The Hu2G10 mAb targets the cleaved-activated form of Trop-2 and exploits vulnerability of multiple human cancers.

Abstract

e14548 Background: FDA granted accelerated approval to the anti-Trop-2 sacituzumab govitecan-hziy (TRODELVY) antibody-drug conjugate (ADC), for patients with metastatic triple-negative breast cancer. Skin rash and mucosal inflammation were, though, dose-limiting toxicities of other high-effectiveness anti-Trop-2 antibodies, and impeded their development. These adverse events appeared as on-target/off-cancer effects, related to the expression of Trop-2 by normal tissues. Trop-2 is a transmembrane glycoprotein (E. Guerra et al., The Trop-2 signalling network in cancer growth. Oncogene 32, 1594-1600 (2013)), that is cleaved and activated by ADAM10 to induce cancer growth and metastatic diffusion (M. Trerotola et al., Trop-2 cleavage by ADAM10 is an activator switch for cancer growth and metastasis. Neoplasia 23, 415-428 (2021); E. Guerra et al., Trop-2 induces ADAM10-mediated cleavage of E-cadherin and drives EMT-less metastasis in colon cancer. Neoplasia 23, 898-911 (2021)). This cleavage causes a spatial rearrangement of the extracellular portion of Trop-2 and exposes hidden, activation-specific epitopes. We succeeded in generating monoclonal antibodies (mAbs) capable of recognizing these cancer-exposed sites. Methods: Most Trop-2-targeted approaches are based on anti-Trop-2 mAbs that recognize a single immunodominant epitope. Such mAbs have limited therapeutic index. We removed the immunodominant epitope by deletion mutagenesis, and optimized breadth of immune response, by immunizing mice with recombinant proteins with diverse end-glycosylation patterns. Screening for cancer-specific mAbs was performed on mutagenized Trop-2, by cell-based ELISA assays, bio-layer interferometry, flow cytometry and fluorescence microscopy. Results: mAbs were obtained that efficiently bound Trop-2 expressing cancer cells and were able to inhibit cell growth in vitro. We humanized the 2G10 mAb into the Hu2G10 IgG1, with an affinity for recombinant, cancer-specific, cleaved Trop-2 that is several orders of magnitude higher than existing competitor mAbs. In vivo the naked Hu2G10 antibody was effective at inhibiting the growth of breast, colon, ovary and prostate cancers. In the ADC setting Hu2G10 drove high anticancer specificity and effectiveness. Conclusions: Hu2G10 shows unprecedented specificity and efficacy, for best-in-class anti-Trop-2 anticancer therapy.