1H NMR (500 MHz, CD3OD) 1.72 (1H, dq), 1.85-2.06 (8H, m), 2.22 (1H, h), 2.35-2.43 (1H, m), 3.01 (1H, td), 3.31-3.38 (1H, m), 3.4-3.47 (1H, m), 3.58 (1H, td), 3.77 (1H, dt), 3.92-3.99 (2H, m), 4.01-4.11 (4H, m), 4.23 (1H, qd), 4.49 (1H, dd), 4.67 (1H, dd), 4.74 (2H, s), is 6.44 (1H, d), 6.62 (1H, dd), 6.80 (1H, t), 7.51 (1H, d), 7.66 (1H, d), 7.88 (1H, dd), 7.97 (1H, d), 8.59 (1H, d).
A solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 54 (31 mg, 0.05 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (17 mg, 0.12 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL), and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (27 mg, 89%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2456; 1H NMR (500 MHz, CD3OD) 1.67 (1H, dq), 1.82-1.97 (4H, m), 1.97-2.02 (1H, m), 2.04 (3H, s), 2.20 (1H, h), 2.35 (1H, dt), 3.16 (1H, td), 3.33-3.44 (2H, m), 3.54 (1H, td), 3.74 (1H, td), 3.91 (2H, ddd), 4.05-4.14 (4H, m), 4.24 (1H, qd), 4.48 (1H, dd), 4.6-4.75 (3H, m), 6.49 (1H, d), 6.64 (1H, d), 6.82 (1H, t), 7.54 (1H, s), 7.68 (1H, d), 7.90 (1H, dd), 8.00 (1H, d), 8.62 (1H, is d).
A solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 56 (74 mg, 0.11 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (62 mg, 0.45 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL) and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (60 mg, 83%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2418; 1H NMR (500 MHz, CD3OD) 1.75 (1H, dq), 1.93-2.14 (8H, m), 2.21-2.31 (2H, m), 3.26-3.33 (2H, m), 3.43-3.54 (1H, m), 3.77 (1H, q), 3.8-3.84 (1H, m), 3.97 (1H, q), 4.04-4.1 (1H, m), 4.11 (3H, s), 4.26-4.38 (2H, m), 4.51 (1H, d), 4.58-4.67 (2H, m), 4.73 (1H, dd), 6.37 (1H, d), 6.55 (1H, d), 6.80 (1H, t), 7.60 (1H, s), 7.69 (1H, d), 7.90 (1H, dd), 8.06 (1H, s), 8.62 (1H, d).
A solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 57 (109 mg, 0.17 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (57 mg, 0.41 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL) and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (93 mg, 87%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2410; 1H NMR (500 MHz, CD3OD) 1.72 (1H, dq), 1.86-2.02 (4H, m), 2.03 (3H, s), 2.05-2.11 (1H, m), 2.17-2.35 (2H, m), 3.12-3.2 (1H, m), 3.19-3.27 (1H, m), 3.44 (1H, t), 3.73-3.85 (2H, m), 3.95 (1H, dt), 4.00-4.08 (1H, m), 4.10 (3H, s), 4.22-4.35 (2H, m), 4.48 (1H, d), 4.54-4.64 (2H, m), 4.71 (1H, dd), 6.33 (1H, d), 6.55 (1H, d), 6.78 (1H, t), 7.59 (1H, s), 7.67 (1H, d), 7.88 (1H, dd), 8.05 (1H, s), 8.60 (1H, d).
A solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 58 (67 mg, 0.10 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (36 mg, 0.26 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was is stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL) and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (67 mg, 102%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2416; 1H NMR (500 MHz, CD3OD) 1.70 (1H, dq), 1.86-1.98 (4H, m), 2.03 (3H, s), 2.08-2.17 (1H, m), 2.21 (1H, dq), 2.35-2.46 (1H, m), 3.08-3.2 (1H, m), 3.33-3.39 (1H, m), 3.46-3.54 (1H, m), 3.75 (1H, dt), 3.85-3.96 (2H, m), 4.07-4.12 (4H, m), 4.19-4.32 (2H, m), 4.47-4.58 (2H, m), 4.61 (1H, d), 4.76 (1H, dd), 6.33 (1H, dd), 6.56 (1H, dd), 6.78 (1H, t), 7.58 (1H, d), 7.67 (1H, dd), 7.88 (1H, dd), 8.04 (1H, d), 8.59 (1H, dd).
A solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-tetrahydrofuran-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 59 (67 mg, 0.10 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (73 mg, 0.52 mmol) in a mixture of MeCN (1 mL) and water (0.2 mL) was stirred at rt overnight. The reaction mixture was concentrated under reduced pressure and the residue was partitioned between DCM (2 mL) and 1M HCl (aq, 2 mL). The aqueous phase was extracted with another portion of DCM (2 mL) and the combined organic layer was concentrated and dried under reduced pressure to give the title compound (62 mg, 95%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H37ClN5O6: 646.2426, found: 646.2414; is 1H NMR (500 MHz, CD3OD) 1.73 (1H, dq), 1.95 (2H, p), 1.99-2.08 (5H, m), 2.11-2.2 (1H, m), 2.23 (1H, h), 2.33-2.41 (1H, m), 3.25-3.32 (1H, m), 3.35-3.41 (1H, m), 3.52-3.6 (1H, m), 3.75 (1H, dt), 3.86-3.96 (2H, m), 4.08-4.17 (4H, m), 4.25-4.34 (2H, m), 4.54-4.6 (2H, m), 4.64 (1H, d), 4.78 (1H, dd), 6.37 (1H, d), 6.56 (1H, d), 6.80 (1H, t), 7.58 (1H, s), 7.69 (1H, d), 7.90 (1H, dd), 8.05 (1H, d), 8.62 (1H, d).
2-2(((R-(4-Chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid. Isomer 1
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 62 (114 mg, 0.18 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (49 mg, 0.35 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod B, (gradient: 5-95%) to give the title compound Isomer 1 (65 mg, 58%); HRMS (ESI) m/z [M+H]+ calcd for C33H32ClF2N4O5: 637.2024, found: 637.2040; 1H NMR (600 MHz, DMSO-d6) 1.5-1.62 (1H, m), 1.63-1.75 (1H, m), 1.88-1.94 (1H, m), 1.97 (3H, s), 2.13-2.23 (1H, m), 2.29-2.42 (2H, m), 2.62-2.74 (2H, m), 3.12-3.22 (1H, m), 3.22-3.3 (1H, m), 3.33-3.45 (1H, m), 3.74 (1H, d), 4.12-4.26 is (3H, m), 4.45 (1H, td), 4.75 (1H, dd), 4.85 (1H, dd), 5.12-5.22 (1H, m), 6.24-6.33 (1H, m), 6.43-6.48 (1H, m), 6.70 (1H, t), 7.30 (1H, dd), 7.47-7.57 (3H, m), 8.15-8.19 (1H, m).
2(((R-(4-Chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid, Isomer 2
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 63 (129 mg, 0.20 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (55 mg, 0.40 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight. The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod B, (gradient: 5-95%) to give the title compound Isomer 2 (88 mg, 70%); HRMS (ESI) m/z [M+H]+ calcd for C33H32ClF2N4O5: 637.2024, found: 637.2030; 1H NMR (600 MHz, DMSO-d6) 1.5-1.59 (2H, m), 1.59-1.65 (1H, m), 2.01 (4H, s), 2.29-2.43 (2H, m), 2.49-2.51 (1H, m), 2.6-2.69 (1H, m), 2.71-2.77 (1H, m), 3.09-3.15 (1H, m), 3.21-3.31 (2H, m), 3.78 (1H, d), 4.11 (1H, d), 4.21-4.26 (1H, m), 4.31 (1H, q), 4.41-4.48 (1H, m), 4.71-4.83 (2H, m), 5.11-5.18 (1H, m), 6.26 (1H, dd), is 6.46-6.5 (1H, m), 6.70 (1H, t), 7.26-7.31 (1H, m), 7.48-7.56 (3H, m), 8.15-8.19 (1H, m).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 64 (177 mg, 0.27 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (76 mg, 0.54 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2(g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight.
The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod B, (gradient: 5-95%) to give the title compound Isomer 3 (102 mg, 59%); HRMS (ESI) m/z [M+H]+ calcd for C33H32ClF2N4O5: 637.2024, found: 637.2062; 1H NMR (600 MHz, DMSO-d6) 1.57-1.64 (1H, m), 1.68-1.74 (2H, m), 2.01 (3H, s), 2.02-2.09 (1H, m), 2.32-2.4 (1H, m), 2.41-2.47 (1H, m), 2.49-2.51 (1H, m), 2.65-2.72 (2H, m), 3.12-3.27 (3H, m), 3.63 (1H, d), 4.18 (1H, d), 4.33-4.44 (2H, m), 4.47-4.54 (1H, m), 4.62 (1H, dd), 4.91 (1H, dd), 5.06-5.13 (1H, m), 6.25-6.29 (1H, m), 6.48 (1H, dd), 6.70 (1H, t), 7.30 (1H, dd), 7.5-7.56 (3H, m), 8.15-8.18 (1H, m).
