Patent Grant
10464955
The present invention relates to the preparation of novel charged phosphinate linkers used for the conjugation of a drug, in particular, a cytotoxic agent to a biological molecule. The present invention also relates to methods of making cell-binding agent-drug (cytotoxic agent) conjugates comprising either modification of drugs with these charged linkers first, followed by reaction with cell-binding agents; or modification of cell-binding agents with these charged linkers first, followed by reaction with drugs.
Targeted drug delivery (Muller, R; Keck, C (2004) J. Biotech. 113, 151) whose objective is to prolong, localize, target and have a protected drug interaction with the diseased tissue has been extensively studied during the past three decades. There are different types of drug delivery vehicles, such as, antibodies, proteins, vitamins, peptides, polymeric micelles, liposomes, lipoprotein-based drug carriers, nano-particle drug carriers, dendrimers etc. An ideal drug delivery vehicle must be non-toxic, biocompatible, non-immunogenic, and biodegradable (Scott, R; Crabbe, D; et al (2008) Expert Opin. Drug Deli. 5, 459) and avoid recognition by the host's defense mechanisms (Saltzman, W.; Torchilin, V. (2008). “Drug delivery systems” Access Science. McGraw-Hill Co.). The link between the delivery vehicles, in particular, antibodies and the cell-killing agent plays a critical role in the development of targeted drug delivery systems, as the nature of the linker significantly affects the potency, selectivity and the pharmacokinetics of the resulting conjugates (Zhao, R.; Wilhelm, S. et al, (2011) J. Med. Chem. 36, 5404; Doronina, S.; Mendelsohn, B.; et al, (2006) Bioconjug Chem, 17, 114; Hamann, P.; Hinman, L; et al. (2005) Bioconjug Chem. 16, 346). Four types of linkers had been used for preparation of cell binding agent-drug conjugates that have entered the clinic: (a) acid-labile linkers, exploiting the acidic endosomal and lysosomal intracellular microenvironment; (b) linkers cleavable by lysosomal proteases; (c) chemically stable thioether linkers that release a lysyl adduct after proteolytic degradation of the antibody inside the cell; and (d) disulfide-containing linkers, which are cleaved upon exposure to an intracellular thiol ((Zhao, R.; Wilhelm, S. et al, (2011) J. Med. Chem. 36, 5404).
Conjugates of cell-binding agents with drugs or modified chemical compounds via different types of linkers have been described (U.S. Pat. Nos. 4,680,338, 5,122,368, 5,141,648, 5,208,020, 5,416,064; 5,475,092, 5,543,390, 5,563,250 5,585,499, 5,880,270, 6,214,345, 6,436,931, 6,372,738, 6,340,701, 6,989,452, 7,129,261, 7,375,078, 7,498,302, 7,507,420, 7,691,962, 7,910,594, 7,968,586, 7,989,434, 7,994,135, 7,999,083, 8,153,768, 8,236,319, Zhao, R.; et al, (2011) J. Med. Chem. 36, 5404; Doronina, S.; et al, (2006) Bioconjug Chem, 17, 114; Hamann, P.; et al. (2005) Bioconjug Chem. 16, 346). Typically, in these conjugates, the cell-binding agents are first modified with a bifunctional agent such as SPDP (N-succinimidyl 3-(2-pyridyldithio) propionate), or SMCC (succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate); or SPDB (N-succinimidyl 4-(2-pyridyldithio)butanoate); to introduce an active disulfide or a maleimido moiety. Reaction with a thiol-containing cytotoxic drug provides a conjugate in which the cell-binding agent, such as a monoclonal antibody, and drug are linked via disulfide bonds or thioether bonds.
However, the use of the cell binding molecule-drug conjugates, such as antibody-drug conjugates (ADCs), in developing therapies for a wide variety of cancers has been limited both by the availability of specific targeting agents (carriers) as well as the conjugation methodologies which result in the formation of protein aggregates when the amount of the drugs that are conjugated to the carrier (i.e., the drug loading) is increased. Normally the tendency for cytotoxic drug conjugates to aggregate is especially problematic when the conjugation reactions are performed with the hydrophobic linkers. Since higher drug loading increases the inherent potency of the conjugate, it is desirable to have as much drug loaded on the carrier as is consistent with retaining the affinity of the carrier protein. The presence of aggregated protein, which may be nonspecifically toxic and immunogenic, and therefore must be removed for therapeutic applications, makes the scale-up process for the production of these conjugates more difficult and decreases the yield of the products.
Consequently, there is a critical need to improve methods for conjugating drugs/cytotoxic drugs to carriers (cell binding molecules) that minimize the amount of aggregation and thereby allow for as high a drug loading as possible through the application of a charged crosslinker.
The present invention provides charged linkers containing phosphinate, group to link drugs to a cell-binding agent (e.g., an antibody). The preferred formula of the cell binding molecule-charged linker-drug conjugates can be represented as: Cb-(-L-Drug)n, wherein Cb is a cell-binding agent, L is a charged linker, Drug is a drug molecule, and n is an integer from 1 to 20. The advantages in applying the charged linker in the cell molecule-drug conjugate are: a). reducing the aggregation of the conjugates in water based media; b). enabling higher drug-per-cell binding molecule-ratio conjugate, resulting in higher potency; c). being retained inside the target cell after the drug-linker released from the conjugates, which can combat permeability-glycoprotein (Pgp)-expressing multidrug resistant (MDR) cells.
In one aspect of the present invention, the charged linker is represented by formula (I) wherein Y can react with a cell-binding agent and Z can react with a cytotoxic drug:
wherein:
Y represents a functional group that enables reaction with a cell-binding agent;
Z represents a functional group that enables linkage of a cytotoxic drug via a disulfide, thioether, thioester, peptide, hydrazone, ether, ester, carbamate, carbonate, amine (secondary, tertiary, or quartary), inline, cycloheteroalkyane, heteroaromatic, alkoxime or amide bond;
R1, R2, R3, R4, and R5, are the same or different and are H, linear alkyl having from 1-6 carbon atoms, branched or cyclic alkyl having from 3 to 6 carbon atoms, linear, branched or cyclic alkenyl or alkynyl, or 1-6 carbon atoms of esters, ether, amide, or polyethyleneoxy unit of formula (OCH2CH2)p, wherein p is an integer from 0 to about 1000, or combination thereof.
Additionally R1, R7, and R3 are respectively a chain of atoms selected from C, N, O, S, Si, and P that covalently connects the cell-surface binding ligand, the phosphinate group, the conjugated drug and among themselves (R1, R2 and R3). The atoms used in forming the hydrophilic linker may be combined in all chemically relevant ways, such as forming alkylene, alkenylene, and alkynylene, ethers, polyoxyalkylene, esters, amines, imines, polyamines, hydrazines, hydrazones, amides, ureas, semicarbazides, carbazides, alkoxyamines, alkoxylamines, urethanes, amino acids, peptides, acyloxylamines, hydroxamic acids, or combination thereof.
M is H, or Na, or K, or N+R1R2R3 or a pharmaceutical salt. R1. R2 and R3 are described above.
In another aspect, this invention provides a cell-binding agent-drug conjugate of formula (II), in which the cell-binding agent, Cb, and the drug, Drug, have reacted at the two ends of the charged linker:
wherein:
Cb represents a cell-binding agent;
Drug represents the drug linked to the cell-binding agent via the charged linker by a disulfide, thioether, thioester, peptide, hydrazone, ether, ester, carbamate, carbonate, cycloheteroalkyane, heteroaromatic, alkoxime or amide bond;
R1, R2, R3, R4, R5, and M are described the same previously in formula (I).
In a further aspect, the present invention provides a modified cell-binding agent of formula (III), in which the cell-binding agent, Cb, has reacted with the charged linker, which still has Z, a group capable of reacting with a drug:
Wherein the substituents are as defined above.
In an even further aspect, the present invention provides a modified drug of formula (IV), in which the drug, Drug, has reacted with the charged linker, which still has Y, a group capable of reacting with the cell-binding agent:
Wherein the substituents are as defined above.
The present invention further relates to a method of making a cell-binding molecule-drug conjugate of formula (II), wherein the drug is linked to a cell-binding agent via the charged linker.
The present invention also relates to a method of making a modified cell-binding molecule of formula (III), wherein the cell-binding molecule is reacted with the charged linker.
The present invention also relates to a method of making a modified drug of formula (IV), wherein the drug is reacted with the charged linker.
Definitions
“Alkyl” means an aliphatic hydrocarbon group which may be straight or branched having 1 to 8 carbon atoms in the chain or cyclic. “Branched” means that one or more lower alkyl groups such as methyl, ethyl or propyl are attached to a linear alkyl chain. Exemplary alkyl groups include methyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, n-pentyl, 3-pentyl, octyl, nonyl, decyl, cyclopentyl, cyclohexyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl, 3,3-dimethylpentyl, 2,3,4-trimethylpentyl, 3-methylhexyl, 2,2-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 3,5-dimethylhexyl, 2,4-dimethylpentyl, 2-methylheptyl, 3-methylheptyl, n-heptyl, isoheptyl, n-octyl, and isooctyl. A C1-C8 alkyl group can be unsubstituted or substituted with one or more groups including, but not limited to, —C1˜C8 alkyl, —O—(C1˜C8 alkyl), -aryl, —C(O)R, —OC(O)R′, —C(O)OR′, —C(O)NH2, —C(O)NHR′, —C(O)N(R)2—NHC(O)R′, —S(O)2R′, —S(O)R′, —OH, -halogen (F, Cl, Br or I), —N3, —NH2, —NH(R′), —N(R′)2 and —CN; where each R′ is independently selected from —C1˜C8 alkyl and aryl.
A “C3˜C8 carbocycle” means a 3-, 4-, 5-, 6-, 7- or 8-membered saturated or unsaturated non-aromatic carbocyclic ring. Representative C3˜C8 carbocycles include, but are not limited to, -cyclopropyl, -cyclobutyl, -cyclopentyl, -cyclopentadienyl, -cyclohexyl, -cyclohexenyl, -1,3-cyclohexadienyl, -1,4-cyclohexadienyl, -cycloheptyl, -1,3-cycloheptadienyl, -1,3,5-cycloheptatrienyl, -cyclooctyl, and -cyclooctadienyl. A C3-C8 carbocycle group can be unsubstituted or substituted with one or more groups including, but not limited to, —C1˜C8 alkyl, —O—(C1˜C8 alkyl), -aryl, —C(O)R′, —OC(O)R, —C(O)OR′, —C(O)NH2, —C(O)NHR′)2—C(O)N(R′)2—NHC(O)R′, —S(O) 2R, —S(O)R′, —OH, -halogen, —N3, —NH2, —NH(R′), —N(R′)2 and —CN; where each R′ is independently selected from —C1˜C8 alkyl and aryl.
A “C3˜C8 carbocyclo” refers to a C3-C8 carbocycle group defined above wherein one of hydrogen atoms on the carbocycle is replaced with a bond.
“Heterocycle” refers to an aromatic or non-aromatic C3˜C14 carbocycle in which one to four of the ring carbon atoms are independently replaced with a heteroatom from the group of O, N, S Se, and P. Preferable heteroatoms are oxygen, nitrogen and sulphur. Suitable heterocyclics are also disclosed in The Handbook of Chemistry and Physics, 76th Edition, CRC Press, Inc., 1995-1996, p. 2-25 to 2-26, the disclosure of which is hereby incorporated by reference.
Preferred non aromatic heterocyclic include, but are not limited to pyrrolidinyl, pyrazolidinyl, imidazolidinyl, oxiranyl, tetrahydrofuranyl, dioxolanyl, tetrahydro-pyranyl, dioxanyl, dioxolanyl, piperidyl, piperazinyl, morpholinyl, pyranyl, imidazolinyl, pyrrolinyl, pyrazolinyl, thiazolidinyl, tetrahydrothiopyranyl, dithianyl, thiomorpholinyl, dihydro-pyranyl, tetrahydropyranyl, dihydropyranyl, tetrahydro-pyridyl, dihydropyridyl, tetrahydropyrinidinyl, dihydrothiopyranyl, azepanyl, as well as the fused systems resulting from the condensation with a phenyl group.
“Alkyl”, “cycloalkyl”, “alkenyl”, “alkynyl”, “aryl”, “heteroaryl”, “heterocyclic” and the like refer also to the corresponding “alkylene”, “cycloalkylene”, “alkenylene”, “alkynylene”, “arylene”, “heteroarylene”, “heterocyclene” and the likes which are formed by the removal of two hydrogen atoms.
“Halogen atom” refers to fluorine, chlorine, bromine or iodine atom; preferably bromine and chlorine atom.
“Pharmaceutically” or “pharmaceutically acceptable” refer to molecular entities and compositions that do not produce an adverse, allergic or other untoward reaction when administered to an animal, or a human, as appropriate.
“Pharmaceutically acceptable excipient” includes any carriers, diluents, adjuvants, or vehicles, such as preserving or antioxidant agents, fillers, disintegrating agents, wetting agents, emulsifying agents, suspending agents, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. The use of such media and agents for pharmaceutical active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active ingredient, its use in the therapeutic compositions is contemplated. Supplementary active ingredients can also be incorporated into the compositions as suitable therapeutic combinations.
As used herein, “pharmaceutical salts” refer to derivatives of the disclosed compounds wherein the parent compound is modified by making acid or base salts thereof. The pharmaceutically acceptable salts include the conventional non-toxic salts or the quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric and the like; and the salts prepared from organic acids such as acetic, propionic, succinic, tartaric, citric, methanesulfonic, benzenesulfonic, glucoronic, glutamic, benzoic, salicylic, toluenesulfonic, oxalic, fumaric, maleic, lactic and the like. Further addition salts include ammonium salts such as tromethamine, meglumine, epolamine, etc., metal salts such as sodium, potassium, calcium, zinc or magnesium.
The pharmaceutical salts of the present invention can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acidic or basic forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two. Generally, non-aqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418, the disclosure of which is hereby incorporated by reference.
The novel conjugates disclosed herein use hydrophilic phosphinate cross-linkers. Examples of some suitable cross-linkers and their synthesis are shown in
The Charged Linkers
The synthetic routes to produce phosphinate-containing charged crosslinkers as well as the preparation of the antibody-drug conjugates of the present invention are shown in
Preferably, the charged linkers are compounds of the formula (I) below:
wherein:
Y represents a functional group that enables reaction with a cell-binding agent;
Z represents a functional group that enables linkage of a cytotoxic drug via a disulfide, thioether, thioester, peptide, hydrazone, ether, ester, carbamate, carbonate, amine (secondary, tertiary, or quartary), inline, oximine, cycloheteroalkyane, heteroaromatic or amide bond;
M is H, or Na, or K, or N+R1R2R3 or a pharmaceutical salt.
R1, R2, R3, R4, and R5, are the same or different and are H, linear alkyl having from 1-6 carbon atoms, branched or cyclic alkyl having from 3 to 6 carbon atoms, linear, branched or cyclic alkenyl or alkynyl or polyethyleneoxy unit of formula (OCH2CH2)p, wherein p is an integer from 0 to about 1000.
In another embodiment, R1, R2, and R3 can be respectively a chain of atoms selected from C, N, O, S, Si, and P that covalently connects the cell-surface binding ligand, the phosphinate group, the conjugated drug and themselves (R1, R2 and R3). The atoms used in forming the hydrophilic linker may be combined in all chemically relevant ways, such as forming alkylene, alkynylene, and alkynylene, ethers, polyoxyalkylene, esters, amines, imines, polyamines, hydrazines, hydrazones, amides, ureas, semicarbazides, carbazides, alkoxyamines, alkoxylamines, urethanes, amino acids, acyloxylamines, hydroxamic acids, and many others. In addition, it is to be understood that the atoms forming the linker (L) may be either saturated or unsaturated, or may be radicals, or may be cyclized upon each other to form divalent cyclic structures, including cyclo alkanes, cyclic ethers, cyclic amines, arylenes, heteroarylenes, and the like in the linker.
Examples of the functional group, Y, that enables reaction with a cell-binding agent include amine reacting agents such as but not limited to N-hydroxysuccinmide esters, p-nitrophenyl esters, dinitrophenyl esters, pentafluorophenyl esters; thiol reactive agents such as but not limited to pyridyldisulfides, nitropyridyldisulfides, maleimides, haloacetates and carboxylic acid chlorides.
Examples of the functional group, Z, which enables linkage of a cytotoxic drug, include groups that enable linkage via a disulfide, thioether, thioester, peptide, hydrazone, ester, carbamate, carbanate, or amide bond. Such functional groups include, but are not limited to, thiol, disulfide, amino, carboxy, aldehydes, maleimido, haloacetyl, hydrazines, and hydroxy.
In preferred embodiments, R1, R2, and R3, are linear alkyl having from 1-6 carbon atoms, or polyethyleneoxy unit of formula (OCH2CH2)p, p=1-100.
The synthesis of 2-dithio-pyridyl containing cross-linkers of formulae (I) is shown, for example, in
Cell-Binding Agent Drug-Conjugates
The conjugates of the present invention can be represented by the following formula, Cb-(-L-Drug)n, wherein Cb is a cell-binding agent, L is a charged phosphinate linker, Drug is a drug molecule, and n is an integer from 1 to 20.
The charged phosphinate linker L may be composed of one or more linker components. Exemplary linker components include 6-maleimidocaproyl (“MC”), maleimidopropanoyl (“MP”), valine-citrulline (“val-cit” or “vc”), alanine-phenylalanine (“ala-phe” or “af”), p-aminobenzyloxycarbonyl (“PAB”), 4-thiopentanoate (“SPP”), 4-(N-maleimidomethyl)cyclohexane-1 carboxylate (“MCC”), (4-acetyl)aminobenzoate) (“SIAB”), 4-thio-butyrate (SPDB), 4-thio-2-hydroxysulfonyl-butyrate (2-Sulfo-SPDB), ethyleneoxy —CH2CH2O— as one or more repeating units (“EO” or “PEO”). Additional linker components are known in the art and some are described herein.
Example structures of these components containing linkers are:
(MC, 6-maleimidocaproyl containing)
(MP, maleimidopropanoyl containing)
(PAB, p-aminobenzyloxycarbonyl containing)
(valine-citrulline containing)
(MCC, 4-(N-maleimidomethyl)cyclohexane-carboxylate)
((4-acetyl)aminobenzoate containing)
(4-thio-2-hydroxysulfonyl-butyrate, 2-sulfo-SPDB) Preferably, the conjugates have the following formula (II):
wherein:
Cb represents a cell-binding agent;
Drug represents the drug linked to the cell-binding agent via the hydrophilic linkers of this invention by a disulfide, thioether, thioester, peptide, hydrazone, ether, ester, carbamate, carbonate, heterocyclic ring, amine, imine, alkoxime or amide bond;
R1, R2, R3, R4, R5, and M are described the same previously in formula (I).
As described in more detail below, the drug can be any of many small molecule drugs, including, but not limited to, tubulysins, calicheamicins, auristatins, maytansinoids, CC-1065 analogs, morpholinos doxorubicins, taxanes, cryptophycins, epothilones, and benzodiazepine dimers (e.g., dimmers of pyrrolobenzodiazepine (PBD) or tomaymycin), indolinobenzodiazepines, imidazobenzothiadiazepines, or oxazolidinobenzodiazepines).
To synthesize the conjugate, the cell-binding agent can be first modified with the crosslinkers of the present invention to introduce reactive disulfide groups, maleimido, haloacetyl or hydrazide groups. Synthesis of the cell-binding agent-drug conjugates linked via disulfide bonds is achieved by a disulfide exchange between the disulfide bond in the modified cell-binding agent and a drug containing a free thiol group. Synthesis of the cell-binding agent-drug conjugates linked via thioether is achieved by reaction of the maleimido or haloacetyl modified cell-binding agent and a drug containing a free thiol group. Synthesis of conjugates bearing an acid labile hydrazone link can be achieved by reaction of a carbonyl group with the hydrazide moiety in the linker, by methods known in the art (see, for example, P. Hamann et al., Hinman, L. M., et al, Cancer Res. 53, 3336-334, 1993; B. Laguzza et al., J. Med. Chem., 32; 548-555, 1959; P. Trail et al., Cancer Res., 57; 100-105, 1997),
Alternatively, the drug can be modified with the charged crosslinkers of the present invention to give a modified drug of formula (IV) bearing a functionality capable of reacting with a cell binding agent. For example a thiol-containing drug can be reacted with the charged crosslinker of formula (I) bearing a maleimdo substituent at neutral pH in aqueous buffer to give a drug connected to the charged linker via a thioether link. A thiol-containing drug can undergo disulfide exchange with a charged linker bearing a pyrdiyldithio moiety to give a modified drug attached via a disulfide bond to the charged crosslinker. A drug bearing a hydroxyl group or a thiol group can be reacted with a hydrophilic crosslinker bearing a halogen of this invention, in the presence of a mild base, to give a modified drug bearing an ether or thiol ether link. A hydroxyl group containing drug can be condensed with a charged crosslinker of formula (I) bearing a carboxyl group, in the presence of a dehydrating agent, such as EDC or dicyclohexylcarbodimide, to give an ester link. An amino group containing drug can similarly undergo condensation with a carboxyl group on the charged crosslinker of formula (I) to give an amide bond.
The conjugate may be purified by standard biochemical means, such as gel filtration on a Sephadex G25 or Sephacryl S300 column, adsorption chromatography, and ion exchange or by dialysis. In some cases (e.g. folic acid, melanocyte stimulating hormone, EGF etc) the cell-binding agent-drug conjugates can be purified by chromatography such as by HPLC, medium pressure column chromatography or ion exchange chromatography.
Modified Cell-Binding Agents
The cell-binding agent modified by reaction with crosslinkers of the present invention are preferably represented by the formula (III)
wherein the substituents are as described above for the charged linker and the cell-binding agent drug conjugate.
In preferred embodiments, Z is a disulfide substituent, a maleimido, haloacetyl group, or a N-hydroxy succinimide ester, and Cb linked with R1 is through thioether, amide, or disulfide bond. The modified cell-binding agent can be prepared by reacting the cell-binding agent with the charged crosslinkers by methods known in the art for other crosslinkers (U.S. Pat. Nos. 5,846,545, 5,585,499, 5,475,092, 5,414,064, 5,208,020, and 4,563,304; J. Carlsson et al, Biochem. J. (1978) 173, 723-737(1978); Goff, D. A., BioConjugate Chem. (1990), 1, 381-386; L. Delprino et al. J. Pharm. Sci. (1993), 82, 506-512; S. Arpicco et al., Bioconjugate Chem(1997), 8, 327-337).
Advantageously, because the phosphinate groups are soluble in water or require only a small percentage of organic solvent to maintain solubility in aqueous solution, the reaction between the cell-binding agent and the cross-linker can be conducted in aqueous solution. The cross-linking reagent is dissolved in aqueous buffer, optionally containing a small amount (typically <10% by volume) of a polar organic solvent that is miscible with water, for example different alcohols, such as methanol, ethanol, and propanol, dimethyl formamide (DMF), dimethyl acetamide (DMA), or dimethylsulfoxide (DMSO) at a high concentration, for example 1-100 mM, and then an appropriate aliquot is added to the buffered aqueous solution of the cell-binding agent. An appropriate aliquot is an amount of solution that introduces 1-10 cross-linking groups per cell-binding agent, preferably 1-5 groups, and the volume to be added should not exceed 10%, preferably 5%, and most preferably 0-3% of the volume of the cell-binding agent solution. The aqueous solutions for the cell-binding agents are buffered between pH 6 and 9, preferably between 6.5 and 7.5 and can contain any non-nucleophilic buffer salts useful for these pH ranges. Typical buffers include phosphate, triethanolamine HCl, HEPES, and MOPS buffers, which can contain additional components, such as cyclodextiins, sucrose and salts, for example, NaCl. After the addition the reaction is incubated at a temperature of from 4 to 40° C., preferably at ambient temperature. The progress of the reaction can be monitored by measuring the increase in the absorption at 325 nm or another appropriate wavelength. After the reaction is complete, isolation of the modified cell-binding agent can be performed in a routine way, using for example gel filtration chromatography, or adsorptive chromatography.
The extent of modification can be assessed by measuring the absorbance of the nitropyridine thione, dinitropyridine dithione, carboxamidopyridine dithione or dicarboxamidopyridine dithione group released.
Modified Cytotoxic Drugs
The cytotoxic drugs modified by reaction with crosslinkers of the present invention are preferably represented by the formula (IV):
wherein the substituents are as defined above.
In preferred embodiments, Y is a disulfide substituent, a maleimido, haloacetyl group, or a N-hydroxy succinimide ester.
The modified drugs can be prepared by reacting the drug with the crosslinkers of the present invention to give a modified drug of formula (IV) bearing a functionality capable of reacting with a cell binding agent. For example a thiol-containing drug can be reacted with the crosslinker of formula (I) bearing a maleimdo substituent at neutral pH in aqueous buffer to give a drug connected to the charged linker via a thioether link. A thiol-containing drug can undergo disulfide exchange with a hydrophilic linker bearing a pyrdiyldithio moiety to give a modified drug attached via a disulfide bond to the charged crosslinker. A drug bearing a hydroxyl group can be reacted with a crosslinker bearing a halogen, in the presence of a mild base, to give a modified drug bearing an ether link. A hydroxyl group containing drug can be condensed with a crosslinker of formula (I) bearing a carboxyl group, in the presence of a dehydrating agent, such as dicyclohexylcarbodimide, to give an ester link. An amino group containing drug can similarly undergo condensation with a carboxyl group on the charged crosslinker of formula (I) to give an amide bond. The modified drug can be purified by standard methods such as column chromatography over silica gel or alumina, crystallization, preparatory thin layer chromatography, ion exchange chromatography or HPLC.
Cell-Binding Agents
The cell-binding molecule that comprises the conjugates and the modified cell-binding agents of the present invention may be of any kind presently known, or that become known, molecule that binds to, complexes with or reacts with a moiety of a cell population sought to be therapeutically or otherwise biologically modified.
