Patent Application
20230302160
The present disclosure relates to a tumor contrast compound, a preparation method thereof and their application in tumor diagnostic imaging.
Molecular imaging is an interdiscipline formed by the integration of molecular biology and medical imaging. Compared with traditional imaging examinations, molecular imaging technology has the advantages of high sensitivity and quantitativeness. Molecular imaging technology or molecular displaying technology mainly includes magnetic resonance (MR) molecular imaging, nuclear medicine molecular imaging and optical molecular imaging. Among them, near-infrared fluorescence (NIRF) imaging technology in optical molecular imaging technology has been widely used in clinical surgery, and the basic principle is as follows: near-infrared light emitted by fluorescent tracers after excitation is utilized to identify blood vessels, lymph nodes and tumor lesions and the like, and assist clinicians in surgical operations. At present, indocyanine green (ICG) has been widely used in clinical fields as a near-infrared fluorescent tracer. However, because ICG does not specifically identify tumor tissue, the use of ICG in surgical tumor resection surgery can only indirectly label tumor lesions through vascular and tissue differences. Its imaging results are poor, which are manifested as insufficient sensitivity, poor tumor specificity, and short imaging time.
The present disclosure provides a completely new near-infrared fluorescent tumor labeling technology. It is found in the present disclosure that, as shown in
Thus, a first aspect of the present disclosure provides a contrast agent comprising a compound represented by the following formula I and anionic surfactant:
In one or more embodiments, L is a bond, n1 and n2 are the same integers, and R21 and R22 are the same groups.
In one or more embodiments, the group —(CH2)n1-L-R22 is different from the group —(CH2)n2-R21, and the steric hindrance of the group —(CH2)n1-L-R22 is greater than that of the group —(CH2)n2-R21.
In one or more embodiments, R1-R8 are all hydrogen.
In one or more embodiments, L is selected from the group consisting of *-T1-C(O)—, *—C(O)—, *-T1-, *—S(O)2-T2- and *-T2-S(O)2—, wherein T1 is selected from the group consisting of NH, O and S, T2 is selected from the group consisting of O, NH and S, * indicates the location in connection with —(CH2)n1-.
In one or more embodiments, the group —(CH2)n1-L-R22 is the same as the group —(CH2)n2-R21, both of which are C1-6 alkyl groups that are optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH and —NH2. Preferably, the group —(CH2)n1-L-R22 is the same as the group —(CH2)n2-R21, and both are C1-6 alkyl groups that are optionally substituted by —COOH.
In one or more embodiments, the group —(CH2)n1-L-R22 is different from the group —(CH2)n2-R21, the group —(CH2)n1-L-R22 is C1-6 alkyl that is optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH and —NH2, preferably C1-6 alkyl that is optionally substituted by —COOH; the group —(CH2)n2-R21 is C1-6 alkyl that is optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH and —NH2, preferably C1-6 alkyl that is optionally substituted by —COOH, and the carbon chain length of the group —(CH2)n2-R21 is shorter than the carbon chain length of the group —(CH2)n1-L-R22. Preferably, the group —(CH2)n1-L-R22 is C1-6 alkyl that is optionally substituted by —COOH, the group —(CH2)n2-R21 is C1-6 alkyl that is optionally substituted by —COOH, and the carbon chain length of the group —(CH2)n2-R21 is shorter than the carbon chain length of the group —(CH2)n1-L-R22. More preferably, the group —(CH2)n1-L-R22 is C1-6 alkyl that is substituted by —COOH, the group —(CH2)n2-R21 is an unsubstituted C1-6 alkyl, and the carbon chain length of the group —(CH2)n2-R21 is shorter than the carbon chain length of the group —(CH2)n1-L-R22.
In one or more embodiments, the compound of Formula I does not comprise Compound 8.
Preferably, in Formula I, in R1-R8 as well as R21 and R22, said an optionally substituted C3-10 cycloalkyl, an optionally substituted 6-14-membered aryl, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heterocyclic group, an optionally substituted C3-15 cycloalkyl, an optionally substituted C3-15 cycloalkyl C1-6 alkyl, an optionally substituted 6-14-membered arylalkyl group, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heteroaryl C1-6 alkyl, an optionally substituted 5-14-membered heterocyclic group and an optionally substituted 5-14-membered heterocyclic group C1-6 alkyl are each an optionally substituted by 1-3 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, hydroxyl, carboxyl, amino (including —NH2 and —NR′R″), C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, and R″ is selected from H and C1-4 alkyl.
In one or more embodiments, R22 is C6-14 aryl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, amino, C1-6 alkoxy and halogenated C1-6 alkoxy or an optionally substituted C3-15 cycloalkyl C1-6 alkyl.
In one or more embodiments, the group —(CH2)n1-L-R22 is different from the group —(CH2)n2-R21; wherein R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, —NH2, —NR′R″, C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl; R21 is H, halogen, C1-6 alkyl, halogenated C1-6 alkyl, C2-6 alkenyl or C2-6 alkynyl, and when R21 is H, n2 is not 0. Preferably, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; the group —(CH2)n2-R21 is C1-6 alkyl or halogenated C1-6 alkyl.
In one or more embodiments, R1-R8 are hydrogen; L is *-T1-C(O)— or *—C(O)—T1-, wherein T1 is selected from the group consisting of NH, O and S; n1 and n2 are each integer of 1 to 6; the group —(CH2)n2-R21 is C1-6 alkyl; R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
In one or more embodiments, the anion is selected from the group consisting of F−, Cl−, Br−, I−, NO3−, SO42−, PO43−, HPO42−, H2PO4−, CO32−, HCO3−, SO32−, HSO3−, CH3COO− and CH3SO3−.
In one or more embodiments, the compound of Formula I according to the present disclosure has the structure represented by the following formula Ia, Ib or Ic:
In one or more embodiments, the compound of Formula I is one or more selected from the following compounds:
In one or more embodiments, the molar ratio of the compound of Formula I to the anionic surfactant in the contrast agent is 1:(0.1˜20), such as 1:(0.1˜10) or 1:(0.2˜5).
In one or more embodiments, the anionic surfactant is selected from carboxylates, sulfonates, sulfate salts, phosphate ester salts and lipopeptide anionic surfactants; preferably, the anionic surfactant is selected from sulfonates and lipopeptide anionic surfactants; more preferably, the anionic surfactant is selected from the group consisting of surfactin, sodium lauryl sulfonate and sodium lauryl benzene sulfonate.
In one or more embodiments, the final concentration of the compound of Formula I in the contrast agent is in the range of 1˜100 μM, such as 5˜50 μM or 10˜30 μM.