A mixture of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 65 (159 mg, 0.24 mmol) and 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (68 mg, 0.49 mmol) in a sealed flask was evacuated and backfilled with N2(g) (×3). A mixture of MeCN (3 mL) and water (0.6 mL) was degassed by N2 (g) bubbling for 15 min, and then added. The reaction mixture was evacuated and backfilled with N2(g) (×3) and then stirred at rt overnight.
The reaction mixture was concentrated in vacuo, and the residue was dissolved in DMSO, filtered and purified by preparative HPLC, PrepMethod B, (gradient: 5-95%) to give the title compound Isomer 4 (90 mg, 58%); HRMS (ESI) m/z [M+H]+ calcd for C33H32ClF2N405: 637.2024, found: 637.2052; 1H NMR (600 MHz, DMSO-d6) 1.69-1.79 (2H, m), 1.97 (3H, s), 2.22-2.28 (1H, m), 2.34-2.4 (1H, m), 2.44-2.5 (2H, m), 2.63-2.67 (1H, m), 2.68-2.74 (1H, m), 3.13-3.2 (1H, m), 3.2-3.25 (1H, m), 3.39-3.45 (1H, m), 3.61 (1H, d), 4.18-4.26 (2H, m), 4.39-4.46 (1H, m), 4.48-4.55 (1H, m), 4.6-4.66 (1H, m), 4.94 (1H, dd), 5.07-5.15 (1H, m), 6.29-6.34 (1H, m), 6.43-6.48 (1H, m), 6.71 (1H, t), 7.26-7.31 (1H, m), 7.47-7.5 (1H, m), 7.51-7.57 (2H, m), 8.15-8.2 (1H, m).
LiOH·H2O (4 mg, 89 μmol) and rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-ethyl-1H-imidazol-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 70 (39 mg, 0.057 mmol) were dissolved in a mixture of THE and water (1:1, 5 mL), and the reaction mixture was stirred at ambient temperature for 24 h. The reaction mixture was concentrated at reduced pressure to give the lithium salt of the title compound, which was is purified by preprative HPLC, PrepMethod E, (gradient 35-50%) to give the title compound as the lithium salt (28 mg, 72%); HRMS (ESI) m/z [M+H]+ calcd for C34H34Cl2N7O4: 674.2044, found: 674.2080; 1H NMR (600 MHz, DMSO-d6) δ 8.68 (d, 1H), 7.97 (dd, 1H), 7.90 (s, 1H), 7.77 (s, 1H), 7.62 (s, 1H), 7.58 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.42 (s, 1H), 6.27 (d, 1H), 5.72 (d, 1H), 5.62 (d, 1H), 3.98 (dt, 2H), 3.73 (dd, 2H), 3.61 (d, 1H), 2.85 (d, 1H), 2.57 (t, 1H), 2.41 (d, 2H), 2.28 (q, 1H), 2.14 (q, 1H), 1.97 (s, 3H), 1.72-1.55 (m, 2H), 1.34-1.21 (m, 1H), 1.16 (t, 3H).
rel-4-Chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-ethyl-1H-imidazol-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid. Isomer 3
LiOH·H2O (4 mg, 89 μmol) and rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-ethyl-1H-imidazol-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 71 (16 mg, 0.023 mmol) were dissolved in a mixture of THE and water (1:1, 5 mL), and the reaction mixture was stirred at ambient temperature for 24 h. The reaction mixture was concentrated at reduced pressure to give the lithium salt of the title compound, which was purified by preprative HPLC, PrepMethod E, (gradient 35-50%) to give the title compound as the lithium salt (14 mg, 89%); HRMS (ESI) m/z [M+H]+ calcd for C34H34Cl2N7O4: 674.2044, found: 674.2050; 1H NMR (500 MHz, DMSO-d6) δ 8.71 (d, 1H), 8.01 (dd, 1H), 7.90 (s, 1H), 7.76 (s, 1H), 7.67-7.51 (m, 2H), 6.71 (t, 1H), 6.54 (d, 1H), 6.41 (s, 1H), 6.28 (d, 1H), 5.84-5.50 (m, 2H), 3.99 (q, 2H), 3.83-3.56 (m, 3H), 2.88 (d, 1H), 2.54 (d, 2H), 2.32 (m, 1H), 2.25 (q, 1H), 2.06 (d, 1H), 1.97 (s, 3H), 1.60 (dq, 2H), 1.29-1.07 (m, 4H).
LiOH·H2O (4 mg, 89 μmol) and rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-ethyl-1H-imidazol-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 72 (36 mg, 0.052 mmol) were dissolved in a mixture of THE and water (1:1, 5 mL), and the reaction mixture was stirred at ambient temperature for 24 h. The reaction mixture was concentrated at reduced pressure to give the lithium salt of the title compound, which was purified by preprative HPLC, PrepMethod E, (gradient 35-50%) to give the title compound as the lithium salt (13 mg, 37%); HIRMS (ESI) m/z [M+H]+ calcd for C34H34Cl2N7O4: 674.2044, found: 674.2046; 1H NMR (600 MHz, DMSO-d6) δ 8.68 (d, 1H), 7.96 (dd, 1H), 7.90 (d, 1H), 7.76 (d, 1H), 7.67-7.60 (m, 1H), 7.58 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.42 (s, 1H), 6.27 (d, 1H), 5.72 (d, 1H), 5.62 (d, 1H), 3.98 (qd, 2H), 3.73 (dd, 2H), 3.61 (d, 1H), 2.85 (d, 1H), 2.57 (dd, 1H), 2.46-2.37 (m, 2H), 2.28 (q, 1H), 2.14 (q, 1H), 1.97 (s, 3H), 1.73-1.54 (m, 2H), 1.27 (p, 1H), 1.16 (t, 3H).
LiOH monohydrate (9.7 mg, 231 μmol) was added to a solution of methyl 2-(((1RS,6RS)-5-(2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 33 (102 mg, 154 μmol) in THF:H2O (1:1, 5 mL) at 0° C. and the reaction mixture was stirred for 12 h. The reaction mixture was concentrated and the residue was acidified with 1 M citric acid (aq). The solids were collected by filtration to give the title compound (96 mg, 95%) as a white solid; IRMS (ESI) m z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2258; 1H NMR (400 MHz, DMSO-d6) δ 12.54 (s, 1H), 7.88 (d, 1H), 7.62-7.44 (m, 2H), 7.42-7.16 (m, 2H), 6.73-6.68 (m, 1H), 6.54 (d, 1H), 6.32 (t, 1H), 5.20-4.98 (m, 1H), 4.81-4.74 (dt, 1H), 4.67-4.60 (m, 1H), 4.49-4.43 (m, 1H), 4.38-4.27 (m, 1H), 3.96 (s, 4H), 3.91-3.42 (m, 3H), 3.01-2.85 (m, 1H), 2.74-2.56 (m, 4H), 2.45-2.28 (m, 1H), 2.25-2.07 (m, 1H), 1.99 (s, 3H), 1.90-1.44 (m, 2H), 1.28-1.22 (m, 1H).
LiOH monohydrate (5 mg, 0.114 mmol) was added to a solution of methyl 2-(((1RS,6RS)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-5-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate Intermediate 84 (26 mg, 38 μmol) in THF:H2O (2:1, 1 mL) and the reaction mixture was stirred at rt for 16 h. The reaction mixture was purified by preparative HPLC, Prep Method E (gradient: 0-60%) to give the lithium salt of the title compound (9.9 mg, 48%); HRMS (ESI) is m/z [M+H]+ calcd for C33H35ClN5O6: 632.2270, found: 632.2284; 1H NMR (600 MHz, DMSO-d6) δ 8.75-8.63 (m, 1H), 8.06-7.90 (m, 1H), 7.64-7.51 (m, 1H), 7.36 (d, 1H), 6.95 (s, 1H), 6.71-6.69 (m, 1H), 6.52 (dd, 1H), 6.32 (q, 1H), 5.10-5.00 (m, 1H), 4.72-4.50 (m, 1H), 4.48-4.44 (m, 1H), 4.33-4.31 (m, 1H), 3.86-3.70 (m, 2H), 3.69 (s, 3H), 3.68-3.52 (m, 1H), 2.91 (dt, 1H), 2.70-2.64 (m, 2H), 2.55 (s, 2H), 2.28-2.15 (m, 3H), 1.98 (s, 3H), 1.78-0.99 (m, 4H).
rac-2-(((1R,6R)-5-(2-(5-Chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(2-(2,2-difluorocyclopropoxy)ethyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylic acid
LiOH monohydrate (19 mg, 0.43 mmol) was added to a solution of rac-methyl 2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(2-(2,2-difluorocyclopropoxy)ethyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Intermediate 85 (197 mg, 0.28 mmol) in THF:H2O (2:1, 5 mL) and the reaction mixture was stirred at rt for 16 h. The reaction mixture was acidified with NaH2PO4 (aq) solution and diluted with water. The formed precipitate was collected by filtration, washed with water and dried in vacuo to give the title compound (112 mg, 62%); HRMS (ESI) m z [M+H]+ calcd for C34H35ClF2N5O6: 682.2238, found: 682.2252;_1H NMR (500 MHz, DMSO-d6) δ 12.40 (s, 1H), 8.83-8.50 (m, 1H), 8.04-7.89 (m, 1H), 7.81 (s, 1H), 7.58 (t, 1H), 7.24 (s, 1H), 6.70 (q, 1H), 6.52 (dd, 1H), 6.31 (t, 1H), 4.72-4.50 (m, 2H), 3.94 (s, 4H), 3.89-3.81 (m, 1H), 3.81-3.69 (m, 2H), 3.67-3.60 (m, 1H), 2.96-2.79 (m, 1H), 2.72-2.63 (m, 2H), 2.61-2.55 (m, 2H), 2.42-2.36 (m, 1H), 2.36-2.26 (m, 1H), 2.21-2.07 (m, 1H), 1.97 (s, 3H), 1.74-1.55 (m, 2H), 1.39-1.17 (m, 2H).
