Camptothecin-binding moiety conjugates
The invention relates to therapeutic conjugates with improved ability to target various diseased cells containing a targeting moiety (such as an antibody or antibody fragment), a linker and a camptothecin as a therapeutic moiety, and further relates to processes for making and using the said conjugates.
1. A conjugate of a camptothecin drug and a targeting moiety (TM) of the formula: TM-[L]-CPT, where TM is a cancer targeting monoclonal antibody; CPT is selected from the consisting of camptothecin, 10-hydroxy camptothecin, SN-38, topotecan, lurtotecan, 9-aminocamptothecin, and 9-nitrocamptothecin; and L is a linker system of the type X-Y-Z wherein X is an antibody coupling moiety comprising a defined polyethylene glycol (PEG) wherein the defined PEG contains a defined number of monomeric units that is between 1 and 12 monomeric units, Y is a lysosomally cleavable polypeptide, and Z is 4-aminobenzyloxy moiety, which is connected to the CPT drug, wherein Z is connected to a 20-hydroxy group of CPT by a carbonate or carbamate bond and X is connected to TM by a bond between a thiol group and a thiol-reactive moiety selected from the group consisting of maleimide, vinylsulfone, bromoacetamide, and iodoacetamide.
2. The conjugate according to claim 1 , wherein the antibody is a chimeric antibody or a humanized antibody.
3. The conjugate according to claim 1 , wherein X is an antibody coupling moiety comprising a defined polyethylene glycol (PEG) with 12 monomeric units.
4. The conjugate according to claim 1 , wherein the antibody is a human antibody.
5. The conjugate according to claim 1 , wherein the monoclonal antibody is part of a bi-specific or multi-specific antibody.
6. The conjugate according to claim 5 , wherein the monoclonal antibody is part of a trivalent, bi-specific antibody complex.
7. The conjugate according to claim 1 , wherein said CPT is SN-38.
8. The conjugate according to claim 1 , wherein the cleavable polypeptide is selected from the group consisting of Phe-Lys, Val-Cit, Ala-Leu, and Leu-Ala-Leu.
9. The conjugate according to claim 1 wherein ‘Z’ is connected via a carbonate bond to the 20-hydroxy group of CPT.
10. A process for producing the conjugate of claim 1 , wherein the linker is first conjugated to the CPT drug, thereby producing a CPT drug-linker conjugate, wherein said CPT drug-linker conjugate is subsequently conjugated to a monoclonal antibody, wherein the CPT comprises a 10-hydroxyl group that is a t-butyloxycarbonyl (BOC) derivative and the CPT further comprises a 20-hydroxyl group that is a carbonate derivative and the process comprises exposing the derivatized CPT to trifluoroacetic acid (TFA), wherein the TFA removes the BOC from the 10-hydroxyl position but does not remove the carbonate from the 20-hydroxyl position.
11. The process of claim 10 wherein the derivatized CPT is exposed to TFA for two to five minutes.
12. The process of claim 11 , wherein the CPT is SN-38.
13. The process of claim 11 , wherein said CPT drug-linker is not purified prior to conjugation to a monoclonal antibody or fragment.
14. The conjugate according to claim 1 , wherein said antibody is attached to between 1 and 12 CPT moieties.
15. The conjugate according to claim 14 , wherein said antibody is a murine, chimeric, primatized, humanized, or human monoclonal antibody, and said antibody is in intact, fragment (Fab, Fab′, F(ab)2, F(ab′)2), or sub-fragment (single-chain constructs) form.
16. The conjugate according to claim 15 , wherein said antibody is a chimeric antibody.
17. The conjugate according to claim 15 , wherein said antibody is a humanized antibody.
18. The conjugate according to claim 1 , wherein said antibody binds to a B-cell lineage antigen, a T-cell antigen, a myeloid lineage antigen or a HLA-DR antigen.
19. The conjugate according to claim 18 , wherein said antibody binds to a CD20 antigen.
20. The conjugate according to claim 1 , wherein said antibody binds to an antigen selected from the group consisting of CD74, CD22, epithelial glycoprotein-1, carcinoembryonic antigen (CEA or CD66e), colon-specific antigen-p, alpha-fetoprotein, CC49, prostate-specific membrane antigen, carbonic anhydrase IX, HER-2/neu, EGFR (ErbB1), ErbB2, ErbB3, ILGF, BrE3, CD19, CD20, CD21, CD23, CD33, CD45, CD74, CD80, VEGF, ED-B fibronectin, P1GF, other tumor angiogenesis antigens, MUC1, MUC2, MUC3, MUC4, gangliosides, HCG, EGP-2, CD37, HLA-DR, CD30, Ia, A3, A33, Ep-CAM, KS-1, Le(y), S100, PSA, tenascin, folate receptor, Thomas-Friedreich antigens, tumor necrosis antigens, Ga 733, IL-2, IL-6, T101, MAGE, migration inhibition factor (MIF), an antigen that is bound by L243, an antigen that is bound by PAM4, CD66a (BGP), CD66b (CGM6), CD66c (NCA), CD66d (CGM1), and TAC.
21. The conjugate of claim 1 , wherein said conjugate is in a form suitable for parenteral administration.
22. The conjugate of claim 1 , wherein the monoclonal antibody is part of a composite, multispecific antibody.
23. The conjugate of claim 22 , wherein the composite antibody binds to two or more antigens selected from the group consisting of CD74, CD22, epithelial glycoprotein-1, carcinoembryonic antigen (CEA or CD66e), colon-specific antigen-p, alpha-fetoprotein, CC49, prostate-specific membrane antigen, carbonic anhydrase IX, HER-2/neu, BrE3, CD19, CD20, CD21, CD23, CD33, CD45, CD74, CD80, VEGF, EGF receptor (ErbB1), ErbB2, ErbB3, P1GF, VEGF, ED-B fibronectin, MUC1, MUC2, MUC3, MUC4, ILGF, gangliosides, HCG, EGP-2, CD37, HLA-DR, CD30, Ia, A3, A33, Ep-CAM, KS-1, Le(y), S100, PSA, tenascin, folate receptor, Thomas-Friedreich antigens, tumor necrosis antigens, tumor angiogenesis antigens, MIF, Ga 733, IL-2, IL-6, T101, MAGE, an antigen that is bound by L243, an antigen that is bound by PAM4, CD66a (BGP), CD66b (CGM6), CD66c (NCA), CD66d (CGM1) and TAC.
24. The conjugate of claim 22 , wherein the composite antibody binds to CD20 and CD22.
25. The conjugate of claim 22 , wherein the composite antibody contains at least one binding site for EGP-1.
26. The conjugate of claim 22 , wherein the composite antibody contains at least one binding site for CD74.
27. The conjugate of claim 1 , wherein the antibody is a fusion protein.
28. The conjugate of claim 27 , further comprising a second fusion protein, antibody or antibody fragment.
29. The conjugate of claim 28 , wherein the second fusion protein, antibody or antibody fragment is also linked to one or more CPT moieties.
30. The conjugate of claim 27 , wherein said fusion protein comprises a murine, chimeric, primatized, humanized, or human monoclonal antibody, and said antibody is in intact, fragment (Fab, Fab′, F(ab).sub.2, F(ab′).sub.2), or sub-fragment (single-chain constructs) form.
31. The conjugate of claim 28 , wherein the two fusion proteins bind to the same epitope, to different epitopes on the same antigen or to different antigens.