In one or more embodiments, the contrast agent is a lyophilized powder.
In one or more embodiments, the contrast agent is in the form of an aqueous solution.
A second aspect of the present disclosure provides a compound represented by the following Formula I′:
In one or more embodiments of the compound of Formula I′ according to the present disclosure, R1—R8 are all hydrogen.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, L is selected from the group consisting of *-T1-C(O)—, *—C(O)-T1-, *—S(O)2-T2- and *-T2-S(O)2—, wherein T1 is selected from the group consisting of NH, O and S, T2 is selected from the group consisting of O, NH and S, * indicates the location in connection with —(CH2)n1-.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, in R1-R8 as well as R21 and R22, said an optionally substituted C3-10 cycloalkyl, an optionally substituted 6-14-membered aryl, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heterocyclic group, an optionally substituted C3-15 cycloalkyl, an optionally substituted C3-15 cycloalkyl C1-6 alkyl, an optionally substituted 6-14-membered arylalkyl group, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heteroaryl C1-6 alkyl, an optionally substituted 5-14-membered heterocyclic group and an optionally substituted 5-14-membered heterocyclic group C1-6 alkyl are each an optionally substituted by 1-3 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, hydroxyl, carboxyl, amino (including —NH2 and —NR′R″), C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, and R″ is selected from H and C1-4 alkyl.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, amino, C1-6 alkoxy and halogenated C1-6 alkoxy.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, —NH2, —NR′R″, C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; R21 is H, halogen, C1-6 alkyl, halogenated C1-6 alkyl, C2-6 alkenyl or C2-6 alkynyl, and when R21 is H, n2 is not 0. Preferably, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; the group —(CH2)n2-R21 is C1-6 alkyl or halogenated C1-6 alkyl.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, R1—R8 are hydrogen; L is *-T1-C(O)— or *—C(O)-T1-, wherein T1 is selected from the group consisting of NH, O and S; n1 and n2 are each integer of 1 to 6; the group —(CH2)n2-R21 is C1-6 alkyl; R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, the anion is selected from the group consisting of F−, Cl−, Br−, I−, NO3−, SO42−, PO43−, HPO42−, H2PO4−, CO32−, HCO3−, SO32−, HSO3−, CH3COO− and CH3SO3−.
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, the compound of Formula I′ has the structure represented by the following formula Ia′, Ib′ or Ic′:
In one or more preceding embodiments of the compound of Formula I′ according to the present disclosure, the compound of Formula I′ is selected from the group consisting of:
The present disclosure also provides a contrast agent comprising a compound of Formula I′ according to the present disclosure and an optional anionic surfactant. Preferably, the anionic surfactant is selected from carboxylates, sulfonates, sulfate salts, phosphate ester salts and lipopeptide anionic surfactants; preferably, the anionic surfactant is selected from sulfonates and lipopeptide anionic surfactants; more preferably, the anionic surfactant is selected from the group consisting of surfactin, sodium lauryl sulfonate and sodium lauryl benzene sulfonate. Preferably, the final concentration of the compound of Formula I′ in the contrast agent is in the range of 1˜100 μM, such as 5˜50 μM or 10˜30 μM. Preferably, the contrast agent comprises anionic surfactant, and the molar ratio of the compound of Formula I′ to the anionic surfactant in the contrast agent is 1:(0.1˜20), such as 1:(0.1˜10) or 1:(0.2˜5). Preferably, the contrast agent is a lyophilized powder or in the form of an aqueous solution.
The present disclosure also provides a detection kit comprising the compound of Formula I′ according to the present disclosure and an optional anionic surfactant. Preferably, the anionic surfactant is as described in any embodiment herein, preferably surfactin or sodium dodecyl sulfonate (SDS). In some embodiments, the detection kit comprises the contrast agent described in any embodiment herein.
The present disclosure also provides use of the compound of Formula I′ and the optional anionic surfactant as described in any embodiment herein in the manufacture of a tumor contrast agent.
The present disclosure also provides use of the compound of Formula I and the anionic surfactant as described in any embodiment herein in the manufacture of a tumor contrast agent.
The present disclosure also provides a tumor imaging method, which comprises a step of contacting a tissue to be examined with the contrast agent according to the present disclosure.
In one or more embodiments, the method is an in vitro method, said contacting comprises immersing, spraying and/or smearing the tissue to be examined with the contrast agent of the present disclosure.
In one or more embodiments, the method is an in vivo method, said contacting comprises administering the contrast agent to the patient to be examined.
The present disclosure also provides a contrast agent or a kit containing the contrast agent, wherein the contrast agent comprises Compound 4 and/or Compound 6 and anionic surfactant:
The present disclosure also provides a contrast agent or a kit containing the contrast agent, wherein the contrast agent comprises Compound 4 and/or Compound 6, and surfactin or sodium lauryl sulfonate:
Preferably, the contrast agent is in the form of an aqueous solution or a lyophilized powder.
It should be understood that within the scope of the present disclosure, the above technical features of the present disclosure and the technical features specifically described below (such as in examples) may be combined with each other, thereby constituting a preferred technical solution.
The theory or mechanism described and disclosed herein, whether right or wrong, should not limit the scope of the disclosure in any way, i.e., the content of the disclosure may be implemented without being limited by any particular theory or mechanism.
The range of values described herein shall be deemed to have covered and specifically disclosed all possible sub-ranges and any individual values within the range. For example, “comprising 1 to 20 carbon atoms” would include comprising 1 to 10 carbon atoms, comprising 2 to 10 carbon atoms, comprising 5 carbon atoms, etc.
Herein, when describing embodiments, or examples, it should be understood that it is not intended to limit the present disclosure to these embodiments or examples. In contrast, all substitutes, modifications and equivalents of the methods and materials described in the present disclosure may be covered within the scope of the claims. Those skilled in the art will be able to identify a number of methods and materials similar to or equivalent to those described herein that may be used to implement the present disclosure.
In the present disclosure, for the sake of conciseness, not all possible combinations of various technical features in various embodiments or examples are described. Thus, as long as there is no contradiction in the combination of these technical features, each technical feature of each embodiment or example may be arbitrarily combined, and all possible combinations should be considered to be within the scope of the present description.
In the present disclosure, halogens include F, Cl, Br, and I.
The term “alkyl” refers to a linear or branched monovalent saturated hydrocarbon group, typically containing 1-12 carbon atoms (C1-12 alkyl), preferably 1-6 carbon atoms (C1-6 alkyl). Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, and tert-butyl. Alkylene refers to divalent alkyls are such as —CH2—, —CH2CH2—, etc. The number of carbon atoms of alkylene group is usually 1-6.
