Heterogeneity in <i>NECTIN4</i> Expression Across Molecular Subtypes of Urothelial Cancer Mediates Sensitivity to Enfortumab Vedotin.

Abstract

Enfortumab vedotin (EV) is an antibody-drug conjugate (ADC) targeting NECTIN4 (encoded by the <i>PVRL4/NECTIN4</i> gene) approved for treatment-refractory metastatic urothelial cancer. Factors that mediate sensitivity or resistance to EV are unknown. In this study, we sought to (i) examine heterogeneity of <i>NECTIN4</i> gene expression across molecular subtypes of bladder cancer and (ii) determine whether NECTIN4 expression mediates EV sensitivity or resistance. Molecular subtyping and <i>NECTIN4</i> expression data from seven muscle-invasive bladder cancer clinical cohorts (<i>n</i> = 1,915 total specimens) were used to assess <i>NECTIN4</i> expression across molecular subtypes. The outcome of the transcriptomic analysis was relative <i>NECTIN4</i> expression in the consensus molecular subtypes of bladder cancer. Expression of <i>NECTIN4</i> was validated in bladder cancer cell lines. NECTIN4 was stably overexpressed or knocked down in basal and luminal bladder cancer cell lines and EV drug sensitivity assays were performed, as measured by cell proliferation and clonogenic assays. <i>NECTIN4</i> expression is heterogenous across molecular subtypes of bladder cancer and significantly enriched in luminal subtypes. <i>NECTIN4</i> expression is positively correlated with luminal markers <i>GATA3, FOXA1</i>, and <i>PPARG</i> across all cohorts. <i>NECTIN4</i> expression is both necessary and sufficient for EV sensitivity in luminal and basal subtypes of urothelial bladder cancer cells. Downregulation of <i>NECTIN4</i> leads to EV resistance. Sensitivity to EV is mediated by expression of <i>NECTIN4</i>, which is enriched in luminal subtypes of bladder cancer. These findings may have implications for biomarker development, patient selection, and the inclusion of molecular subtyping in ongoing and future EV clinical trials.<i>See related commentary by Teo and Rosenberg, p. 4950</i>.