The cell binding agents include, but are not limited to, large molecular weight proteins such as, for example, full-length antibodies (polyconal antibodies, monoclonal antibodies, dimers, multimers, multispecific antibodies (e.g., bispecific antibodies); single chain antibodies; fragments of antibodies such as Fab, Fab′, F(ab′)2, Fv, [Parham, J. Immunol. 131, 2895-2902 (1983)], fragments produced by a Fab expression library, anti-idiotypic (anti-Id) antibodies, CDR's, and epitope-binding fragments of any of the above which immuno-specifically bind to cancer cell antigens, viral antigens, microbial antigens or a protein generated by the immune system that is capable of recognizing, binding to a specific antigen or exhibiting the desired biological activity (Miller et al (2003) Jour. of Immunology 170:4854-4861); interferons (such as type I, II, III); peptides; lymphokines such as IL-2, IL-3, IL-4, IL-6, GM-CSF, interferon-gamma (IFN-γ); hormones such as insulin, TRH (thyrotropin releasing hormones), MSH (melanocyte-stimulating hormone), steroid hormones, such as androgens and estrogens, melanocyte-stimulating hormone (MSH); growth factors and colony-stimulating factors such as epidermal growth factors (EGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), transforming growth factors (TGF), such as TGFα, TGFβ, insulin and insulin like growth factors (IGF-I, IGF-II) G-CSF, M-CSF and GM-CSF [Burgess, Immunology Today, 5, 155-158 (1984)]; vaccinia growth factors (VGF); fibroblast growth factors (FGFs); smaller molecular weight proteins, poly-peptide, peptides and peptide hormones, such as bombesin, gastrin, gastrin-releasing peptide; platelet-derived growth factors; interleukin and cytokines, such as interleukin-2 (IL-2), interleukin-6 (IL-6), leukemia inhibitory factors, granulocyte-macrophage colony-stimulating factor (GM-CSF); vitamins, such as folate; apoproteins and glycoproteins, such as transferrin {O'Keefe et al, 260 J. Biol. Chem. 932-937 (1985)}; sugar-binding proteins or lipoproteins, such as lectins; cell nutrient-transport molecules; and small molecular inhibitors, such as prostate-specific membrane antigen (PSMA) inhibitors and small molecular tyrosine kinase inhibitors (TKI), non-peptides or any other cell binding molecule or substance, such as bioactive polymers (Dhar, et al, Proc. Natl. Acad., Sci. 2008, 105, 17356-61); dendrimers (Lee, et al, Nat. Biotechnol. 2005, 23, 1517-26; Almutairi, et al; Proc. Natl. Acad. Sci. 2009, 106, 685-90); nanoparticles (Liong, et al, ACS Nano, 2008, 19, 1309-12; Medarova, et al, Nat. Med. 2007, 13, 372-7; Javier, et al, Bioconjugate Chem. 2008, 19, 1309-12); liposomes (Medinai, et al, Curr. Phar. Des, 2004, 10, 2981-9); viral capsides (Flenniken, et al, Viruses Nanotechnol. 2009, 327, 71-93). In general monoclonal antibodies are preferred as a cell-surface binding agent if an appropriate one is available. And antibodies may be murine, human, humanized, chimeric, or derived from other species.
Production of antibodies used in the present invention involves in vivo or in vitro procedures or combinations thereof. Methods for producing polyclonal anti-receptor peptide antibodies are well-known in the art, such as in U.S. Pat. No. 4,493,795 (to Nestor et al). A monoclonal antibody is typically made by fusing myeloma cells with the spleen cells from a mouse that has been immunized with the desired antigen (Köhler, G.; Milstein, C. (1975). Nature 256: 495-497). The detailed procedures are described in “Antibodies—A Laboratory Manual”, Harlow and Lane, eds., Cold Spring Harbor Laboratory Press, New York (1988), which is incorporated herein by reference. Particularly monoclonal antibodies are produced by immunizing mice, rats, hamsters or any other mammal with the antigen of interest such as the intact target cell, antigens isolated from the target cell, whole virus, attenuated whole virus, and viral proteins. Splenocytes are typically fused with myeloma cells using polyethylene glycol (PEG) 6000. Fused hybrids are selected by their sensitivity to HAT (hypoxanthine-aminopterin-thymine), Hybridomas producing a monoclonal antibody useful in practicing this invention are identified by their ability to immunoreact specified receptors or inhibit receptor activity on target cells.
A monoclonal antibody used in the present invention can be produced by initiating a monoclonal hybridoma culture comprising a nutrient medium containing a hybridoma that secretes antibody molecules of the appropriate antigen specificity. The culture is maintained under conditions and for a time period sufficient for the hybridoma to secrete the antibody molecules into the medium. The antibody-containing medium is then collected. The antibody molecules can then be further isolated by well-known techniques, such as using protein-A affinity chromatography; anion, cation, hydrophobic, or size exclusive chromatographies (particularly by affinity for the specific antigen after Protein A, and sizing column chromatography); centrifugation, differential solubility, or by any other standard technique for the purification of proteins.
Media useful for the preparation of these compositions are both well-known in the art and commercially available and include synthetic culture media. An exemplary synthetic medium is Dulbecco's minimal essential medium (DMEM; Dulbecco et al., Virol. 8:396 (1959)) supplemented with 4.5 gm/l glucose, 20 mm glutamine, 20% fetal calf serum and with an anti-foaming agent, such as polyoxyethylene-polyoxypropylene block copolymer.
In addition, antibody-producing cell lines can also be created by techniques other than fusion, such as direct transformation of B lymphocytes with oncogenic DNA, or transfection with an oncovirus, such as Epstein-Barr virus (EBV, also called human herpesvirus 4 (HHV-4)) or Kaposi's sarcoma-associated herpesvirus (KSHV). See, U.S. Pat. Nos. 4,341,761; 4,399,121; 4,427,783; 4,444,887; 4,451,570; 4,466,917; 4,472,500; 4,491,632; 4,493,890. A monoclonal antibody may also be produced via an anti-receptor peptide or peptides containing the carboxyl terminal as described well-known in the art. See Niman et al., Proc. Natl. Acad. Sci. USA, 80: 4949-4953 (1983); Geysen et al., Proc. Natl. Acad. Sci. USA, 82: 178-182 (1985); Lei et al. Biochemistry 34(20): 6675-6688, (1995). Typically, the anti-receptor peptide or a peptide analog is used either alone or conjugated to an immunogenic carrier, as the immunogen for producing anti-receptor peptide monoclonal antibodies.
There are also a number of other well-known techniques for making monoclonal antibodies as binding molecules in this invention. Particularly useful are methods of making fully human antibodies. One method is phage display technology which can be used to select a range of human antibodies binding specifically to the antigen using methods of affinity enrichment. Phage display has been thoroughly described in the literature and the construction and screening of phage display libraries are well known in the art, see, e.g., Dente et al, Gene. 1.48(1):7-13 (1994); Little et al, Biotechnol Adv. 12(3):539-55 (1994); Clackson et al., Nature 352:264-628 (1991); Huse et al., Science 246:1275-1281 (1989).
Moncolonal antibodies derived by hybridoma technique from another species than human, such as mouse, can be humanized to avoid human anti-mouse antibodies when infused into humans. Among the more common methods of humanization of antibodies are complementarity-determining region grafting and resurfacing. These methods have been extensively described, see e.g. U.S. Pat. Nos. 5,859,205 and 6,797,49:2; Liu et al, Immunol Rev. 222:9-27 (2008); Almagro et al, Front Biosci. 1; 13:1619-33 (2008); Lazar et al, Mol Immunol. 44(8):1986-98 (2007); Li et al, Proc. Natl. Acad. Sci. USA. 103(10):3557-62 (2006) each incorporated herein by reference. Fully human antibodies can also be prepared by immunizing transgenic mice, rabbits, monkeys, or other mammals, carrying large portions of the human immunoglobulin heavy and light chains, with an immunogen. Examples of such mice are: the Xenomouse. (Abgenix, Inc.), the HuMAb-Mouse (Medarex/BMS), the VelociMouse (Regeneron), see also U.S. Pat. Nos. 6,596,541, 6,207,418, 6,150,584, 6,111,166, 6,075,181, 5,922,545, 5,661,016, 5,545,806, 5,436,149 and 5,569,825. In human therapy, murine variable regions and human constant regions can also be fused to construct called “chimeric antibodies” that are considerably less immunogenic in man than murine mAbs (Kipriyanov et al, Mol Biotechnol. 26:39-60 (2004); Houdebine, Curr Opin Biotechnol. 13:625-9 (2002) each incorporated herein by reference). In addition, site-directed mutagenesis in the variable region of an antibody can result in an antibody with higher affinity and specificity for its antigen (Brannigan et al, Nat Rev Mol Cell Biol. 3:964-70, (2002)); Adams et al, J. Immunol Methods. 231:249-60 (1999)) and exchanging constant regions of a mAb can improve its ability to mediate effector functions of binding and cytotoxicity.
Antibodies immunospecific for a malignant cell antigen can also be obtained commercially or produced by any method known to one of skill in the art such as, e.g., chemical synthesis or recombinant expression techniques. The nucleotide sequence encoding antibodies immunospecific for a malignant cell antigen can be obtained commercially, e.g., from the GenBank database or a database like it, the literature publications, or by routine cloning and sequencing.
Apart from an antibody, a peptide or protein that bind/block/target or in some other way interact with the epitopes or corresponding receptors on a targeted cell can be used as a binding molecule. These peptides or proteins could be any random peptide or proteins that have an affinity for the epitopes or corresponding receptors and they don't necessarily have to be of the immunoglobulin family. These peptides can be isolated by similar techniques as for phage display antibodies (Szardenings, J Recept Signal Transduct Res. 2003; 23(4):307-49). The use of peptides from such random peptide libraries can be similar to antibodies and antibody fragments. The binding molecules of peptides or proteins may be conjugated on or linked to a large molecules or materials, such as, but is not limited, an albumin, a polymer, a liposome, a nano particle, as long as such attachment permits the peptide or protein to retain its antigen binding specificity.
Examples of antibodies used for conjugation of drugs via the hydrophilic linkers of this prevention for treating cancer, autoimmune disease, and infectious disease include, but are not limited to, 3F8 (anti-GD2), Abagovomab (anti CA-125), Abciximab (anti CD41 (integrin alpha-IIb), Adalimumab (anti-TNF-α), Adecatumumab (anti-EpCAM, CD326), Afelimomab (anti-TNF-α); Afutuzumab (anti-CD20), Alacizumab pegol (anti-VEGFR2), ALD518 (anti-IL-6), Alemtuzumab (Campath, MabCampath, anti-CD52), Altumomab (anti-CEA), Anatumomab (anti-TAG-72), Amukinzumab (IMA-638, anti-IL-13), Apolizumab (anti-HLA-DR), Arcitumomab (anti-CEA), Aselizumab (anti-L-selectin (CD62L), Atlizumab (tocilizumab, Actemra, RoActemra, anti-IL-6 receptor), Atorolimumab (anti-Rhesus factor), Bapineuzumab (anti-beta amyloid), Basiliximab (Simulect, antiCD25 (a chain of IL-2 receptor), Bavituximab (anti-phosphatidylserine), Bectumomab (LymphoScan, anti-CD22), Belimumab (Benlysta, LymphoStat-B, anti-BAFF), Benralizumab (anti-CD125), Bertilimumab (anti-CCL11 (eotaxin-1)), Besilesomab (Scintimun, anti-CEA-related antigen), Bevacizumab (Avastin, anti-VEGF-A), Biciromab (FibriScint, anti-fibrin II beta chain), Bivatuzumab (anti-CD44 v6), Blinatumomab (BITE, anti-CD19), Brentuximab (cAC10, anti-CD30 TNFRSF8), Biiakinumab (anti-IL-12, IL-23) Canakinumab (Ilaris, anti-IL-1), Cantuzumab (C242, anti-CanAg), Capromab, Catumaxomab (Reinovab, anti-EpCAM, anti-CD3), CC49 (anti-TAG-72), Cedelizumab (anti-CD4), Certolizumab pegol (Cimzia anti-TNF-α), Cetuximab (Erbitux, IMC-C225, anti-EGFR), Citatuzumab bogatox (anti-EpCAM), Cixutumumab (anti-IGF-1), Clenoliximab (anti-CD4), Clivatuzumab (anti-MUC1), Conatumumab (anti-TRAIL-R2), CR6261 (anti-Influenza A hemagglutinin), Dacetuzumab (anti-CD40), Daclizumab (Zenapax, anti-CD25 (a chain of IL-2 receptor)), Daratumumab (anti-CD38 (cyclic ADP ribose hydrolase), Denosumab (Prolia, anti-RANKL), Detumomab (anti-B-lymphoma cell), Dorlimomab, Dorlixizumab, Ecromeximab (anti-GD3 ganglioside), Eculizumab (Soliris, anti-C5), Edobacomab (anti-endotoxin), Edrecolomab (Panorex, MAb17-1A, anti-EpCAM), Efalizumab (Raptiva, anti-LFA-1 (CD11a), Efungumab (Mycograb, anti-Hsp90), Elotuzumab (anti-SLAMF7), Elsilimomab (anti-IL-6), Enlimomab pegol (anti-ICAM-1 (CD54)), Epitumomab (anti-episialin), Epratuzumab (anti-CD22), Erlizumab (anti-ITGB2 (CD1.8)), Ertumaxomab (Rexomun, anti-HER2/neu, CD3), Etaracizumab (Abegrin, anti-integrin αvβ3), Exbivirumab (anti-hepatitis B surface antigen), Fanolesomab (NeutroSpec, anti-CD15), Faralimomab (anti-interferon receptor), Farletuzumab (anti-folate receptor 1), Felvizumab (anti-respiratory syncytial virus), Fezakinumab (anti-IL-22), Figitumumab (anti-IGF-1 receptor), Fontolizumab (anti-IFN-γ), Foravirumab (anti-rabies virus glycoprotein), Fresolimumab (anti-TGF-β), Galiximab (anti-CD80), Gantenerumab (anti-beta amyloid), Gavilimomab (anti-CD147 (basigin)), Gemtuzumab (anti-CD33), Girentuximab (anti-carbonic anhydrase 9), Glembatumumab (CR011, anti-GPNMB), Golimumab (Simponi, anti-TNF-α), Gomiliximab (anti-CD23 (IgE receptor)), Ibalizumab (anti-CD4), Ibritumomab (anti-CD20), Igovomab (Indimacis-125, anti-CA-125), Imciromab (Myoscint, anti-cardiac myosin), Infliximab (Remicade, anti-TNF-α), Intetumumab (anti-CD51), Inolimomab (anti-CD25 (a chain of IL-2 receptor)), Inotuzumab (anti-CD22), Ipilimumab (anti-CD152), Iratumumab (anti-CD30 (TNFRSF8)), Keliximab (anti-CD4), Labetuzumab (CEA-Cide, anti-CEA), Lebrikizumab (anti-IL-13), Lemalesomab (anti-NCA-90 (granulocyte antigen)), Lerdelimumab (anti-TGF beta 2), Lexatumumab (anti-TRAIL-R2), Libivirumab (anti-hepatitis B surface antigen), Lintuzumab (anti-CD33), Lucatumumab (anti-CD40), Lumiliximab (anti-CD23 (IgE receptor), Mapatumumab (anti-TRAIL-R1), Maslimomab (anti-T-cell receptor), Matuzumab (anti-EGFR), Mepolizumab (Bosatria, anti-IL-5), Metelimumab (anti-TGF beta 1), Milatuzumab (anti-CD74), Minretumomab (anti-TAG-72), Mitumomab (BEC-2, anti-GD3 ganglioside), Morolimumab (anti-Rhesus factor), Motavizumab (Numax, anti-respiratory syncytial virus), Muromonab-CD3 (Orthoclone OKT3, anti-CD3), Nacolomab (anti-C242), Naptumomab (anti-5T4), Natalizumab (Tysabri, anti-integrin α4), Nebacumab (anti-endotoxin), Necitumumab (anti-EGFR), Nerelimomab (anti-TNF-α), Nimotuzumab (Theracim., Theraloc, anti-EGFR), Nofetumomab, Ocrelizumab (anti-CD20), Odulimomab (Afolimomab, anti-LFA-1 (CD11a)), Ofatumumab (Arzerra, anti-CD20), Olaratumab (anti-PDGF-R α), Omalizumab (Xolair, anti-IgE Fe region), Oportuzumab (anti-EpCAM), Oregovomab (OvaRex, anti-CA-125), Otelixizumab (anti-CD3), Pagibaximab (anti-lipoteichoic acid), Palivizumab (Synagis, Abbosynagis, anti-respiratory syncytial virus), Panitumumab (Vectibix, ABX-EGF, anti-EGFR), Panobacumab (anti-Pseudomonas aeruginosa), Pascolizumab (anti-IL-4), Pemtumomab (Theragyn, anti-MUC1), Pertuzumab (Omnitarg, 2C4, anti-HER2/neu), Pexelizumab (anti-CS), Pintumomab (anti-adenocarcinoma antigen), Priliximab (anti-CD4), Pritumumab (anti-vimentin), PRO 140 (anti-CCRS), Racotumomab (1E10, anti-(N-glycolylneuraminic acid (NeuGc, NGNA)-gangliosides GM3)), Rafivirumab (anti-rabies virus glycoprotein), Ramucirumab (anti-VEGFR2), Ranibizumab (Lucentis, anti-VEGF-A), Raxibacumab (anti-anthrax toxin, protective antigen), Regavirumab (anti-cytomegalovirus glycoprotein B), Reslizumab (anti-Rilotumumab (anti-HGF), Rituximab (MabThera, Rituxanmab, anti-CD20), Robatumumab (anti-IGF-1 receptor), Rontalizumab (anti-IFN-α), Rovelizumab (LeukArrest, anti-CD11, CD18), Ruplizumab (Antova, anti-CD154 (CD40L)), Satumomab (anti-TAG-72), Sevirumab (anti-cytomegalovirus), Sibrotuzumab (anti-FAP), Sifalimumab (anti-IFN-α), Siltuximab (anti-IL-6), Siplizumab (anti-CD2), (Smart) M195 (anti-CD33), Solanezumab (anti-beta amyloid), Sonepcizumab (anti-sphingosine-1-phosphate), Sontuzumab (anti-episialin), Stamulumab (anti-myostatin), Sulesomab (LeukoScan, (anti-NCA-90 (granulocyte antigen), Tacatuzumab (anti-alpha-fetoprotein), Tadocizumab (anti-integrin αIIbβ3), Talizumab (anti-IgE), Tanezumab (anti-NGF), Taplitumomab (anti-CD19), Tefibazumab (Aurexis, (anti-clumping factor A), Telimomab, Tenatumomab (anti-tenascin C), Teneliximab (anti-CD40), Teplizumab (anti-CD3), TGN1412 (anti-CD28), Ticilimumab (Tremelimumab, (anti-CTLA-4), Tigatuzumab (anti-TRAIL-R2), TNX-650 (anti-IL-13), Tocilizumab (Atlizumab, Actemra, RoActemra, (anti-IL-6 receptor), Toralizumab (anti-CD154 (CD40L)), Tositumomab (anti-CD20), Trastuzumab (Herceptin, (anti-HER2/neu), Tremelimumab (anti-CTLA-4), Tucotuzumab celmoleukin (anti-EpCAM), Tuvirumab (anti-hepatitis B virus), Urtoxazumab (anti-Escherichia coli), Ustekinumab (Stelara, anti-IL-12, IL-23), Vapaliximab (anti-AOC3 (VAP-1)), Vedolizumab, (anti-integrin α4β7), Veltuzumab (anti-CD20), Vepalimoinab (anti-AOC3 (VAP-1), Visilizumab (Nuvion, anti-CD3), Vitaxin (anti-vascular integrin axb1), Volociximab (anti-integrin α5β1), Votumumab (HumaSPECT, anti-tumor antigen CTAA16.88), Zalutumumab (HuMax-EGFr, (anti-EGFR), Zanolimumab (HuMax-CD4, anti-CD4), Ziralimumab (anti-CD147 (basigin)), Zolimomab (anti-CD5), Etanercept (Enbre®), Alefacept (Amevive®), Abatacept (Orencia®), Rilonacept (Arcalyst), 14F7 [anti-IRP-2 (Iron Regulatory Protein 2)], 14G2a (anti-GD2 ganglioside, from Nat. Cancer Inst. for melanoma and solid tumors), J591 (anti-PSMA, Weill Cornell Medical School for prostate cancers), 225.28S [anti-HMW-MAA (High molecular weight-melanoma-associated antigen), Sorin Radiofarmaci S. R. L. (Milan, Italy) for melanoma], COL-1 (anti-CEACAM3, CGM1, from Nat. Cancer Inst. USA for colorectal and gastric cancers), CYT-356 (Oncoltad®, for prostate cancers), HNK20 (OraVax Inc. for respiratory syncytial virus), ImmuRAIT (from Immunomedics for NHL), Lym-1 (anti-HLA-DR10, Peregrine Pharm. for Cancers), MAK-195F [anti-TNF (tumor necrosis factor; TNFA, TNF-alpha; TNFSF2), from Abbott/Knoll for Sepsis toxic shock], MEDI-500 [T10B9, anti-CD3, TR.αβ (T cell receptor alpha/beta), complex, from MedImmune Inc for Graft-versus-host disease], RING SCAN [ anti-TAG 72 (tumour associated glycoprotein 72), from Neoprobe Corp. for Breast, Colon and Rectal cancers], Avicidin (anti-EPCAM (epithelial cell adhesion molecule), anti-TACSTD1 (Tumor-associated calcium signal transducer 1), anti-GA733-2 (gastrointestinal tumor-associated protein 2), anti-EGP-2 (epithelial glycoprotein 2); anti-KSA; KS1/4 antigen; M4S; tumor antigen 17-1A; CD326, from NeoRx Corp. for Colon, Ovarian, Prostate cancers and NHL]; LymphoCide (Immunomedics, NJ), Smart ID10 (Protein Design Labs), Oncolym (Techniclone Inc, CA), Allomune (BioTransplant, CA), anti-VEGF (Genentech, CA); CEAcide (Immunomedics, NJ), IMC-1C11 (ImClone Systems, NJ) and Cetuximab (ImClone, NJ)
Other antibodies as binding ligands include, but are not limited to, are antibodies against the following antigens: Aminopeptidase N (CD13), Annexin A1, B7-H3 (CD276, various cancers), CA125 (ovarian), CA15-3 (carcinomas), CA19-9 (carcinomas), L6 (carcinomas), Lewis Y (carcinomas), Lewis X (carcinomas), alpha fetoprotein (carcinomas), CA242 (colorectal), placental alkaline phosphatase (carcinomas), prostate specific antigen (prostate), prostatic acid phosphatase (prostate), epidermal growth factor (carcinomas), CD2 (Hodgkin's disease, NHL lymphoma, multiple myeloma), CD3 epsilon (T cell lymphoma, lung, breast, gastric, ovarian cancers, autoimmune diseases, malignant ascites), CD19 (B cell malignancies), CD20 (non-Hodgkin's lymphoma), CD22 (leukemia, lymphoma, multiple myeloma, SLE), CD30 (Hodgkin's lymphoma), CD33 (leukemia, autoimmune diseases), CD38 (multiple myeloma), CD40 (lymphoma, multiple myeloma, leukemia (CLL)), CD51 (Metastatic melanoma, sarcoma), CD52 (leukemia), CD56 (small cell lung cancers, ovarian cancer, Merkel cell carcinoma, and the liquid tumor, multiple myeloma), CD66e (cancers), CD70 (metastatic renal cell carcinoma and non-Hodgkin lymphoma), CD74 (multiple myeloma), CD80 (lymphoma), CD98 (cancers), mucin (carcinomas), CD221 (solid tumors), CD227 (breast, ovarian cancers), CD262 (NSCLC and other cancers), CD309 (ovarian cancers), CD326 (solid tumors), CEACAM3 (colorectal, gastric cancers), CEACAM5 (carcinoembryonic antigen; CEA, CD66e) (breast, colorectal and lung cancers), DLL4 (Δ-like-4), EGFR (Epidermal Growth Factor Receptor, various cancers), CTLA4 (melanoma), CXCR4 (CD184, Heme-oncology, solid tumors), Endoglin (CD105, solid tumors), EPCAM (epithelial cell adhesion molecule, bladder, head, neck, colon, NHL prostate, and ovarian cancers), ERBB2 (Epidermal Growth Factor Receptor 2; lung, breast, prostate cancers), FCGRI (autoimmune diseases), FOLR (folate receptor, ovarian cancers), GD2 ganglioside (cancers), G-28 (a cell surface antigen glyvolipid, melanoma), GD3 idiotype (cancers), Heat shock proteins (cancers), HER1 (lung, stomach cancers), HER2 (breast, lung and ovarian cancers), HLA-DR.10 (NHL), HLA-DRB (NHL, B cell leukemia), human chorionic gonadotropin (carcinoma), IGF1R (insulin-like growth factor 1 receptor, solid tumors, blood cancers), IL-2 receptor (interleukin 2 receptor, T-cell leukemia and lymphomas), IL-6R (interleukin 6 receptor, multiple myeloma, RA, Castleman's disease, IL6 dependent tumors), Integrins (αvβ3, α5β1, α6β4, α11β3, α5β5, αvβ5, for various cancers), MAGE-1 (carcinomas), MAGE-2 (carcinomas), MAGE-3 (carcinomas), MAGE 4 (carcinomas), anti-transferrin receptor (carcinomas), p97 (melanoma), MS4A1 (membrane-spanning 4-domains subfamily A member 1, Non-Hodgkin's B cell lymphoma, leukemia), MUC1 or MUC1-KLH (breast, ovarian, cervix, bronchus and gastrointestinal cancer), MUC16 (CA125) (Ovarian cancers), CEA (colorectal), gp100 (melanoma), MART1 (melanoma), MPG (melanoma), MS4A1 (membrane-spanning 4-domains subfamily A, small cell lung cancers, NHL), Nucleolin, Neu oncogene product (carcinomas), P21 (carcinomas), Paratope of anti-(N-glycolylneuraminic acid, Breast, Melanoma cancers), PLAP-like testicular alkaline phosphatase (ovarian, testicular cancers), PSMA (prostate tumors), PSA (prostate), ROBO4, TAG 72 (tumour associated glycoprotein 72, AML, gastric, colorectal, ovarian cancers), T cell transmembrane protein (cancers), Tie (CD202b), TNFRSHOB (tumor necrosis factor receptor superfamily member 10B, cancers), TNFRSF13B (tumor necrosis factor receptor superfamily member 13B, multiple myeloma, NHL, other cancers, RA and SLE), TPBG (trophoblast glycoprotein, Renal cell carcinoma), TRAIL-R1 (Tumor necrosis apoprosis Inducing ligand Receptor 1, lymphoma, NHL, colorectal, lung cancers), VCAM-1 (CD106, Melanoma), VEGF, VEGF-A, VEGF-2 (CD309) (various cancers). Some other tumor associated antigens recognized by antibodies have been reviewed (Gerber, et al, mAbs 1:3, 247-253 (2009); Novellino et al, Cancer Immunol Immunother. 54(3), 187-207 (2005). Franke, et al, Cancer Biother Radiopharm. 2000, 15, 459-76). Examples of these antigens that antibodies against are: Many other Cluster of Differentiations (CD4, CD5, CD6, CD7, CD8, CD9, CD10, CD11a, CD11b, CD11c, CD12w, CD14, CD15, CD16, CDw17, CD18, CD21, CD23, CD24, CD25, CD26, CD27, CD28, CD29, CD31, CD32, CD34, CD35, CD36, CD37, CD41, CD42, CD43, CD44, CD45, CD46, CD47, CD48, CD49b, CD49c, CD53, CD54, CD55, CD58, CD59, CD61, CD62E, CD62L, CD62P, CD63, CD68, CD69, CD71, CD72, CD79, CD81, CD82, CD83, CD86, CD87, CD88, CD89, CD90, CD91, CD95, CD96, CD100, CD103, CD105, CD106, CD109, CD117, CD120, CD127, CD133, CD134, CD135, CD138, CD141, CD142, CD143, CD144, CD147, CD151, CD152, CD154, CD156, CD158, CD163, CD166, CD168, CD184, CDw186, CD195, CD202 (a, b), CD209, CD235a, CD271, CD303, CD304), Annexin A1, Nucleolin, Endoglin (CD105), ROBO4, Amino-peptidase N, □-like-4 (DLL4), VEGFR-2 (CD309), CXCR4 9CD184), Tie2, B7-H3, WT1, MUC1, LMP2, HPV E6 E7, EGFRvIII, 1-IER-2/neu, Idiotype, MAGE A3, p53 nonmutant, NY-ESO-1, GD2, CEA, MelanA/MART1, Ras mutant, gp100, p53 mutant, Proteinase3 (PRI), bcr-abl, Tyrosinase, Survivin, hTERT, Sarcoma translocation breakpoints, EphA2, PAP, ML-IAP, AFP, EpCAM, ERG (TMPRSS2 ETS fusion gene), NA17, PAX3, ALK, Androgen receptor, Cyclin B1, Polysialic acid, MYCN, RhoC, TRP-2, GD3, Fucosyl GNU, Mesothelin, PSCA, MAGE A1, sLe(a), CYP1B1, PLAC1, GM3, BORIS, Tn, GloboH, ETV6-AML, NY-BR-1, RGS5, SART3, STn, Carbonic anhydrase IX, PAXS, OY-TES1, Sperm protein 17, LCK, HMWMAA, AKAP-4-SSX2, XAGE 1, B7H3, Legumain, Tie 2, Page4, VEGFR2, MAD-CT-1, FAP, PDGFR-P, MAD-CT-2, Fos-related antigen 1.