The term “alkenyl” refers to a straight-chain or branched monovalent hydrocarbon group containing one or more double bonds, typically containing 2-12 carbon atoms (C2-12 alkenyl), preferably containing 2-6 carbon atoms (C2-6 alkenyl). Examples of alkenyl include, but are not limited to, vinyl, propenyl, allyl and 1,4-butadienyl.
The term “alkynyl” refers to a straight or branched monovalent hydrocarbon group containing one or more triple bonds, typically containing 2-12 carbon atoms (C2-12 alkynyl), preferably containing 2-6 carbon atoms (C2-6 alkynyl). Examples of alkynyl include, but are not limited to, ethynyl, 1-propynyl, 1-butynyl, 2-butynyl, and 1-methyl-2-butynyl.
The term “cyclic hydrocarbon group” or “carbon ring group” means a stable non-aromatic monocyclic or polycyclic hydrocarbon group composed only of carbon atoms and hydrogen atoms, which may include a fused ring system, a bridge ring system or a spiro ring system having 3 to 15 carbon atoms, preferably 3 to 10 carbon atoms, and which are saturated or unsaturated and may be connected to the rest of the molecule via any suitable carbon atom by a single bond. Unless otherwise specified in the specification, carbon atoms in cyclic hydrocarbon groups may optionally be oxidized. Examples of cyclic hydrocarbon groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexadienyl, cycloheptyl, cyclooctyl, 1H-indenyl, indanyl, 1,2,3,4-tetrahydro-naphthyl, 5,6,7,8-tetrahydro-naphthyl, 8,9-dihydro-7H-benzocyclohepten-6-yl, 6,7,8,9-tetrahydro-5H-benzocycloheptenyl, 5,6,7,8,9,10-hexahydro-benzocyclooctenyl, fluorenyl, dicyclo[2.2.1]heptyl, 7,7-dimethyl-dicyclo[2.2.1]heptyl, dicyclo[2.2.1]heptenyl, dicyclo[2.2.2]octyl, dicyclo[3.1.1]heptyl, dicyclo[3.2.1]octyl, dicyclo[2.2.2]octenyl, dicyclo[3.2.1]octenyl, adamantyl, octahydro-4,7-methylene-1H-indenyl and octahydro-2,5-methylene-cyclopentadienyl, etc. Exemplary cyclic hydrocarbon groups or carbon ring groups are cycloalkyl groups usually having a number of carbon atoms in the ring of 3-10, preferably 3-8. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and adamantyl. Exemplary cyclic hydrocarbon groups or carbon ring groups can be cycloalkenyl groups, i.e. monovalent hydrocarbon groups containing non-aromatic double-bonded hydrocarbon rings. The number of carbon atoms in the ring of the cycloalkenyl group is usually 3-10, such as 3-8. Examples of cycloalkenyl group include, but are not limited to, cyclopentenyl, cyclohexenyl, and cycloheptenyl.
The term “aryl” or “aromatic ring” refers to a monovalent hydrocarbon group containing an aromatic hydrocarbon ring and is usually C6-14 aryl. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, and anthracenyl.
The term “heteroaryl” or “heteroaromatic ring” refers to a group containing 5-14 ring atoms and 6, 10 or 14 electrons that are shared on the ring system, wherein the ring atoms contained are carbon atoms and 1-3 heteroatoms selected from the group consisting of oxygen, nitrogen, and sulfur. Examples of heteroaryl groups include, but are not limited to, thienyl, benzisothiazolyl, benzothienyl, naphtho[2,3-b]thienyl, thianthrenyl, furanyl, pyranyl, isobenzofuranyl, chromenyl, xanthenyl, pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, indolizinyl, isoindolyl, 3H-indolyl, indolyl, indazolyl, purinyl, 4H-quinazinyl, isoquinolinyl, quinolinyl, phthalazinyl, naphthyridinyl, quinazolinyl, cinnolinyl, pteridinyl, carbazolyl, O-carbolinyl, phenanthridinyl, acridinyl, perimidinyl, phenanthrolinyl, phenazinyl, isothiazolyl, phenothiazinyl and isoxazolyl, etc.
The term “heterocyclic group” or “hetero-ring” refers to a saturated or partially saturated 3-7-membered single ring or 7-10-membered bicyclic system consisting of carbon atoms and 1-3 heteroatoms selected from the group consisting of O, N, and S. Heterocyclic groups include, but are not limited to, tetrahydrofuranyl, pyrranyl, piperidinyl, piperazinyl, pyrrolidinyl, imidazolidyl, imidazolidinyl, dihydroindolyl, isodihydroindolyl, quinuclidinyl, morpholinyl, isochromanyl, chromanyl, pyrazolidinyl and pyrazolinyl and the like.
The groups described herein may be substituted. The number of substituents can be one or more, for example 1-4. Unless otherwise indicated, when a group is referred to herein as being “an optionally substituted”, its substituents typically include, but are not limited to, one or more (for example, 1, 2, 3 or 4) of halogen, C1-6 alkyl, halogenated C1-6 alkyl, C1-6 alkoxy, halogenated C1-6 alkoxy, C2-6 alkenyl, C2-6 alkynyl, C6-14 aryl, C5-14 heteroaryl, C3-10 heterocyclic group, C3-8 cycloalkyl, hydroxyl, nitro, cyano, mercapto, amino, C1-6 acyl. In a preferred embodiment, the substituent is one or more selected from the group consisting of halogen, hydroxyl, amino, C1-6 alkyl and C1-6 alkoxy.
In the present disclosure, “amino” includes —NH2 and —NR′R″, wherein R′ and R″ are each independently H, C1-4 alkyl and C1-6 alkoxycarbonyl; preferably, R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
The contrast agent of the present disclosure comprises one or more compounds represented by Formula I:
In some embodiments of the compound of Formula I, L is the bond, n1 and n2 are the same integers, and R21 and R22 are the same groups. Therefore, in these embodiments, the group —(CH2)n1-L-R22 is the same as the group —(CH2)n2-R21, for example, both of which are C1-6 alkyl groups that are optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH and —NH2. Preferably, the group —(CH2)n1-L-R22 is the same as the group —(CH2)n2-R21, and both are C1-6 alkyl groups that are optionally substituted by —COOH. More preferably, in these embodiments, R1—R8 are all hydrogen.