LiOH monohydrate (7 mg, 0.16 mmol) was added to a solution of rac-methyl 2-(((1R,6R)-5-(2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(2-(2,2-difluorocyclopropoxy)ethyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Intermediate 86 (74 mg, 0.11 mmol) in THF:H2O (2:1, 5 mL) and the reaction mixture was stirred at rt for 16 h. The reaction mixture was acidified with NaH2PO4 (aq) and diluted with water. The formed precipitate was collected by filtration, washed with water and dried in vacuo to give the title compound (53 mg, 73%); HRMS (ESI) m/z [M+H]+ calcd for C33H32ClF3N5O5: 670.2038, found: 670.2042; 1H NMR (500 MHz, DMSO-d6) δ 12.42 (s, 1H), 8.80-8.55 (m, 1H), 8.15-7.86 (m, 2H), 7.67-7.54 (m, 1H), 7.48 (d, 1H), 6.71 (q, 1H), 6.53 (dd, 1H), 6.34-6.28 (m, 1H), 4.78-4.59 (m, 2H), 3.96 (s, 2H), 3.90-3.60 (m, 3H), 3.02-2.79 (m, 1H), 2.69 (d, 2H), 2.62-2.58 (m, 2H), 2.38-2.30 (m, 1H), 2.21-2.07 (m, 1H), 1.97 (s, 3H), 1.78-1.55 (m, 3H), 1.41-1.07 (m, 2H).
1,3,4,6,7,8-Hexahydro-2H-pyrimido[1,2-a]pyrimidine (38 mg, 0.28 mmol) was added to a solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 93 (120 mg, 0.18 mmol) in MeCN (12 mL) and water (3 mL) and the reaction mixture was stirred at 20° C. for 4 h. The reaction mixture was concentrated and the residue was diluted with EtOAc (100 mL). The organic layer was washed with sat brine (25 mL), dried over Na2SO4, filtered and evaporated. The crude product was purified by preparative HPLC, PrepMethod H (gradient: 35-60%) to give the title compound Isomer 1 (64 mg, 55%) as a white solid; MS (ESI) m/z [M+H]+ 640.3; 1H NMR (400 MHz, DMSO-d6) δ 1.50-1.71 (m, 3H), 1.90-2.02 (m, 1H), 2.05 (s, 3H), 2.30-2.39 (m, 2H), 2.59-2.79 (m, 2H), 3.13 (d, 1H), 3.21-3.29 (m, 2H), 3.72 (d, 1H), 3.97 (s, 3H), 4.10 (d, 1H), 4.22 (dt, 1H), 4.31 (q, 1H), 4.44 (td, 1H), 4.64-4.78 (m, 2H), 5.12 (d, 1H), 6.29 (dd, 1H), 6.51 (dd, 1H), 6.73 (t, 1H), 7.27 (d, 1H), 7.66-7.80 (m, 2H), 7.89 (d, 1H), 7.98 (dd, 1H), 12.82 (s, 1H).
The title compound was prepared as described for Example 15a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 94 (130 mg, 0.20 mmol) to give the title compound Isomer 2 (87 mg, 68%) as a white solid; MS (ESI) m/z [M+H]+ 640.3; 1H NMR (400 MHz, DMSO-d6) δ 1.73-1.76 (m, 2H), 2.02 (s, 4H), 2.21-2.39 (m, 2H), 2.42-is 2.49 (m, 1H), 2.61-2.75 (m, 2H), 3.12-3.16 (m, 2H), 3.40-3.50 (m, 1H), 3.56 (d, 1H), 3.97 (s, 3H), 4.13-4.29 (m, 2H), 4.40-4.46 (m, 1H), 4.49-4.62 (m, 2H), 4.88 (dd, 1H), 5.06-5.15 (m, 1H), 6.30-6.40 (m, 1H), 6.48 (dd, 1H), 6.73 (t, 1H), 7.28 (d, 1H), 7.67-7.78 (m, 2H), 7.86-8.04 (m, 2H), 12.85 (s, 1H).
1,3,4,6,7,8-Hexahydro-2H-pyrimido[1,2-a]pyrimidine (35 mg, 0.25 mmol) was added to a solution of methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 97 (110 mg, 0.17 mmol) in MeCN (5 mL) and water (1.6 mL) and the reaction mixture was stirred at 20° C. for 2 h. The pH of the reaction mixture was adjusted to 7 by citric acid (0.5 M) and water (20 mL) was added. The aqueous layer was extracted with EtOAc (3×20 mL) and the combined organic layer was dried over Na2SO4, filtered and evaporated. The crude product was purified by preparative HPLC, PrepMethod H (gradient: 48-70%) to give the title compound Isomer 1 (66 mg, 61%) as a white solid; MS (ESI) m/z [M+H]+ 644.2; 1H NMR (300 MHz, DMSO-d6) δ 1.39-1.81 (m, 3H), 1.99 (t, 1H), 2.06 (s, 3H), 2.37 (dt, 2H), 2.58-2.85 (m, 2H), 3.15 (d, 1H), 3.31-4.10 (m, 2H), 3.80 (d, 1H), 4.15 (d, 1H), 4.21-4.42 (m, 2H), 4.46 (q, 1H), 4.67-4.93 (m, 2H), 5.10-5.21 (m, 1H), 6.29 (d, 1H), 6.52 (d, 1H), 6.74 (t, 1H), 7.73-7.80 (m, is 2H), 7.82 (d, 1H), 7.89-8.04 (m, 1H), 8.27 (d, 1H).
The title compound was prepared from methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 98 (140 mg, 0.21 mmol) as described for Example 16a to give the title compound Isomer 2 (74 mg, 54%) as a white solid; MS (ESI) m/z [M+H]+ 644.1; 1H NMR (300 MHz, DMSO-d6) δ 1.76 (d, 2H), 2.02 (s, 4H), 2.26 (q, 1H), 2.41 (q, 2H), 2.60-2.81 (m, 2H), 3.17 (t, 2H), 3.45 (d, 1H), 3.65 (d, 1H), 4.24 (d, 2H), 4.38-4.51 (m, 1H), 4.55 (dd, 1H), 4.66 (d, 1H), 4.97 (dd, 1H), 5.11 (q, 1H), 6.35 (d, 1H), 6.49 (dd, 1H), 6.74 (t, 1H), 7.71-7.77 (m, 2H), 7.82 (d, 1H), 7.91-8.05 (m, 1H), 8.29 (d, 1H), 13.21 (s, 1H).
The title compound was prepared from methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 99 (140 mg, 0.21 mmol) as described for Example 16a to give the title compound Isomer 3 (86 mg, 63%) as a white solid; MS (ESI) m/z [M+H]+ 644.1; 1H NMR (300 MHz, DMSO-d6) δ 1.60 (d, 1H), 1.70 (s, 1H), 1.85-1.98 (m, 1H), 2.02 (s, 3H), 2.21 (t, 1H), 2.38 (dt, 2H), 2.60-2.85 (m, 2H), 3.20 (t, 1H), 3.39-3.45 (m, 2H), 3.76 (d, 1H), 4.07-4.36 (m, 3H), 4.47 (q, 1H), 4.77 (dd, 1H), 4.88 (dd, 1H), 5.15-5.25 (m, 1H), 6.17-6.40 (m, 1H), 6.48 (d, 1H), 6.74 (t, 1H), 7.72-7.78 (m, 2H), 7.83 (d, 1H), 7.89-8.05 (m, 1H), 8.28 (d, 1H), 13.26 (s, 1H).