In another specific embodiment, the cell-binding-drug conjugates via the hydrophilic likers of this invention are used for the treatment of cancers. The cancers include, but are not limited, Adrenocortical Carcinoma, Anal Cancer, Bladder Cancer, Brain Tumor (Adult, Brain Stem Glioma, Childhood, Cerebellar Astrocytoma, Cerebral Astrocytoma, Ependymoma, Medulloblastoma, Supratentorial Primitive Neuroectodermal and Pineal Tumors, Visual Pathway and Hypothalamic Glioma), Breast Cancer, Carcinoid Tumor, Gastrointestinal, Carcinoma of Unknown Primary, Cervical Cancer, Colon Cancer, Endometrial Cancer, Esophageal Cancer, Extrahepatic Bile Duct Cancer, Ewings Family of Tumors (PNET), Extracranial Germ Cell Tumor, Eye Cancer, Intraocular Melanoma, Gallbladder Cancer, Gastric Cancer (Stomach), Germ Cell Tumor, Extragonadal, Gestational Trophoblastic Tumor, Head and Neck Cancer, Hypopharyngeal Cancer, Islet Cell Carcinoma, Kidney Cancer (renal cell cancer), Laryngeal Cancer, Leukemia (Acute Lymphoblastic, Acute Myeloid, Chronic Lymphocytic, Chronic Myelogenous, Hairy Cell), Lip and Oral Cavity Cancer, Liver Cancer, Lung Cancer (Non-Small Cell, Small Cell, Lymphoma (AIDS-Related, Central Nervous System, Cutaneous T-Cell, Hodgkin's Disease, Non-Hodgkin's Disease, Malignant Mesothelioma, Melanoma, Merkel Cell Carcinoma, Metasatic Squamous Neck Cancer with Occult Primary, Multiple Myeloma, and Other Plasma Cell Neoplasms, Mycosis Fungoides, Myelodysplastic Syndrome, Myeloproliferative Disorders, Nasopharyngeal Cancer, Neuroblastoma, Oral Cancer, Oropharyngeal Cancer, Osteosarcoma, Ovarian Cancer (Epithelial, Germ Cell Tumor, Low Malignant Potential Tumor), Pancreatic Cancer (Exocrine, islet Cell Carcinoma), Paranasal Sinus and Nasal Cavity Cancer, Parathyroid Cancer, Penile Cancer, Pheochromocytoma Cancer, Pituitary Cancer, Plasma Cell Neoplasm, Prostate Cancer Rhabdomyosarcoma, Rectal Cancer, Renal Cell Cancer (kidney cancer), Renal Pelvis and Ureter (Transitional Cell), Salivary Gland Cancer, Sezary Syndrome, Skin Cancer, Skin Cancer (Cutaneous T-Cell Lymphoma, Kaposi's Sarcoma, Melanoma), Small Intestine Cancer, Soft Tissue Sarcoma, Stomach Cancer, Testicular Cancer, Thymoma (Malignant), Thyroid Cancer, Urethral Cancer, Uterine Cancer (Sarcoma), Unusual Cancer of Childhood, Vaginal Cancer, Vulvar Cancer, Wilms' Tumor.
In another specific embodiment, the cell-binding-drug conjugates via the hydrophilic likers of this invention are used in accordance with the compositions and methods for the treatment or prevention of an autoimmune disease. The autoimmune diseases include, but are not limited, Achlorhydra Autoimmune Active Chronic Hepatitis, Acute Disseminated Encephalomyelitis, Acute hemorrhagic leukoencephalitis, Addison's Disease, Agammaglobulinemia, Alopecia areata, Amyotrophic Lateral Sclerosis, Ankylosing Spondylitis, Anti-GBM/TBM Nephritis, Antiphospholipid syndrome, Antisynthetase syndrome, Arthritis, Atopic allergy, Atopic Dermatitis, Autoimmune Aplastic Anemia, Autoimmune cardiomyopathy, Autoimmune hemolytic anemia, Autoimmune hepatitis, Autoimmune inner ear disease, Autoimmune lymphoproliferative syndrome, Autoimmune peripheral neuropathy, Autoimmune pancreatitis, Autoimmune polyendocrine syndrome Types I, II, & III, Autoimmune progesterone dermatitis, Autoimmune thrombocytopenic purpura, Autoimmune uveitis, Balo disease/Balo concentric sclerosis, Bechets Syndrome, Berger's disease, Bickerstaffs encephalitis, Blau syndrome, Bullous Pemphigoid, Castleman's disease, Chagas disease, Chronic Fatigue Immune Dysfunction Syndrome, Chronic inflammatory demyelinating polyneuropathy, Chronic recurrent multifocal ostomyelitis, Chronic lyme disease, Chronic obstructive pulmonary disease, Churg-Strauss syndrome, Cicatricial Pemphigoid, Coeliac Disease, Cogan syndrome, Cold agglutinin disease, Complement component 2 deficiency, Cranial arteritis, CREST syndrome, Crohns Disease (a type of idiopathic inflammatory bowel diseases), Cushing's Syndrome, Cutaneous leukocytoclastic angiitis, Dego's disease, Dercum's disease, Dermatitis herpetiformis, Dermatomyositis, Diabetes mellitus type 1, Diffuse cutaneous systemic sclerosis, Dressler's syndrome, Discoid lupus erythematosus, Eczema, Endometriosis, Enthesitis-related arthritis, Eosinophilic fasciitis, Epidermolysis bullosa acquisita, Erythema nodosum, Essential mixed cryoglobulinemia, Evan's syndrome, Fibrodysplasia ossificans progressiva, Fibromyalgia, Fibromyositis, Fibrosing aveolitis, Gastritis, Gastrointestinal pemphigoid, Giant cell arteritis, Glomerulonephritis, Goodpasture's syndrome, Graves' disease, Guillain-Barré syndrome, Hashimoto's encephalitis, Hashimoto's thyroiditis, Haemolytic anaemia, Henoch-Schonlein purpura, Herpes gestationis, Hidradenitis suppurativa, Hughes syndrome (See Antiphospholipid syndrome), Hypogammaglobulinemia, Idiopathic Inflammatory Demyelinating Diseases, Idiopathic pulmonary fibrosis, Idiopathic thrombocytopenic purpura (See Autoimmune thrombocytopenic purpura), IgA nephropathy (Also Berger's disease), inclusion body myositis, Inflammatory demyelinating polyneuopathy, Interstitial cystitis, Irritable Bowel Syndrome, Juvenile idiopathic arthritis, Juvenile rheumatoid arthritis, Kawasaki's Disease, Lambert-Eaton myasthenic syndrome, Leukocytoclastic vasculitis, Lichen planus, Lichen sclerosus, Linear IgA disease (LAD), Lou Gehrig's Disease (Also Amyotrophic lateral sclerosis), Lupoid hepatitis, Lupus erythematosus, Majeed syndrome, Ménière's disease, Microscopic polyangiitis, Miller-Fisher syndrome, Mixed Connective Tissue Disease, Morphea, Mucha-Habermann disease, Muckle-Wells syndrome, Multiple Myeloma, Multiple Sclerosis, Myasthenia gravis, Myositis, Narcolepsy, Neuromyelitis optica (Devic's Disease), Neuromyotonia, Occular cicatricial pemphigoid, Opsoclonus myoclonus syndrome, Ord thyroiditis, Palindromic rheumatism, PANDAS (Pediatric Autoimmune Neuropsychiatric Disorders Associated with Streptococcus), Paraneoplastic cerebellar degeneration, Paroxysmal nocturnal hemoglobinuria, Parry Romberg syndrome, Parsonnage-Turner syndrome, Pars planitis, Pemphigus, Pemphigus vulgaris, Pernicious anaemia, Perivenous encephalomyelitis, POEMS syndrome, Polyarteritis nodosa, Polymyalgia rheumatica, Polymyositis, Primary biliary cirrhosis, Primary sclerosing cholangitis, Progressive inflammatory neuropathy, Psoriasis, Psoriatic Arthritis, Pyoderma gangrenosum, Pure red cell aplasia, Rasmussen's encephalitis, Raynaud phenomenon, Relapsing polychondritis, Reiter's syndrome, Restless leg syndrome, Retroperitoneal fibrosis, Rheumatoid arthritis, Rheumatoid fever, Sarcoidosis, Schizophrenia, Schmidt syndrome, Schnitzler syndrome, Scleritis, Scleroderma, Sjögren's syndrome, Spondyloarthropathy, Sticky blood syndrome, Still's Disease, Stiff person syndrome, Subacute bacterial endocarditis, Susac's syndrome, Sweet syndrome, Sydenham Chorea, Sympathetic ophthalmia, Takayasu's arteritis, Temporal arteritis (giant cell arteritis), Tolosa-Hunt syndrome, Transverse Myelitis, Ulcerative Colitis (a type of idiopathic inflammatory bowel diseases), Undifferentiated connective tissue disease, Undifferentiated spondyloarthropathy, Vasculitis, Vitiligo, Wegener's granulomatosis, Wilson's syndrome, Wiskott-Aldrich syndrome.
In another specific embodiment, a binding molecule used for the conjugate via the hydrophilic linkers of this invention for the treatment or prevention of an autoimmune disease includes, but are not limited to, anti-elastin antibody; Abys against epithelial cells antibody; Anti-Basement Membrane Collagen Type IV Protein antibody; Anti-Nuclear Antibody; Anti cls DNA; Anti ss DNA, Anti Cardiolipin Antibody IgM, IgG; anti-celiac antibody; Anti Phospholipid Antibody IgK, IgG; Anti SM Antibody; Anti Mitochondrial Antibody; Thyroid Antibody; Microsomal Antibody, T-cells antibody; Thyroglobulin Antibody, Anti SCL-70; Anti-Jo; Anti-U.sub.1RNP; Anti-La/SSB; Anti SSA; Anti SSB; Anti Perital Cells Antibody; Anti Histones; Anti RNP; C-ANCA; P-ANCA; Anti centromere; Anti-Fibrillarin, and Anti GBM Antibody, Anti-ganglioside antibody; Anti-Desmogein 3 antibody; Anti-p62 antibody; Anti-sp100 antibody; Anti-Mitochondrial(M2) antibody; Rheumatoid factor antibody; Anti-MCV antibody; Anti-topoisomerase antibody; Anti-neutrophil cytoplasmic(cANCA) antibody;
In certain preferred embodiments, the binding molecule for the conjugate in the present invention, can bind to both a receptor or a receptor complex expressed on an activated lymphocyte which is associated with an autoimmune disease. The receptor or receptor complex can comprise an immunoglobulin gene superfamily member (e.g. CD2, CD3, CD4, CD8, CD19, CD22, CD28, CD79, CD90, CD152/CTLA-4, PD-1, or ICOS), a TNF receptor superfamily member (e.g. CD27, CD40, CD95/Fas, CD134/0X40, CD137/4-1BB, INF-R1, TNFR-2, RANK, TACI, BCMA, osteoprotegerin, Apo2/TRAIL-R1, TRAIL-R:2, TRAIL-R3, TRAIL-R4, and APO-3), an integrin, a cytokine receptor, a chemokine receptor, a major histocompatibility protein, a lectin (C-type, S-type, or I-type), or a complement control protein.
In another specific embodiment, useful binding ligands that are immunospecific for a viral or a microbial antigen are humanized or human monoclonal antibodies. As used herein, the term “viral antigen” includes, but is not limited to, any viral peptide, polypeptide protein (e.g. HIV gp120, HIV nef, RSV F glycoprotein, influenza virus neuramimidase, influenza virus hemagglutinin, HTLV tax, herpes simplex virus glycoprotein (e.g. gB, gC, gD, and gE) and hepatitis B surface antigen) that is capable of eliciting an immune response. As used herein, the term “microbial antigen” includes, but is not limited to, any microbial peptide, polypeptide, protein, saccharide, polysaccharide, or lipid molecule (e.g., a bacterial, fungi, pathogenic protozoa, or yeast polypeptide including, e.g., LPS and capsular polysaccharide 5/8) that is capable of eliciting an immune response. Examples of antibodies available 1 for the viral or microbial infection include, but are not limited to, Palivizumab which is a humanized anti-respiratory syncytial virus monoclonal antibody for the treatment of RSV infection; PRO542 which is a CD4 fusion antibody for the treatment of HIV infection; Ostavir which is a human antibody for the treatment of hepatitis B virus; PROTVIR which is a humanized IgG.sub,1 antibody for the treatment of cytomegalovirus; and anti-LPS antibodies.
The cell binding molecules—drug conjugates via the hydrophilic linkers of this invention can be used in the treatment of infectious diseases. These infectious diseases include, but are not limited to, Acinetobacter infections, Actinomycosis, African sleeping sickness (African trypanosomiasis), AIDS (Acquired immune deficiency syndrome), Amebiasis, Anaplasmosis, Anthrax, Arcanobacterium haemolyticum infection, Argentine hemorrhagic fever, Ascariasis, Aspergillosis, Astrovirus infection, Babesiosis, Bacillus cereus infection, Bacterial pneumonia, Bacterial vaginosis, Bacteroides infection, Balantidiasis, Baylisascaris infection, BK virus infection, Black piedra, Blastocystis hominis infection, Blastomycosis, Bolivian hemorrhagic fever, Borrelia infection, Botulism (and Infant botulism), Brazilian hemorrhagic fever, Brucellosis, Burkholderia infection, Buruli ulcer, Calicivirus infection (Norovirus and Sapovirus), Campylobacteriosis, Candidiasis (Moniliasis; Thrush), Cat-scratch disease, Cellulitis, Chagas Disease (American trypanosomiasis), Chancroid, Chickenpox, Chlamydia, Chlamydophila pneumoniae infection, Cholera, Chromoblastomycosis, Clonorchiasis, Clostridium difficile infection, Coccidioidomycosis, Colorado tick fever, Common cold (Acute viral rhinopharyngitis; Acute coryza), Creutzfeldt-Jakob disease, Crimean-Congo hemorrhagic fever, Cryptococcosis, Cryptosporidiosis, Cutaneous larva migrans, Cyclosporiasis, Cysticercosis, Cytomegalovirus infection, Dengue fever, Dientamoebiasis, Diphtheria, Diphyllobothriasis, Dracunculiasis, Ebola hemorrhagic fever, Echinococcosis, Ehrlichiosis, Enterobiasis (Pinworm infection), Enterococcus infection, Enterovirus infection, Epidemic typhus, Erythema infectiosum (Fifth disease), Exanthem subitum, Fasciolopsiasis, Fasciolosis, Fatal familial insomnia, Filariasis, Food poisoning by Clostridium perfringens, Free-living amebic infection, Fusobacterium infection, Gas gangrene (Clostridial myonecrosis), Geotrichosis, Gerstmann-Sträussler-Scheinker syndrome, Giardiasis, Glanders, Gnathostomiasis, Gonorrhea, Granuloma inguinale (Donovanosis), Group A streptococcal infection, Group B streptococcal infection, Haemophilus influenzae infection, Hand, foot and mouth disease (HFMD), Hantavirus Pulmonary Syndrome, Helicobacter pylon infection, Hemolytic-uremic syndrome, Hemorrhagic fever with renal syndrome, Hepatitis A, Hepatitis B, Hepatitis C, Hepatitis D, Hepatitis E, Herpes simplex, Histoplasmosis, Hookworm infection, Human bocavirus infection, Human ewingii ehrlichiosis, Human granulocytic anaplasmosis, Human metapneumovirus infection, Human monocytic ehrlichiosis, Human papillomavirus infection, Human parainfluenza virus infection, Hymenolepiasis, Epstein-Barr Virus Infectious Mononucleosis (Mono), Influenza, Isosporiasis, Kawasaki disease, Keratitis, Kingella kingae infection, Kuru, Lassa fever, Legionellosis (Legionnaires' disease), Legionellosis (Pontiac fever), Leishmaniasis, Leprosy, Leptospirosis, Listeriosis, Lyme disease (Lyme borreliosis), Lymphatic filariasis (Elephantiasis), Lymphocytic choriomeningitis, Malaria, Marburg hemorrhagic fever, Measles, Melioidosis (Whitmore's disease), Meningitis, Meningococcal disease, Metagonimiasis, Microsporidiosis, Molluscum contagiosum, Mumps, Murine typhus (Endemic typhus), Mycoplasma pneumonia, Mycetoma, Myiasis, Neonatal conjunctivitis (Ophthalmia neonatorum), (New) Variant Creutzfeldt-Jakob disease (vCJD, nvCJD), Nocardiosis, Onchocerciasis (River blindness), Paracoccidioidomycosis (South American blastomycosis), Paragonimiasis, Pasteurellosis, Pediculosis capitis (Head lice), Pediculosis commis (Body lice), Pediculosis pubis (Pubic lice, Crab lice), Pelvic inflammatory disease, Pertussis (Whooping cough), Plague, Pneumococcal infection, Pneumocystis pneumonia, Pneumonia, Poliomyelitis, Prevotella infection, Primary amoebic meningoencephalitis, Progressive multifocal leukoencephalopathy, Psittacosis, Q fever, Rabies, Rat-bite fever, Respiratory syncytial virus infection, Rhinosporidiosis, Rhinovirus infection, Rickettsial infection, Rickettsialpox, Rift Valley fever, Rocky mountain spotted fever, Rotavirus infection, Rubella, Salmonellosis, SARS (Severe Acute Respiratory Syndrome), Scabies, Schistosomiasis, Sepsis, Shigellosis (Bacillary dysentery), Shingles (Herpes zoster), Smallpox (Variola), Sporotrichosis, Staphylococcal food poisoning, Staphylococcal infection, Strongyloidiasis, Syphilis, Taeniasis, Tetanus (Lockjaw), Tinea barbae (Barber's itch), Tinea capitis (Ringworm of the Scalp), Tinea corporis (Ringworm of the Body), Tinea cruris (Jock itch), Tinea manuum (Ringworm of the Hand), Tinea nigra, Tinea pedis (Athlete's foot), Tinea unguium (Onychomycosis), Tinea versicolor (Pityriasis versicolor), Toxocariasis (Ocular Larva Migrans), Toxocariasis (Visceral Larva Migrans), Toxoplasmosis, Trichinellosis, Trichomoniasis, Trichuriasis (Whipworm infection), Tuberculosis, Tularemia, Ureaplasma urealyticum infection, Venezuelan equine encephalitis, Venezuelan hemorrhagic fever, Viral pneumonia, West Nile Fever, White piedra (Tinea blanca), Yersinia pseudotuberculosis infection, Yersiniosis, Yellow fever, Zygomycosis.
The cell binding molecules, which are more proffered to be antibodies described in this patent that are against pathogenic strains include, but are not limit, Acinetobacter baumannii, Actinomyces israelii, Actinomyces gerencseriae and Propionibacterium propionicus, Trypanosoma brucei, HIV (Human immunodeficiency virus), Entamoeba histolytica, Anaplasma genus, Bacillus anthracis, Arcanobacterium haemolyticum, Junin virus, Ascaris lumbricoides, Aspergillus genus, Astroviridae family, Babesia genus, Bacillus cereus, multiple bacteria, Bacteroides genus, Balantidium coli, Baylisascaris genus, BK virus, Piedraia hortae, Blastocystis hominis, Blastomyces dermatitides, Machupo virus, Borrelia genus, Clostridium botulinum, Sabia, Brucella genus, usually Burkholderia cepacia and other Burkholderia species, Mycobacterium ulcerans, Caliciviridae family, Campylobacter genus, usually Candida albicans and other Candida species, Bartonella henselae, Group A Streptococcus and Staphylococcus, Trypanosoma cruzi, Haemophilus ducreyi, Varicella zoster virus (VZV), Chlamydia trachomatis, Chlamydophila pneumoniae, Vibrio cholerae, Fonsecaea pedrosoi, Clonorchis sinensis, Clostridium difficile, Coccidioides immitis and Coccidioides posadasii, Colorado tick fever virus, rhinoviruses, coronaviruses, CJD prion, Crimean-Congo hemorrhagic fever virus, Cryptococcus neoformans, Cryptosporidium genus, Ancylostoma braziliense; multiple parasites, Cyclospora cayetanensis, Taenia solium, Cytomegalovirus, Dengue viruses (DEN-1, DEN-2, DEN-3 and DEN-4)-Flaviviruses, Dientamoeba fragilis, Corynebacterium diphtheriae, Diphyllobothrium, Dracunculus medinensis, Ebolavirus, Echinococcus genus, Ehrlichia genus, Enterobius vermicularis, Enterococcus genus, Enterovirus genus, Rickettsia prowazekii, Parvovirus B19, Human herpesvirus 6 and Human herpesvirus 7, Fasciolopsis buski, Fasciola hepatica and Fasciola gigantica, FPI prion, Filarioidea superfamily, Clostridium perfringens, Fusobacterium genus, Clostridium perfringens; other Clostridium species, Geotrichum candidum, GSS prion, Giardia intestinalis, Burkholderia mallei, Gnathostoma spinigerum and Gnathostoma hispidum, Neisseria gonorrhoeae, Klebsiella granulomatis, Streptococcus pyogenes, Streptococcus agalactiae, Haemophilus influenzae, Enteroviruses, mainly Coxsackie A virus and Enterovirus 71, Sin Nombre virus, Helicobacter pylori, Escherichia coli O157:H7, Bunyaviridae family, Hepatitis A Virus, Hepatitis B Virus, Hepatitis C Virus, Hepatitis D Virus, Hepatitis E Virus, Herpes simplex virus 1, Herpes simplex virus 2, Histoplasma capsulatum, Ancylostoma duodenale and Necator americanus, Hemophilus influenzae, Human bocavirus, Ehrlichia ewingii, Anaplasma phagocytophilum, Human metapneumovirus, Ehrlichia chaffeensis, Human papillomavirus, Human parainfluenza viruses, Hymenolepis nana and Hymenolepis diminuta, Epstein-Barr Virus, Orthomyxoviridae family, Isospora belli, Kingella kingae, Klebsiella pneumoniae, Klebsiella ozaenas, Klebsiella rhinoscleromotis, Kuru prion, Lassa virus, Legionella pneumophila, Legionella pneumophila, Leishmania genus, Mycobacterium leprae and Mycobacterium lepromatosis, Leptospira genus, Listeria monocytogenes, Borrelia burgdorferi and other Borrelia species, Wuchereria bancrofti and Brugia malayi, Lymphocytic choriomeningitis virus (LCMV), Plasmodium genus, Marburg virus, Measles virus, Burkholderia pseudomallei, Neisseria meningitides, Metagonimus yokagawai, Microsporidia phylum, Molluscum contagiosum virus (MCV), Mumps virus, Rickettsia typhi, Mycoplasma pneumoniae, numerous species of bacteria (Actinomycetoma) and fungi (Eumycetoma), parasitic dipterous fly larvae, Chlamydia trachomatis and Neisseria gonorrhoeae, vCJD prion, Nocardia asteroides and other Nocardia species, Onchocerca volvulus, Paracoccidioides brasiliensis, Paragonimus westermani and other Paragonimus species, Pasteurella genus, Pediculus humanus capitis, Pediculus humanus corporis, Phthirus pubis, Bordetella pertussis, Yersinia pestis, Streptococcus pneumoniae, Pneumocystis jirovecii, Poliovirus, Prevotella genus, Naegleria fowleri, JC virus, Chlamydophila psittaci, Coxiella burnetii, Rabies virus, Streptobacillus moniliformis and Spirillum minus, Respiratory syncytial virus, Rhinosporidium seeberi, Rhinovirus, Rickettsia genus, Rickettsia akari, Rift Valley fever virus, Rickettsia rickettsii, Rotavirus, Rubella virus, Salmonella genus, SARS coronavirus, Sarcoptes scabiei, Schistosoma genus, Shigella genus, Varicella zoster virus, Variola major or Variola minor, Sporothrix schenckii, Staphylococcus genus, Staphylococcus genus, Staphylococcus aureus, Streptococcus pyogenes, Strongyloides stercoralis, Treponema pallidum, Taenia genus, Clostridium tetani, Trichophyton genus, Trichophyton tonsurans, Trichophyton genus, Epidermophyton floccosum, Trichophyton rubrum, and Trichophyton mentagrophytes, Trichophyton rubrum, Hortaea werneckii, Trichophyton genus, Malassezia genus, Toxocara canis or Toxocara cati, Toxoplasma gondii, Trichinella spiralis, Trichomonas vaginalis, Trichuris trichiura, Mycobacterium tuberculosis, Francisella tularensis, Ureaplasma urealyticum, Venezuelan equine encephalitis virus, Vibrio colerae, Guanarito virus, West Nile virus, Trichosporon beigelii, Yersinia pseudotuberculosis, Yersinia enterocolitica, Yellow fever virus, Mucorales order (Mucormycosis) and Entomophthorales order (Entomophthoramycosis), Pseudomonas aeruginosa, Campylobacter (Vibrio) fetus, Aeromonas hydrophila, Edwardsiella tarda, Yersinia pestis, Shigella dysenteriae, Shigella flexneri, Shigella sonnei, Salmonella typhimurium, Treponema pertenue, Treponema carateneum, Borrelia vincentii, Borrelia burgdorferi, Leptospira icterohemorrhagiae, Pneumocystis carinii, Brucella abortus, Brucella suis, Brucella melitensis, Mycoplasman spp., Rickettsia prowazeki, Rickettsia tsutsugumushi, Clamydia spp.; pathogenic fungi (Aspergillus fumigatus, Candida albicans, Histoplasma capsulatum); protozoa (Entomoeba Trichomonas tenas, Trichomonas hominis, Tryoanosoma gambiense, Trypanosoma rhodesiense, Leishmania donovani, Leishmania tropica, Leishmania braziliensis, Pneumocystis pneumonia, Plasmodium vivax, Plasmodium falciparum, Plasmodium malaria); or Helminiths (Schistosoma japonicum, Schistosoma mansoni, Schistosoma haematobium, and hookworms).