In some other embodiments of the compound of Formula I, the group —(CH2)n1-L-R22 is different from the group —(CH2)n2-R21, and the steric hindrance of the group —(CH2)n1-L-R22 is greater than that of the group —(CH2)n2-R21. For example, R22 is selected from the group consisting of an optionally substituted C3-15 cycloalkyl, an optionally substituted C3-15 cycloalkyl C1-6 alkyl, an optionally substituted 6-14-membered aryl group, an optionally substituted 6-14-membered arylalkyl group, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heteroaryl C1-6 alkyl, an optionally substituted 5-14-membered heterocyclic group and an optionally substituted 5-14-membered heterocyclic group C1-6 alkyl, while R21 is not selected from these groups, for example, R21 is H, halogen, nitro, cyano, —SO3, —COOH, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 acyl, —NH2 or C1-6 alkoxy, preferably H, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 acyl, —NH2 or C1-6 alkoxy, more preferably H or C1-6 alkyl. Preferably, in these embodiments, R1-R8 are all hydrogen. Preferably, in these embodiments, L is selected from the group consisting of *-T1-C(O)—, *—C(O)-T1-, *—S(O)2-T2- and *-T2-S(O)2—, wherein T1 is selected from the group consisting of NH, O and S, T2 is selected from the group consisting of O, NH and S, * indicates the location in connection with —(CH2)n1-. More preferably, L is selected from *-T1-C(O)— and *—C(O)-T1-, wherein T1 is selected from NH and O.
In some embodiments, the group —(CH2)n1-L-R22 is different from the group —(CH2)n2-R21, and the group —(CH2)n1-L-R22 is C1-6 alkyl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH, —NH2, preferably C1-6 alkyl that is optionally substituted by —COOH; the group —(CH2)n2-R21 is C1-6 alkyl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH, —NH2, preferably C1-6 alkyl that is optionally substituted by —COOH, and the carbon chain length of the group —(CH2)n2-R21 is shorter than the carbon chain length of the group —(CH2)n1-L-R22. Preferably, the group —(CH2)n1-L-R22 is C1-6 alkyl that is optionally substituted by —COOH, the group —(CH2)n2-R21 is C1-6 alkyl that is optionally substituted by —COOH, and the carbon chain length of the group —(CH2)n2-R21 is shorter than the carbon chain length of the group —(CH2)n1-L-R22. More preferably, the group —(CH2)n1-L-R22 is C1-6 alkyl that is substituted by —COOH, the group —(CH2)n2-R21 is an unsubstituted C1-6 alkyl, and the carbon chain length of the group —(CH2)n2-R21 is shorter than the carbon chain length of the group —(CH2)n-L-R22. Preferably, in these embodiments, L is a bond.
Preferably, the compound of Formula I does not comprise Compound 8.
Preferably, in the compound of Formula I, R1-R8 are each independently H, halogen or C1-C4 alkyl.
Preferably, in the compound of Formula I, L is selected from the group consisting of *-T1-C(O)—, *—C(O)-T1-, *—S(O)2-T2- and *-T2-S(O)2—, wherein T1 is selected from the group consisting of NH, O and S, T2 is selected from the group consisting of O, NH and S, * indicates the location in connection with —(CH2)n1-. More preferably, L is *-T1-C(O)— or *—C(O)-T1-, T1 is preferably NH or O.
Preferably, in Formula I, in R1-R8 as well as R21 and R22, said an optionally substituted C3-10 cycloalkyl, an optionally substituted 6-14-membered aryl, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heterocyclic group, an optionally substituted C3-15 cycloalkyl, an optionally substituted C3-15 cycloalkyl C1-6 alkyl, an optionally substituted 6-14-membered arylalkyl group, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heteroaryl C1-6 alkyl, an optionally substituted 5-14-membered heterocyclic group and an optionally substituted 5-14-membered heterocyclic group C1-6 alkyl are each an optionally substituted by 1-3 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, hydroxyl, carboxyl, amino (including —NH2 and —NR′R″), C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, and R″ is selected from H and C1-4 alkyl. For said C3-15 cycloalkyl, 6-14-membered aryl, 5-14-membered heteroaryl and 5-14-membered heterocyclic group in R21 and R22, further preferred substituent is —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
Preferably, in the compound of Formula I, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, —NH2, —NR′R″, C1-6 alkoxy and halogenated C1-6 alkoxy; more preferably C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by one —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
In some embodiments, the group —(CH2)n1-L-R22 is different from the group —(CH2)n2-R21, wherein R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, —NH2, —NR′R″, C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; R21 is H, halogen, C1-6 alkyl, halogenated C1-6 alkyl, C2-6 alkenyl or C2-6 alkynyl, and when R21 is H, n2 is not 0. Preferably, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; the group —(CH2)n2-R21 is C1-6 alkyl or halogenated C1-6 alkyl.
In one or more embodiments, R1-R8 are hydrogen; L is *-T1-C(O)— or *—C(O)-T1-, wherein T1 is selected from NH or O; n1 and n2 are each integer of 1 to 6; the group —(CH2)n2-R21 is C1-6 alkyl; R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
Preferably, in the compound of Formula I, the anion is selected from the group consisting of F−, Cl−, Br−, I−, NO3−, SO42−, PO43−, HPO42−, H2PO4−, CO32−, HCO3−, SO32−, HSO3−, CH3COO− and CH3SO3−.
In one or more embodiments, the compound of Formula I according to the present disclosure has the structure represented by the following formula Ia, Ib or Ic:
Preferably, the compound of Formula I in the contrast agent is one or more selected from the group consisting of
Preferably, the contrast agent of the present disclosure further comprises anionic surfactant. Suitable anionic surfactants include carboxylates, sulfonates, sulfate salts, phosphate ester salts and lipopeptide anionic surfactants. Preferable anionic surfactants include sulfonates and lipopeptide anionic surfactants, such as surfactin, sodium lauryl sulfonate and sodium lauryl benzene sulfonate, etc. In a particularly preferred embodiment, the anionic surfactant in the contrast agent of the present disclosure is surfactin (CAS No.: 24730-31-2). In the contrast agent of the present disclosure, the molar ratio of the compound of Formula I to the anionic surfactant in the contrast agent of the present disclosure is 1: (20˜0.1), such as 1: (10˜0.1) or 1:(5˜0.2).
In the contrast agent of the present disclosure, the final concentration of the compound of Formula I in the contrast agent is in the range of 1˜100 μM, such as 5˜50 μM or 10˜30 μM.