1,3,4,6,7,8-Hexahydro-2H-pyrimido[1,2-a]pyrimidine (59 mg, 0.42 mmol) was added to a solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 102 (180 mg, 0.28 mmol) in MeCN (6 mL) and water (1.5 mL) and the reaction mixture was stirred at 20° C. for 2 h. The reaction mixture was concentrated and the residue was diluted with EtOAc (50 mL). The organic layer was washed with sat brine (25 mL), dried over Na2SO4, filtered and evaporated. The crude product was purified by preparative HPLC, PrepMethod I (gradient: 40-65%), to give the title compound Isomer 1 (102 mg, 58%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H32F2N5O5: 628.2366, found: 628.2400; 1H NMR (400 MHz, DMSO-d6) δ 1.50-1.61 (m, 2H), 1.61-1.69 (m, 1H), 1.93 (dd, 1H), 2.01 (s, 3H), 2.28-2.46 (m, 2H), 2.62-2.78 (m, 2H), 3.11-3.18 (m, 1H), 3.25-3.40 (m, overlapped with solvent), 3.75 (d, 1H), 4.12 (d, 1H), 4.15-4.28 (m, 2H), 4.44-4.52 (m, 1H), 4.70-4.86 (m, is 2H), 5.14-5.24 (m, 1H), 6.33 (dd, 1H), 6.48 (dd, 1H), 6.73 (t, 1H), 7.53 (dd, 1H), 7.69-7.77 (m, 2H), 7.94-8.02 (m, 1H), 8.18 (d, 1H), 13.04 (s, 1H).
The title compound was prepared in analogy with the description for Example 17a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 103 (150 mg, 0.23 mmol). The crude product was purified by preparative HPLC, PrepMethod H (gradient: 40-70%), to give the title compound Isomer 2 (100 mg, 68%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H32F2N5O5: 628.2366, found: 628.2404; 1H NMR (400 MHz, DMSO-d6) δ 1.68-1.89 (m, 2H), 2.02 (s, 4H), 2.26 (p, 1H), 2.33-2.48 (m, 1H), 2.58-2.79 (m, 2H), 3.18 (t, 1H), 3.28-3.35 (m, 1H), 3.44 (dt, 1H), 3.62 (d, 1H), 4.23 (dd, 2H), 4.43-4.51 (m, 1H), 4.52-4.58 (m, 1H), 4.65 (dd, 1H), 4.95 (dd, 1H), 5.05-5.20 (m, 1H), 6.35 (dd, 1H), 6.48 (dd, 1H), 6.74 (t, 1H), 7.53 (dd, 1H), 7.67-7.78 (m, 2H), 7.92-8.02 (m, 1H), 8.21 (d, 1H), 13.01 (s, 1H).
2-((1R*,6S*)-5-((R*)-2-(4-C3ano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid Isomer 3
The title compound was prepared in analogy with the description for Example 17a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-cyano-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-fluoro-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 104 (180 mg, 0.28 mmol). The crude product was purified by preparative HPLC, PrepMethod H (gradient: 40-70%), to give the title compound Isomer 3 (97 mg, 55%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H32F2N5O5: 628.2366, found: 628.2394; 1H NMR (400 MHz, DMSO-d6) δ 1.55-1.62 (m, 1H), 1.65-1.76 (m, 1H), 1.90-2.00 (m, 1H), 2.01 (s, 3H), 2.17-2.22 (m, 1H), 2.31-2.45 (m, 2H), 2.60-2.77 (m, 2H), 3.20 (t, 1H), 3.21-3.27 (s, 1H), 3.39-3.43 (m, 1H), 3.75 (d, 1H), 4.15-4.28 (m, 3H), 4.46 (td, 1H), 4.73-4.93 (m, 2H), 5.12-5.28 (m, 1H), 6.33 (dd, 1H), 6.48 (dd, 1H), 6.73 (t, 1H), 7.53 (dd, 1H), 7.67-7.83 (m, 2H), 7.91-8.04 (m, 1H), 8.18 (d, 1H), 13.04 (s, 1H).
NaOH (38 mg, 0.95 mmol) was added to a solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 106 (120 mg, 0.19 mmol) in MeOH (10 mL) and water (2 mL) and the reaction mixture was stirred at 25° C. for 12 h. The reaction mixture was concentrated and the pH was adjusted to 5-6 with 0.1 M HCl (1.5 mL). The aqueous layer was concentrated and the residue is was diluted with EtOAc (20 mL). The organic layer was washed with sat brine (2×50 mL), dried over Na2SO4, filtered and evaporated. The residue was purified by preparative HPLC, PrepMethod J (gradient: 30-60%) to give the title compound Isomer 1 (62 mg, 53%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C33H33ClFN4O5: 619.2118, found: 619.2142; 1H NMR (400 MHz, DMSO-d6) δ 1.51-1.71 (m, 3H), 1.91-2.00 (m, 1H), 2.02 (s, 3H), 2.36 (q, 2H), 2.65 (dt, 1H), 2.71-2.78 (m, 1H), 3.06-3.18 (m, 1H), 3.15-3.30 (m, 2H), 3.75 (d, 1H), 4.10 (d, 1H), 4.19-4.37 (m, 2H), 4.45 (q, 1H), 4.74 (qd, 2H), 5.10-5.20 (m, 1H), 6.28 (d, 1H), 6.49 (d, 1H), 6.72 (t, 1H), 7.32 (dd, 1H), 7.54 (dd, 2H), 7.64 (d, 1H), 7.82 (d, 1H), 8.25 (s, 1H).
The title compound was prepared as described for Example 18a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 107 (80 mg, 0.13 mmol) to give the title compound Isomer 2 (47 mg, 60%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C33H33ClFN4O5: 619.2118, found: 619.2156; 1H NMR (400 MHz, DMSO-d6) δ 1.59 (d, 1H), 1.69 (s, 1H), 1.90-1.98 (m, 1H), 1.98 (s, 3H), 2.20 (p, 1H), 2.38 (t, 2H), 2.63-2.77 (m, 2H), 3.19 (t, 1H), 3.41 (m, overlapping with solvent), 3.71 (d, 1H), 4.11-4.28 (m, 3H), 4.46 (q, 1H), 4.72 (dd, 1H), 4.83 (d, 1H), 5.14-5.25 (m, 1H), 6.32 (d, 1H), 6.46 (d, 1H), 6.71 (t, 1H), 7.30-7.37 (m, 1H), 7.51-7.60 (m, 2H), 7.66 (d, 1H), 7.82 (d, 1H), 8.27 (s, 1H).
2-2(((R-(4-Chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid Isomer 4
The title compound was prepared as described for Example 18a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 108 (80 mg, 0.13 mmol) to give the title compound Isomer 4 (45 mg, 57%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C33H33ClFN4O5: 619.2118, found: 619.2132; 1H NMR (400 MHz, DMSO-d6) δ 1.76 (d, 2H), 1.99 (s, 4H), 2.21-2.40 (m, 2H), 2.62-2.72 (m, 2H), 3.16 (t, 1H), 3.25 (s, 2H), 3.44 (d, 1H), 3.58 (d, 1H), 4.16-4.30 (m, 2H), 4.44 (q, 1H), 4.49-4.53 (m, 1H), 4.60 (d, 1H), 4.91 (dd, 1H), 5.05-5.16 (m, 1H), 6.34 (d, 1H), 6.47 (d, 1H), 6.72 (t, 1H), 7.33 (dd, 1H), 7.51-7.59 (m, 2H), 7.63 (d, 1H), 7.83 (d, 1H), 8.26 (s, 1H).
NaOH (30 mg, 0.75 mmol) was added to a solution of methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 110 (100 mg, 0.15 mmol) in MeOH (10 mL) and water (2 mL) and the reaction mixture was stirred at 20° C. for 3 h. The reaction mixture was concentrated and acidified with 0.1 M HCl. The solvent was removed under reduced pressure and the residue was purified by preparative HPLC, PrepMethod K (gradient: 18-55%) to give the title compound Isomer 1 (14 mg, 14%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2238; 1H NMR (400 MHz, DMSO-d6) δ 1.58 (s, 1H), 1.69 (s, 1H), 1.90-1.97 (m, 1H), 1.99 (s, 3H), 2.21 (t, 1H), 2.27-2.42 (m, 2H), 2.67 (t, 2H), 3.15-3.15 (m, 2H), 3.38 (m, overlapped with solvent), 3.67 (d, 1H), 3.97 (s, 3H), 4.11-4.28 (m, 3H), 4.45 (q, 1H), 4.59-4.72 (m, 1H), 4.78-4.85 (m, 1H), 5.11-5.21 (m, 1H), 6.32 (d, 1H), 6.46 (d, 1H), 6.72 (t, 1H), 7.28 (s, 1H), 7.31-7.40 (m, 1H), 7.51-7.62 (m, 2H), 7.88 (s, 1H).
The title compound was prepared as described for Example 19a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 111 (180 mg, 0.27 mmol). The crude product was purified by preparative HPLC, PrepMethod L (gradient: 24-54%) to give the title compound Isomer 2 (58 mg, 33%) as a yellow solid; HRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2262; 1H NMR (400 MHz, DMSO-d6) δ 1.58 (s, 3H), 2.02 (s, 4H), 2.25-2.41 (m, 2H), 2.658-2.78 (m, 2H), 3.13 (s, 1H), 3.20-3.30 (m, 2H), 3.70 (d, 1H), 3.95 (s, 3H), 4.09 (d, 1H), 4.20 (s, 1H), 4.32 (q, 1H), 4.44 (q, 1H), 4.63 (d, 1H), 4.73 (d, 1H), 5.13 (s, 1H), 6.28 (d, 1H), 6.49 (d, 1H), 6.72 (t, 1H), 7.32 (d, 2H), 7.54 (t, is 2H), 7.85 (s, 1H).