Other antibodies as cell binding ligands used in this invention for treatment of viral disease include, but are not limited to, antibodies against antigens of pathogenic viruses, including as examples and not by limitation: Poxyiridae, Herpesviridae, Adenoviridae, Papovaviridae, Enteroviridae, Picornaviridae, Parvoviridae, Reoviridae, Retroviridae, influenza viruses, parainfluenza viruses, mumps, measles, respiratory syncytial virus, rubella, Arboviridae, Rhabdoviridae, Arenaviridae, Non-A/Non-B Hepatitis virus, Rhinoviridae, Coronaviridae, Rotoviridae, Oncovirus [such as, HBV (Hepatocellular carcinoma), HPV (Cervical cancer, Anal cancer), Kaposi's sarcoma-associated herpesvirus (Kaposi's sarcoma), Epstein-Barr virus (Nasopharyngeal carcinoma, Burkitt's lymphoma, Primary central nervous system lymphoma), MCPyV (Merkel cell cancer), SV40 (Simian virus 40), HCV (Hepatocellular carcinoma), HTLV-I (Adult T-cell leukemia/lymphoma)], Immune disorders caused virus: [such as Human Immunodeficiency Virus (AIDS)]; Central nervous system virus: [such as, JCV (Progressive multifocal leukoencephalopathy), MeV (Subacute sclerosing panencephalitis), LCV (Lymphocytic choriomeningitis), Arbovirus encephalitis, Orthomyxoviridae (probable) (Encephalitis lethargica), RV (Rabies), Chandipura virus, Herpesviral meningitis, Ramsay Hunt syndrome type II; Poliovirus (Poliomyelitis, Post-polio syndrome), HTLV-I (Tropical spastic paraparesis)]; Cytomegalovirus (Cytomegalovirus retinitis, HSV (Herpetic keratitis)); Cardiovascular virus [such as CBV (Pericarditis, Myocarditis)]; Respiratory system/acute viral nasopharyngitis/viral pneumonia: [Epstein-Barr vials (EBV infection/Infectious mononucleosis), Cytomegalovirus; SARS coronavirus (Severe acute respiratory syndrome) Orthomyxoviridae: Influenzavirus A/B/C (InfluenzaJAvian influenza), Paramyxovirus: Human parainfluenza viruses (Parainfluenza), RSV (Human respiratory syncytial virus), hMPV]; Digestive system virus [MuV (Mumps), Cytomegalovirus (Cytomegalovirus esophagitis); Adenovirus (Adenovirus infection); Rotavirus, Norovirus, Astrovirus, Coronavirus; HBV (Hepatitis B virus), CBV, HAV (Hepatitis A virus), HCV (Hepatitis C virus), HDV (Hepatitis D virus), HEV (Hepatitis E virus), HGV (Hepatitis G virus)]; Urogenital virus [such as, BK virus, MuV (Mumps)].
According to a further object, the present invention also concerns pharmaceutical compositions comprising the conjugate via the hydrophilic linkers of the invention together with a pharmaceutically acceptable carrier for treatment of cancer and autoimmune disorders. The method for treatment of cancer and autoimmune disorders can be practiced in vitro, in vivo, or ex vivo. Examples of in vitro uses include treatments of cell cultures in order to kill all cells except for desired variants that do not express the target antigen; or to kill variants that express undesired antigen. Examples of ex vivo uses include treatments of hematopoietic stem cells (HSC) prior to the performance of the transplantation (HSCT) into the same patient in order to kill diseased or malignant cells. For instance, clinical ex vivo treatment to remove tumour cells or lymphoid cells from bone marrow prior to autologous transplantation in cancer treatment or in treatment of autoimmune disease, or to remove T cells and other lymphoid cells from allogeneic bone marrow or tissue prior to transplant in order to prevent graft-versus-host disease, can be carried out as follows. Bone marrow is harvested from the patient or other individual and then incubated in medium containing serum to which is added the conjugate of the invention, concentrations range from about 1 pM to 0.1 mM, for about 30 minutes to about 48 hours at about 37° C. The exact conditions of concentration and time of incubation (=dose) are readily determined by the skilled clinicians. After incubation the bone marrow cells are washed with medium containing serum and returned to the patient by i.v. infusion according to known methods. In circumstances where the patient receives other treatment such as a course of ablative chemotherapy or total-body irradiation between the time of harvest of the marrow and reinfusion of the treated cells, the treated marrow cells are stored frozen in liquid nitrogen using standard medical equipment.
For clinical in vivo use, the conjugate via the linkers of the invention will be supplied as solutions or as a lyophilized solid that can be redisolved in sterile water for injection. Examples of suitable protocols of conjugate administration are as follows. Conjugates are given weekly for 8 weeks as an i.v. bolus. Bolus doses are given in 50 to 500 ml of normal saline to which human serum albumin (e.g. 0.5 to 1 mL of a concentrated solution of human serum albumin, 100 mg/mL) can be added. Dosages will be about 50 μg to 20 mg/kg of body weight per week, i.v. (range of 10 μg to 200 mg/kg per injection). 8 weeks after treatment, the patient may receive a second course of treatment. Specific clinical protocols with regard to route of administration, excipients, diluents, dosages, times, etc., can be determined by the skilled clinicians.
Examples of medical conditions that can be treated according to the in vivo or ex vivo methods of killing selected cell populations include malignancy of any types of cancer, autoimmune diseases, graft rejections, and infections (viral, bacterial or parasite).
The amount of a conjugate which is required to achieve the desired biological effect, will vary depending upon a number of factors, including the chemical characteristics, the potency, and the bioavailability of the conjugates, the type of disease, the species to which the patient belongs, the diseased state of the patient, the route of administration, all factors which dictate the required dose amounts, delivery and regimen to be administered.
In general terms, the conjugates via the linkers of this invention may be provided in an aqueous physiological buffer solution containing 0.1 to 10% w/v conjugates for parenteral administration. Typical dose ranges are from 1 μg/kg to 0.1 g/kg of body weight per day; a preferred dose range is from 0.01 mg/kg to 20 mg/kg of body weight per day or an equivalent dose in a human child. The preferred dosage of drug to be administered is likely to depend on such variables as the type and extent of progression of the disease or disorder, the overall health status of the particular patient, the relative biological efficacy of the compound selected, the formulation of the compound, the route of administration (intravenous, intramuscular, or other), the pharmacokinetic properties of the compound by the chosen delivery route, and the speed (bolus or continuous infusion) and schedule of administrations (number of repetitions in a given period of time).
The conjugates via the linkers of the present invention are also capable of being administered in unit dose forms, wherein the term “unit dose” means a single dose which is capable of being administered to a patient, and which can be readily handled and packaged, remaining as a physically and chemically stable unit dose comprising either the active conjugate itself, or as a pharmaceutically acceptable composition, as described hereinafter. As such, typical total daily dose ranges are from 0.01 to 100 mg/kg of body weight. By way of general guidance, unit doses for humans range from 1 mg to 3000 mg per day. Preferably the unit dose range is from 1 to 500 mg administered one to four times a day, and even more preferably from 10 mg to 500 mg, once a day. Conjugatess provided herein can be formulated into pharmaceutical compositions by admixture with one or more pharmaceutically acceptable excipients. Such unit dose compositions may be prepared for use by oral administration, particularly in the form of tablets, simple capsules or soft gel capsules; or intranasally, particularly in the form of powders, nasal drops, or aerosols; or dermally, for example, topically in ointments, creams, lotions, gels or sprays, or via trans-dermal patches.
Drugs/Cytotoxic Agents
Drugs that can be conjugated to a cell-binding molecule in the present invention are small molecule drugs including cytotoxic agents, which can be linked to or after they are modified for linkage to the cell-binding agent. A “small molecule drug” is broadly used herein to refer to an organic, inorganic, or organometallic compound that may have a molecular weight of for example 100 to 1800, more suitably from 120 to 1400. Small molecule drugs are well characterized in the art, such as in WO05058367A2, and in U.S. Pat. No. 4,956,303, among others and are incorporated in their entirety by reference. The drugs includes known drugs and those that may become known drugs.
Drugs that are known include, but not limited to, 1). Chemotherapeutic agents: a). Alkylating agents: such as Nitrogen mustards: chlorambucil, chlornaphazine, cyclophosphamide, dacarbazine, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, mannomustine, mitobronitol, melphalan, mitolactol, pipobroman, novembichin, phenesterine, prednimustine, thiotepa, trofosfamide, uracil mustard; CC-1065 (including its adozelesin, carzelesin and bizelesin synthetic analogues); duocarmycin (including the synthetic analogues, KW-2189 and CBI-TMI); benzodiazepine dimers (e.g., dimmers of pyrrolobenzodiazepine (PBD) or tomaymycin, indolinobenzodiazepines, imidazobenzothiadiazepines, or oxazolidinobenzodiazepines); Nitrosoureas: (carmustine, lomustine, chlorozotocin, fotemustine, nimustine, ranimustine); Alkylsulphonates: (busulfan, treosulfan, improsulfan and piposulfan); Triazenes: (dacarbazine); Platinum containing compounds; (carboplatin, cisplatin, oxaliplatin); aziridines, such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines including altretamine, triethylenemelamine, trietylenephosphoramide, triethylenethiophosphaoramide and trimethylolomelamine]; b). Plant Alkaloids: such as Vinca alkaloids: (vincristine, vinblastine, vindesine, vinorelbine, navelbin); Taxoids: (paclitaxel, docetaxol) and their analogs, Maytansinoids (DM1, DM2, DM3, DM4, maytansine and ansamitocins) and their analogs, cryptophycins (particularly cryptophycin 1 and cryptophycin 8); epothilones, eleutherobin, discodermolide, bryostatins, dolostatins, auristatins, tubulysins, cephalostatins; pancratistatin; a sarcodictyin; spongistatin; c). DNA Topoisomerase Inhibitors: such as [Epipodophyllins: (9-aminocamptothecin, camptothecin, crisnatol, daunomycin, etoposide, etoposide phosphate, irinotecan, mitoxantrone, novantrone, retinoic acids (retinols), teniposide, topotecan, 9-nitrocamptothecin (RFS 2000)); mitomycins: (mitomycin C)]; d). Anti-metabolites: such as {[Anti-folate: DHFR inhibitors: (methotrexate, trimetrexate, denopterin, pteropterin, aminopterin (4-acid) or the other folic acid analogues); IMP dehydrogenase Inhibitors: (mycophenolic acid, tiazofurin, ribavirin, EICAR); Ribonucleotide reductase Inhibitors: (hydroxyurea, deferoxamine)]; [Pyrimidine analogs: Uracil analogs: (ancitabine, azacitidine, 6-azauridine, capecitabine (Xeloda), carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, 5-Fluorouracil, floxuridine, ratinexed(Tomuclex)); Cytosine analogs: (cytarabine, cytosine arabinoside, fludarabine); Purine analogs: (azathioprine, fludarabine, mercaptopurine, thiamiprine, thioguanine)]; folic acid replenisher, such as frolinic acid}; e). Hormonal therapies: such as {Receptor antagonists: [Anti-estrogen; (megestrol, raloxifene, tamoxifen); LHRH agonists: (goscrclin, leuprolide acetate); Anti-androgens: (bicalutamide, flutamide, calusterone, dromostanolone propionate, epitiostanol, goserelin, leuprolide, mepitiostane, nilutamide, testolactone, trilostane and other androgens inhibitors)]; Retinoids/Deltoids: [Vitamin D3 analogs: (CB 1093, EB 1089 KH 1060, cholecalciferol, ergocalciferol); Photodynamic therapies: (verteporfin, phthalocyanine, photosensitizer Pc4, demethoxy-hypocrellin A); Cytokines: (Interferon-alpha, Interferon-gamma, tumor necrosis factor (TNFs), human proteins containing a TNF domain)]}; f). Kinase inhibitors, such as BIBW 2992 (anti-EGFR/Erb2), imatinib, gefitinib, pegaptanib, sorafenib, dasatinib, sunitinib, erlotinib, nilotinib, lapatinib, axitinib, pazopanib. vandetanib, E7080 (anti-VEGFR2), mubritinib, ponatinib (AP24534), bafetinib (INNO-406), bosutinib (SKI-606), cabozantinib, vismodegib, iniparib, ruxolitinib, CYT387, axitinib, tivozanib, sorafenib, bevacizumab, cetuximab, Trastuzumab, Ranibizumab, Panitumumab, ispinesib; g). antibiotics, such as the enediyne antibiotics (e.g. calicheamicins, especially calicheamicin γ1, δ1 and β1, see, e.g., J. Med. Chem., 39 (11), 2103-2117 (1996), Angew Chem Intl. Ed, Engl, 33:183-186 (1994); dynemicin, including dynemicin A and deoxydynemicin; esperamicin, kedarcidin, C-1027, maduropeptin, as well as neocarzinostatin chromophore and related chromoprotein enediyne antiobiotic chromomophores), aclacinomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, carabicin, carminomycin, carzinophilin; chromomycins, dactinomycin, daunorubicin, doxorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin, epirubicin, epirubicin, idarubicin, marcellomycin, nitomycins, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; f). Others: such as Polyketides (acetogenins), especially bullatacin and bullatacinone; gemcitabine, epoxomicins (e. g. carfilzomib), bortezomib, thalidomide, lenalidomide, pomalidomide, tosedostat, zybrestat, PLX4032, STA-9090, Stimuvax, allovectin-7, Xegeva, Provenge, Yervoy, Isoprenylation inhibitors (such as Lovastatin), Dopaminergic neurotoxins (such as 1-methyl-4-phenylpyridinium ion), Cell cycle. inhibitors (such as staurosporine), Actinomycins (such as Actinomycin D, dactinomycin), Bleomycins (such as bleomycin A2, bleomycin B2, peplomycin), Anthracyclines (such as daunorubicin, doxorubicin (adriamycin), idarubicin, epirubicin, pirarubicin, zorubicin, mtoxantrone, MDR inhibitors (such as verapamil), Ca2+ ATPase inhibitors (such as thapsigargin), Histone deacetylase inhibitors (Vorinostat, Romidepsin, Panobinostat, Valproic acid, Mocetinostat (MGCD0103), Belinostat, PC1-24781, Entinostat, SB939, Resminostat, Givinostat, AR-42, CUDC-101, sulforaphane, Trichostatin A); Thapsigargin, Celecoxib, glitazones, epigallocatechin gallate, Disulfiram, Salinosporamide A.; Anti-adrenals, such as aminoglutethimide, mitotane, trilostane; aceglatone; aldophosphamide glycoside; aminolevulinic acid; amsacrine; arabinoside, bestrabucil; bisantrene; edatraxate; defofamine; demecolcine; diaziquone; eflornithine (DFMO), elfomithine; elliptinium acetate, etoglucid; gallium nitrate; gacytosine, hydroxyurea; ibandronate, lentinan; lonidamine; mitoguazone; mitoxantrone; mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; podophyllinic acid; 2-ethylhydrazide; procarbazine; PSL®; razoxane; rhizoxin; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2,2′,2″-trichlorotriethylamine; trichothecenes (especially T-2 toxin, verrucarin A, roridin A and anguidine); urethane, siRNA, antisense drugs, and a nucleolytic enzyme.
2). An anti-autoimmune disease agent includes, but is not limited to, cyclosporine, cyclosporine A, aminocaproic acid, azathioprine, bromocriptine, chlorambucil, chloroquine, cyclophosphamide, corticosteroids (e.g. amcinonide, betamethasone, budesonide, hydrocortisone, flunisolide, fluticasone propionate, fluocortolone danazol, dexamethasone, Triamcinolone acetonide, beclometasone dipropionate), DHEA, enanercept, hydroxychloroquine, infliximab, meloxicam, methotrexate, mofetil, mycophenylate, prednisone, sirolimus, tacrolimus.
3). An anti-infectious disease agent includes, but is not limited to, a). Aminoglycosides: amikacin, astromicin, gentamicin (netilmicin, sisomicin, isepamicin), hygromycin B, kanamycin (amikacin, arbekacin, bekanamycin, dibekacin, tobramycin), neomycin (framycetin, paromomycin, ribostamycin), netilmicin, spectinomycin, streptomycin, tobramycin, verdamicin; b). Amphenicols: azidamfenicol, chloramphenicol, florfenicol, thiamphenicol; c). Ansamycins: geldanamycin, herbimycin; d). Carbapenems: biapenem, doripenem, ertapenem, imipenem/cilastatin, meropenem, panipenem; e). Cephems: carbacephem (loracarbef), cefacetrile, cefaclor, cefradine, cefadroxil, cefalonium, cefaloridine, cefalotin or cefalothin, cefalexin, cefaloglycin, cefamandole, cefapirin, cefatrizine, cefazaflur, cefazedone, cefazolin, cefbuperazone, cefcapene, cefdaloxime, cefepime, cefminox, cefoxitin, cefprozil, cefroxadine, ceftezole, cefuroxime, cefixime, cefdinir, cefditoren, cefepime, cefetamet, cefmenoxime, cefodizime, cefonicid, cefoperazone, ceforanide, cefotaxime, cefotiam, cefozopran, cephalexin, cefpimizole, cefpiramide, cefpirome, cefpodoxime, cefprozil, cefquinome, cefsulodin, ceftazidime, cefteram, ceftibuten, ceftiolene, ceftizoxime, ceftobiprole, ceftriaxone, cefuroxime, cefuzonam, cephamycin (cefoxitin, cefotetan, cefinetazole), oxacephem (flomoxef, latamoxef); Glycopeptides: bleomycin, vancomycin (oritavancin, telavancin), teicoplanin (dalbavancin), ramoplanin; g). Glycylcyclines: e. g. tigecycline; g). β-Lactamase inhibitors: penam (sulbactam, tazobactam), clavam (clavulanic acid); i). Lincosamides: clindamycin, lincomycin; j). Lipopeptides: daptomycin, A54145, calcium-dependent antibiotics (CDA); k). Macrolides: azithromycin, cethromycin, clarithromycin, dirithromycin, erythromycin, flurithromycin, josamycin, ketolide (telithromycin, cethromycin), midecamycin, miocamycin, oleandomycin, rifamycins (rifampicin, rifampin, rifabutin, rifapentine), rokitamycin, roxithromycin, spectinomycin, spiramycin, tacrolimus (FK506), troleandomycin, telithromycin; l). Monobactams: aztreonam, tigemonam; m). Oxazolidinones: linezolid; n). Penicillins: amoxicillin, ampicillin (pivampicillin, bacampicillin, metampicillin, azidocillin, azlocillin, benzylpenicillin, benzathine benzylpenicillin, benzathine phenoxymethylpenicillin, clometocillin, procaine benzylpenicillin, carbenicillin (carindacillin), cloxacillin, dicloxacillin, epicillin, flucloxacillin, mecillinam (pivmecillinam), mezlocillin, meticillin, nafcillin, penamecillin, penicillin, pheneticillin, phenoxymethylpenicillin, piperacillin, propicillin, sulbenicillin, temocillin, ticarcillin; o). Polypeptides: bacitracin, colistin, polymyxin B; p). Quinolones: alatrofloxacin, balofloxacin, ciprofloxacin, clinafloxacin, danofloxacin, difloxacin, enoxacin, enrofloxacin, floxin, garenoxacin, gatifloxacin, gemifloxacin, grepafloxacin, kano trovafloxacin, levofloxacin, lomefloxacin, marbofloxacin, moxifloxacin, nadifloxacin, norfloxacin, orbifloxacin, ofloxacin, pefloxacin, trovafloxacin, grepafloxacin, sitafloxacin, sparfloxacin, temafloxacin, tosufloxacin, trovafloxacin; q). Streptogramins: pristinamycin, quinupristinidalfopristin); r). Sulfonamides: mafenide, prontosil, sulfacetamide, sulfamethizole, sulfanilimide, sulfasalazine, sulfisoxazole, trimethoprim, trimethoprim-sulfamethoxazole (co-trimoxazole); s). Steroid antibacterials: e.g. fusidic acid; t). Tetracyclines: doxycycline, chlortetracycline, clomocycline, demeclocycline, lymecycline, meclocycline, metacycline, minocycline, oxytetracycline, penimepicycline, rolitetracycline, tetracycline, glycylcyclines (e.g. ti.gecycline); u). Other types of antibiotics: annonacin, arsphenamine, bactoprenol inhibitors (Bacitracin), DADAL/AR inhibitors (cycloserine), dictyostatin, discodermolide, eleutherobin, epothilone, ethambutol, etoposide, faropenem, fusidic acid, furazolidone, isoniazid, laulimalide, metronidazole, mupirocin, mycolactone, NAM synthesis inhibitors (e. g. fosfomycin), nitrofurantoin, paclitaxel, platensimycin, pyrazinamide, quinupristin/dalfopristin, rifampicin (rifampin), tazobactam tinidazole, uvaricin;
4). Anti-viral drugs: a). Entry/fusion inhibitors: aplaviroc, maraviroc, vicriviroc, gp41 (enfuvirtide), PRO 140, CD4 (ibalizumab); b). Integrase inhibitors: raltegravir, elvitegravir, globoidnan A; c). Maturation inhibitors: bevirimat, vivecon; d). Neuraminidase inhibitors: oseltamivir, zanamivir, peramivir; e). Nucleosides & nucleotides: abacavir, aciclovir, adefovir, amdoxovir, apricitabine, brivudine, cidofovir, clevudine, dexelvucitabine, didanosine (ddI), elvucitabine, emtricitabine (FTC), entecavir, famciclovir, fluorouracil (5-FU), 3′-fluoro-substituted 2′,3′-dideoxynucleoside analogues (e.g. 3′-fluoro-2′,3′-dideoxythymidine (FLT) and 3′-fluoro-2′,3′-dideoxyguanosine (FLG), fomivirsen, ganciclovir, idoxuridine, lamivudine (3TC), 1-nucleosides (e.g. β-1-thymidine and β-1-2′-deoxycytidine), penciclovir, racivir, ribavirin, stampidine, stavudine (d4T), taribavirin (viramidine), telbivudine, tenofovir, trifluridine valaciclovir, valganciclovir, zalcitabine (ddC), zidovudine (AZT); f). Non-nucleosides: amantadine, ateviridine, capravirine, diarylpyrimidines (etravirine, rilpivirine), delavirdine, docosanol, emivirine, efavirenz, foscarnet (phosphonoformic acid), imiquimod, interferon alfa, loviride, lodenosine, methisazone, nevirapine, NOV-205, peginterferon alfa, podophyllotoxin, rifampicin, rimantadine, resiquimod (R-848), tromantadine; g). Protease inhibitors: amprenavir, atazanavir, boceprevir, darunavir, fosamprenavir, indinavir, lopinavir, nelfinavir, pleconaril, ritonavir, saquinavir, telaprevir (VX-950), tipranavir; h). Other types of anti-virus drugs: abzyme, arbidol, calanolide a, ceragenin, cyanovirin-n, diarylpyrimidines, epigallocatechin gallate (EGCG), foscarnet, griffithsin, taribavirin (viramidine), hydroxyurea, KP-1461, miltefosine, pleconaril, portmanteau inhibitors, ribavirin, seliciclib.