Therefore, in a particularly preferred embodiment, the contrast agent of the present disclosure comprises the compound of Formula I and surfactin, wherein the compound of Formula I is as described in any of the preceding embodiments. Preferably, in the contrast agent, R1-R8 are all hydrogen, L is a bond, the group —(CH2)n1-L-R22 is the same as the group —(CH2)n2-R21, for example, both of which are C1-6 alkyl groups that are optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, nitro, cyano, —SO3, —COOH and —NH2, preferably C1-6 alkyl groups that are optionally substituted by —COOH. Preferably, in the compound of Formula I of the contrast agent, R1-R8 are all hydrogen, L is *-T1-C(O)— or *—C(O)-T1-, wherein T1 is selected from NH or O; n1 and n2 are each integer of 1 to 6; R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substituents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, —NH2, —NR′R″, C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; R21 is H, halogen, C1-6 alkyl, halogenated C1-6 alkyl, C2-6 alkenyl or C2-6 alkynyl, and when R21 is H, n2 is not 0. Preferably, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; the group —(CH2)n2-R21 is C1-6 alkyl or halogenated C1-6 alkyl.
Preferably, in the compound of Formula I in the contrast agent, R1—R8 are hydrogen; L is *-T1-C(O)— or *—C(O)-T1-, wherein T1 is selected from NH or O; n1 and n2 are each integer of 1 to 6; the group —(CH2)n2-R21 is C1-6 alkyl; R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl. Preferably, in the contrast agent, the compound of Formula I is any one or more of Compounds 1-7, more preferably any one or more of Compounds 3-7, more preferably Compound 4 and/or Compound 6. Preferably, the molar ratio of the compound of Formula I to the surfactant in the contrast agent is 1:(10˜0.1), preferably 1:(5-0.2). Preferably, the final concentration of the compound of Formula I in the contrast agent is in the range of 5˜50 μM, preferably 10˜30 μM.
Preferably, the contrast agent of the present disclosure further comprises a solvent. The solvent should be able to dissolve the compound of Formula I of the present disclosure. Preferably, the solvent is water. In some embodiments, the contrast agent of the present disclosure is an aqueous solution of the compound of Formula I comprising the surfactant according to the present disclosure.
In a particularly preferred embodiment, the particle size of the contrast agent solution according to the present disclosure is 10-200 nm, more preferably 30-150 nm, more preferably 50-100 nm.
The contrast agent of the present disclosure may be in the form of a solution, or may be in the form of a lyophilized powder.
In some embodiments, the present disclosure also provides a detection kit comprising the compound of Formula I according to any embodiment of the present disclosure. The compound of Formula I according to the present disclosure in the kit may be provided as an aqueous solution or as a solid powder. Preferably, the kit also contains anionic surfactant according to any embodiment herein. The ingredients in the kit can be individually packaged or supplied as a mixture. When individually packaged, the compound of Formula I of the present disclosure and an optional surfactant may be mixed in water to form a contrast agent before use. In some embodiments, the detection kit of the present disclosure comprises a contrast agent of the compound of Formula I according to the present disclosure. Preferably, the contrast agent containing the compound of Formula I of the present disclosure is the contrast agent as described in any embodiment of the present disclosure. The kit may also contain instructions instructing the technician to use the ingredients in the kit to make a contrast agent and/or instructing the technician to use the contrast agent for tumor tissue angiography.
In some embodiments, the present disclosure provides the compound of Formula I′ as defined below:
Preferably, in Formula I′, R1—R8 are each independently hydrogen, halogen or C1-C4 alkyl, preferably are all H.
Preferably, in Formula I′, L is selected from the group consisting of *-T1-C(O)—, *—C(O)-T1-, *—S(O)2-T2- and *-T2-S(O)2—, wherein T1 is selected from the group consisting of NH, O and S, T2 is selected from the group consisting of O, NH and S, * indicates the location in connection with —(CH2)n1-. More preferably, L is selected from *-T1-C(O)— and *—C(O)-T1-, wherein T1 is selected from NH and O.
Preferably, in R1-R8 as well as R21 and R22 of Formula I′, said an optionally substituted C3-10 cycloalkyl, an optionally substituted 6-14-membered aryl, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heterocyclic group, an optionally substituted C3-15 cycloalkyl, an optionally substituted C3-15 cycloalkyl C1-6 alkyl, an optionally substituted 6-14-membered arylalkyl group, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heteroaryl C1-6 alkyl, an optionally substituted 5-14-membered heterocyclic group and an optionally substituted 5-14-membered heterocyclic group C1-6 alkyl are each an optionally substituted by 1-3 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, hydroxyl, carboxyl, amino (including —NH2 and —NR′R″), C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, and R″ is selected from H and C1-4 alkyl.
Preferably, in Formula I′, R22 is selected from the group consisting of an optionally substituted C3-15 cycloalkyl, an optionally substituted C3-15 cycloalkyl C1-6 alkyl, an optionally substituted 6-14-membered aryl group, an optionally substituted 6-14-membered arylalkyl group, an optionally substituted 5-14-membered heteroaryl group, an optionally substituted 5-14-membered heteroaryl C1-6 alkyl, an optionally substituted 5-14-membered heterocyclic group and an optionally substituted 5-14-membered heterocyclic group C1-6 alkyl. Preferably, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, amino, C1-6 alkoxy and halogenated C1-6 alkoxy.
Preferably, in Formula I′, R21 is H, halogen, nitro, cyano, —SO3, —COOH, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 acyl, —NH2 or C1-6 alkoxy, preferably H, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 acyl, —NH2 or C1-6 alkoxy, more preferably H or C1-6 alkyl.
Preferably, in Formula I′, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by 1 or 2 substitutents selected from the group consisting of halogen, C1-6 alkyl, halogenated C1-6 alkyl, —NH2, —NR′R″, C1-6 alkoxy and halogenated C1-6 alkoxy, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; R21 is H, halogen, C1-6 alkyl, halogenated C1-6 alkyl, C2-6 alkenyl or C2-6 alkynyl, and when R21 is H, n2 is not 0. Preferably, R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl; the group —(CH2)n2-R21 is C1-6 alkyl or halogenated C1-6 alkyl.
Preferably, in Formula I′, R1—R8 are hydrogen; L is *-T1-C(O)— or *—C(O)-T1-, wherein T1 is selected from the group consisting of NH, O and S; n1 and n2 are each integer of 1 to 6; the group —(CH2)n2-R21 is C1-6 alkyl; R22 is C6-14 aryl or C3-15 cycloalkyl C1-6 alkyl that is optionally substituted by —NR′R″, wherein R′ is selected from C1-4 alkyl and C1-6 alkoxycarbonyl, R″ is selected from H and C1-4 alkyl.
Preferably, in Formula I′, the anion is selected from the group consisting of F−, Cl−, Br−, I−, NO3−, SO42−, PO43−, HPO42−, H2PO4−, CO32−, HCO3−, SO32−, HSO3−, CH3COO− and CH3SO3.