2-2(((R-(4-Chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylic acid Isomer 3
The title compound was prepared as described for Example 19a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 112 (140 mg, 0.21 mmol). The crude product was purified by preparative HPLC, PrepMethod J (gradient: 30-60%) to give the title compound Isomer 3 (81 mg, 59%) as a white solid; HIRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2256; 1H NMR (400 MHz, DMSO-d6) δ 1.61 (s, 1H), 1.71 (s, 2H), 2.03 (s, 4H), 2.34 (t, 1H), 2.40-2.48 (m, 1H), 2.63-2.73 (m, 2H), 3.11-3.29 (m, 3H), 3.58 (d, 1H), 3.95 (s, 3H), 4.14 (d, 1H), 4.38 (p, 2H), 4.44-4.61 (m, 2H), 4.84 (dd, 1H), 5.07 (q, 1H), 6.29 (d, 1H), 6.50 (d, 1H), 6.72 (t, 1H), 7.28 (s, 1H), 7.33 (dd, 1H), 7.50-7.61 (m, 2H), 7.87 (s, 1H).
The title compound was prepared as described for Example 19a from methyl 2-(((1R*,6S*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-4-methoxy-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 113 (70 mg, 0.11 mmol) to give the title compound Isomer 4 (35 mg, 52%) as a white solid; HIRMS (ESI) m/z [M+H]+ calcd for C34H35ClFN4O6: 649.2224, found: 649.2252; 1H NMR (400 MHz, DMSO-d6) δ 1.75 (d, 2H), 1.99 (s, 4H), 2.27 (t, 1H), 2.35 (t, 1H), 2.46 (d, 1H), 2.59-2.78 (m, 2H), 3.15 (t, 1H), 3.23 (s, 1H), 3.43 (d, 1H), 3.55 (d, 1H), 3.96 (s, 3H), 4.12-4.28 (m, 2H), 4.37-4.46 (m, 1H), 4.46-4.64 (m, 2H), 4.87 (dd, 1H), 5.01-5.14 (m, 1H), 6.34 (d, 1H), 6.47 (d, 1H), 6.72 (t, 1H), 7.28 (s, 1H), 7.33 (dd, 1H), 7.49-7.62 (m, 2H), 7.89 (d, 1H).
NaOH (37 mg, 0.92 mmol) was added to a solution of methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 120 (120 mg, 0.18 mmol) in MeOH (10 mL) and water (2 mL) and the reaction mixture was stirred at 20° C. for 3 h. The reaction mixture was concentrated at reduced pressure and the residue was diluted with water (15 mL) and the pH was adjusted to 7 with 2 M HCl. The aqueous layer was extracted with EtOAc (3×15 mL) and the combined organic layer was dried over Na2SO4, filtered and concentrated at reduced pressure. The crude compound was purified by preparative HPLC, PrepMethod K (gradient: 17-51%) to give the title compound Isomer 1 (88 mg, 75%) as a white solid; HIRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1810; 1H NMR (300 MHz, CD3OD) δ 1.77 (q, 2H), 1.87-2.01 (m, 1H), 2.02 (s, 3H), 2.25 (p, 1H), 2.39-2.59 (m, 2H), 2.70-2.81 (m, 2H), 3.25-3.45 (m, overlapped with solvent), 3.87 (d, 1H), 4.27 (d, 1H), 4.31-4.46 (m, 2H), 4.49-4.68 (m, 1H), 4.78 (dd, 1H), 5.00 (dd, 1H), 5.25-5.36 (m, 1H), 6.36 (dd, 1H), 6.44 (dd, 1H), 6.74 (t, 1H), 7.68 (dd, 1H), 7.89 (dd, 1H), 7.99 (d, 1H), 8.26 (d, 1H), 8.61 (dd, 1H).
The title compound was prepared as described for Example 20a from methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 121 (120 mg, 0.18 mmol) to give the title compound Isomer 2 (82 mg, 70%) as a white solid; HIRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1820; 1H NMR (300 MHz, DMSO-d6) δ 1.72 (s, 3H), 2.01 (s, 3H), 2.05-2.19 (m, 1H), 2.30-2.50 (m, 2H), 2.60-2.78 (m, 2H), 3.12-3.30 (m, 3H), 3.64 (d, 1H), 4.19 (d, 1H), 4.30-4.55 (m, 3H), 4.65 (d, 1H), 4.83-4.99 (m, 1H), 5.05-5.19 (m, 1H), 6.32 (d, 1H), 6.48 (d, 1H), 6.74 (t, 1H), 7.61 (d, 1H), 7.83 (s, 1H), 8.02 (dd, 1H), 8.23 (d, 1H), 8.72 (d, 1H).
The title compound was prepared as described for Example 20a from methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 122 (130 mg, 0.20 mmol) to give the title compound Isomer 3 (52 mg, 41%) as a white solid; HIRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1810; 1H NMR (400 MHz, DMSO-d6) δ 1.61 (s, 3H), 2.00 (s, 4H), 2.30-3.41 (m, 2H), 2.60-2.72 (m, 2H), 3.12-3.17 (m, 1H), 3.23-3.38 (m, overlapped with solvent), 3.77 (d, 1H), 4.14 (d, 1H), 4.22-4.30 (m, 2H), 4.45 (q, 1H), 4.68-4.87 (m, 2H), 5.15 (s, 1H), 6.30 (d, 1H), 6.47 (d, 1H), 6.72 (t, 1H), 7.60 (d, 1H), 7.81 (s, 1H), 8.00 (dd, 1H), 8.22 (s, 1H), 8.71 (d, 1H).
The title compound was prepared as described for Example 20a from methyl 4-chloro-2-(((1R*,6S*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 123 (130 mg, 0.20 mmol) to give the title compound is Isomer 4 (38 mg, 30%) as a white solid; HRMS (ESI) m/z [M+H]+ calcd for C32H32Cl2N5O5: 636.1774, found: 636.1818; 1H NMR (400 MHz, DMSO-d6) δ 1.74 (d, 2H), 1.98 (s, 4H), 2.26 (t, 1H), 2.34 (t, 1H), 2.43 (s, 1H), 2.59-2.79 (m, 2H), 3.15-3.21 (m, 1H), 3.19 (s, 1H), 3.24 (s, 1H), 3.61 (d, 1H), 4.25-4.34 (m, 2H), 4.38-4.48 (m, 1H), 4.53 (q, 1H), 4.63 (d, 1H), 4.94 (dd, 1H), 5.10-5.16 (m, 1H), 6.35 (d, 1H), 6.47 (d, 1H), 6.73 (t, 1H), 7.62 (d, 1H), 7.80 (s, 1H), 8.00 (dd, 1H), 8.22 (s, 1H), 8.71 (d, 1H).
LiOH monohydrate (13 mg, 0.31 mmol) was added to a solution of methyl 2-(((1R*,6R*)-5-((R*)-2-(4-chloro-2-fluorophenyl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 125 (78 mg, 0.12 mmol) in THF:water (1:1, 2 mL) and the reaction mixture was stirred at rt overnight. The reaction mixture was concentrated in vacuo, the residue was diluted with water (3 mL) and acidified to pH 3 with 10% NaH2PO4 (aq). The precipitate was filtered off and dried in vacuo to give the title compound Isomer 1 (30 mg, 32%) as a solid; MS (ESI) m/z [M+H]+ 618.2; 1H NMR (600 MHz, DMSO-d6) δ 12.75 (s, 1H), 8.67 (d, 1H), 8.25 (s, 1H), 7.99 (dd, 1H), 7.80 (dd, 1H), 7.67-7.64 (d, 1H), 7.61-7.57 (d, 1H), 6.72 (t, 1H), 6.51 (d, 1H), 6.39 (d, 1H), 5.21-5.05 (m, 1H), 4.84-4.75 (m, 1H), 4.66 (dd, 1H), 4.51 (q, 1H), 4.43 (q, 1H), 4.38 (d, 1H), 4.16 (dt, 1H), 4.05 (d, 1H), 3.73 (dd, 1H), 3.62-3.56 (m, 2H), 3.37-3.32 (m, overlapped with solvent), 3.08 (d, 1H), 2.73 (d, 1H), 2.65-2.60 (m, overlapped with solvent), 2.43-2.37 (m, 2H), 2.31-2.21 (m, is 1H), 2.00 (s, 3H)
The title compound was prepared as described for Example 21a from methyl 2-(((1R*,6R*)-5-((R*1-2-(4-chloro-2-fluoronhenyl)-2-methylbenzo[d][1.31dioxol-4-yll-2.5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 126 (28 mg, 44 μmol) to give the title compound Isomer 2 (21 mg, 93%) as a solid; MS (ESI) m/z [M+H]+ 618.2; 1H NMR (600 MHz, DMSO-d6) δ 12.76 (s, 1H), 8.70 (d, 1H), 8.25 (s, 1H), 7.99 (d, 1H), 7.80 (d, 1H), 7.67-7.64 (d, 1H), 7.62-7.59 (d, 1H), 6.72 (t, 1H), 6.50 (d, 1H), 6.41 (d, 1H), 5.15 (s, 1H), 4.83 (d, 1H), 4.66 (dd, 1H), 4.49-4.53 (m, 1H), 4.47-4.37 (m, 2H), 4.15 (d, 1H), 4.07 (d, 1H), 3.76 (dd, 1H), 3.72 (t, 1H), 3.66 (t, 1H), 3.57 (d, 1H), 3.21 (d, 2H), 3.12-3.00 (m, 2H), 2.76-2.70 (m, 1H), 2.60 (s, overlapped with solvent), 2.30-2.25 (m, 1H), 1.98 (s, 3H).
