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Last Updated: March 29, 2024

Claims for Patent: 9,428,543


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Summary for Patent: 9,428,543
Title:Preparation of maytansinoid antibody conjugates by a one-step process
Abstract: The invention provides a one-step process for preparing a cell-binding agent cytotoxic agent conjugate comprising contacting a cell-binding agent with a cytotoxic agent to form a first mixture comprising the cell-binding agent and the cytotoxic agent and contacting the first mixture comprising the cell-binding agent and the cytotoxic agent with a bifunctional crosslinking reagent, which provides a linker, in a solution having a pH of about 4 to about 9 to provide a second mixture comprising the cell-binding agent cytotoxic agent conjugate, wherein the cell-binding agent is chemically coupled through the linker to the cytotoxic agent, free cytotoxic agent, and reaction by-products. The second mixture is then optionally subjected to purification to provide a purified cell-binding agent cytotoxic agent conjugate.
Inventor(s): Li; Xinfang (Newton, MA), Worful; Jared M. (Pittsburgh, PA)
Assignee: ImmunoGen, Inc. (Waltham, MA)
Application Number:14/313,868
Patent Claims:1. A process for preparing an antibody maytansinoid conjugate comprising the step of: (a) contacting an antibody with a maytansinoid to form a first mixture comprising the antibody and the maytansinoid, then contacting the first mixture with a bifunctional crosslinking reagent comprising a linker, in a solution having a pH of about 7 to about 9 to provide a second mixture comprising (i) the antibody maytansinoid conjugate, wherein the antibody is chemically coupled through the linker to the maytansinoid, (ii) free maytansinoid, and (iii) reaction by-products, wherein the maytansinoid is N2'-deacetyl-N2'-(3-mercapto-1-oxopropyl)-maytansine (DM1) or N2'-deacetyl-N2'-(4-methyl-4-mercapto-1-oxopentyl)-maytansine (DM4).

2. The process of claim 1, wherein the contacting in step (a) is effected by providing the antibody in a reaction vessel, adding the maytansinoid to the reaction vessel to form the first mixture comprising the antibody and the maytansinoid, and then adding the bifunctional crosslinking reagent to the first mixture.

3. The process of claim 1, wherein the contacting in step (a) occurs at a temperature of about 16.degree. C. to about 24.degree. C.

4. The process of claim 1, wherein the contacting in step (a) occurs at a temperature of about 0.degree. C. to about 15.degree. C.

5. The process of claim 1, wherein the antibody is selected from the group consisting of huN901, huMy9-6, huB4, huC242, trastuzumab, bivatuzumab, sibrotuzumab, the anti-integrin .alpha.v.beta.6 antibody CNTO95, huDS6, rituximab, an antibody that binds to Her2, an antibody that binds to epidermal growth factor receptor (EGFR), an antibody that binds to CD27L, an antibody that binds to EGFRvIII, an antibody that binds to Cripto, an antibody that binds to CD138, an antibody that binds to EphA2, an integrin targeting antibody, an antibody that binds to CD37, an antibody that binds to a folate receptor, an antibody that binds to Her3, and an antibody that binds to insulin-like growth factor 1 receptor (IGF1R).

6. The process of claim 1, wherein the solution in step (a) comprises sucrose.

7. The process of claim 1, wherein the solution in step (a) comprises a buffering agent selected from the group consisting of a citrate buffer, an acetate buffer, a succinate buffer, and a phosphate buffer.

8. The process of claim 1, wherein the solution in step (a) comprises a buffering agent selected from the group consisting of HEPPSO (N-(2-hydroxyethyl)piperazine-N'-(2-hydroxypropanesulfonic acid)), POPSO (piperazine-1,4-bis-(2-hydroxy-propane-sulfonic acid)dehydrate), HEPES (4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid), HEPPS (EPPS) (4-(2-hydroxyethyl)piperazine-1-propanesulfonic acid), TES (N-[tris(hydroxymethyl)methyl]-2-aminoethanesulfonic acid), and a combination thereof.

9. The process of claim 1, wherein the process further comprises the step of: (b) purifying the second mixture comprising the antibody maytansinoid conjugate to provide a purified antibody maytansinoid agent conjugate.