In some embodiments, the compound of Formula I′ according to the present disclosure has the structure represented by the following formula Ia′, Ib′ or Ic′:
In one or more preceding embodiments of the compound according to the present disclosure, the compound of Formula I′ is selected from the group consisting of
The present disclosure also provides a method for preparing the compound of Formula I′, which comprises reacting the compound of Formula 1 with a compound Z—R22 to obtain the compound of Formula I′:
The reaction may be carried out at low temperatures (e.g. below 20° C.) and under the protection of inert gases. The compound of Formula 1 and Z—R22 may be dissolved in an organic solvent such as methylene chloride, tetrahydrofurand/or ethyl ether, and then slowly added with 1-equivalent of a salt of a condensing agent formed with an acid (such as 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide salt) and a catalyst (such as 4-dimethylaminopyridine) dissolved in the same organic solvent. The mixture is stirred at low speed. After the end of the reaction, dichloromethane is added, and the resultant is washed, dried, and purified when needed.
The contrast agent of the present disclosure may be used for tumor tissue angiography to distinguish tumor tissue from normal tissue. Unlike existing contrast agents, the contrast agent of the present disclosure can be used to treat tissues in vitro to develop tumor tissues. In some embodiments, the contrast agent of the present disclosure may also be used for in vivo administration.
Accordingly, the present disclosure provides a method for contrasting tumor tissues or cancer tissues in vitro, which comprises a step of contacting the tissues to be examined with a contrast agent or a compound of Formula I′ of the present disclosure. Preferably, said contrast agent is the contrast agent described in any embodiment of the present disclosure. Contacting comprises immersing, smearing and/or spraying the tissue to be examined with the contrast agent of the present disclosure, so that the imaging of the tumor lesion is completed. Generally, after the tumor tissue is exposed to a solution containing the contrast agent or the compound of Formula I′ of the present disclosure for a period of time, the tumor tissue or cancer tissue can be developed by imaging with a fluorescence imager.
The present disclosure also provides a use of a compound of Formula I and anionic surfactant in the preparation of a contrast agent for tumor tissues. The present disclosure also provides a use of the compound of Formula I′ of the present disclosure in the preparation of a contrast agent for tumor tissues.
As confirmed by the Examples of the present disclosure, the targeted recognition of tumor tissues by other compounds of the present disclosure is achieved by organic anion transporting polypeptides (OATP). Therefore, any tumor or cancer tissue (especially solid tumors) expressing organic anion transporting polypeptides can be contrasted with the contrast agent or the compound of Formula I′ and distinguished from the surrounding normal tissues. More particularly, the compounds of Formulas I and I′ of the present disclosure, the contrast agents thereof, and the methods of the present disclosure may be used to contrast tumor tissues or cancer tissues selected from the group consisting of liver cancer, melanoma, neuroblastoma, breast cancer, ovarian cancer, lung cancer (small cell lung cancer and non-small cell lung cancer), Wilms tumor, cervical cancer, testicular cancer, soft tissue sarcoma, bladder cancer, primary brain cancer, gastric cancer, colon cancer, malignant pancreatic insulinoma, malignant carcinoid cancer, choriocarcinoma, mycosis fungoides, head and neck cancer, osteogenic sarcoma, pancreatic cancer, rhabdomyosarcoma, Kaposi sarcoma, thyroid cancer, esophageal cancer, kidney cancer, endometrial cancer, adrenal cortical adenocarcinoma, skin cancer and prostate cancer, etc.
In some embodiments, the compounds, the contrast agents and the methods of the present disclosure may be particularly useful for (1) early screening of cavity tumors, such as oral cavity cancer, esophageal cancer, gastric cancer, bowel cancer, colorectal cancer, bladder cancer, etc.; (2) intraoperative imaging of all known solid tumors, such as breast cancer, kidney cancer, brain cancer, etc., to check whether the surgical resection is complete; (3) photodynamic therapy of tumor lesions (mainly cavity tumors, such as bladder cancer, colorectal cancer, etc.).
The present disclosure has the following advantages:
The present disclosure will be described below referring to the specific examples. It should be understood that these examples are merely illustrative and are not intended to limit the present disclosure. The methods and reagents used in the examples, unless otherwise stated, are methods and reagents conventional in the art.
The compound of Formula II may be prepared by the following steps:
At low temperature, under the protection of inert gas, Compound 3 and Compound 9 were dissolved in dichloromethane in an equivalent amount with stirring at low speed, and slowly added with 1 equivalent of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and a catalytic amount of a solution of 4-dimethylaminopyridine in dichloromethane within 2 hours. The system was stirred at low speed for 24 hours. After the end of the reaction as monitored by silica gel thin layer chromatography, dichloromethane was added. The resultant was washed with saline, and dried with anhydrous sodium sulfate overnight. Silica gel column chromatography (methanol/dichloromethane) was used for purification, and the final purified product was a green powdery solid.
Compound 4 having the structure shown below was prepared according to the above method.
The chemical structure of Compound 4 was characterized as follows: 1H NMR (500 MHz, CD3Cl, δ): 8.37-8.31 (m, 2H), 7.54-7.53 (m, 1H), 7.41-7.35 (m, 7H), 7.25-7.12 (m, 4H), 7.03-7.01 (m, 1H), 6.32-6.31 (m, 1H), 6.30-6.29 (m, 1H), 4.32-4.26 (m, 4H), 2.76-2.73 (m, 4H), 2.63-2.60 (m, 2H), 1.96-1.83 (m, 8H), 1.72-1.71 (m, 12H), 1.47 (t, J=7.0 Hz, 3H).
13C NMR (125 MHz, CD3Cl, δ): 172.1, 171.9, 150.6, 150.2, 144.5, 143.9, 129.4, 128.9, 127.6, 125.8, 125.3, 122.2, 121.5, 110.8, 101.5, 49.4, 49.2, 44.6, 39.9, 33.9, 31.9, 31.4, 30.2, 29.7, 29.4, 28.2, 28.0, 27.2, 26.6, 26.4, 24.5, 22.7, 20.7, 14.1, 12.5.
HRMS: m/z [M]+ calc for C44H50ClN2O2+: 673.3555; found: 673.3550.
Compound 5 having the structure shown below was prepared according to the above method.
The chemical structure of Compound 5 was characterized as follows:
1H NMR (500 MHz, CD3Cl, δ): 8.38-8.29 (m, 2H), 7.43-7.37 (m, 7H), 7.25-7.21 (m, 2H), 7.15-7.14 (m, 1H), 6.95-6.93 (m, 2H), 6.38-6.35 (m, 1H), 6.26-6.23 (m, 1H), 4.36-4.32 (m, 2H), 4.24-4.21 (m, 2H), 2.76-2.70 (m, 4H), 2.60-2.57 (m, 2H), 1.93-1.85 (m, 8H), 1.72-1.71 (m, 12H), 1.51 (s, 9H), 1.47 (t, J=7.0 Hz, 3H).