The title compound was prepared as described for Example 21a from methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 128 (41 mg, 0.066 mmol) to give the title compound Isomer 3 (35 mg, 86%) as a solid; MS (ESI) m/z [M+H]+ 602.2; 1H NMR (600 MHz, DMSO-d6) δ 12.69 (s, 1H), 8.72 (d, 1H), 8.25 (s, 1H), 8.01 (dd, 1H), 7.78 (dd, 1H), 7.65-7.56 (m, 2H), 6.71 (t, 1H), 6.55 (d, 1H), 6.32 (d, 1H), 5.14-5.11 (m, 1H), 4.84 (dd, 1H), 4.63 (dd, 1H), 4.50-4.43 (m, 1H), 4.33 (dt, 1H), 3.86 (s, 1H), 3.79 (d, 1H), 3.72 (s, 1H), 3.03 (s, 1H), 2.81-2.60 (m, 4H), 2.41-2.35 (m, 1H), 2.31 (s, 1H), 2.09 (q, 1H), 1.97 (s, 3H), 1.64 (p, 1H), 1.55-1.49 (m, 1H), 1.16 (s, 1H).
The title compound was prepared as described for Example 21a from methyl 2-(((1R*,6R*)-5-((R*)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(((S)-oxetan-2-yl)methyl)-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 129 (42 mg, 0.68 mmol) to give the title compound Isomer 4 (35 mg, 84%) as a solid; MS (ESI) m/z [M+H]+ 602.2; 1H NMR (600 MHz, DMSO-d6) δ 12.72 (s, 1H), 8.67 (d, 1H), 8.24 (s, 1H), 7.93 (dd, 1H), 7.78 (dd, 1H), 7.68-7.47 (m, 2H), 6.71 (t, 1H), 6.51 (d, 1H), 6.32 (d, 1H), 5.09-5.05 (m, 1H), 4.81 (dd, 1H), 4.68 (dd, 1H), 4.48 (q, 1H), 4.37 (dt, 1H), 3.98 (d, 1H), 3.78 (d, 1H), 3.62 (s, 1H), 2.79-2.52 (m, 5H), 2.45-2.38 (m, overlaps with solvent), 2.23 (q, 1H), 1.97 (s, 3H), 1.86-1.68 (m, 2H), 1.54-1.38 (m, 1H).
rel-2-(((1R,6R)-5-((R)-2-(5-Chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylic acid Isomer 1
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate, Isomer 1 Intermediate 134 (68 mg, 0.10 mmol) to give the title compound Isomer 1 (65 mg, 94%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H33ClFN6O4: 643.2230, found: 643.2268; 1H NMR (500 MHz, DMSO-d6) δ 13.09 (s, 1H), 8.68 (d, 1H), 8.13 (s, 1H), 7.95 (dd, 1H), 7.58 (d, 1H), 7.50 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.30 (d, 1H), 4.74 (d, 1H), 4.55 (d, 1H), 3.86-3.72 (m, 3H), 2.97 (d, 1H), 2.74-2.66 (m, 1H), 2.66-2.54 (m, 3H), 2.39-2.29 (m, overlapped with solvent), 2.21-2.15 (m, 1H), 1.97 (s, 3H), 1.72-1.56 (m, 2H), 1.33-1.30 (m, 1H), 0.79-0.63 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate, Isomer 2 Intermediate 135 (62 mg, 94 μmol) to give the title compound Isomer 2 (50 mg, 93%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H33ClFN6O4: 643.2230, found: 643.2246; 1H NMR (500 MHz, DMSO-d6) δ 13.08 (s, 1H), 8.72 (d, 1H), 8.13 (s, 1H), 8.01 (dd, 1H), 7.59 (d, 1H), 7.50 (d, 1H), 6.71 (t, 2H), 6.55 (d, 1H), 6.32 (d, 1H), 4.78 (d, 1H), 4.54 (d, 1H), 3.84-3.77 (m, 3H), 3.07-2.90 (m, 1H), 2.74-2.52 (m, 4H), 2.43-2.38 (m, overlapped with solvent), 2.35-2.27 (m, 1H), 2.11-2.08 (m, lfH), 1.97 (s, 3H), 1.64 (dt, 1H), 1.57 (d, 1H), 1.26 (t, 1H), 0.78-0.65 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate, Isomer 3 Intermediate 136 (84 mg, 0.13 mmol) to give the title compound Isomer 3 (52 mg, 73%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H33ClFN6O4: 643.2230, found: 643.2244; 1H NMR (500 MHz, DMSO-d6) δ 13.08 (s, 1H), 8.68 (d, 1H), 8.13 (s, 1H), 7.95 (dd, 1H), 7.58 (d, 1H), 7.50 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.30 (d, 1H), 4.74 (d, 1H), 4.55 (d, 1H), 3.85-3.71 (m, 3H), 2.97 (d, 1H), 2.75-2.66 (m, 1H), 2.66-2.53 (m, 3H), 2.42-2.29 (m, overlapped with solvent), 2.25-2.12 (m, 1H), 1.97 (s, 3H), 1.72-1.65 (m, 1H), 1.62 (d, 1H), 1.32 (t, 1H), 0.79-0.65 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate, Isomer 4 Intermediate 137 (59 mg, 90 μmol) to give the title compound Isomer 4 (18 mg, 31%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H33ClFN6O4: 643.2230, found: 643.2280; 1H NMR (500 MHz, DMSO-d6) δ 13.07 (s, 1H), 8.72 (d, 1H), 8.12 (d, 1H), 8.01 (dd, 1H), 7.59 (d, 1H), 7.50 (d, 1H), 6.71 (t, 1H), 6.54 (d, 1H), 6.31 (d, 1H), 4.78 (d, 1H), 4.54 (d, 1H), 3.86-3.71 (m, 3H), 3.01 (d, 1H), 2.72-2.57 (m, 4H), 2.44-2.39 (m, overlapped with solvent), 2.35-2.29 (m, 1H), 2.12-2.05 (m, 1H), 1.97 (s, 3H), 1.69-1.62 (m, 1H), 1.61-1.54 (m, 1H), 1.31-1.19 (m, 1H), 0.79-0.67 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Isomer 2 Intermediate 140 (65 mg, 0.10 mmol) to give the title compound Isomer 2 (56 mg, 87%) as a solid; HIRMS (ESI) m/z [M+H]+ calcd for C33H31ClFN6O4: 629.2074, found: 629.2102; 1H NMR (500 MHz, DMSO-d6) δ 13.02 (s, 1H, exchange), 8.72 (d, 1H), 8.23 (s, 1H), 8.01 (dd, 1H), 7.59 (d, 1H), 7.53 (d, 1H), 6.71 (t, 1H), 6.55 (d, 1H), 6.32 (d, 1H), 4.90 (d, 1H), 4.72 (d, 1H), 3.92-3.81 (m, 2H), 3.80-3.72 (m, 1H), 2.99 (dd, 1H), 2.75-2.62 (m, 3H), 2.38-2.27 (m, 1H), 2.13-2.05 (m, 1H), 1.97 (s, 3H), 1.70-1.63 (m, 1H), 1.58 (dd, 1H), 1.47-1.22 (m, 5H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-fluoro-1H-benzo[d]imidazole-6-carboxylate Isomer 4 Intermediate 141 (99 mg, 0.15 mmol) to give the title compound Isomer 4 (73 mg, 87%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C33H31ClFN6O4: 629.2074, found: 629.2110; 1H NMR (500 MHz, DMSO-d6) δ 13.1 (s, 1H, exchange), 8.68 (d, 1H), 8.23 (d, 1H), 7.95 (dd, 1H), 7.67-7.41 (m, 2H), 6.70 (t, 1H), 6.52 (d, 1H), 6.30 (d, 1H), 4.87 (d, 1H), 4.74 (d, 1H), 3.86 (q, 2H), 3.77 (d, 1H), 2.96 (dd, 1H), 2.76-2.66 (m, 1H), 2.63-2.58 (m, 1H), 2.38-2.32 (m, 1H), 2.22-2.12 (m, 1H), 1.97 (s, 3H), 1.76-1.63 (m, 2H), 1.46-1.28 (m, 6H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Isomer 1 Intermediate 146 (16 mg, 24 μmol) to give the title compound Isomer 1 (15 mg, 94%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H34ClN605: 642.2274, found: 642.2324; 1H NMR (500 MHz, DMSO-d6) δ 12.85 (s, 1H), 8.67 (d, 1H), 7.97-7.93 (m, 2H), 7.57 (d, 1H), 7.28 (s, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.30 (d, 1H), 4.80 (d, 1H), 4.67 (d, 1H), 3.95 (s, 3H), 3.87-3.73 (m, 3H), 2.93 (d, 1H), 2.71 (q, 1H), 2.45-2.39 (m, overlapped with solvent), 2.33 (q, 1H), 2.17 (q, 1H), 1.97 (s, 3H), 1.73-1.60 (m, 2H), 1.44-1.18 (m, 5H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Isomer 2 Intermediate 147 (20 mg, 31 μmol) to give the title compound Isomer 2 (20 mg, 93%) as a solid; HIRMS (ESI) m/z [M+H]+ calcd for C34H34ClN605: 642.2274, found: 642.2310; 1H NMR (500 MHz, DMSO-d6) δ 12.85 (s, 1H), 8.72 (d, 1H), 8.01 (dd, 1H), 7.96 (d, 1H), 7.59 (d, 1H), 7.27 (d, 1H), 6.71 (t, 1H), 6.60-6.52 (m, 1H), 6.32 (d, 1H), 4.83 (d, 1H), 4.64 (d, 1H), 3.95 (s, 3H), 3.87-3.72 (m, 3H), 2.97 (d, 1H), 2.78-2.62 (m, 2H), 2.45-2.38 (m, 1H), 2.30 (q, 1H), 2.15-2.05 (m, 1H), 1.97 (s, 3H), 1.65 (p, 1H), 1.56 (q, 1H), 1.44-1.20 (m, 5H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Isomer 3 Intermediate 148 (19 mg, 29 μmol) to give the title compound Isomer 3 (18 mg, 96%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H34ClN605: 642.2274, found: 642.2284; 1H NMR (500 MHz, DMSO-d6) δ 12.85 (s, 1H), 8.67 (d, 1H), 7.97-7.93 (m, 2H), 7.57 (dd, 1H), 7.28 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.30 (d, 1H), 4.80 (d, 1H), 4.67 (d, 1H), 3.95 (s, 3H), 3.89-3.73 (m, 3H), 2.93 (d, 1H), 2.71 (q, 1H), 2.59-2.39 (m, overlapped with solvent), 2.37-2.29 (m, 1H), 2.18 (q, 1H), 1.97 (s, 3H), 1.70 (p, 1H), 1.63 (q, 1H), 1.43-1.18 (m, 5H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-cyanocyclopropyl)methyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Isomer 4 Intermediate 149 (20 mg, 31 μmol) to give the title compound Isomer 4 (18 mg, 92%) as a solid; HRMS (ESI) m/z [M+H]+ calcd for C34H34ClN605: 642.2274, found: 642.2278; 1H NMR (500 MHz, DMSO-d6) δ 12.86 (s, 1H), 8.72 (d, 1H), 8.01 (dd, 1H), 7.95 (d, 1H), 7.59 (dd, 1H), 7.27 (d, 1H), 6.71 (t, 1H), 6.54 (d, 1H), 6.32 (d, 1H), 4.83 (d, 1H), 4.64 (d, 1H), 3.95 (s, 3H), 3.87-3.72 (m, 3H), 2.97 (d, 1H), 2.77-2.63 (m, 2H), 2.45-2.38 (m, 1H), 2.30 (q, 1H), 2.09 (q, 1H), 1.97 (s, 3H), 1.65 (p, 1H), 1.56 (q, 1H), 1.43-1.20 (m, 5H).
The title compound was prepared as described for Example 21a from rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-1H-benzo[d]imidazole-6-carboxylate Isomer 1 Intermediate 154 (29 mg, 43 μmol) to give the title compound Isomer 1 (28 mg, 92%) as a solid; MS (ESI) m/z [M+H]+ 659.2; 1H NMR (500 is MHz, DMSO-d6) δ 13.12 (s, 1H), 8.72 (d, 1H), 8.23 (s, 1H), 8.01 (dd, 1H), 7.78 (s, 1H), 7.59 (d, 1H), 6.71 (t, 1H), 6.54 (d, 1H), 6.31 (d, 1H), 4.78 (d, 1H), 4.55 (d, 1H), 3.90-3.75 (m, 3H), 3.01 (d, 1H), 2.74-2.57 (m, 4H), 2.32 (q, 1H), 2.09 (q, 1H), 1.97 (s, 3H), 1.66 (p, 1H), 1.56 (q, 1H), 1.29-1.20 (m, 2H), 0.80-0.65 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-1H-benzo[d]imidazole-6-carboxylate Isomer 2 Intermediate 155 (32 mg, 46 μmol) to give the title compound Isomer 2 (27 mg, 87%) as a solid; MS (ESI) m/z [M+H]+ 659.2; 1H NMR (500 MHz, DMSO-d6) δ 13.12 (s, 1H), 8.68 (d, 1H), 8.23 (d, 1H), 7.95 (dd, 1H), 7.79 (d, 1H), 7.58 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.29 (d, 1H), 4.75 (d, 1H), 4.56 (d, 1H), 3.92-3.73 (m, 3H), 2.96 (d, 1H), 2.71-2.59 (m, 4H), 2.47-2.31 (m, 2H), 2.20-2.13 (m, 1H), 1.97 (s, 3H), 1.74-1.65 (m, 1H), 1.65-1.58 (m, 1H), 1.31 (p, 1H), 0.80-0.62 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-1H-benzo[d]imidazole-6-carboxylate Isomer 3 Intermediate 156 (27 mg, 40 μmol) to give the title compound Isomer 3 (26 mg, 94%) as a solid; MS (ESI) m/z [M+H]+ 659.0; 1H NMR (500 MHz, DMSO-d6) δ 13.12 (s, 1H), 8.68 (d, 1H), 8.23 (d, 1H), 7.95 (dd, 1H), 7.79 (d, 1H), 7.58 (d, 1H), 6.70 (t, 1H), 6.51 (d, 1H), 6.29 (d, 1H), 4.75 (d, 1H), 4.56 (d, 1H), 3.93-3.68 (m, 3H), 2.96 (d, 1H), 2.74-2.66 (m, 1H), 2.63 (d, 2H), 2.61-2.54 (m, 2H), 2.39-2.32 (m, 1H), 2.17 (q, 1H), 1.97 (s, 3H), 1.76-1.55 (m, 2H), 1.35-1.26 (m, 1H), 0.79-0.65 (m, 4H
The title compound was prepared as described for Example 21a from rel-methyl 4-chloro-2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-((1-(cyanomethyl)cyclopropyl)methyl)-1H-benzo[d]imidazole-6-carboxylate Isomer 4 Intermediate 157 (29 mg, 43 μmol) to give the title compound Isomer 4 (27 mg, 93%) as a solid; MS (ESI) m/z [M+H]+ 659.0; 1H NMR (500 MHz, DMSO-d6) δ 13.12 (s, 1H), 8.72 (d, 1H), 8.23 (d, 1H), 8.01 (dd, 1H), 7.78 (d, 1H), 7.59 (d, 1H), 6.71 (t, 1H), 6.54 (d, 1H), 6.31 (d, 1H), 4.78 (d, 1H), 4.55 (d, 1H), 3.87-3.76 (m, 3H), 3.01 (d, 1H), 2.72-2.58 (m, 4H), 2.46-2.40 (d, 1H, overlapped with solvent), 2.32 (q, 1H), 2.09 (q, 1H), 1.97 (s, 3H), 1.71-1.62 (m, 1H), 1.60-1.53 (m, 1H), 1.29-1.20 (m, 1H), 0.81-0.63 (m, 4H).
The title compound was prepared as described for Example 21a from rel-methyl 2-(((1R,6R)-5-((R)-2-(5-chloropyridin-2-yl)-2-methylbenzo[d][1,3]dioxol-4-yl)-2,5-diazabicyclo[4.2.0]octan-2-yl)methyl)-1-(2-cyclopropoxyethyl)-4-methoxy-1H-benzo[d]imidazole-6-carboxylate Isomer 3 Intermediate 162 (46 mg, 70 μmol) to give the title compound Isomer 3 (32 mg, 96%) as a solid; MS (ESI) m/z [M+H]+ 646.4; 1H NMR (500 MHz, DMSO-d6) δ 12.75 (s, 1H), 8.72 (d, 1H), 8.02 (dd, 1H), 7.79 (s, 1H), 7.59 (d, 1H), 7.23 (s, 1H), 6.71 (t, 1H), 6.61-6.45 (m, 1H), 6.32 (d, 1H), 4.66-4.46 (m, 2H), 3.93 (s, 3H), 3.84-3.81 (m, 2H), 3.78-3.72 (m, 1H), 3.72-3.64 (m, 2H), 3.25-3.19 (m, 1H), 2.92 (d, 1H), 2.71-2.62 (m, 2H), 2.28 (q, 1H), 2.12-2.09 (m, 1H), 1.97 (s, 3H), 1.69-1.54 (m, 2H), 1.25-1.22 (m, 2H), 0.34-0.32 (m, 2H), 0.27-0.19 (m, 2H).