10. The process of claim 9, wherein the second mixture is purified by subjecting the mixture to tangential flow filtration, selective precipitation, adsorptive filtration, adsorptive chromatography, non-absorptive chromatography, or a combination thereof, to purify the antibody maytansinoid conjugate from the free cytotoxic agent and reaction by-products.

11. The process of claim 10, wherein the second mixture is purified by subjecting the mixture to tangential flow filtration.

12. The process of claim 9, further comprising holding the second mixture between steps (a)-(b) to release the unstably bound linkers from the antibody.

13. The process of claim 12, wherein the second mixture is held for about 20 hours at a temperature of about 2.degree. C. to about 8.degree. C.

14. The process of claim 9, further comprising quenching the second mixture between steps (a)-(b) to quench any unreacted maytansinoid and/or unreacted bifunctional crosslinking reagent.

15. The process of claim 14, wherein the mixture is quenched by contacting the second mixture with a quenching reagent that reacts with the free maytansinoid.

16. The process of claim 15, wherein the quenching reagent is selected from the group consisting of 4-maleimidobutyric acid, 3-maleimidopropionic acid, N-ethylmaleimide, iodoacetamide, and iodoacetamidopropionic acid.

17. The process of claim 1, wherein the antibody is a monoclonal antibody.

18. The process of claim 17, wherein the antibody is a humanized monoclonal antibody.

19. The process of claim 1, wherein the maytansinoid is N.sup.2'-deacetyl-N.sup.2'-(3-mercapto-1-oxopropyl)-maytansine (DM1).

20. The method of claim 19, wherein the bifunctional crosslinking reagent is N-succinimidyl 4-(maleimidomethyl)cyclohexanecarboxylate (SMCC).

21. The process of claim 1, wherein the maytansinoid is N.sup.2'-deacetyl-N.sup.2'-(4-methyl-4-mercapto-1-oxopentyl)-maytansine (DM4).

22. The method of claim 21, wherein the bifunctional crosslinking reagent is N-succinimidyl 4-(2-pyridyldithio)butanoate (SPDB), N-succinimidyl-4-(2-pyridyldithio)2-sulfo butanoate (sulfo-SPDB).

23. The process of claim 1, wherein the antibody is chemically coupled to the maytansinoid via chemical bonds selected from the group consisting of disulfide bonds, acid labile bonds, photolabile bonds, peptidase labile bonds, thioether bonds, and esterase labile bonds.

24. The process of claim 1, wherein the bifunctional crosslinking reagent comprises an N-succinimidyl ester moiety, an N-sulfosuccinimidyl ester moiety, a maleimido-based moiety, or a haloacetyl-based moiety.

25. The process of claim 24, wherein the bifunctional crosslinking reagent is selected from the group consisting of N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP), N-succinimidyl 4-(2-pyridyldithio)pentanoate (SPP), N-succinimidyl 4-(2-pyridyldithio)butanoate (SPDB), N-succinimidyl-4-(2-pyridyldithio)2-sulfo butanoate (sulfo-SPDB), N-succinimidyl 4-(maleimidomethyl)cyclohexanecarboxylate (SMCC), PEG-mal, sulfo-Mal, and CX1-1.

Details for Patent 9,428,543

Applicant Tradename Biologic Ingredient Dosage Form BLA Approval Date Patent No. Expiredate
Genentech, Inc. RITUXAN rituximab Injection 103705 11/26/1997 ⤷  Try a Trial 2031-03-29
Idec Pharmaceuticals Corp. RITUXAN rituximab Injection 103737 02/19/2002 ⤷  Try a Trial 2031-03-29
Genentech, Inc. HERCEPTIN trastuzumab For Injection 103792 09/25/1998 ⤷  Try a Trial 2031-03-29
Genentech, Inc. HERCEPTIN trastuzumab For Injection 103792 02/10/2017 ⤷  Try a Trial 2031-03-29
>Applicant >Tradename >Biologic Ingredient >Dosage Form >BLA >Approval Date >Patent No. >Expiredate

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