13C NMR (125 MHz, CD3Cl, δ): 172.4, 172.0, 171.6, 152.9, 150.2, 145.8, 144.8, 143.6, 136.4, 128.9, 128.0, 127.6, 125.6, 125.0, 122.2, 121.7, 110.8, 110.5, 101.9, 101.1, 49.5, 49.1, 44.4, 40.0, 33.8, 31.2, 29.7, 28.3, 28.1, 28.0, 27.1, 26.6, 26.3, 24.5, 22.7, 20.7, 14.4, 12.5.
HRMS: m/z [M]+ calc for C49H59ClN3O4+: 788.4189; found: 788.4185.
Compound 6 having the structure shown below was prepared according to the above method.
The chemical structure of Compound 6 was characterized as follows:
1H NMR (500 MHz, CD3Cl, δ): 10.69 (s, 1H), 8.42-8.39 (m, 1H), 8.28-8.24 (m, 1H), 7.91-7.89 (m, 2H), 7.43-7.35 (m, 5H), 7.31-7.29 (m, 1H), 7.24-7.17 (m, 3H), 7.05-7.03 (m, 1H), 6.43-6.40 (m, 1H), 5.99-5.96 (m, 1H), 4.28-4.25 (m, 2H), 4.07-4.02 (m, 2H), 2.75-2.73 (m, 2H), 2.71-2.68 (m, 2H), 2.62-2.59 (m, 2H), 1.90-1.82 (m, 8H), 1.71-1.70 (m, 12H), 1.50 (s, 9H), 1.43 (t, J=7.0 Hz, 3H).
13C NMR (125 MHz, CD3Cl, δ): 174.4, 172.7, 169.7, 150.7, 146.5, 142.5, 142.1, 141.7, 141.1, 140.7, 129.2, 128.7, 128.3, 127.1, 126.2, 126.1, 124.5, 122.3, 122.2, 120.6, 111.9, 109.6, 103.3, 98.8, 49.5, 48.8, 45.0, 38.9, 36.7, 31.2, 28.4, 28.1, 28.0, 26.9, 26.6, 26.4, 26.0, 25.2, 20.6, 12.4, 12.0.
HRMS: m/z [M]+ calc for C49H60ClN4O3+: 787.4348; found: 787.4346.
Compound 7 having the structure shown below was prepared according to the above method.
The chemical structure of Compound 7 was characterized as follows:
1H NMR (500 MHz, CD3Cl, δ): 10.69 (s, 1H), 8.42-8.39 (m, 1H), 8.28-8.24 (m, 1H), 7.91-7.89 (m, 2H), 7.43-7.35 (m, 5H), 7.31-7.29 (m, 1H), 7.24-7.17 (m, 3H), 7.05-7.03 (m, 1H), 6.43-6.40 (m, 1H), 5.99-5.96 (m, 1H), 4.28-4.25 (m, 2H), 4.07-4.02 (m, 2H), 2.75-2.73 (m, 2H), 2.71-2.68 (m, 2H), 2.62-2.59 (m, 2H), 1.90-1.82 (m, 8H), 1.71-1.70 (m, 12H), 1.50 (s, 9H), 1.43 (t, J=7.0 Hz, 3H).
13C NMR (125 MHz, CD3Cl, δ): 174.4, 172.7, 169.7, 150.7, 146.5, 142.5, 142.1, 141.7, 141.1, 140.7, 129.2, 128.7, 128.3, 127.1, 126.2, 126.1, 124.5, 122.3, 122.2, 120.6, 111.9, 109.6, 103.3, 98.8, 49.5, 48.8, 45.0, 38.9, 36.7, 31.2, 28.4, 28.1, 28.0, 26.9, 26.6, 26.4, 26.0, 25.2, 20.6, 12.4, 12.0.
HRMS: m/z [M]+ calc for C49H60ClN4O3+: 787.4348; found: 787.4346.
Compounds 1, 2, 3 and 8 used in the examples have the following structures, respectively:
Compound 1 is reported in a reference, Near-infrared fluorescence imaging of cancer mediated by tumor hypoxia and HIF1Ialpha/OATPs signaling axis. Biomaterials 2014, 35, 8175-8185;
Compound 2 is reported in a reference, A self-assembled “albumin-conjugate” nanoprobe for near infrared optical imaging of subcutaneous and metastatic tumors. ACS Appl. Bio Mater. 2020, 3, 327-334;
Compounds 3 and 8 are reported in a reference, A novel “mosaic-type” nanoparticle for selective drug release targeting hypoxic cancer cells. Nanoscale 2019, 11, 2211-2222.
786o cells (3×105/well) were seeded in a 6-well plate in advance and placed in a cell culture incubator for 48 h. After the culture medium was blotted, a culture medium comprising the corresponding compound (5 μM) and Mito-tracker (0.2 nM) was added to the experimental well, incubated for 30 min, cleaned three times with PBS solution, and imaged using a fluorescence microscope equipped with an infrared camera and a continuous LED light source. The near-infrared photos were taken under Ex=745 nm, Em=820 nm; the Mito-tracker photos were taken under Ex=488 nm and Em=520 nm.
The experimental results for Compounds 1-3, 4, 5, and 7 are shown in
The results showed that Compounds 1-3, 4, 5 and 7 could be taken up by tumor cells, and their intracellular distribution could basically be colocalized with the Mito-tracker distribution, which could label the mitochondria of cells, so this experiment proved at the cellular level that such compounds could enter tumor cells and be enriched in mitochondria.
The same method as Example 2 was employed except that the human renal cancer cell line A498 was used to detect the localization of Compounds 4, 5 within the cell.
The experimental results for Compounds 4 and 5 are shown in
Surfactin in different concentrations was added to the aqueous solution of the corresponding compound (final concentration of 20 μM), and the molar ratio of the compound to be tested to surfactin was 1:5, 1:1 and 1:0.2. A methanol solution of the corresponding compound (final concentration of 20 μM) was used as a positive control. The test solution was shaken for 5 min and added to a black 96-well plate with 50 ul of test solution per experimental well. Then a fluorescence imager was used for imaging. The photos were taken under Ex=745 nm, Em=820 nm. The ROI (Region of Interest) value for each well was quantitatively calculated.
The results are shown in
Surfactin in different concentrations was added to the aqueous solution of the corresponding compound (final concentration of 20 μM), and the molar ratio of the compound to be tested to surfactin was 1:5, 1:1, 1:0.2. A methanol solution of the corresponding compound (final concentration of 20 μM) was used as a positive control. The test solution was shaken for 5 minutes and its fluorescence emission spectra (Ex=730 nm, Em=760-960 nm) were detected using a fluorescence spectrophotometer.