5). The drugs used for conjugates via a charged linker of the present invention also include radioisotopes. Examples of radioisotopes (radionuclides) are 3H, 11C, 14C, 18F, 32P, 35S, 64Cu, 68Ga, 86Y, 99Tc, 111In, 123I, 124I, 125I, 131I, 133Xe, 177Lu, 211At, or 213Bi. Radioisotope labeled antibodies are useful in receptor targeted imaging experiments or can be for targeted treatment such as with the antibody-drug conjugates of the invention (Wu et al (2005) Nature Biotechnology 23(9): 1137-1146). The cell binding molecules, e.g. an antibody can be labeled with ligand reagents through the charged linkers of the present patent that bind, chelate or otherwise complex a radioisotope metal, using the techniques described in Current Protocols in Immunology, Volumes 1 and 2, Coligen et al, Ed. Wiley-Interscience, New York, N.Y., Pubs. (1991). Chelating ligands which may complex a metal ion include DOTA, DOTP, DOTMA, DTPA and TETA (Macmcyclics, Dallas, Tex.).
6). The pharmaceutically acceptable salts, acids or derivatives of any of the above drugs.
Preferred cytotoxic agents that conjugated to a cell-binding molecule via a charged linker of this patent are tubulusins, maytansinoids, taxanoids (taxanes), CC-1065 analogs, daunorubicin and doxorubicin compounds, benzodiazepine dimers (e.g., dimers of pyrrolobenzodiazepine (PBD), tomaymycin, anthramycin, indolinobenzodiazepines, imidazobenzothiadiazepines, or oxazolidinobenzodiazepines), calicheamicins and the enediyne antibiotics, actinomycin, azaserines, bleomycins, epirubicin, tamoxifen, idarubicin, dolastatins/auristatins (e.g. monomethyl auristatin E, MMAE, MMAF, auristatin PYE, auristatin TP, Auristatins 2-AQ, 6-AQ, EB (AEB), and EFP (AEFP)), duocarmycins, thiotepa, vincristine, hemiasterlins, esperanlicins, and their analogues and derivatives thereof.
Tubulysins that are preferred for conjugation in the present invention are well known in the art and can be isolated from natural sources according to known methods or prepared synthetically according to known methods (e. g. Balasubramanian, R.; et al. J. Med. Chem., 2009, 52, 238-240. Wipf, P.; et al. Org. Lett., 2004, 6, 4057-4060. Pando, O.; et al. J. Am. Chem. Soc., 2011, 133, 7692-7695. Reddy, J. A.; et al. Mol. Pharmaceutics, 2009, 6, 1518-1525. Raghavan, B.; et al. J. Med. Chem., 2008, 51, 1530-1533. Patterson, A. W.; et al. J. Org. Chem., 2008, 73, 4362-4369. Pando, O.; et al. Org. Lett., 2009, 11 (24), pp 5567-5569. Wipf, P.; et al. Org. Lett., 2007, 9 (8), 1605-1607. Friestad, G. K.; Org. Lett., 2004, 6, pp 3249-3252. Hillary M. Peltier, H. M.; et al. J. Am. Chem. Soc., 2006, 128, 16018-16019. Chandrasekhar, S.; et al. J. Org. Chem., 2009, 74, 9531-9534. Liu, Y.; et al. Mol. Pharmaceutics, 2012, 9, 168-175. Friestad, G. K.; et al. Org. Lett., 2009, 11, 1095-1098. Kubicek, K.; et al., Angew Chem Int Ed Engl, 2010. 49: p. 4809-12, Chai, Y.; et al., Chem Biol, 2010, 17: 296-309. Ulrich, A.; et al., Angew Chem int Ed Engl, 2009, 48, 4422-5. Sani, M.; et al. Angew Chem Int Ed Engl, 2007, 46, 3526-9. Domling, A.; et al., Angew Chem Int Ed Engl, 2006, 45, 7235-9. Patent applications: Zanda, M.; et al, Can. Pat. Appl. CA 2710693 (2011). Chai, Y.; et al. Eur. Pat. Appl. 2174947 (2010), PCT WO 2010034724. Leamon, C.; et al, PCT WO 2010033733, WO 2009002993. Ellman, J.; et al, PCT WO 2009134279; PCT WO 2009012958, US appl. 20110263650, 20110021568, Matschiner, G.; et al, PCT WO 2009095447. Vlahov, I.; et al, PCT WO 2009055562, WO 2008112873. Low, P.; et al, PCT WO 2009026177. Richter, W., PCT WO 2008138561. Kjems, J.; et al, PCT WO 2008125116. Davis, M.; et al, PCT WO 2008076333. Diener, J.; et al, U.S. Pat. Appl. 20070041901, WO 2006096754. Matschiner, G.; et al, PCT WO 2006056464. Vaghefi, F.; et al, 5 PCT WO 2006033913. Doemling, A., Ger. Offen. DE 102004030227; PCT WO 2004005327; WO 2004005326; WO2004005269. Stanton, M.; et al, U.S. Pat. Appl. Publ. 20040249130. Hoene, G.; et al, Ger. Offen. DE 10:254439; DE 1024115:2; DE 10008089. Leung, D.; et al, PCT WO 2002077036. Reichenbach, H.; et al, Ger. Offen. DE 19638870; Wolfgang, R.; US 20120129779, Chen, H., US appl. 20110027274. The preferred structure of tubulysins for conjugation of cell binding molecules are described in the patent application of PCT/IB2012/053554.
Calicheamicins and their related enediyne antibiotics that are preferred for cell-binding molecule-drug conjugates of this patent are described in: Nicolaou, K. C. et al, Science 1992, 256, 1172-1178; Proc. Natl. Acad. Sci USA. 1993, 90, 5881-5888), U.S. Pat. Nos. 4,970,198; 5,053,394; 5,108,912; 5,264,586; 5,384,412; 5,606,040; 5,712,374; 5,714,586; 5,739,116; 5,770,701; 5,770,710; 5,773,001; 5,877,296; 6,015,562; 6,124,310; 8,153,768.
Maytansinoids that are preferred to be used in the present invention including maytansinol and maytansinol analogues are described in U.S. Pat. Nos. 4,256,746, 4,361,650 and 4,307,016, 4,294,757, 4,294,757, 4,371,533, 4,424,219, 4,331,598, 4,450,254, 4,364,866, 4,313,946, 4,315,929 4,362,663, 4,32:2,348, 4,371,533, 4,424,219, 5,208,020, 5,416,064, 5,208,020; 5,416,064; 6,333.410; 6,441,163; 6,716,821, 7,276,497, 7,301,019, 7,303,749, 7,368,565, 7,411,063, 7,851,432, 8,163,888.
Taxanes, which includes Paclitaxel (Taxol), a cytotoxic natural product, and docetaxel (Taxotere), a semi-synthetic derivative, and their analogs which are preferred for conjugation via the charged linkers of the present patent are exampled in: K C. Nicolaou et al., J. Am. Chem. Soc. 117, 2409-2420, (1995); Ojima et al, J. Med. Chem. 39:3889-3896 (1996); 40:267-278 (1997); 45, 56:20-5623 (2002); Ojima et al., Proc, Natl. Acad. Sci., 96:4256-4261 (1999; Kim et al., Bull. Korean Chem. Soc., 20, 1389-1390 (1999); Miller, et al. J. Med. Chem., 47, 4802-4805(2004); U.S. Pat. Nos. 5,475,011 5,728,849, 5,811,45:2; 6,340,701; 6,372,738; 6,391,913, 6,436,931; 6,589,979; 6,596,757; 6,706,708; 7,008,942; 7,186,851; 7,217,819; 7,276,499; 7,598,290; 7,667,054.
CC-1065 analogues and doucarmycin analogs are also preferred to be used for a conjugate with the charged linker of the present patent. The examples of the CC-1065 analogues and doucarmycin analogs as well as their synthesis are described in: e.g. Warpehoski et al, J. Med. Chem. 31:590-603 (1988), D. Boger et al., J. Org. Chem; 66; 6654-6661, 2001; U.S. Pat. Nos. 4,169,888, 4,391,904, 4,671,958, 4,816,567, 4,912,227, 4,923,990, 4,952,394, 4,975,278, 4,978,757, 4,994,578, 5,037,993, 5,070,092, 5,084,468, 5,101,038, 5,117,006, 5,137,877, 5,138,059, 5,147,786, 5,187,186, 5,223,409, 5,225,539, 5,288,514, 5,324,483, 5,332,740, 5,332,837, 5,334,528, 5,403,484, 5,427,908, 5,475,092, 5,495,009, 5,530,101, 5,545,806, 5,547,667, 5,569,825, 5,571,698, 5,573,922, 5,580,717, 5,585,089, 5,585,499, 5,587,161, 5,595,499, 5,606,017, 5,622,929, 5,625,126, 5,629,430, 5,633,425, 5,641,780, 5,660,829, 5,661,016, 5,686,237, 5,693,762, 5,703,080, 5,712,374, 5,714,586, 5,739,116, 5,739,350, 5,770,429, 5,773,001, 5,773,435, 5,786,377, 5,786,486, 5,789,650, 5,814,318, 5,846,545, 5,874,299, 5,877,296, 5,877,397, 5,885,793, 5,939,598, 5,962,216, 5,969,108, 5,985,908, 6,060,608, 6,066,742, 6,075,181, 6,103,236, 6,114,598, 6,130,237, 6,132,722, 6,143,901, 6,150,584, 6,162,963, 6,172,197, 6,180,370, 6,194,612, 6,214,345, 6,262,271, 6,281,354, 6,310,209, 6,329,497, 6,342,480, 6,486,326, 6,512,101, 6,521,404, 6,534,660, 6,544,731, 6,548,530, 6,555,313, 6,555,693, 6,566,336, 6,586,618, 6,593,081, 6,630,579, 6,756,397, 6,759,509, 6,762,179, 6,884,869, 6,897,034, 6,946,455, 7,049,316, 7,087,600, 7,091,186, 7,115,573, 7,129,261, 7,214,663, 7,223,837, 7,304,032, 7,329,507, 7,329,760, 7,388,026, 7,655,660, 7,655,661, 7,906,545, 8,012,978.
Daunorubicin/Doxorubicin Analogues are also preferred for conjugation via the charged linker of the present patent. The preferred structures and their synthesis are exampled in: Hurwitz, E., et al., Cancer Res. 35, 1175-1181 (1975). Yang, H. M., and Reisfeld, R. A., Proc. Natl. Acad, Sci. 85, 1189-1193 (1988); Pietersz, C. A., E., et al., E., et al.,” Cancer Res. 48, 926-9311 (1988); Trouet, et al., 79, 626-629 (1982); Z. Brich et al., J. Controlled Release, 19, 245-258 (1992); Chen et al., Syn. Comm., 33, 2377-2390, 2003; King et al., Bioconj. Chem., 10, 279-288, 1999; King et al., J. Med. Chem., 45, 4336-4343, 2002; Kratz et al., J Med Chem., 45, 5523-33. 2002; Kratz et al., Biol Pharm. Bull. January 21, 56-61, 1998; Lau et al., Bioorg. Med. Chem. 3, 1305-1312, 1995; Scott et al., Bioorg. Med.1 Chem. Lett. 6, 1491-1496; 1996; Watanabe et al., Tokai J. Experimental Clin. Med. 15, 327-334, 1990; Zhou et al., J. Am. Chem. Soc, 126, 15656-7, 2004; WO 01/38318; U.S. Patent No.). U.S. Pat. Nos. 5,106,951; 5,122,368; 5,146,064; 5,177,016; 5,208,323; 5,824,805; 6,146,658; 6,214,345; 7569358; 7,803,903; 8,084,586; 8,053,205. Auristatins and dolastatins are preferred in conjugation via the charged linker of this patent. The auristatins (e. g. auristain E (AE) auristatin EB (AEB), auristatin EFP (AEFP), monomethyl auristatin E (MMAE), Monomethylauristatin (MMAF), Auristatin F phenylene diamine (AFP) and a phenylalanine variant of MMAE) which are synthetic analogs of dolastatins, are described in Int, J. Oncol. 15:367-7:2 (1999); Molecular Cancer Therapeutics, vol. 3, No. 8, pp, 921-932 (2004); U.S. Application Nos. 11134826, 20060074008, 2006022925. U.S. Pat. Nos. 4,414,205, 4,753,894, 4,764,368, 4,816,444, 4,879,278, 4,943,628, 4,978,744, 5,122,368, 5,165,923, 5,169,774, 5,286,637, 5,410,024, 5,521,284, 5,530,097, 5,554,725, 5,585,089, 5,599,902, 5,629,197, 5,635,483, 5,654,399, 5,663,149, 5,665,860, 5,708,146, 5,714,586, 5,741,892, 5,767,236, 5,767,237, 5,780,588, 5,821,337, 5,840,699, 5,965,537, 6,004,934, 6,033,876, 6,034,065, 6,048,720, 6,054,297, 6,054,561, 6,124,431, 6,143,721, 6,162,930, 6,214,345, 6,239,104, 6,323,315, 6,342,219, 6,342,221, 6,407,213, 6,569,834, 6,620,911, 6,639,055, 6,884,869, 6,913,748, 7,090,843, 7,091,186, 7,097,840, 7,098,305, 7,098,308, 7,498,298, 7,375,078, 7,462,352, 7,553,816, 7,659,241, 7,662,387, 7,745,394, 7,754,681, 7,829,531, 7,837,980, 7,837,995, 7,902,338, 7,964,566, 7,964,567, 7,851,437, 7,994,135.
The benzodiazepine dimers (e g dimmers of pyrrolobenzodiazepine (PBD) or (tomaymycin), indolinobenzodiazepines, imidazobenzothiadiazepines, or oxazolidinobenzodiazepines) which are preferred cytotoxic agents according to the present invention are exampled in the art: U.S. Pat. Nos. 8,163,736; 8,153,627; 8,034,808; 7,834,005; 7,741,319; 7,704,924; 7,691,848; 7,678,787; 7,612,062; 7,608,615; 7,557,099; 7,528,128; 7,528,126; 7,511,032; 7,429,658; 7,407,951; 7,326,700; 7,312,210; 7,265,105; 7,202,239; 7,189,710; 7,173,026; 7,109,193; 7,067,511; 7,064,120; 7,056,913; 7,049,311; 7,022,699; 7,015,215; 6,979,684; 6,951,853; 6,884,799; 6,800,622; 6,747,144; 6,660,856; 6,608,192; 6,562,806; 6,977,254; 6,951,853; 6,909,006; 6,344,451; 5,880,122; 4,935,362; 4,764,616; 4,761,412; 4,723,007; 4,723,003; 4,683,230; 4,663,453; 4,508,647; 4,464,467; 4,427,587; 4,000,304; US patent appl. 20100203007, 20100316656, 20030195196.
Analogues and derivatives of the cytotoxic drugs/agents described in the present patent can be conjugated via a charged linker of the present patent. One skilled in the art of drugs/cytotoxic agents will readily understand that each of the drugs/cytotoxic agents described herein can be modified in such a manner that the resulting compound still retains the specificity and/or activity of the starting compound. The skilled artisan will also understand that many of these compounds can be used in place of the drugs/cytotoxic agents described herein. Thus, the drugs/cytotoxic agents of the present invention include analogues and derivatives of the compounds described herein.
All references cited herein and in the examples that follow are expressly incorporated by reference in their entireties.
The invention is further described in the following examples, which are not intended to limit the scope of the invention. Cell lines described in the following examples were maintained in culture according to the conditions specified by the American Type Culture Collection (ATCC) or Deutsche Sammlung von Mikroorganismen and Zellkulturen GmbH, Braunschweig, Germany (DMSZ), unless otherwise specified. Cell culture reagents were obtained from Invitrogen Corp., unless otherwise specified. All anhydrous solvents were commercially obtained and stored in Sure-seal bottles under nitrogen. All other reagents and solvents were purchased as the highest grade available and used without further purification. NMR spectra were recorded on Varian Mercury 300 MHz Instrument. Chemical shifts (.delta.) are reported in parts per million (ppm) referenced to tetramethylsilane at 0.00 and coupling constants (J) are reported in Hz. Low resolution mass spectral data were acquired on a Waters Micromass ZMD mass spec with Waters 2795 HPLC separations module and the 2996 photodiode array detector.
A mixture of ammonium hypophosphite (8.00 g, 96 mmol) and hexamethydisilazane (20.0 mL, 96 mmol) was heated at 120° C. for 1 h under argon. After the mixture was cooled to 0° C., ethyl acrylate (10.4 mL, 96 mmol) was carefully added dropwise, and the resulting mixture was stirred at 50° C. for 2 h. Then the mixture was cooled to room temperature, dibromoethane (40.0 mL) was added, and the mixture was stirred for 5 h at 120° C. The formed trimethylbromosilane and excess dibromoethane were removed under vacuum. Then 100 mL of aqueous ethanol (1:1) were added dropwise to the residue and refluxed for 0.5 h. Then the solvent was removed under vacuum and extracted with ethyl acetate. The organic layer was dried over magnesium sulfate and the solvent was removed under vacuum to give the title compound 4 (10.85 g, 41% yield). 1H NMR (300 MHz, CD3OD): δ 1.26 (t, J=7.1 Hz, 3H), 2.07 (m, 2H), 2.42 (m, 2H), 2.62 (m, 2H), 3.59 (m, 2H), 4.15 (q, J=7.1 Hz, 2H). 31P NMR (100 MHz, CD3OD): δ 49.5; ESI MS m/z−C7H13BrO4P (M-H), cacld. 271.98, found 271.97.
An amount of 10.84 g of 4 (20 mmol) was treated with 100.0 mL of triethyl orthoformate, and the mixture was refluxed with a Dean-Stark trap to remove ethanol and ethyl formate. Excess triethyl orthoformate was removed under vacuum to give 5 and 6 ([39.2:60.8 31P NMR ratio], 11.83 g). 6: 1H NMR (300 MHz, CD3OD) δ 1.27 (m, 611), 2.19 (n, 21-1), 2.57 (m, 2H), 4.11 (m, 4H), 6.36 (n, 3H). 31P NMR (100 MHz, CD3OD): δ 44.9; 5: 31P NMR (100 MHz, CD3OD) 8. 53.3; ESI MS m/z+, 5: 323.01 (M+Na), 6: 243.09 (M+Na).
The mixture of compound 5 and 6 (10.0 g, 38.4 mmol estimated from above ratio) in 100 ml of THF at 20° C. was added drop wise the mixture of thiolacetic acid (3.0 ml, 42.0 mmol) and DIPEA (8.5 ml, 48.9 mmol) in50 ml of dry THF in 1.5 hour. After 24 h under Ar, the mixture was concentrated, diluted with EtOAc/Hexane, washed with 1.0 M NaH2PO4, dried over MgSO4, filtered, evaporated, and SiO2 chromatographic purification (1:12 to 1:10 EtAc/Hexane) to afford the title compound 7 (10.01 g (88%% yield). ESI MS m/z+319.08 (M+Na).
Compound 7 (5.00 g, 16.89 mmol) in 100 ml of methanol was added 50 ml of 3 M NaOH. After being stirred under Ar for 3 h, the mixture was neutralized with 3 M H3PO4 to pH 7.2 under Ar. The mixture was added dropwise to the solution of 1,2-bis(5-nitropyridin-2-yl)disulfane (20.0 g, 64.4 mmol) in 200 ml of methanol. After being stirred for 4 h under Ar, the mixture was concentrated, diluted with EtOAc/Hexane (1:1), separated, and the organic layer was washed with pure water (3×25 ml) while the generated each of aqueous layer was washed with EtOAc/Hexane (1:1, 35 ml). The aqueous layers were combined, acidified with HCl/HOAc to pH 3˜4, concentrated to ˜1.0 ml, diluted with MeCN (60 ml), sonicated (or quickly stirred) for 1 h, filtered, washed the pellet with water/MeCN (1:10). The solution was then concentrated and purified on a SiO2 column eluted with water/MeCN/HOAc (1:10:0.01), pooled the fraction, added DMF (˜5 ml), evaporated to dryness to afford the title compound 8 (4.41 g, 85% yield). ESI MS, m/z−306.01 (M-H).
Compound 8 (2.20 g, 7.16 mmol) in DMA (50 ml) was added 0.2 ml of HCl (cone) and the mixture was evaporated to dryness. Then the compound redissolved in dry DMA (60 ml) was added, NHS (0.90 g, 7.82 mmol) and EDC (3.00 g, 15.62 mmol). The mixture was stirred under Ar overnight, evaporated and purified on SiO2 chromatography eluted with 4:1:1% Acetone/DCM/HOAc, pooled the fractions, evaporated and solidified with EtOH/Tol/Hexane to afford the title compound (2.11 g, 73% yield). 1H NMR (DMSO-d6, 300 MHz) 8.39 (dd, 1171, J=3.5, 4.7 Hz), 7.84 (m, 2H), 7.24 (in, 1H), 2.93˜2.89 (m, 2H), 2.74 (s, 4H), 2.41˜2.37 (m, 2H), 2.09˜2.03 (m, 4H); MS m/z−403.2 (M-H).
Maleimide (10.0 g, 103.0 mmol) in ethylether (350 ml) was added furan (11.0 ml, 151.2 mmol). The mixture was heated inside a 1 L of autoclave bomb at 100° C. for 8 h. The bomb was cooled down to room temperature, and the inside solid was rinsed with methanol, concentrated and crystallized in ethyl acetate/hexane to afford 16.9 g (99%) of the title compound. 1H NMR (DMF-d7, 300 MHz): 11.06 (s, 1H) (NH), 6.61 (m, 2H), 5.15 (m, 2H), 2.97 (m, 2H). 13C NMR 178.86, 137.72, 82.05, 49.93. MS m/z+188.4 (M+Na). Example 7. Ethyl ((3, 6-endoxo-Δ-tetrahydrophthalido)-ethyl)(ethoxy)phosphoryl)propanoate or ethyl 34(24(3aR,4R,7S)-1,3-dioxo-3a,4,7,7a-tetrahydro-1H-4,7-epoxyisoindo1-2(3H)-yl)ethyl)(ethoxy)phosphoryl)propanoate (16)
3, 6-Endoxo-Δ-tetrahydrophthalhide (2.40 g, 14.55 mmol) in DMA (60 ml) was added K2CO3 (4.2 g, 30.39 mmol) and KI (0.40 g, 3.45 mmol). After stirring under Ar for 1 hr, the mixture of compound 5 and 6 (2.6 g, 10.0 mmol estimated from above ratio) in DMA (10 ml) was added. The mixture was stirred under Ar overnight, evaporated, re-dissolved in EtOAc (100 me, washed with water (2×50 ml) and 1.0 M NaH2PO4 (2×50 nil), dried over Na2SO4, filtered, evaporated and purified by SiO2 chromatography and eluted with EtOAc/hexane (1:15-1:8) to afford the title compound (3.11 g, 81% yield). ESI MS m/z+408.20 (M+Na).
Compound 16 (3.00 g, 7.79 mmol), in the mixture of DMA (20 ml), toluene (20 ml) and HCl (8N,10 ml) was heated at 120-140° C. for 8 h. During the reaction time, 5×10 ml of water was gradually added to keep the reaction volume around 40 ml. The mixture was concentrated and purified by SiO2 chromatography eluted with (1:10:0.01 to 1:8:0.01) water/CH3CN/HOAc to afford the title compound (1.55 g, 76% yield). ESI MS m/z−260.10 (M-H).
Compound 17 (1.50 g, 5.74 mmol) in DMA (50 ml) was added 0.1 ml of HCl (cone) and the mixture was evaporated to dryness. Then the compound redissolved in dry DMA (40 ml) was added, NHS (0.71 g, 6.01 mmol) and EDC (3.00 g, 15.62 mmol). The mixture was stirred under Ar overnight, evaporated and purified on SiO2 chromatography eluted with 4:1:1% Acetone/DCM/HOAc, pooled the fractions, evaporated and solidified with EtOH/Tol/Hexane to afford the title compound (1.56 g, 76% yield). ESI MS m/z−357.20 (M-H).
A mixture of ammonium hypophosphite (8.00 g, 96 mmol) and hexamethydisilazane (20.0 mL, 96 mmol) was heated at 120° C. for 1 h under argon. After the mixture was cooled to 0° C., ethyl acrylate (10.4 mL, 96 mmol) was carefully added dropwise, and the resulting mixture was stirred at 50° C. for 2 h. Then the mixture was cooled to room temperature, 1,3-dibromobutane (40.0 mL) was added, and the mixture was stirred for 5 h at 120° C. The formed trimethylbromosilane was removed under vacuum. Then 100 mL of aqueous ethanol (1:1) were added dropwise to the residue, refluxed for 0.5 h and then concentrated under vacuum. The mixture was diluted with water, carefully neutralized to pH 7 with 0.1 NI NaOH, extracted with hexane (2×80 ml). The aqueous layer was acidified to pH ˜3, extracted with ethyl acetate (3×80 ml). The organic layer (EtOAc) was dried over magnesium sulfate, concentrated under vacuum, and purified on SiO2 eluted with acetone/CH2Cl2 (4:1) to give the title compound 29 (12.38 g, 43% yield). ESI MS m/z−299.20 (M H).
Compound 29 (12.20 g, 40.66 mmol) was treated with 100.0 mL of triethyl orthoformate, and the mixture was refluxed with a Dean-Stark trap to remove ethanol and ethyl formate. Excess triethyl orthoformate was removed under vacuum to give 30 and 31 ([21.8:79.2 31P NMR ratio], 10.13 g, 93% yield). ESI MS m/z+30: cacld. for C11H22BrNaO4P 351.04, found 351.20; 31: cacld for C11H21NaO4P 271.12, found 271.20.
The mixture of compound 30 and 31 (6.1 g, 22.76 mmol estimated from above ratio) in 100 ml of THF was added drop wise the mixture of thiolacetic acid (3.0 ml, 42.0 mmol) and DIPEA (8.5 ml, 48.9 mmol) in 50 ml of THF. After stirred at 50° C. under Ar for 24 h, the mixture was concentrated, diluted with EtAc/Hexane, washed with 1.0 M NaH2PO4, dried over MgSO4, filtered, evaporated, and SiO2 chromatographic purification (1:12 to 1:10 EtAc/Hexane) to afford the title compound 32 (5.23 g, 71% yield). ESI MS adz+347.20 (M+Na).