CHOK1 GLP-1R cAMP Assay
A cell line stably expressing the human GLP-1R receptor (NM_002062.5, including the naturally-occurring variant Leu260Phe) in a CHO-K1 (ATCC® CCL-61™) was used for assay.
GLP-1 receptor mediated agonist activity was determined in a cell-based assay measuring cyclic adenosine monophosphate (cAMP) levels in cells using Homogeneous Time-Resolved Fluorescence (HTRF) cAMP detection kit (CisBio catalog #62AM4PEC, cAMP Gs Dynamic range kit). The cAMP detection method is based on a competitive immunoassay, in which cAMP produced by the cells and cAMP labeled with the dye d2 compete for binding to a Europium-Cryptate-labeled anti-cAMP antibody. The specific HTRF signal is inversely proportional to the concentration of cAMP.
Compounds were added to individual well in 384 well-assay plates (Greiner #784076) using an Echo (LabCyte) dispenser from 10 mM stocks. Varying concentration of compounds were added to wells, and DMSO was used to normalize each well to a volume of 100 nL. A dose response curve of GLP-1(7-36)NH2 (Bachem H-6795) was included in each run. 5 μL of cAMP concentration response standards are applied in specified wells in the assay plates. Cryo-preserved cells are thawed and resuspended in assay buffer pre-heated to 37° C. (20 mM 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid (HEPES) pH 7.4, 1× Hank's Balanced Salt Solution (HBSS, Life Technologies #14065) supplemented with 0.1% (w/v) bovine serum albumin (Sigma, A-7030)). Cells were centrifuged at 250*g for 5 min at rt, and resuspended in room tempered assay buffer to a final density of 0.16*106 cell/mL, to deliver 800 cells/well. 5 μL of assay buffer with 1 mM 3-isobutyl-1-methylxanthin (IBMX; Sigma cat I-7018) was dispensed per well in assay plates using a multidrop combi (Thermo Scientific) subsequently 5 μL of cell suspension was distributed to relevant wells in the assay using a multidrop dispenser. Assay plates were incubated 20 min at rt.
Detection reagents, Europium-Cryptate-labeled anti-cAMP antibody and cAMP labeled with the dye d2, are diluted in lysis buffer, provided by the manufacturer. 5 μL of each detection reagent is supplemented to each assay well using a multidrop dispenser. Assay plates are incubated in the dark for at least one h. The HTRF signal is measured using the HTRF module (excitation: 337 nm, emission A: 665 nm and emission B: 620 nm) in Pherastar FSX (BMG Labtech).
Raw data were converted to pM cAMP using the cAMP standard curve included in each run. Converted data were further analyzed in Genedata Screener (Genedata) and EC50 determinations were made from agonist dose-response curves analyzed with a curve fitting is program using a 4-parameter logistic dose response equation (Equation y=A+((B-A)/1+((C/x){circumflex over ( )}D))) where A is no stimulation, B is full stimulation, C is the EC50 and D is the Hill slope). The percent effect was determined relative to a saturating concentration of a full GLP-1R agonist (GLP-1(7-36)NH2 has 100% effect in this assay setup).
The GLP-1R EC50 values for the Example compounds are set forth in Table 1 herein below.
A HTRF cAMP assay (cAMP Gs dynamic kit; CisoBio Cat #62AM4PEJ) was used to identify agonists of GLP-1R in a pancreatic insulinoma cell line (EndoC-PH1). The EndoC-PH1 cell line was sourced from Univercell Biosolutions and is a genetically engineered human pancreatic R cell line which exhibits glucose-inducible insulin secretion. EndoC-PHlcells have detectable GLP-1R messenger ribonucleic acid (mRNA) as detected by quantitative polymerase chain reaction (qPCR). The functionality of GLP-1R signalling in EndoC-PH1 has been demonstrated by Exendin-4 treatment leading to augmented insulin secretion; an effect which is blunted with short hairpin ribonucleic acid (shRNA)-mediated knockdown of GLP-1R. The EndoC-PH1 cell line is a valid model of human beta cells and applicable for screenings to identify novel drug target candidates (Mol. Metab., 2018, 8, 144-157). CisBio HTRF cAMP kits are based on a competitive immunoassay using cryptate-labelled anti-cAMP antibody and d2-labeled cAMP. The detection kit is intended for the direct is quantitative determination of cAMP. The specific signal (i.e. energy transfer) is inversely proportional to the concentration of cAMP in the standard or sample. Test compounds (10 mM in DMSO) were diluted into assay buffer (HBSS (Sigma #H8264) supplemented with 25 mM HEPES (Gibco #15630, pH 7.4), 0.1% BSA (Sigma #A3059) and 0.5 mM IBMX (Sigma #I7018) included fresh on the day of the assay) into 96 well U-bottom plates (Greiner #650201). Diluted compounds were transferred to ECHO source polypropelene plates (Labcyte #P-05525) and dose response curves were dispensed acoustically using ECHO 550 into black shallow-well u-bottom 384-well HTRF Assay Plates (Corning 4514). Cryovials of EndoC-H1 (supplied at 1×10e7 cells/vial) were used directly for screening. The cryovials and were removed from N2(1) and thawed rapidly in a 37° C. water bath. The cells were resuspended in assay buffer and centrifuged at 300 g for 5 min. Cells were resuspended in assay buffer at the appropriate concentration, typically at 12e5 cells per mL (3000 cells per well, dependent on cell batch) and 2.5 μL diluted cells were added to all wells of destination plate by Multidrop combi reagent dispenser (Thermofisher). The plates were incubated at rt for 30 min. The assay was stopped by adding 2.5 μL anti-cAMP cryptate solution to all wells and 2.5 μL cAMP-d2 solution (both diluted 1:20 in lysis buffer) to columns 1-22 by Combi drop. A volume of 2.5 μL cAMP-d2 solution was added to wells E23 to P24 and 2.5 μL lysis buffer added to wells A23 to D24 by multichannel pipette. The plates were incubated at rt for 1 h and read on an Envision plate reader using excitation wavelength of 320 nm and emission of 590 nm and 660 nm.
Raw data from Envision is converted to % DeltaF according to the manufacturer's instructions. Dose response curves are analysed via 4-Parameter Logistical Analysis and assay plate Z’ values obtained. Samples are graphed as percentage (%) activation plots compared to GIP (1-42, Bachem H-5645) with assay window defined by negative control as basal cell is cAMP levels and positive control are defined by maximum GIP (82.5 nM) signal. GLP-1 (7-36 amide, Bachem H-6795) dose response curve was included on all plates.
The EndoC EC50 values for the Example compounds are set forth in Table 2 herein below.
Inhibition of phosphodiesterase-3 (PDE3) has been shown to result in an increase in cardiovascular mortality in clinical trials (Movsesian M.A., Kukreja R.C. (2011) Phosphodiesterase Inhibition in Heart Failure. In: Francis S., Conti M., Houslay M. (eds) Phosphodiesterases as Drug Targets. Handbook of Experimental Pharmacology, vol 204. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-17969-3 10). Chronic treatment with PDE3 inhibitors has been shown to result in increased mortality, primarily as a result of arrhythmias and sudden death (Expert Opinion on InvestigationalDrugs, 2002, 11, 1529-1536; J. of Cardiovasc. Trans. Res., 2010, 3, 507-515) and it may therefore be an advantage to as far as possible avoid PDE3 inhibitory activity.
Evaluation of the effects of compounds on the activity of the human phosphodiesterase-3A is quantified by measuring the formation of 5′AMP from cAMP using a human recombinant is enzyme expressed in a clonal isolate of Spodoptera frugiperda cells (Sf9) cells.
The test compound, reference compound or water (control) are added to a buffer containing 40 mM tris(hydroxymethyl)aminomethane (Tris)/HCl (pH 7.4) and 8 mM MgCl2, 450 nMcAMP and 0.25 μCi [3H]cAMP.
Thereafter, the reaction is initiated by addition of the enzyme (about 1U) and the mixture is incubated for 20 min at 22° C.
For basal control measurements, the enzyme is omitted from the reaction mixture.
Following incubation SPA beads are added. After 30 min at 22° C. under shaking, the amount of [3H]5′AMP is quantified with a scintillation counter (Topcount, Packard).
The results are expressed as a percent inhibition of the control enzyme activity. The standard inhibitory reference compound is milrinone (CAS number 78415-72-2), which is tested in each experiment at several concentrations to obtain an inhibition curve from which its IC50 value is calculated.
The PDE3 IC50 values for Example compounds and reference compounds are set forth in Table 3 herein below.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/EP2022/077530 | 10/4/2022 | WO |
| Number | Date | Country | |
|---|---|---|---|
| 63262105 | Oct 2021 | US | |
| 63264441 | Nov 2021 | US |
