Preparation,Pharmacodynamics And Pharmacokinetices Of A Novel C-Met ADC SHR-A1403

Emerging evidence demonstrates that a c-Met antibody-drug conjugate?ADC?has superior efficacy and managable safety profiles compared with those of currently available small molecular inhibitors or antibodies for the treatment of c-Met-overexpressed cancers.SHR-A1403 is a novel c-Met ADC composed of a humanized IgG2 monoclonal antibody against c-Met conjugated to a novel cytotoxic microtubule inhibitor via an uncleavable ATPPA linker.Here we describ the preparation technology,in vitro and in vivo pharmacodynamics,and pharmacokinetics of SHR-A1403.As components of SHR-A1403,anti-c-Met monoclonal antibody SHR-A1403 mAb and conjugated toxin SHR153024 were prepared.SHR-A1403 mAb was expressed in CHO cells using DNA recombinant technology and purified by affinity and ion exchange chromatography,followed by inactivation and remove of viruses.Synthesis route of SHR153024 was further optimized after retrosynthesis analysis,so as to fit for industrial manufacture.The crude product was purified using liquid phase method,and its structure was confirmed using methods of mass spectrum,infrared and ultraviolet spectroscopy,and nuclear magnetic resonance.SHR-A1403was synthesized via three-step conjugation and then mixed with excipients to achieve final concentrations of 10±0.4 g/L.Altogether,the preparation technologies of SHR-A1403 and its components were stable and controllable,and the products obtained met all the criteria of investigational products used in preclincal and clinial studies.In efficacy studies,SHR-A1403 showed high affinity to c-Met proteins derived from human or monkey,but rarely had affinity to mice c-Met.SHR-A1403 exhibited potent inhibitory effects on proliferation of multiple cancer cell lines,and the activities were basically proportional to protein expression of c-Met on the cell membrane.In mice bearing tumors derived from human cancer cell lines?hepatic HCCLM3 cell,gastric MKN-45 cell,and lung NCI-H1993 cell?or patient hepatocellular carcinoma?HCC?tissues,SHR-A1403 showed excellent antitumor efficacy.All these models were confirmed with high c-Met protein expression.The molecular mechanisms of antitumor efficacy were further investigated in in vitro settings.The results showed that antibody binding with c-Met contributed to SHR-A1403 endocytosis,and subsequently the complex translocated to lysosomes.In lysosomes,the complex was hydrolized by lysosomal enzymes which led to release of the conjugated toxin.The released toxin caused cell cycle arrest by inhibition on microtubulin aggregation and finally displayed cell killing effect.In conclusion,our work demonstrate that SHR-A1403 has significant antitumor activity in cancer cell lines,xenograft mouse models and an HCC PDX model,which all have high c-Met levels.Furthermore,the molecular mechanisms mediating the antitumor efficacy are clearly elucidated.These data provide references for potential clinical efficacy of SHR-A1403to treat cancers with c-Met overexpression.The in vivo pharmacokinetics profiles of SHR-A1403 were investigated and characterized in normal cynomolgus monkeys.Serum concentrations of ADC and total antibody,and anti-drug antibody?ADA?were detected using validated ELISA methods,and concentrations of free toxin were detected using validated LC-MS/MS method.The results showed low systemic clearance of both ADC and total antibody in monkeys as reflected by gradual decrease in serum concentrations.Half-life(t_1/2)of ADC ranged from 4 to 7.5 days in monkeys.Almost no free toxin was detected in plasma,indicating limited possibility of SHR-A1403 to cause toxicitity due to exposure of free toxin in circulation.Relatively low incidence of ADA in monkeys were observed,which had no impact on pharmacokinetics profile of the ADC.During early discovery stage,undesirable exposure and ADA incidence were observed for SHR-A1403 with high or low drug-antibody ratio?DAR?,which was DAR=5 to 6 and DAR=1,respectively,and therefore prompted selection of an appropriate DAR=2 for SHR-A1403 used in our work.Collectively,we explore and confirm the preparation technology of SHR-A1403 which is applicable for industrial manufacture,fully demonstrate the potent antitumor efficacy in both in vitro and in vivo models,and reveal the favourable pharmacokinetics properties in monkeys.These results provide technical support for investigational product preparation for preclinical and clinical studies,and also rational for study design of clinical trials of SHR-A1403.