Compound 32 (5.20 g, 16.04 mmol) in 100 ml of methanol was added 50 ml of 3 M NaOH. After being stirred under Ar for 3 h, the mixture was neutralized with 3 M H3PO4 to pH 7.2 under Ar. The mixture was added dropwise to the solution of 1,2-bis(5-nitropyridin-2-yl)disulfane (20.0 g, 64.4 mmol) in 200 ml of methanol. After being stirred for 4 h under Ar, the mixture was concentrated, diluted with EtOAc/Hexane (1:1), separated, and the organic layer was washed with pure water (3×25 ml) while the generated each of aqueous layer was washed with EtOAc/Hexane (1:1, 35 ml). The aqueous layers were combined, acidified with HCl/HOAc to pH 3˜4, concentrated to ˜10 ml, diluted with MeCN (60 ml), sonicated (or quickly stirred) for 1 h, filtered, washed the pellet with water/MeCN (1:10). The solution was then concentrated and purified on a SiO2 column eluted with water/MeCN/HOAc (1:10:0.01), pooled the fraction, added DMF (˜5 ml), evaporated to dryness to afford the title compound 33 (4.40 g, 81% yield). ESI MS, m/z−334.10 (M-H).
Compound 33 (2.20 g, 6.56 mmol) in DMA (50 ml) was added 0.2 ml of HCl (cone) and the mixture was evaporated to dryness. Then the compound redissolved in dry DMA (60 ml) was added NHS (0.85 g, 7.39 mmol) and EDC (3.00 g, 15.62 mmol). The mixture was stirred under Ar overnight, evaporated and purified on SiO2 chromatography eluted with 4:1:1% Acetone/DCM/HOAc, pooled the fractions, evaporated and solidified with EtOH/Tol/Hexane to afford the title compound (L98 g, 70% yield). ESI MS m/z−431.2 (M-H).
A mixture of ammonium hypophosphite (8.00 g, 96 mmol) and hexamethydisilazane (20.0 mL, 96 mmol) was heated at 120° C. for 1 h under argon. After the mixture was cooled to 0° C., ethyl acrylate (10.4 mL, 96 mmol) was carefully added dropwise, and the resulting mixture was stirred at 50° C. for 2 h. Then the mixture was cooled to room temperature, 1,3-dibromo-3-methylbutane (44.0 mL) was added, and the mixture was stirred for 5 h at 120° C. The formed trimethylbromosilane was removed under vacuum. Then 100 mL of aqueous ethanol (1:1) were added dropwise to the residue, refluxed for 0.5 h and then concentrated under vacuum. The mixture was diluted with water, carefully neutralized to pH 7 with 0.1 M NaOH, extracted with hexane (2×80 ml). The aqueous layer was acidified to pH ˜3, extracted with ethyl acetate (3×80 ml). The organic layer (EtOAc) was dried over sodium sulfate, concentrated under vacuum, and purified on SiO2 eluted with acetone/CH2Cl2 (4:1) to give the title compound 37 (12.95 g, 43% yield). ESI MS m/z−313.10 (M H).
Compound 37 (12.90 g, 41.07 mmol) was treated with 100.0 mL of triethyl orthoformate, and the mixture was refluxed with a Dean-Stark trap to remove ethanol and ethyl formate. Excess triethyl orthoformate was removed under vacuum to give 38 and 39 ([12.5:87.5 31P NMR ratio], 9.89 g, 89% yield). ESI MS m/z+38: cacid. for C12H24BrNaO4P 365.06, found 365.10; 39: cacld for C12H23NaO4P 283.13, found 285.20.
The mixture of compound 38 and 39 (8.5 g, 31.25 mmol estimated from above ratio) in 100 ml of THF was added drop wise the mixture of thiolacetic acid (5.0 ml, 70.0 mmol) and DIPEA (12.5 ml, 71.9 mmol) in 80 ml of THF. After stirred at 70° C. under Ar for 35 h, the mixture was concentrated, diluted with EtAc/Hexane, washed with 1.0 M NaH2PO4, dried over MgSO4, filtered, evaporated, and SiO2 chromatographic purification (1:12 to 1:10 EtAc/Hexane) to afford the title compound 40 (6.55. g, 62% yield). ESI MS m/z+361.20 (M+Na).
Compound 40 (6.50 g, 19.22 mmol) in 100 ml of methanol was added 50 ml of 3 M NaOH, After being stirred under Ar for 3 h, the mixture was neutralized with 3 M H3PO4 to pH 7.2 under Ar. The mixture was added dropwise to the solution of 1,2-bis(5-nitropyridin-2-yl)disulfane (20.0 g, 64.4 mmol) in 200 ml of methanol. After being stirred for 24 h under Ar, the mixture was concentrated, diluted with EtOAc/Hexane (1:1), separated, and the organic layer was washed with pure water (3×25 ml) while the generated each of aqueous layer was washed with EtOAc/Hexane (1:1, 35 ml). The aqueous layers were combined, acidified with HCl/HOAc to pH 3˜4, concentrated to ˜10 diluted with MeCN (60 ml), sonicated (or quickly stirred) for 1 h, filtered, washed the pellet with water/MeCN (1:10). The solution was then concentrated and purified on a SiO2 column eluted with water/MeCN/HOAc (1:10:0.01), pooled the fraction, added DMF (˜5 ml), evaporated to dryness to afford the title compound 41 (5.16 g, 77% yield). ESI MS, m/z−348.12 (M-H).
Compound 41(2.10 g, 6.01 mmol) in DMA (50 ml) was added 0.2 ml of HCl (conc) and the mixture was evaporated to dryness. Then the compound redissolved in dry DMA (60 ml) was added NHS (0.80 g, 6.96 mmol) and EDC (3.00 g, 15.6:2 mmol). The mixture was stirred under Ar overnight, evaporated and purified on SiO2 chromatography eluted with 4:1:1% Acetone/DCM/HOAc, pooled the fractions, evaporated and solidified with EtOH/Tol/Hexane to afford the title compound (1.93 g, 72% yield). ESI MS m/z−445.10 (M-H).
The antiHer2 antibody is modified with phosphinate linker at 8 mg/mL antibody, a 10 fold molar excess of phosphinate linker (˜30 mM stock solution in DMA). The reaction is carried out in 100 mM NaH2PO4, pH7.4 buffer with DMA (5% v/v) for 15, 30, 60, 120, and 240 minutes at 25° C. The modified antiHer2 was purified by G25 column with 50 mM NaH2PO4, 50 mM NaCl, and 2 mM EDTA, pH6.5 to remove the excess phosphinate linker.
A phosphinate linker containing thiopyridine (SPP) linker was dissolved in DMA at a concentration of approximately 10 mM. An antibody was dialyzed into buffer A (50 mM NaH2PO4, 50 mM NaCl, 2 mM EDTA, pH 6.5). For the linker reaction, the antibody was at 8 mg/ml, and 4 equivalents of linker were added while stirring in the presence of 5% (v/v) DMA. The reaction was allowed to proceed at ambient temperature for 90 minutes. Unreacted linker was removed from the antibody by Sephadex G25 gel filtration using a Sephadex G25 column equilibrated with Buffer A at pH 6.5 or 150 mM potassium phosphate buffer containing 100 mM NaCl, pH 7.4 as indicated. For the SPP containing linker, the extent of modification was assessed by release of pyridine-2-thione using 50 mM DTT and measuring the absorbance at 343 nm as described below (ε343=8080 M−1 CM−1 for free pyridine-2-thione). For the conjugation reaction, thiol-containing drug (such tubulysin TZ041) was dissolved in DMA (N, N-dimethylacetamide) at a concentration of approximately 10 mM. The drug (1-1.5-fold molar excess relative to the number of linker molecules per antibody as indicated) was slowly added with stirring to the antibody which was at a concentration of 2.5 mg/ml in buffer A (pH 6.5 or pH 7.4) in a final concentration of 3% (v/v) DMA. The reaction was allowed to proceed at ambient temperature for the indicated times. Drug-conjugated antibody was purified using a Sephadex G25 column equilibrated with buffer B (PBS (NaH2PO4), pH 6.5). The extent of drug conjugation to antibody was assessed by measuring A254 and A280 of the conjugate.
The targeted cells (e.g. Ramos cells, 20,000 cells) were cultured in the presence of various concentrations of unconjugated antibody or the antibody conjugate for 96 hours after which cell viability was measured by propidium iodide exclusion and analyzed by flow cytometry using a Becton Dickinson FACSort (Becton Dickinson, Franklin Lakes, N.J.). Red fluorescent intensity (emission at 617 nm in the FL2 channel) of the cells excited at 488 nm was measured. The regions for viable cells were also set using both the forward light scatter and right-angle light scatter properties of the cells. The loss of viability was determined by the loss of cells from within the gated region defining viable cells. The average number of viable cells per 6 replicate cultures was calculated. The survival fraction was plotted versus conjugate concentration to determine the IC50 value (50% cell killing concentration) of the conjugate.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/CN2014/072769 | 2/28/2014 | WO | 00 |
| Publishing Document | Publishing Date | Country | Kind |
|---|---|---|---|
| WO2015/127685 | 9/3/2015 | WO | A |
| Number | Name | Date | Kind |
|---|---|---|---|
| 4000304 | Wu et al. | Dec 1976 | A |
| 4150584 | Theijsmeijer | Apr 1979 | A |
| 4169888 | Hanka et al. | Oct 1979 | A |
| 4256746 | Miyashita et al. | Mar 1981 | A |
| 4294757 | Asai | Oct 1981 | A |
| 4307016 | Asai et al. | Dec 1981 | A |
| 4313946 | Powell et al. | Feb 1982 | A |
| 4315929 | Freedman et al. | Feb 1982 | A |
| 4322348 | Asai et al. | Mar 1982 | A |
| 4331598 | Hasegawa et al. | May 1982 | A |
| 4341761 | Ganfield et al. | Jul 1982 | A |
| 4361650 | Asai et al. | Nov 1982 | A |
| 4362663 | Kida et al. | Dec 1982 | A |
| 4364866 | Asai et al. | Dec 1982 | A |
| 4371533 | Akimoto et al. | Feb 1983 | A |
| 4391904 | Litman et al. | Jul 1983 | A |
| 4399121 | Albarella et al. | Aug 1983 | A |
| 4414205 | Pettit | Nov 1983 | A |
| 4424219 | Hashimoto et al. | Jan 1984 | A |
| 4427587 | Kaneko et al. | Jan 1984 | A |
| 4427783 | Newman et al. | Jan 1984 | A |
| 4444887 | Hoffmann | Apr 1984 | A |
| 4450254 | Isley et al. | May 1984 | A |
| 4451570 | Royston et al. | May 1984 | A |
| 4464467 | Hatori et al. | Aug 1984 | A |
| 4466917 | Nussenzweig et al. | Aug 1984 | A |
| 4472500 | Milstein et al. | Sep 1984 | A |
| 4491632 | Wands et al. | Jan 1985 | A |
| 4493795 | Nestor, Jr. et al. | Jan 1985 | A |
| 4493890 | Morris | Jan 1985 | A |
| 4508647 | Hatori et al. | Apr 1985 | A |
| 4563304 | Carlsson et al. | Jan 1986 | A |
| 4663453 | Glamkowski et al. | May 1987 | A |
| 4671958 | Rodwell et al. | Jun 1987 | A |
| 4680338 | Sundoro | Jul 1987 | A |
| 4683230 | Tsunakawa et al. | Jul 1987 | A |
| 4723003 | Glamkowski et al. | Feb 1988 | A |
| 4723007 | Glamkowski et al. | Feb 1988 | A |
| 4753894 | Frankel et al. | Jun 1988 | A |
| 4761412 | Glamkowski et al. | Aug 1988 | A |
| 4764368 | Blattler et al. | Aug 1988 | A |
| 4764616 | Glamkowski et al. | Aug 1988 | A |
| 4816444 | Pettit et al. | Mar 1989 | A |
| 4816567 | Cabilly et al. | Mar 1989 | A |
| 4879278 | Pettit et al. | Nov 1989 | A |
| 4912227 | Kelly et al. | Mar 1990 | A |
| 4923990 | Nakano et al. | May 1990 | A |
| 4935362 | Tsunakawa et al. | Jun 1990 | A |
| 4943628 | Rosen et al. | Jul 1990 | A |
| 4952394 | Senter | Aug 1990 | A |
| 4956303 | Self | Sep 1990 | A |
| 4970198 | Lee et al. | Nov 1990 | A |
| 4975278 | Senter et al. | Dec 1990 | A |
| 4978744 | Pettit et al. | Dec 1990 | A |
| 4978757 | Kelly et al. | Dec 1990 | A |
| 4994578 | Ohba et al. | Feb 1991 | A |
| 5006651 | Broadhurst | Apr 1991 | A |
| 5037993 | Ohba et al. | Aug 1991 | A |
| 5053394 | Ellestad et al. | Oct 1991 | A |
| 5070092 | Kanda et al. | Dec 1991 | A |
| 5084468 | Saito et al. | Jan 1992 | A |
| 5094848 | Brixner | Mar 1992 | A |
| 5101038 | Nakano et al. | Mar 1992 | A |
| 5106951 | Morgan, Jr. et al. | Apr 1992 | A |
| 5108912 | Lee et al. | Apr 1992 | A |
| 5117006 | Saito et al. | May 1992 | A |
| 5122368 | Greenfield et al. | Jun 1992 | A |
| 5137877 | Kaneko et al. | Aug 1992 | A |
| 5138059 | Takahashi et al. | Aug 1992 | A |
| 5141648 | Hylarides et al. | Aug 1992 | A |
| 5146064 | Poirier | Sep 1992 | A |
| 5147786 | Feng et al. | Sep 1992 | A |
| 5165923 | Thorpe et al. | Nov 1992 | A |
| 5169774 | Frankel et al. | Dec 1992 | A |
| 5177016 | Balsari et al. | Jan 1993 | A |
| 5187186 | Kanda et al. | Feb 1993 | A |
| 5208020 | Chari et al. | May 1993 | A |
| 5208323 | Page et al. | May 1993 | A |
| 5223409 | Ladner et al. | Jun 1993 | A |
| 5225539 | Winter | Jul 1993 | A |
| 5264586 | Nicolaou et al. | Nov 1993 | A |
| 5286637 | Veronese et al. | Feb 1994 | A |
| 5288514 | Ellman | Feb 1994 | A |
| 5324483 | Cody et al. | Jun 1994 | A |
| 5332740 | Saito et al. | Jul 1994 | A |
| 5332837 | Kelly et al. | Jul 1994 | A |
| 5334528 | Stanker et al. | Aug 1994 | A |
| 5384412 | Nicolaou et al. | Jan 1995 | A |
| 5403484 | Ladner et al. | Apr 1995 | A |
| 5410024 | Pettit et al. | Apr 1995 | A |
| 5414064 | Lux et al. | May 1995 | A |
| 5416064 | Chari et al. | May 1995 | A |
| 5427908 | Dower et al. | Jun 1995 | A |
| 5436149 | Barnes | Jul 1995 | A |
| 5475011 | Ojima et al. | Dec 1995 | A |
| 5475092 | Chari et al. | Dec 1995 | A |
| 5495009 | Matteucci et al. | Feb 1996 | A |
| 5521284 | Pettit et al. | May 1996 | A |
| 5530097 | Pettit et al. | Jun 1996 | A |
| 5530101 | Queen et al. | Jun 1996 | A |
| 5543390 | Yatvin et al. | Aug 1996 | A |
| 5545806 | Lonberg et al. | Aug 1996 | A |
| 5547667 | Angelucci et al. | Aug 1996 | A |
| 5554725 | Pettit | Sep 1996 | A |
| 5563250 | Hylarides et al. | Oct 1996 | A |
| 5569825 | Lonberg et al. | Oct 1996 | A |
| 5571698 | Ladner et al. | Nov 1996 | A |
| 5573922 | Hoess et al. | Nov 1996 | A |
| 5580717 | Dower et al. | Dec 1996 | A |
| 5585089 | Queen et al. | Dec 1996 | A |
| 5585499 | Chari et al. | Dec 1996 | A |
| 5587161 | Burke et al. | Dec 1996 | A |
| 5595499 | Zander et al. | Jan 1997 | A |
| 5599902 | Pettit et al. | Feb 1997 | A |
| 5606017 | Willner et al. | Feb 1997 | A |
| 5606040 | McGahren et al. | Feb 1997 | A |
| 5622929 | Willner et al. | Apr 1997 | A |
| 5625126 | Lonberg et al. | Apr 1997 | A |
| 5629197 | Ring et al. | May 1997 | A |
| 5629430 | Terashima et al. | May 1997 | A |
| 5633425 | Lonberg et al. | May 1997 | A |
| 5635483 | Pettit et al. | Jun 1997 | A |
| 5641780 | Amishiro et al. | Jun 1997 | A |
| 5654399 | Sakakibara et al. | Aug 1997 | A |
| 5660829 | Burke et al. | Aug 1997 | A |
| 5661016 | Lonberg et al. | Aug 1997 | A |
| 5663149 | Pettit et al. | Sep 1997 | A |
| 5665860 | Pettit et al. | Sep 1997 | A |
| 5679700 | Caldwell | Oct 1997 | A |
| 5686237 | Al-Bayati | Nov 1997 | A |
| 5693762 | Queen et al. | Dec 1997 | A |
| 5703080 | Nakakura et al. | Dec 1997 | A |
| 5708146 | Willner et al. | Jan 1998 | A |
| 5712374 | Kuntsmann et al. | Jan 1998 | A |
| 5714586 | Kuntsmann et al. | Feb 1998 | A |
| 5728849 | Bouchard et al. | Mar 1998 | A |
| 5739116 | Hamann et al. | Apr 1998 | A |
| 5739350 | Kelly et al. | Apr 1998 | A |
| 5741892 | Barlozzari et al. | Apr 1998 | A |
| 5767236 | Kim et al. | Jun 1998 | A |
| 5767237 | Sakakibara et al. | Jun 1998 | A |
| 5770429 | Lonberg et al. | Jun 1998 | A |
| 5770701 | McGahren et al. | Jun 1998 | A |
| 5770710 | McGahren et al. | Jun 1998 | A |
| 5773001 | Hamann et al. | Jun 1998 | A |
| 5773435 | Kadow et al. | Jun 1998 | A |
| 5780588 | Pettit et al. | Jul 1998 | A |
| 5786377 | Garcia et al. | Jul 1998 | A |
| 5786486 | Fukuda et al. | Jul 1998 | A |
| 5789650 | Lonberg et al. | Aug 1998 | A |
| 5811452 | Ojima et al. | Sep 1998 | A |
| 5814318 | Lonberg et al. | Sep 1998 | A |
| 5821337 | Carter et al. | Oct 1998 | A |
| 5824805 | King et al. | Oct 1998 | A |
| 5840699 | Sakakibara et al. | Nov 1998 | A |
| 5846545 | Chari et al. | Dec 1998 | A |
| 5859205 | Adair et al. | Jan 1999 | A |
| 5874299 | Lonberg et al. | Feb 1999 | A |
| 5877296 | Hamann et al. | Mar 1999 | A |
| 5877397 | Lonberg et al. | Mar 1999 | A |
| 5880122 | Trybulski et al. | Mar 1999 | A |
| 5880270 | Beminger et al. | Mar 1999 | A |
| 5885793 | Griffiths et al. | Mar 1999 | A |
| 5922545 | Mattheakis et al. | Jul 1999 | A |
| 5939598 | Kucherlapati et al. | Aug 1999 | A |
| 5962216 | Trouet et al. | Oct 1999 | A |
| 5965537 | Ritter et al. | Oct 1999 | A |
| 5969108 | McCafferty et al. | Oct 1999 | A |
| 5985908 | Boger | Nov 1999 | A |
| 6004934 | Sakakibara et al. | Dec 1999 | A |
| 6015562 | Hinman et al. | Jan 2000 | A |
| 6033876 | Lemke et al. | Mar 2000 | A |
| 6034065 | Pettit et al. | Mar 2000 | A |
| 6048720 | Dalborg et al. | Apr 2000 | A |
| 6054297 | Carter et al. | Apr 2000 | A |
| 6054561 | Ring | Apr 2000 | A |
| 6060608 | Boger | May 2000 | A |
| 6066742 | Fukuda et al. | May 2000 | A |
| 6075181 | Kucherlapati et al. | Jun 2000 | A |
| 6103236 | Suzawa et al. | Aug 2000 | A |
| 6111166 | van de Winkel | Aug 2000 | A |
| 6114598 | Kucherlapati et al. | Sep 2000 | A |
| 6124310 | Denny et al. | Sep 2000 | A |
| 6124431 | Sakakibara et al. | Sep 2000 | A |
| 6130237 | Denny et al. | Oct 2000 | A |
| 6132722 | Siemers et al. | Oct 2000 | A |
| 6143721 | Janssen et al. | Nov 2000 | A |
| 6143901 | Dervan | Nov 2000 | A |
| 6146658 | Bosslet et al. | Nov 2000 | A |
| 6150584 | Kucherlapati et al. | Nov 2000 | A |
| 6162930 | Pinney et al. | Dec 2000 | A |
| 6162963 | Kucherlapati et al. | Dec 2000 | A |
| 6172197 | McCafferty et al. | Jan 2001 | B1 |
| 6180370 | Queen et al. | Jan 2001 | B1 |
| 6194612 | Boger et al. | Feb 2001 | B1 |
| 6207418 | Hori et al. | Mar 2001 | B1 |
| 6214345 | Firestone et al. | Apr 2001 | B1 |
| 6239104 | Pettit et al. | May 2001 | B1 |
| 6262271 | Boger | Jul 2001 | B1 |
| 6281354 | Boger | Aug 2001 | B1 |
| 6309646 | Lees | Oct 2001 | B1 |
| 6310209 | Boger | Oct 2001 | B1 |
| 6323315 | Pettit et al. | Nov 2001 | B1 |
| 6329497 | Boger | Dec 2001 | B1 |
| 6333410 | Chari et al. | Dec 2001 | B1 |
| 6340701 | Chari et al. | Jan 2002 | B1 |
| 6342219 | Thorpe et al. | Jan 2002 | B1 |
| 6342221 | Thorpe et al. | Jan 2002 | B1 |
| 6342480 | Trouet et al. | Jan 2002 | B1 |
| 6344451 | Steffan et al. | Feb 2002 | B1 |
| 6372738 | Chari et al. | Apr 2002 | B2 |
| 6391913 | Page et al. | May 2002 | B1 |
| 6407213 | Carter et al. | Jun 2002 | B1 |
| 6436931 | Chari et al. | Aug 2002 | B1 |
| 6441163 | Chari et al. | Aug 2002 | B1 |
| 6486326 | Boger | Nov 2002 | B2 |
| 6512101 | King et al. | Jan 2003 | B1 |
| 6521404 | Griffiths et al. | Feb 2003 | B1 |
| 6534660 | Yougxin et al. | Mar 2003 | B1 |
| 6544731 | Griffiths et al. | Apr 2003 | B1 |
| 6548530 | Boger | Apr 2003 | B1 |
| 6555313 | Griffiths et al. | Apr 2003 | B1 |
| 6555693 | Ge et al. | Apr 2003 | B2 |
| 6562806 | Thurston et al. | May 2003 | B1 |
| 6566336 | Sugiyama et al. | May 2003 | B1 |
| 6569834 | Pettit et al. | May 2003 | B1 |
| 6586618 | Zhao et al. | Jul 2003 | B1 |
| 6589979 | Bombardelli et al. | Jul 2003 | B2 |
| 6593081 | Griffiths et al. | Jul 2003 | B1 |
| 6596541 | Murphy et al. | Jul 2003 | B2 |
| 6596757 | Chari et al. | Jul 2003 | B1 |
| 6608192 | Thurston et al. | Aug 2003 | B1 |
| 6620911 | Pettit et al. | Sep 2003 | B1 |
| 6630579 | Chari et al. | Oct 2003 | B2 |
| 6639055 | Carter et al. | Oct 2003 | B1 |
| 6660856 | Wang | Dec 2003 | B2 |
| 6706708 | Chari et al. | Mar 2004 | B2 |
| 6716821 | Zhao et al. | Apr 2004 | B2 |
| 6747144 | Thurston et al. | Jun 2004 | B1 |
| 6756397 | Zhao et al. | Jun 2004 | B2 |
| 6759509 | King et al. | Jul 2004 | B1 |
| 6762179 | Cochran et al. | Jul 2004 | B2 |
| 6797492 | Daugherty et al. | Sep 2004 | B2 |
| 6800622 | Kamal et al. | Oct 2004 | B1 |
| 6884799 | Kamal et al. | Apr 2005 | B2 |
| 6884869 | Senter et al. | Apr 2005 | B2 |
| 6897034 | Bebbington et al. | May 2005 | B2 |
| 6909006 | Thurston et al. | Jun 2005 | B1 |
| 6913748 | Widdison | Jul 2005 | B2 |
| 6946272 | Powell | Sep 2005 | B1 |
| 6946455 | Sugiyama et al. | Sep 2005 | B2 |
| 6951853 | Kamal et al. | Oct 2005 | B1 |
| 6977254 | Failli et al. | Dec 2005 | B2 |
| 6979684 | Kamal et al. | Dec 2005 | B1 |
| 6989452 | Ng et al. | Jan 2006 | B2 |
| 7008942 | Chari et al. | Mar 2006 | B2 |
| 7015215 | Kamal et al. | Mar 2006 | B2 |
| 7022699 | Failli et al. | Apr 2006 | B2 |
| 7049311 | Thurston et al. | May 2006 | B1 |
| 7049316 | Zhao et al. | May 2006 | B2 |
| 7056913 | Kamal et al. | Jun 2006 | B2 |
| 7064120 | Failli et al. | Jun 2006 | B2 |
| 7067511 | Thurston et al. | Jun 2006 | B2 |
| 7071311 | Radka | Jul 2006 | B2 |
| 7087600 | Ng et al. | Aug 2006 | B2 |
| 7090843 | Francisco et al. | Aug 2006 | B1 |
| 7091186 | Senter et al. | Aug 2006 | B2 |
| 7097840 | Erickson et al. | Aug 2006 | B2 |
| 7098305 | Deghenghi et al. | Aug 2006 | B2 |
| 7098308 | Senter et al. | Aug 2006 | B2 |
| 7109193 | Failli et al. | Sep 2006 | B2 |
| 7115573 | Pickford et al. | Oct 2006 | B2 |
| 7129261 | Ng et al. | Oct 2006 | B2 |
| 7173026 | Kamal et al. | Feb 2007 | B2 |
| 7186851 | Baloglu | Mar 2007 | B2 |
| 7189710 | Kamal et al. | Mar 2007 | B2 |
| 7202239 | Failli et al. | Apr 2007 | B2 |
| 7214663 | Bebbington et al. | May 2007 | B2 |
| 7217819 | Chari et al. | May 2007 | B2 |
| 7223837 | De Groot et al. | May 2007 | B2 |
| 7265105 | Thurston et al. | Sep 2007 | B2 |
| 7276497 | Chari et al. | Oct 2007 | B2 |
| 7276499 | Chari et al. | Oct 2007 | B2 |