The experimental results of Compounds 1-8 are shown in
The results showed that compared with the methanol solution, obvious fluorescence quenching of Compound 1 in the aqueous solution could occur under the action of surfactin (the molar ratio of Compound 1 to surfactin was 1:5); when the amount of surfactin was reduced, the fluorescence quenching effect was significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 2 in the aqueous solution could occur under the action of surfactin (the molar ratio of Compound 2 to surfactant was 1:5); when the amount of surfactin was reduced, the fluorescence quenching effect was significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 3 in the aqueous solution could occur under the action of surfactin (the molar ratio of Compound 3 to surfactant was 1:5); when the amount of surfactin was reduced, the fluorescence quenching effect was significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 4 in the aqueous solution could occur under the action of surfactin; when the amount of surfactin was reduced, the fluorescence quenching effect was not significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 5 in the aqueous solution could occur under the action of surfactin; when the amount of surfactin was reduced, the fluorescence quenching effect was not significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 6 in the aqueous solution could occur under the action of surfactin; when the amount of surfactin was reduced, the fluorescence quenching effect was not significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 7 in the aqueous solution could occur under the action of surfactin; when the amount of surfactin was reduced, the fluorescence quenching effect was not significantly reduced. Compared with the methanol solution, obvious fluorescence quenching of Compound 8 in the aqueous solution could occur under the action of surfactin; when the amount of surfactin was reduced, the fluorescence quenching effect was not significantly reduced.
Different surfactants were added to the aqueous solution of Compound 4 (final concentration of 20M), and the molar ratio of the compound to be tested to surfactin was 1:1. In addition, a methanol solution of the compound (final concentration of 20 μM) was used as a positive control. The test solution was shaken for 5 minutes and its fluorescence emission spectra (Ex=730 nm, Em=760-960 nm) were detected using a fluorescence spectrophotometer.
The experimental results are shown in
The results showed that human serum albumin, nonionic surfactants (Kolliphor, P123) and cationic surfactants (dodecyltrimethylammonium chloride, tetrabutylammonium iodide) mixed with Compound 4 in the aqueous solution could reduce their fluorescence signal intensity, but there was still fluorescence signal in the solution. A fluorescence quenching effect occurred after anionic surfactants (surfactin, sodium dodecyl sulfonate, sodium dodecyl benzene sulfonate) were mixed with Compound 4 in the aqueous solution, and there was substantially no fluorescence signal in the solution.
Surfactin (final concentration of 100 μM) was added to the aqueous solution of the corresponding compound (final concentration of 20 μM), and the molar ratio of the compound to be tested to surfactin was 1:5. The test solution was shaken for 5 minutes and its micromorphology was detected using a Transmission Electron Microscope (TEM).
The experimental results are shown in
Female nude mice around 5 weeks of age were ordered and subjected to subcutaneous renal clear cell cancer cell 786o inoculation after one week of barrier environmental adaptation at a specific position above the right lower limb (3×106/pce, suspended in 200 μl PBS solution). When the tumor volume reached 50-150 mm3 (about 12-22 days), nude mice were randomly grouped with 3 mice in each experimental group. Mice were dissected, and tumor tissues were taken.
Compounds 1-8 were added to the aqueous solution of surfactin. The final concentration of surfactin was 100 μM, and the final concentration of each compound was 20 μM (the molar ratio of the compound to surfactant was 1:5).
The tumor tissue was immersed in the above liquid. A fluorescence imager was used for imaging after a specified time.
The experimental results of Compounds 1-8 are shown in
Using the method described in Example 8, a mouse subcutaneous xenograft breast cancer tumor model was constructed, and then the mouse model was used to detect the near-infrared imaging of Compounds 4,6 (in aqueous solution, the molar ratio of the compound to surfactin is 1:5) in the tumor tissue.
The experimental results for Compounds 4 and 6 are shown in
Using the method described in Example 8, a mouse subcutaneous xenograft renal clear cell carcinoma tumor model was constructed, and then the mouse model was used to detect the near-infrared imaging of Compound 4 (in aqueous solution, the molar ratio of the compound to surfactin was 1:5 with a final concentration of 1 mM of sodium bromosulfophthalein (50 equivalent) added) in the tumor tissue.
The experimental results are shown in
Establishment of the mouse primary bowel cancer tumor model of APCmin/+ mouse model: parent C57BL/6J-APC min/+ mice and C57BL/6J wt mice were caged and propagated. The genotype of the offspring was detected and the offspring C57BL/6J-APC min/+ mice (diseased mice) were screened. The offspring C57BL/6J-APC min/+ mice were killed at week 20. Mice were dissected and diseased colon tissue was taken. Colon was cut longitudinally for subsequent tests.
Compounds 1-8 were added to the aqueous solution of surfactin. The final concentration of surfactin was 100 μM, and the final concentration of the compound was 20 μM (the molar ratio of the compound to surfactin was 1:5).
The diseased colon tissue of mice was immersed in the above fluid. A fluorescence imager was used for imaging after 12 min.
The experimental results of Compounds 1-8 are shown in
The experimental results showed that, after short-term immersion contact, the mixtures of Compounds 1, 2 and 3 and surfactin as anionic surfactant (the molar ratio of the compound to surfactin was 1:5) had a recognition effect on tumor lesions in mouse primary bowel cancer tumor models. The mixtures of Compounds 4, 5, 6 and surfactant as anionic surfactant (the molar ratio of the compound to surfactant was 1:5) have a high specific recognition on tumor lesions in mouse primary bowel cancer tumor models. The tumor lesion tissues had significantly higher fluorescence intensity than that of normal intestinal tissues, with excellent signal-to-noise ratio and clearly distinguishable imaging results. Compound 4 and Compound 6 were best imaged. The mixture of Compound 7 and surfactin as anionic surfactant (the molar ratio of the compound to surfactin was 1:5) had a recognition effect on tumor lesion tissues in mouse primary bowel cancer tumor models. The mixture of Compound 8 and surfactin as anionic surfactant (the molar ratio of the compound to surfactin was 1:5) had substantially no recognition effect on tumor lesion tissues in mouse primary bowel cancer tumor models. Both of tumor lesion tissues and normal intestinal tissues had relatively weak fluorescence signals. The signal-to-noise ratio of the imaging results was very poor with no obvious imaging effect.
| Number | Date | Country | Kind |
|---|---|---|---|
| 202010885318.8 | Aug 2020 | CN | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/CN2021/114672 | 8/26/2021 | WO |
