Disulfide-linked antibody-maytansinoid conjugates: optimization of in vivo activity by varying the steric hindrance at carbon atoms adjacent to the disulfide linkage.

In this report, we describe the synthesis of a panel of disulfide-linked huC242 (anti-CanAg) antibody maytansinoid conjugates (AMCs), which have varying levels of steric hindrance around the disulfide bond, in order to investigate the relationship between stability to reduction of the disulfide linker and antitumor activity of the conjugate in vivo. The conjugates were first tested for stability to reduction by dithiothreitol in vitro and for plasma stability in CD1 mice. It was found that the conjugates having the more sterically hindered disulfide linkages were more stable to reductive cleavage of the maytansinoid in both settings. When the panel of conjugates was tested for in vivo efficacy in two human colon cancer xenograft models in SCID mice, it was found that the conjugate with intermediate disulfide bond stability having two methyl groups on the maytansinoid side of the disulfide bond and no methyl groups on the linker side of the disulfide bond (huC242-SPDB-DM4) displayed the best efficacy. The ranking of in vivo efficacies of the conjugates was not predicted by their in vitro potencies, since all conjugates were highly active in vitro, including a huC242-SMCC-DM1 conjugate with a noncleavable linkage which showed only marginal activity in vivo. These data suggest that factors in addition to intrinsic conjugate potency and conjugate half-life in plasma influence the magnitude of antitumor activity observed for an AMC in vivo. We provide evidence that bystander killing of neighboring nontargeted tumor cells by diffusible cytotoxic metabolites produced from target cell processing of disulfide-linked antibody-maytansinoid conjugates may be one additional factor contributing to the activity of these conjugates in vivo.

Alphav integrin-targeted immunoconjugates regress established human tumors in xenograft models.

PURPOSE: Targeted delivery of cytotoxic agents to solid tumors through cell surface antigens can potentially reduce systemic toxicity and increase the efficacy of the targeted compounds. The purpose of this study was to show the feasibility of treating solid tumors by targeting alpha(v) integrins with antibody-maytansinoid conjugates and to test the relative in vivo activities of several linker-maytansinoid chemistries. EXPERIMENTAL DESIGN: CNTO 364, CNTO 365, and CNTO 366 are targeted cytotoxic agents created by conjugating the CNTO 95 anti-alpha(v) integrin antibody with three distinct maytansinoid-linker structures. These structures were designed to have varying degrees of chemical substitution surrounding the disulfide bond linking the cytotoxic agent to the antibody. A model conjugate was shown to be specifically cytotoxic in vitro and highly active against established human tumor xenografts in immunocompromised rats. The in vivo antitumor activities of CNTO 364, CNTO 365, and CNTO 366 were compared in rat xenograft models. RESULTS: CNTO 365, with a linker chemistry of expected intermediate stability, was shown to be substantially more active than the other two conjugates with lesser or greater substitution around the disulfide linkage. CONCLUSION: CNTO 95-maytansinoid immunoconjugates are potent antitumor agents against alpha(v) integrin-expressing human carcinomas. These studies show for the first time the feasibility of targeting alpha(v) integrins on solid tumors with tumor-activated prodrugs. The DM4 linker-maytansinoid configuration of CNTO 365 was substantially more active in the models tested here when compared with alternative configurations with greater or lesser chemical substitution surrounding the linker.

Semisynthetic maytansine analogues for the targeted treatment of cancer.

Maytansine, a highly cytotoxic natural product, failed as an anticancer agent in human clinical trials because of unacceptable systemic toxicity. The potent cell killing ability of maytansine can be used in a targeted delivery approach for the selective destruction of cancer cells. A series of new maytansinoids, bearing a disulfide or thiol substituent were synthesized. The chain length of the ester side chain and the degree of steric hindrance on the carbon atom bearing the thiol substituent were varied. Several of these maytansinoids were found to be even more potent in vitro than maytansine. The targeted delivery of these maytansinoids, using monoclonal antibodies, resulted in a high, specific killing of the targeted cells in vitro and remarkable antitumor activity in vivo.

Prostate stem cell antigen as therapy target: tissue expression and in vivo efficacy of an immunoconjugate.

We conducted an expression analysis of prostate stem cell antigen (PSCA)in normal urogenital tissues, benign prostatic hyperplasia (n = 21), prostatic intraepithelial neoplasia (n = 33), and primary (n = 137) and metastatic (n = 42) prostate adenocarcinoma, using isotopic in situ hybridization on tissue microarrays. In normal prostate, we observe PSCA expression in the terminally differentiated, secretory epithelium; strong expression was also seen in normal urothelium. Forty-eight percent of primary and 64% of metastatic prostatic adenocarcinomas expressed PSCA RNA. Our studies did not confirm a positive correlation between level of PSCA RNA expression and high Gleason grade. We characterized monoclonal anti-PSCA antibodies that recognize PSCA expressed on the surface of live cells, are efficiently internalized after antigen recognition, and kill tumor cells in vitro in an antigen-specific fashion upon conjugation with maytansinoid. Unconjugated anti-PSCA antibodies demonstrated efficacy against PSCA-positive tumors by delaying progressive tumor growth in vivo. Maytansinoid-conjugated antibodies caused complete regression of established tumors in a large proportion of animals. Our results strongly suggest that maytansinoid-conjugated anti-PSCA monoclonal antibodies should be evaluated as a therapeutic modality for patients with advanced prostate cancer.