| 7301019 | Widdison et al. | Nov 2007 | B2 |
| 7303749 | Chari | Dec 2007 | B1 |
| 7304032 | Bebbington et al. | Dec 2007 | B2 |
| 7312210 | Kamal et al. | Dec 2007 | B2 |
| 7326700 | Failli et al. | Feb 2008 | B2 |
| 7329507 | Pickford et al. | Feb 2008 | B2 |
| 7329760 | Zhao et al. | Feb 2008 | B2 |
| 7368565 | Chari et al. | May 2008 | B2 |
| 7375078 | Feng | May 2008 | B2 |
| 7388026 | Zhao et al. | Jun 2008 | B2 |
| 7407951 | Thurston et al. | Aug 2008 | B2 |
| 7411063 | Widdison et al. | Aug 2008 | B2 |
| 7429658 | Howard et al. | Sep 2008 | B2 |
| 7462352 | Hansen et al. | Dec 2008 | B2 |
| 7498298 | Doronina et al. | Mar 2009 | B2 |
| 7498302 | Ng et al. | Mar 2009 | B2 |
| 7507420 | Ng et al. | Mar 2009 | B2 |
| 7511032 | Liu et al. | Mar 2009 | B2 |
| 7528126 | Howard et al. | May 2009 | B2 |
| 7528128 | Ahmed et al. | May 2009 | B2 |
| 7553816 | Senter et al. | Jun 2009 | B2 |
| 7557099 | Howard et al. | Jul 2009 | B2 |
| 7569358 | Salamone et al. | Aug 2009 | B2 |
| 7598290 | Miller et al. | Oct 2009 | B2 |
| 7608615 | Ahmed et al. | Oct 2009 | B2 |
| 7612062 | Gregson et al. | Nov 2009 | B2 |
| 7655660 | Zhao et al. | Feb 2010 | B2 |
| 7655661 | Zhao et al. | Feb 2010 | B2 |
| 7659241 | Senter et al. | Feb 2010 | B2 |
| 7662387 | Law et al. | Feb 2010 | B2 |
| 7667054 | Miller et al. | Feb 2010 | B2 |
| 7678787 | Failli et al. | Mar 2010 | B2 |
| 7691848 | Failli et al. | Apr 2010 | B2 |
| 7691962 | Boyd et al. | Apr 2010 | B2 |
| 7704924 | Thurston et al. | Apr 2010 | B2 |
| 7741319 | Howard et al. | Jun 2010 | B2 |
| 7745394 | Doronina et al. | Jun 2010 | B2 |
| 7754681 | Feng | Jul 2010 | B2 |
| 7754885 | Hoefle et al. | Jul 2010 | B2 |
| 7776814 | Domling et al. | Aug 2010 | B2 |
| 7803903 | Kratz | Sep 2010 | B2 |
| 7816377 | Domling et al. | Oct 2010 | B2 |
| 7829531 | Senter et al. | Nov 2010 | B2 |
| 7833992 | Vargeese | Nov 2010 | B2 |
| 7834005 | Liu et al. | Nov 2010 | B2 |
| 7837980 | Alley et al. | Nov 2010 | B2 |
| 7837995 | Goldenberg | Nov 2010 | B2 |
| 7843005 | Nowak | Nov 2010 | B2 |
| 7851432 | Chari et al. | Dec 2010 | B2 |
| 7851437 | Senter et al. | Dec 2010 | B2 |
| 7902338 | Hansen et al. | Mar 2011 | B2 |
| 7906545 | Zhao et al. | Mar 2011 | B2 |
| 7910594 | Vlahov et al. | Mar 2011 | B2 |
| 7937980 | Hessberger et al. | May 2011 | B2 |
| 7939434 | Tseng et al. | May 2011 | B2 |
| 7964566 | Doronina et al. | Jun 2011 | B2 |
| 7964567 | Doronina et al. | Jun 2011 | B2 |
| 7968586 | Gangwar et al. | Jun 2011 | B2 |
| 7989434 | Feng | Aug 2011 | B2 |
| 7994135 | Doronina et al. | Aug 2011 | B2 |
| 7999083 | Govindan et al. | Aug 2011 | B2 |
| 8012978 | Zhao et al. | Sep 2011 | B2 |
| 8034808 | Delavault et al. | Oct 2011 | B2 |
| 8053205 | Salamone et al. | Nov 2011 | B2 |
| 8084586 | Salamone et al. | Dec 2011 | B2 |
| 8153627 | Kamal et al. | Apr 2012 | B2 |
| 8153768 | Kunz et al. | Apr 2012 | B2 |
| 8163736 | Gauzy et al. | Apr 2012 | B2 |
| 8163888 | Steeves et al. | Apr 2012 | B2 |
| 8236319 | Chari et al. | Aug 2012 | B2 |
| 10131682 | Zhao | Nov 2018 | B2 |
| 20030195196 | Thurston et al. | Oct 2003 | A1 |
| 20040249130 | Stanton et al. | Dec 2004 | A1 |
| 20060022925 | Hara et al. | Feb 2006 | A1 |
| 20060074008 | Senter et al. | Apr 2006 | A1 |
| 20060217360 | Hoefle et al. | Sep 2006 | A1 |
| 20070041901 | Diener et al. | Feb 2007 | A1 |
| 20090088390 | Nishizawa | Apr 2009 | A1 |
| 20090176253 | Bieniarz et al. | Jul 2009 | A1 |
| 20090274713 | Chari | Nov 2009 | A1 |
| 20100041872 | DeFrees | Feb 2010 | A1 |
| 20100062008 | Senter et al. | Mar 2010 | A1 |
| 20100074840 | Griffiths et al. | Mar 2010 | A1 |
| 20100104589 | Govindan et al. | Apr 2010 | A1 |
| 20100136652 | Bieniarz et al. | Jun 2010 | A1 |
| 20100189727 | Rodeck et al. | Jul 2010 | A1 |
| 20100203007 | Li | Aug 2010 | A1 |
| 20100316656 | Bouchard et al. | Dec 2010 | A1 |
| 20100323973 | Leamon et al. | Dec 2010 | A1 |
| 20110021568 | Ellman et al. | Jan 2011 | A1 |
| 20110027274 | Cheng et al. | Feb 2011 | A1 |
| 20110064666 | Ogawa et al. | Mar 2011 | A1 |
| 20110064752 | Hutchinson et al. | Mar 2011 | A1 |
| 20110064753 | Senter et al. | Mar 2011 | A1 |
| 20110076722 | Takahashi | Mar 2011 | A1 |
| 20110134826 | Yang et al. | Jun 2011 | A1 |
| 20110263650 | Ellman et al. | Oct 2011 | A1 |
| 20110293513 | Govindan et al. | Dec 2011 | A1 |
| 20120034295 | Spiegel et al. | Feb 2012 | A1 |
| 20120082617 | Govindan et al. | Apr 2012 | A1 |
| 20120129779 | Richter | May 2012 | A1 |
| 20120165537 | Li | Jun 2012 | A1 |
| 20120201809 | Bhat et al. | Aug 2012 | A1 |
| 20130095123 | Lerchen et al. | Apr 2013 | A1 |
| 20150250896 | Zhao | Sep 2015 | A1 |
| 20160207949 | Zhao | Jul 2016 | A1 |
| 20170327486 | Li | Nov 2017 | A1 |
| 20190127399 | Zhao et al. | May 2019 | A1 |
| 20190127400 | Zhao et al. | May 2019 | A1 |
| 20190127401 | Zhao et al. | May 2019 | A1 |
| Number | Date | Country |
|---|---|---|
| 2 710 693 | Jan 2011 | CA |
| 101784565 | Jul 2010 | CN |
| 196 38 870 | Mar 1998 | DE |
| 100 08 089 | Oct 2001 | DE |
| 102 41 152 | Mar 2004 | DE |
| 102 54 439 | Jun 2004 | DE |
| 103 09 005 | Sep 2004 | DE |
| 10 2004 030 227 | Jan 2006 | DE |
| 2 174 947 | Apr 2010 | EP |
| 2000-511889 | Sep 2000 | JP |
| 2011-519864 | Jul 2011 | JP |
| 8910961 | Nov 1989 | WO |
| 0136003 | May 2001 | WO |
| 0138318 | May 2001 | WO |
| 02077036 | Oct 2002 | WO |
| WO-03001968 | Jan 2003 | WO |
| WO-03077834 | Sep 2003 | WO |
| 2004005269 | Jan 2004 | WO |
| 2004005326 | Jan 2004 | WO |
| 2004005327 | Jan 2004 | WO |
| 2005058367 | Jun 2005 | WO |
| WO-2005110455 | Nov 2005 | WO |
| 2006033913 | Mar 2006 | WO |
| 2006056464 | Jun 2006 | WO |
| 2006096754 | Sep 2006 | WO |
| 2007052169 | May 2007 | WO |
| 2008002993 | Jan 2008 | WO |
| 2008034019 | Mar 2008 | WO |
| 2008070291 | Jun 2008 | WO |
| 2008076333 | Jun 2008 | WO |
| 2008112873 | Sep 2008 | WO |
| 2008125116 | Oct 2008 | WO |
| 2008138561 | Nov 2008 | WO |
| 2009002993 | Dec 2008 | WO |
| 2009012958 | Jan 2009 | WO |
| 2009026177 | Feb 2009 | WO |
| 2009055562 | Apr 2009 | WO |
| 2009095447 | Aug 2009 | WO |
| 2009134279 | Nov 2009 | WO |
| 2009139863 | Nov 2009 | WO |
| 2010033733 | Mar 2010 | WO |
| 2010034724 | Apr 2010 | WO |
| 2012095347 | Jul 2012 | WO |
| 2012143495 | Oct 2012 | WO |
| 2014009774 | Jan 2014 | WO |
| 2014080251 | May 2014 | WO |
| WO-2014080251 | May 2014 | WO |
| Entry |
|---|
| C-H Lin et al., 4 Chemistry & Biology, 859-866 (1997). |
| CAS Abstract Di Sabato, J Am Chem Soc (1961). |
| G. Di Sabato et al., 83 Journal of the American Chemical Society, 4400-4405 (1961). |
| S.L. Shames et al., 83 Journal of the American Chemical Society, 6177-6184 (1981). |
| M. Borloo et al., 9 Synthesis, 1074-1076 (1995). |
| V. Kubí{hacek over (c)}ek et al., 20 Dalton Transactions, 3927-3938 (2003). |
| CAS Abstract of M. Borloo et al., 9 Synthesis, 1074-1076 (1995). |
| IUPAC. Compendium of Chemical Terminology, 2nd ed. “Azides” (the “Gold Book”) (1997) (Downloaded from http://goldbook.iupac.org/plain/A00555-plain.html on Feb. 17, 2019) (Year: 1997). |
| IUPAC. Compendium of Chemical Terminology, 2nd ed. “Ketones” (the “Gold Book”) (1997) (Downloaded from http://goldbook.iupac.org/html/K/K03386.html on Feb. 17, 2019) (Year: 1997). |
| Compound CAS Registry No. 371233-32-8 (2001) (Year: 2001). |
| J.L. Goulet et al., 4 Bioorganic & Medicinal Chemistry Letters (1994) (Year: 1994). |
| C.G. Caldwall et al., 6 Bioorganic & Medicinal Chemistry Letters (1996) (Year: 1996). |
| Adams et al., “Generating Improved Single-Chain Fv Molecules for Tumor Targeting,” Journal of Immunological Methods, (Dec. 10, 1999), vol. 231, Issue 1-2, pp. 249-260. |
| Afar et al., “Preclinical Validation of Anti-TMEFF2-Auristatin E-Conjugated Antibodies in the Treatment of Prostate Cancer,” Molecular Cancer Therapeutics, (Aug. 2004), vol. 3, No. 8, pp. 921-931. |
| Albrecht et al., “Monospecific Bivalent scFv-SH: Effects of Linker Length and Location of an Engineered Cysteine on Production, Antigen Binding Activity and Free SH Accessiblity,” Journal of Immunological Methods, (Mar. 20, 2006), vol. 310, Issue 1-2, pp. 100-116. |
| Alley et al., “Antibody-Drug Conjugates: Targeted Drug Delivery for Cancer,” Current Opinion in Chemical Biology, (Aug. 2010), vol. 14, Issue 4, pp. 529-537. |
| Almagro et al., “Humanization of Antibodies,” Frontiers in Bioscience, (Jan. 1, 2008), vol. 13, pp. 1619-1633. |
| Almutairi et al., “Biodegradable Dendritic Positron-Emitting Nanoprobes for the Noninvasive Imaging of Angiogenesis,” PNAS, (Jan. 20, 2009), vol. 106, No. 3, pp. 685-690. |
| Anderson et al., “Enhanced in Vitro Tumor Cell Retention and Internalization of Antibody Derivatized with Synthetic Peptides,” Bioconjugate Chemistry, (Jan. 1993), vol. 4, No. 1, pp. 10-18. |
| Antczak et al., “Influence of the Linker on the Biodistribution and Catabolism of Actinium-225 Self-Immolative Tumor-Targeted Isotope Generators,” Bioconjugate Chemistry, (Nov.-Dec. 2006), vol. 17, No. 6, pp. 1551-1560. |
| Aoki et al., “Design and Synthesis of a Photocleavable Biotin-Linker for the Photoisolation of Ligand-Receptor Complexes Based on the Photolysis of 8-Quinolinyl Sulfonates in Aqueous Solution,” Bioorganic & Medicinal Chemistry, (May 1, 2009), vol. 17, Issue 9, pp. 3405-3413. |
| Arpicco et al., “New Coupling Reagents for the Preparation of Disulfide Cross-Linked Conjugates with Increased Stability,” Bioconjugate Chemistry, (May-Jun. 1997), vol. 8, No. 3, pp. 327-337. |
| Austin et al. “Oxidizing Potential of Endosomes and Lysosomes Limits Intracellular Cleavage of Disulfide-Based Antiboby-Drug Conjugates,” PNAS, (Dec. 13, 2005), vol. 102, No. 50, pp. 17987-17992. |
| Balasubramanian et al., “Total Synthesis and Biological Evaluation of Tubulysin U, Tubulysin V, and Their Analogues,” Journal of Medicinal Chemistry, (Jan. 22, 2009), vol. 52, No. 2, pp. 238-240. |
| Barbour et al., “Stabilization of Chimeric BR96-Doxorubicin Immunoconjugate,” Pharmaceutical Research, (Feb. 1995), vol. 12, No. 2, pp. 215-222. |
| Beeson et al., “Conditionally Cleavable Radioimmunoconjugates: A Novel Approach for the Release of Radioisotopes from Radioimmunoconjugates,” Bioconjugate Chemistry, (Sep.-Oct. 2003), vol. 14, No. 5, pp. 927-933. |
| Bickel et al., “In Vivo Cleavability of a Disulfide-Based Chimeric Opioid Peptide in Rat Brain,” Bioconjugate Chemistry, (Mar.-Apr. 1995), vol. 6, No. 2, pp. 211-218. |
| Boger et al., “Parallel Synthesis and Evaluation of 132 (+)-1,2,9,9a-Tetrahydrocyclopropa[c]benz[e]indol-4-one (CBI) Analogues of CC-1065 and the Duocarmycins Defining the Contribution of the DNA-Binding Domain,” The Journal of Organic Chemistry, (Oct. 5, 2001), vol. 66, No. 20, pp. 6654-6661. |
| Brannigan et al., “Protein Engineering 20 Years On,” Nature Reviews Molecular Cell Biology, (Dec. 2002), vol. 3, No. 12, pp. 964-970. |
| Brich et al., “Preparation and Characterization of a Water Soluble Dextran Immunoconjugate of Doxorubicin and the Monoclonal Antibody (ABL 364),” Journal of Controlled Release, (Mar. 1992), vol. 19, Issues 1-3, pp. 245-257. |
| Burgess, “The Complex Mediators of Cell Growth and Differentiation,” Immunology Today, (Jun. 1984), vol. 5, No. 6, pp. 155-158. |
| Carlsson et al., “Protein Thiolation and Reversible Protein-Protein Conjugation. N-Succinimidyl 3-(2-pyridyldithio) Propionate, a new Heterobifunctional Reagent,” Biochemical Journal, (Oct. 1978), vol. 173, No. 3, pp. 723-737. |
| Chai et al., “Discovery of 23 Natural Tubulysins from Angiococcus disciformis an d48 and Cystobacter SBCb004,” Chemistry & Biology (Mar. 26, 2010), vol. 17, issue 3, pp. 296-309. |
| Chandrasekhar et al., “Toward Tubulysin: Gram-Scale Synthesis of Tubuvaline-Tubuphenylalanine Fragment,” The Journal of Organic Chemistry, (Dec. 18, 2009), vol. 74, No. 24, pp. 9531-9534. |
| Chen et al., “Antibody-Cytotoxic Agent Conjugates for Cancer Therapy,” Expert Opinion on Drug Delivery, (Sep. 2005), vol. 2, No. 5, pp. 873-890. |
| Chen et al., “Synthesis of Doxorubicin Conjugates Through Hydrazone Bonds to Melanotransferrin P97,” Synthetic Communications: An International Journal for Rapid Communication of Synthetic Organic Chemistry, (2003), vol. 33, No. 14, pp. 2377-2390. |
| Delprino et al., “Toxin-Targeted Design for Anticancer Therapy. I: Synthesis and Biological Evaluation of New Thioimidate Heterobifunctional Reagents,” Journal of Pharmaceutical Sciences, (May 1993), vol. 82, No. 5, pp. 506-512. |
| Dente et al., “Monoclonal Antibodies that Recognise Filamentous Phage: Tools for Phage Display Technology,” Gene, (Oct. 11, 1994), vol. 148, Issue 1, pp. 7-13. |
| Dhar et al., “Targeted Delivery of Cisplatin to Prostate Cancer Cells by Aptamer Functionalized Pt(IV) Prodrug-PLGA-PEG Nanoparticles,” PNAS, (Nov. 11, 2008), vol. 105, No. 45, pp. 17356-17361. |
| Dijoseph et al., “Antibody-Targeted Chemotherapy with CMC-544: a CD22-Targeted Immunoconjugate of Calicheamicin for the Treatment of B-Lymphoid Malignancies,” Blood, (Mar. 1, 2004), vol. 103, No. 5, pp. 1807-1814. |
| Domling et al., “Total Synthesis of Tubulysin U and V,” Angew Chem Int Ed Engl., (Nov. 6, 2006), vol. 45, No. 43, pp. 7235-7239. |
| Doronina et aL, “Enhanced Activity of Monomethylauristatin F through Monoclonal Antibody Delivery: Effects of Linker Technology on Efficacy and Toxicity,” Bioconjugate Chemistry, (Jan.-Feb. 2006), vol. 17, No. 1, pp. 114-124. |
| Doronina et al., “Novel Peptide Linkers for Highly Potent Antibody-Auristatin Conjugate,” Bioconjugate Chemistry, (Oct. 2008), vol. 19, No. 10, pp. 1960-1963. |
| Dulbecco et al., “A Plaque Assay for the Polyoma Virus,” Letters to the Editors, (1959), Virol. 8, pp. 396-397. |
| Ebner et al., “A New, Simple Method for Linking of Antibodies to Atomic Force Microscopy Tips,” Bioconjugate Chemistry, (Jul.-Aug. 2007), vol. 18, No. 4, pp. 1176-1184. |
| Erickson et al., “Tumor Delivery and In Vivo Processing of Disulfide-Linked and Thioether-Linked Antibody-Maytansinoid Conjugates,” Bioconjugate Chemistry, (Jan. 2010), vol. 21, No. 1, pp. 84-92. |
| Flenniken et al., “A Library of Protein Cage Architectures as Nanomaterials,” Viruses and Nanotechnology, (2009), vol. 327 of the series Current Topics in Microbiology and Immunology, pp. 71-93. |
| Frankel et al., “Cell Surface Receptor-Targeted Therapy of Acute Myeloid Leukemia: A Review,” Cancer Biotherapy & Radiopharmaceuticals, (Oct. 2000), vol. 15, No. 5, pp. 459-476. |
| Friestad et al., “Stereoselective Mn-Mediated Coupling of Functionalized Iodides and Hydrazones: A Synthetic Entry to the Tubulysin γ-Amino Acids,” Organic Letters, (Sep. 2004), vol. 6, No. 19, pp. 3249-3252. |
| Friestad et al., “Synthesis of γ-Amino Esters via Mn-Mediated Radical Addition to Chiral γ-Hydrazonoesters,” Organic Letters, (2009), vol. 11, No. 5, pp. 1095-1098. |
| Garsky et al., “The Synthesis of a Prodrug of Doxorubicin Designed to Provide Reduced Systemic Toxicity and Greater Target Efficacy,” Journal of Medicinal Chemistry, (Nov. 22, 2001), vol. 44, No. 24, pp. 4216-4224. |
| Geysen et al., “Small Peptides Induce Antibodies with a Sequence and Structural Requirement for Binding Antigen Comparable to Antibodies Raised Against the Native Protein,” PNAS, (Jan. 1985), vol. 82, No. 1, pp. 178-182. |
| Goff et al., “Substituted 2-Iminothiolanes: Reagents for the Preparation of Disulfide Cross-Linked Conjugates with Increased Stability,” Bioconjugate Chemistry, (Nov.-Dec. 1990), vol. 1, No. 6, pp. 381-386. |
| Greenwald et al., “Drug Delivery Systems Employing 1,4- or 1,6-Elimination: Poly(ethylene glycol) Prodrugs of Amine-Containing Compounds,” Journal of Medicinal Chemistry, (Sep. 9, 1999), vol. 42, No. 18, pp. 3657-3667. |
| Haenseler et al., “Activation of Methotrexate-α-Alanine by Carboxypeptidase A-Monoclonal Antibody Conjugate,” Biochemistry, (Jan. 28, 1992), vol. 31, No. 3, pp. 891-897. |
| Hamann et al., “An Anti-CD33 Antibody-Calicheamicin Conjugate for Treatment of Acute Myeloid Leukemia. Choice of Linker,” Bioconjugate Chemistry, (Jan.-Feb. 2002), vol. 13, No. 1, pp. 40-46. |
| Hamann et al., “An Anti-MUC1 Antibody-Calicheamicin Conjugate for Treatment of Solid Tumors. Choice of Linker and Overcoming Drug Resistance,” Bioconjugate Chemistry, (Mar.-Apr. 2005), vol. 16, No. 2, pp. 346-353. |
| Hamann et al., “A Calicheamicin Conjugate with a Fully Humanized Anti-MUC1 Antibody Shows Potent Antitumor Effects in Breast and Ovarian Tumor Xenografts,” Bioconjugate Chemistry, (Mar.-Apr. 2005), vol. 16, No. 2, pp. 354-360. |
| Hinman et al., “Preparation and Characterization of Monoclonal Antibody Conjugates of the Calicheamicins: A Novel and Potent Family of Antitumor Antibiotics,” Cancer Research, (Jul. 15, 1993), vol. 53, No. 14, pp. 3336-3342. |
| Houdebine, “Antibody Manufacture in Transgenic Animals and Comparisons with other Systems,” Current Opinion in Biotechnology, (Dec. 1, 2002), vol. 13, Issue 6, pp. 625-629. |
| Huse et al., “Generation of a Large Combinatorial Library of the Immunoglobulin Repertoire in Phage Lambda,” Science, (Dec. 8, 1989), vol. 246, No. 4935, pp. 1275-1281. Only Have p. 1275. |
| Javier et al., “Aptamer-Targeted Gold Nanoparticles As Molecular-Specific Contrast Agents for Reflectance Imaging,” Bioconjugate Chemistry, (Jun. 2008), vol. 19, No. 6, pp. 1309-1312. |
| Jeffrey et al., “Design, Synthesis, and in Vitro Evaluation of Dipeptide-Based Antibody Minor Groove Binder Conjugates,” Journal of Medicinal Chemistry, (Mar. 10, 2005), vol. 48, No. 5, pp. 1344-1358. |
| Johnson et al., “Induction of Immunogenicity of Monoclonal Antibodies by Conjugation with Drugs,” Cancer Research, (Oct. 15, 1991), vol. 15, No. 20, pp. 5774-5776. |
| Jones et al., “Conjugates of Double-Stranded Oligonucleotides with Poly(ethylene glycol) and Keyhole Limpet Hemocyanin: A Model for Treating Systemic Lupus Erythematosus,” Bioconjugate Chemistry, (Sep.-Oct. 1994), vol. 5, No. 5, pp. 390-399. |
| Jones et al., “Multivalent Thioether-Peptide Conjugates: B Cell Tolerance of an Anti-Peptide Immune Response,” Bioconjugate Chemistry, (May-Jun. 1999), vol. 10, No. 3, pp. 480-488. |
| Jones et al., “Synthesis of LJP 993, a Multivalent Conjugate of the N-Terminal Domain of β2GPI and Suppression of an Anti-β2GPI Immune Response,” Bioconjugate Chemistry, (Nov.-Dec. 2001), vol. 12, No. 6, pp. 1012-1020. |
| Kashef et al., “Synthesis and Characterization of Pseudomonas Aeruginosa Alginate-Tetanus Toxoid Conjugate,” Journal of Medical Microbiology, (Oct. 2006), vol. 55, Pt. 10, pp. 1441-1446. |
| Kellogg et al., “Disulfide-Linked Antibody-Maytansinoid Conjugates: Optimization of In Vivo Activity by Varying the Steric Hindrance at Carbon Atoms Adjacent to the Disulfide Linkage,” Bioconjugate Chemistry, (Apr. 20, 2011), vol. 22, No. 4, pp. 717-727. |
| Kelly et al., “An Antibody-Cytotoxic Conjugate, BIIB015, is a new Targeted Therapy for Cripto Positive Tumours,” European Journal of Cancer, (Jul. 2011), vol. 47, No. 11, pp. 1736-1746. |
| Kim et al., “C-2 Modified Taxol Analogs with Improved Aqueous Solubility,” Bulletin of the Korean Chemical Society, (1999), vol. 20, No. 12, pp. 1389-1390. |
| King et al., “Monoclonal Antibody Conjugates of Doxorubicin Prepared with Branched Linkers: A Novel Method for Increasing the Potency of Doxorubicin Immunoconjugates,” Bioconjugate Chemistry, (Mar.-Apr. 1999), vol. 10, No. 2, pp. 279-288. |
| King et al., “Monoclonal Antibody Conjugates of Doxorubicin Prepared with Branched Peptide Linkers: Inhibition of Aggregation by Methoxytriethyleneglycol Chains,” Journal of Medicinal Chemistry, (2002), vol. 45, No. 19, pp. 4336-4343. |
| Klussman et al., “Secondary mAb-vcMMAE Conjugates Are Highly Sensitive Reporters of Antibody Internalization via the Lysosome Pathway,” Bioconjugate Chemistry, (2004), vol. 15, No. 4, pp. 765-773. |
| Kovar et al., “Star Structure of Antibody-Targeted HPMA Copolymer-Bound Doxorubicin: A Novel Type of Polymeric Conjugate for Targeted Drug Delivery with Potent Antitumor Effect,” Bioconjugate Chemistry, (2002), vol. 13, No. 2, pp. 206-215. |
| Kratz et al., “Preparation, Characterization and in Vitro Efficacy of Albumin Conjugates of Doxorubicin,” Biological & Pharmaceutical Bulletin, (1998), vol. 21, No. 1, pp. 56-61. |
| Kratz et al., “Probing the Cysteine-34 Position of Endogenous Serum Albumin with Thiol-Binding Doxorubicin Derivatives. Improved Efficacy of an Acid-Sensitive Doxorubicin Derivative with Specific Albumin-Binding Properties Compared to That of the Parent Compound,” Journal of Medicinal Chemistry, (2002), vol. 45, No. 25, pp. 5523-5533. |
