Payload-recycling ADCs (PR-ADCs): A novel ADC platform for reducing free-payload toxicity and enhancing therapeutic index

Background: Antibody-drug conjugates (ADCs) have emerged as an important class of cancer therapeutics in clinical application. Many ADC drugs, however, failed during clinical studies due to excessive toxicity. Even approved ADC drugs also often face challenges in balancing clinical efficacy and safety. A major cause of ADC toxicity is due to free-payload (small molecule toxin) released from ADC drug into circulation. Free-payload often appears rapidly in plasma after ADC administration which causes systemic toxicity. To overcome ADC toxicity caused by free-payload, we have developed a novel technology, payload-recycling ADC (PR-ADC), that allows ADC drugs to re-capture, or recycle, small molecule toxin in circulation, thus reducing free-drug concentration in plasma. Methods: To develop PR-ADC technology, we first generated a high affinity anti-MMAE monoclonal antibody through hybridoma screening. This anti-MMAE antibody exhibits exceptional specificity to free-MMAE with no detectable cross-reactivity to linker-MMAE or conjugated MMAE. Then an anti-mesothelin (MSLN) single domain antibody was attached to the anti-MMAE antibody to create a bispecific antibody that can simultaneously bind to both tumor antigen MSLN and free-MMAE. This bispecific antibody was then conjugated with MMAE to become the PR-ADC molecule which is basically a mesothelin-directing ADC with an additional ability to capture free-MMAE. Results: This PR-ADC can effectively kill MSLN-expressing tumor cells in vitro in a potency similar to or even higher than a conventional MSLN-targeting ADC. On the other hand, this PR-ADC can prevent MMAE-induced cytotoxicity in cell-protection assays, highlighting its ability to effectively shield normal cells or tissues from free-MMAE-induced damage, thus increasing safety of ADC drugs. In vivo , this PR-ADC molecule, compared to conventional ADC, significantly reduced free-MMAE concentration in serum after administrated in mice or rats. In toxicity studies in rats, the PR-ADC significantly improved safety profile, showing larger therapeutic window than the conventional ADC. The increased safety profile of PR-ADC in vivo was further supported by favorable tissue distribution, drug metabolism and pharmacokinetics (DMPK) profiles. In xenograft tumor model studies, the MSLN-targeting PR-ADC achieved robust tumor growth inhibition, outperforming the conventional ADC. Conclusions: The PR-ADC platform improves ADC safety by minimizing free payload-mediated toxicity while maintaining or enhancing anti-tumor efficacy. By enabling active payload recapture, PR-ADCs may support higher clinical dosing, increased effective DAR through captured payload, and an overall superior therapeutic index. This technology represents a promising next-generation strategy for advancing ADC development.