| Kubicek et al., “The Tubulin-Bound Structure of the Antimitotic Drug Tubulysin,” Angewandte Chemie International Edition, (2010), vol. 49, Issue. 28, pp. 4809-4812. |
| Kumaresan et al., “Development of Tissue Plasminogen Activator Specific “On Demand Cleavable” (ODC) Linkers or Radioimmunotherapy by Screening One-Bead-One-Compound Combinatorial Peptide Libraries,” Bioconjugate Chemistry, (2007), vol. 18, No. 1, pp. 175-182. |
| Kumaresan et al., Evaluation of Ketone-Oxime Method for Developing Therapeutic On-Demand Cleavable Immunoconjugates, Bioconjugate Chemistry, (2008), vol. 19, No. 6, pp. 1313-1318. |
| Laguzza et al., “New Antitumor Monoclonal Antibody-Vinca Conjugates LY203725 and Related Compounds: Design, Preparation, and Representative in Vivo Activity,” Journal of Medicinal Chemistry, (1989), vol. 32, No. 3, pp. 548-555. |
| Lau et al.,“Novel Doxorubicin-Monoclonal Anti-carcinoembryonic Antigen Antibody Immunoconjugate Activity in Vitro,” Bioorganic & Medicinal Chemistry, (1995), vol. 3, No. 10, pp. 1305-1312. |
| Lee et al., “Prolonged Circulating Lives of Single-Chain Fv Proteins Conjugated with Polyethylene Glycol: A comparison of Conjugation Chemistries and Compounds,” Bioconjugate Chemistry, (1999), vol. 10, No. 6, pp. 973-981. |
| Lei et al., “Binding of Monoclonal Antibodies against the Carboxyl Terminal Segment of the Nicotinic Receptor δ Subunit Suggests an Unusual Transmembrane Disposition of This Sequence Region,” Biochemistry, (1995), vol. 34, vol. 20, pp. 6675-6688. |
| Li et al., “Reduction of Kidney Uptake in Radiometal Labeled Peptide Linkers Conjugated to Recombinant Antibody Fragments. Site-Specific Conjugation of DOTA-Peptides to a Cys-Diabody,” Bioconjugate Chemistry, (2002), vol. 13, No. 5, pp. 985-995. |
| Li et al., “Human Antibodies for Immunotherapy Development Generated via a Human B Cell Hybridoma Technology,” PNAS, (Mar. 7, 2006), vol. 103, No. 10, pp. 3557-3562. |
| Liong et al., “Multifunctional Inorganic Nanoparticles for Imaging, Targeting, and Drug Delivery,” ACS Nano, (2008), vol. 2, No. 5, pp. 889-896. |
| Lipinski et al., “A structurally Diversified Linker Enhances the Immune Response to a Small Carbohydrate Hapten,” Glycoconjugate Journal, (2011), vol. 28, No. 3-4, pp. 149-164. |
| Little et al., “Surface Display of Antibodies,” Biotechnology Advances, (1994), vol. 12, Issue 3, pp. 539-555. |
| Liu et al., “Engineering Therapeutic Monoclonal Antibodies,” Immunological Reviews, (2008), vol. 222, No. 1, pp. 9-27. |
| Liu et al., “Targeting Cell Surface Alpha(v)beta(3) Integrin Increases Therapeutic Efficacies of a Legumain Protease-Activated Auristatin Prodrug,” Molecular Pharmaceutics, (Oct. 2011), vol. 9, No. 1, pp. 168-175. |
| Medarova et al., “In vivo Imaging of siRNA Delivery and Silencing in Tumors,” Nature Medicine, (Mar. 2007), vol. 13, No. 3, pp. 372-377. |
| Medina et al., “Targeted Liposomal Drug Delivery in Cancer,” Current Pharmaceutical Design, (2004), vol. 10, No. 24, pp. 2981-2989. |
| Meyer-Losic et al., “Improved Therapeutic Efficacy of Doxorubicin through Conjugation with a Novel Peptide Drug Delivery Technology (Vectocell),” Journal of Medicinal Chemistry, (2006), vol. 49, No. 23, pp. 6908-6916. |
| Mikolajczyk et al., “High Yield, Site-Specific Coupling of N-Terminally Modified β-Lactamase to a Proteolytically Derived Single-Sulfhydryl Murine Fab',” Bioconjugate Chemistry, (1994), vol. 5, No. 6, pp. 636-646. |
| Miller et al., “Design, Construction, and In Vitro Analyses of Multivalent Antibodies,” The Journal of Immunology, (2003), vol. 170, No. 9, pp. 4854-4861. |
| Miller et al., “Synthesis of Taxoids with Improved Cytotoxicity and Solubility for Use in Tumor-Specific Delivery,” Journal of Medicinal Chemistry, (2004), vol. 47, No. 20, pp. 4802-4805. |
| Mitchell et al., “Direct Ring Conjugation of Catecholamines and Their Immunological Interactions,” Bioconjugate Chemistry, (2007), vol. 18, No. 1, pp. 268-274. |
| Mohammad et al., “A new Tubulin Polymerization Inhibitor, Auristatin PE, Induces Tumor Regression in a Human Waldenstrom's Macroglobulinemia Xenograft Model,” International Journal of Oncology, (1999), vol. 15, No. 2, pp. 367-372. |
| Moon et al., “Antibody Conjugates of 7-Ethyl-10-hydroxycamptothecin (SN-38) for Targeted Cancer Chemotherapy,” Journal of Medicinal Chemistry, (2008), vol. 51, No. 21, pp. 6916-6926. |
| Nicolaou et al., “Calicheamicin θ1 I: A Rationally Designed Molecule with Extremely Potent and Selective DNA Cleaving Properties and Apoptosis Inducing Activity,” Angewandte Chemie (International Edition in English), (1994), vol. 33, No. 2, pp. 183-186. |
| Nicolaou et al., “Chemical Synthesis and Biological Evaluation of C-2 Taxoids,” Journal of the American Chemical Society, (1995), vol. 117, No. 9, pp. 2409-2420. |
| Niman et al., “Generation of Protein-Reactive Antibodies by Short Peptides is an Event of High Frequency: Implications for the Structural Basis of Immune Recognition,” Proceedings of the National Academy of Sciences, (Aug. 1983), vol. 80, No. 16, pp. 4949-4953. |
| Ojima et al., “Syntheses and Structure-Activity Relationships of the Second-Generation Antitumor Taxoids: Exceptional Activity against Drug-Resistant Cancer Cells,” Journal of Medicinal Chemistry, (1996), vol. 39, No. 20, pp. 3889-3896. |
| Ojima et al., “Syntheses and Structure-Activity Relationships of Taxoids Derived from 14β-Hydroxy-10-deacetylbaccatin III,” Journal of Medicinal Chemistry, (1997), vol. 40, No. 3, pp. 267-278. |
| Ojima et al., “A Common Pharmacophore for Cytotoxic Natural Products that Stabilize Microtubules,” Proceedings of the National Academy of Sciences, (Apr. 1999), vol. 96, No. 8, pp. 4256-4261. |
| Ojima et al., “Tumor-Specific Novel Taxoid-Monoclonal Antibody Conjugates,” Journal of Medicinal Chemistry, (2002), vol. 45, No. 26, pp. 5620-5623. |
| Pando et al., “First Total Synthesis of Tubulysin B,” Organic Letters, (2009), vol. 11, No. 24, pp. 5567-5569. |
| Pando et al., “The Multiple Multicomponent Approach to Natural Product Mimics: Tubugis, N-Substituted Anticancer Peptides with Picomolar Activity,” Journal of the American Chemical Society, (2011), vol. 133, No. 20, pp. 7692-7695. |
| Parham, “On the Fragmentation of Monoclonal IgG1, IgG2a, and IgG2b from BALB/c mice,” The Journal of Immunology, (Dec. 1983), vol. 131, No. 6, pp. 2895-2902. |
| Patterson et al., “Expedient Synthesis of N-Methyl Tubulysin Analogues with High Cytotoxicity,” The Journal of Organic Chemistry, (2008), vol. 73, No. 12, pp. 4362-4369. |
| Peltier et al., “The Total Synthesis of Tubulysin D,” Journal of the American Chemical Society, (2006), vol. 128, No. 50, pp. 16018-16019. |
| Abstract of Chelliah et al., “A Virtual Screening Hit Reveals New Possibilities for Developing Group III Metabotropic Glutamate Receptor Agonists,” Journal of Medicinal Chemistry, (Apr. 8, 2010), vol. 53, No. 7, pp. 2797-2813. (1 page). |
| Abstract of Govindan, S. V. et al., “Designing immunoconjugates for cancer therapy,” Expert Opinion on Biological, (Jul. 1, 2012), vol. 12, No. 7, pp. 873-890. (1 page). |
| Summary of Rozhko, L.F. et al., “Method for synthesis of phosphinic acids from hypophosphites V. The synthesis of pseudo-α,α-dipeptides,” Amino Acids, (Aug. 2005), vol. 29, No. 2, pp. 139-143. (1 page). |
| Office Action dated Apr. 12, 2017, by the State Intellectual Property Office of People's Republic of China in corresponding Chinese Patent Application No. 201480076320.3 and an English Translation of the Office Action. (9 pages). |
| Search Report dated Nov. 28, 2017, by the State Intellectual Property Office of People's Republic of China in corresponding Chinese Patent Application No. 201480076320.3. (1 page). |
| Partial Supplemental European search report dated Sep. 22, 2017, by the European Patent Office in corresponding European Patent Application No. 14883832.9. (12 pages). |
| The extended European search report dated Jan. 9, 2018, by the European Patent Office in corresponding European Patent Application No. 14883832.9. (24 pages). |
| Office Action (Notification of Reasons for Refusal) dated Jul. 18, 2017, by the Japanese Patent Office in corresponding Japanese Patent Application No. 2016-554198 and an English Translation of the Office Action. (7 pages). |
| Office Action (Notification of Reasons for Refusal) dated Dec. 21, 2017, by the Japanese Patent Office in corresponding Japanese Patent Application No. 2016-554198 and an English Translation of the Office Action. (9 pages). |
| Raghavan et al., “Cytotoxic Simplified Tubulysin Analogues,” Journal of Medicinal Chemistry, (2008), vol. 51, No. 6, pp. 1530-1533. |
| Ruppert et al., “Chemical Coupling of a Monoclonal Antisurfactant Protein-B Antibody to Human Urokinase for Targeting Surfactant-Incorporating Alveolar Fibrin,” Bioconjugate Chemistry, (2002), vol. 13, No. 4, pp. 804-811. |
| Safavy et al., “Synthesis and Biological Evaluation of Paclitaxel-C225 Conjugate as a Model for Targeted Drug Delivery,” Bioconjugate Chemistry, (2003), vol. 14, No. 2, pp. 302-310. |
| Safavy et al., “Site-Specifically Traced Drug Release and Biodistribution of a Paclitaxel-Antibody Conjugate toward Improvement of the Linker Structure,” Bioconjugate Chemistry, (2004), vol. 15, No. 6, pp. 1264-1274. |
| Sani et al., “Total Synthesis of Tubulysins U and V,” Angewandte Chemie International Edition, (2007), vol. 46, No. 19, pp. 3526-3529. |
| Scott, Jr., “An Immunotoxin Composed of a Monoclonal Antitransferrin Receptor Antibody Linked by a Disulfide Bond to the Ribosome- Inactivating Protein Gelonin: Potent In Vitro and In Vivo Effects Against Human Tumors,” Journal of the National Cancer Institute, (Nov. 1987), vol. 79, No. 5, pp. 1163-1172. |
| Scott et al., “Synthesis of Reagents for the One Step Incorporation of Hydrazide Functionality onto the Lysine Residues of Proteins, and their use as Linkers for Carbonyl Containing Molecules,” Bioorganic & Medicinal Chemistry Letters, (1996), vol. 6, No. 13, pp. 1491-1496. |
| Scott et al., “Aiming for the heart: Targeted Delivery of Drugs to Diseased Cardiac Tissue,” Expert Opinion on Drug Delivery, (2008), vol. 5, No. 4, pp. 459-470. |
| Senter et al., “Novel Photocleavable Protein Crosslinking Reagents and their use in the Preparation of Antibody-Toxin Conjugates,” Photochemistry and Photobiology, (1985), vol. 42, No. 3, pp. 231-237. |
| Sharkey et al., “Epratuzumab-SN-38: A New Antibody-Drug Conjugate for the Therapy of Hematologic Malignancies,” Molecular Cancer Therapeutics, (Jan. 2012), vol. 11, No. 1, pp. 224-234. |
| Siiman et al., “Tris(3-mercaptopropyl)-N-glycylaminomethane as a New Linker to Bridge Antibody with Metal Particles for Biological Cell Separations,” Bioconjugate Chemistry, (2000), vol. 11, No. 4, pp. 549-556. |
| Skwarczynski et al., “Paclitaxel Prodrugs: Toward Smarter Delivery of Anticancer Agents,” Journal of Medicinal Chemistry, (Dec. 14, 2006), vol. 49, No. 25, pp. 7253-7269. |
| Smith et al., “The Enediyne Antibiotics,” Journal of Medicinal Chemistry, (May 24, 1996), vol. 39, No. 11, pp. 2103-2117. |
| Srinivasachar et al., “New Protein Cross-Linking Reagents that are Cleaved by Mild Acid,” Biochemistry, (1989), vol. 28, No. 6, pp. 2501-2509. |
| Studer et al., “Influence of a Peptide Linker on Biodistribution and Metabolism of Antibody-Conjugated Benzyl-EDTA. Comparison of Enzymatic Digestion in Vitro and in Vivo,” Bioconjugate Chemistry, (1992), vol. 3, No. 5, pp. 424-429. |
| Suzawa et al., “Synthesis of a Novel Duocarmycin Derivative DU-257 and its Application to Immunoconjugate Using Poly(ethylene glycol)-dipeptidyl Linker Capable of Tumor Specific Activation,” Bioorganic & Medicinal Chemistry, (2000), vol. 8, No. 8, pp. 2175-2184. |
| Szardenings, “Phage Display of Random Peptide Libraries: Applications, Limits, and Potential,” Journal of Receptors and Signal Transduction, (2003), vol. 23, No. 4, pp. 307-349. |
| Tadayoni et al., “Synthesis, in Vitro Kinetics, and in Vivo Studies on Protein Conjugates of AZT: Evaluation as a Transport System to Increase Brain Delivery,” Bioconjugate Chemistry, (1993), vol. 4, No. 2, pp. 139-145. |
| Ten Hoeve et al., “Syntheses of Haptens Containing Dioxaphosphorinan Methoxyacetic Acid Linker Arms for the Production of Antibodies to Organophosphate Pesticides,” Bioconjugate Chemistry, (May/Jun. 1997), vol. 8, No. 3, pp. 257-266. |
| Trail et al., “Effect of Linker Variation on the Stability, Potency, and Efficacy of Carcinoma-reactive BR64-Doxorubicin Immunoconjugates,” Cancer Research, (Jan. 1, 1997), vol. 57, No. 1, pp. 100-105. |
| Trouet et al., “A Covalent Linkage between Daunorubicin and Proteins that is Stable in Serum and Reversible by Lysosomal Hydrolases, as Required for a Lysosomotropic Drug-Carrier Conjugate: In Vitro and In Vivo Studies,” Proceedings of the National Academy of Sciences, (Jan. 1982), vol. 79, No. 2, pp. 626-629. |
| Tsai et al., “Sensitive Measurement of Polyethylene Glycol-Modified Proteins,” BioTechniques, (Feb. 2001), vol. 30, No. 2, pp. 396-402. |
| Ullrich et al., “Pretubulysin, a Potent and Chemically Accessible Tubulysin Precursor from Angiococcus disciformis,” Angewandte Chemie International Edition, (2009), vol. 48, No. 24, pp. 4422-4425. |
| Walker et al., “Monoclonal Antibody Mediated Intracellular Targeting of Tallysomycin S10b,” Bioorganic & Medicinal Chemistry Letters, (Aug. 16, 2004), vol. 14, No. 16, pp. 4323-4327. |
| Warpehoski et al., “Stereoelectronic Factors Influencing the Biological Activity and DNA Interaction of Synthetic Antitumor Agents Modeled on CC-1065,” Journal of Medicinal Chemistry, (1988), vol. 31, No. 3, pp. 590-603. |
| Watanabe et al., “Measurement of Cross-Reactive Properties of Adriamycin Derivatives by the Inhibition Enzyme-Linked Immunosorbent Assay for Adriamycin,” Tokai Journal of Experimental and Clinical Medicine, (1990), vol. 15, No. 4, pp. 327-334. |
| Widdison et al., “Semisynthetic Maytansine Analogues for the Targeted Treatment of Cancer,” Journal of Medicinal Chemistry, (2006), vol. 49, No. 14, pp. 4392-4408. |
| Wilbur et al., “Reagents for Astatination of Biomolecules. 5. Evaluation of Hydrazone Linkers in 211At- and 125I-Labeled closo-Decaborate(2-) Conjugates of Fab' as a Means of Decreasing Kidney Retention,” Bioconjugate Chemistry, (2011), vol. 22, No. 6, pp. 1089-1102. |
| Wipf et al., “Synthesis of the Tubuvaline-Tubuphenylalanine (Tuv-Tup) Fragment of Tubulysin,” Organic Letters, (2004), vol. 6, No. 22, pp. 4057-4060. |
| Wipf et al., “Total Synthesis of N14-Desacetoxytubulysin H,” Organic Letters, (2007), vol. 9, No. 8, pp. 1605-1607. |
| Zhao et al., “Synthesis and Evaluation of Hydrophilic Linkers for Antibody-Maytansinoid Conjugates,” Journal of Medicinal Chemistry, (2011), vol. 54, No. 10, pp. 3606-3623. |
| Zhao et al., “Synthesis and Biological Evaluation of Antibody Conjugates of Phosphate Prodrugs of Cytotoxic DNA Alkylators for the Targeted Treatment of Cancer,” Journal of Medicinal Chemistry, (2012), vol. 55 No. 2, pp. 766-782. |
| Zhou et al., “Cell-Specific Delivery of a Chemotherapeutic to Lung Cancer Cells,” Journal of the American Chemical Society, (2004), vol. 126, No. 48, pp. 15656-15657. |
| International Search Report (From PCT/ISA/210) and the Written Opinion of the International Searching Authority (From PCT/ISA/237) dated Dec. 8, 2014, by the State Intellectual Property Office, the P.R. China, in corresponding International Application No. PCT/CN2014/072769. (7 pages). |
| International Preliminary Report on Patentability (Form PCT/IB/373) dated Sep. 6, 2016, by the International Bureau of WIPO, in corresponding International Application No. PCT/CN2014/072769. (1 page). |
| Office Action (Notice of grant for your patent) dated Feb. 16, 2017, by the Australian Patent Office in corresponding Australian Patent Application No. 2014384434. (1 page). |
| Clackson et al., “Making antibody fragments using phage display libraries,” Nature, (Aug. 15, 1991), vol. 352, pp. 624-628. (5 pages). |
| Harlow et al., “Antibodies: A Laboratory Manual,” Cold Spring Harbor, New York: Cold Spring Harbor Laboratory, (1988). (105 pages). |
| Hurwitz et al., “The Covalent Binding of Daunomycin and Adriamycin to Antibodies, with Retention of Both Drug and Antibody Activities,” Cancer Research, (May 1975), vol. 35, No. 5, pp. 1175-1181. (7 pages). |
| Jain, Kewal K., “Drug Delivery Systems,” Methods in Molecular Biology, Humana Press, (2008). (59 pages). |
| Köhler et al., “Continuous cultures of fused cells secreting antibody of predefined specificity,” Nature, (Aug. 7, 1975), vol. 256, No. 5517, pp. 495-497. (3 pages). |
| Lee et al., “Designing dendrimers for biological applications,” Nature Biotechnology, (Dec. 2005), vol. 23, No. 12, pp. 1517-1526. (10 pages). |
| Ojima et al., “Tumor-Specific Novel Taxoid-Monoclonal Antibody Conjugates,” Journal of Medicinal Chemistry, (2002), vol. 45, pp. 5620-5623. (4 pages). |
| “Organic Substituent Groups and Ring Systems,” CRC Handbook of Chemistry and Physics, p. 2-25 and p. 2-26. (2 pages). |
| Pietersz et al., “Immunochemotherapy of a Murine Thymoma with the Use of Idarubicin Monoclonal Antibody Conjugates,” Cancer Research, (Feb. 15, 1988), vol. 48, No. 4, pp. 926-931. (6 pages). |
| Reddy et al., “In Vivo Structural Activity and Optimization Studies of Folate-Tubulysin Conjugates,” Molecular Pharmaceutics, (Jul. 24, 2009), vol. 6, No. 5, pp. 1518-1525. (8 pages). |
| Sun et al., “Design of Antibody-Maytansinoid Conjugates Allows for Efficient Detoxification via Liver Metabolism,” Bioconjugate Chemistry, (2011), vol. 22, No. 4, pp. 728-735. (8 pages). |
| Türk et al., “Relevance of multidrug resistance in the age of targeted therapy,” Current Opinion in Drug Discovery & Development, (2009), vol. 22, No. 2, pp. 246-252. (7 pages). |
| Yang et al. “Doxorubicin conjugated with a monoclonal antibody directed to a human melanoma-associated proteoglycan suppresses the growth of established tumor xenografts in nude mice,” Proceedings of the National Academy of Sciences of the United States of America, (Feb. 1988), vol. 85, No. 4, pp. 1189-1193. (5 pages). |
| Yauch et al., “Recent advances in pathway-targeted cancer drug therapies emerging from cancer genome analysis,” Current Opinion in Genetics & Development, (Feb. 7, 2012), vol. 22, No. 1, pp. 45-49. (5 pages). |
| Zhao et al., “Synthesis and Evaluation of Hydrophilic Linkers for Antibody-Maytansinoid Conjugates,” Journal of Medicinal Chemistry, (Apr. 25, 2011), vol. 54, No. 10, pp. 3606-3623. (18 pages). |
| Office Action dated May 6, 2018 by the State Intellectual Property Office of People's Republic of China in corresponding Chinese Patent Application No. 201480076320.3 and an English translation of the Office Action. (10 pages). |
| Notification of Reasons for Refusal dated Aug. 15, 2018, by the Japanese Patent Office in corresponding Japanese Patent Application No. 2016-554198 and an English translation of the Notification. (5 pages). |
| Decision to Grant a Patent dated Sep. 6, 2018, by the Japanese Patent Office in corresponding Japanese Patent Application No. 2016-554198 and an English translation of the Decision. (5 pages). |
| Third Office Action dated Nov. 18, 2018, by the State Intellectual Property Office of the People's Republic of China in corresponding Chinese Patent Application No. 201480078320.3 and an English translation of the Office Action. (10 pages). |
| Office Action dated Jun. 25, 2019, by the U.S. Patent and Trademark Office in related U.S. Appl. No. 16/228,130. (14 pages). |
| Office Action dated Jun. 26, 2019, by the U.S. Patent and Trademark Office in related U.S. Appl. No. 16/228,310. (15 pages). |
| Office Action dated Jun. 26, 2019, by the U.S. Patent and Trademark Office in related U.S. Appl. No. 16/228,437. (16 pages). |
| Office Action dated Sep. 25, 2017, by the Canadian Patent Office in corresponding Canadian Application No. 2,938,919. (7 pages). |
| Office Action dated Jun. 15, 2018, by the Canadian Patent Office in corresponding Canadian Application No. 2,938,919. (4 pages). |
| Office Action dated Apr. 5, 2019, by the Canadian Patent Office in corresponding Canadian Application No. 2,938,919. (4 pages). |
| Number | Date | Country | |
|---|---|---|---|
| 20170152274 A1 | Jun 2017 | US |
