Triple-negative breast cancer PDX models with acquired resistance to Enhertu ® : A preclinical platform to uncover mechanisms of escape

Triple-negative breast cancer (TNBC) remains an aggressive and heterogeneous subtype with a poor prognosis. Although the antibody-drug conjugate Enhertu® (trastuzumab deruxtecan, T-DXd) has shown clinical activity in a subset of TNBCs with low HER2 expression, the rapid emergence of resistance limits its long-term efficacy. Robust preclinical models are therefore essential to investigate resistance mechanisms and evaluate novel therapeutic combinations. Within our panel of 40 TNBC patient-derived xenografts (PDXs), 14 were exposed in vivo to Enhertu®, of which 11 were sensitive. Among these, some PDX models were treated until the development of acquired resistance. To date, we have established five resistant derivatives from three initially sensitive models through continuous T-DXd exposure. Transcriptomic profiling was performed by RNA sequencing (HTSeq), followed by DESeq-based normalization and differential expression analysis. Functional enrichment analyses focused on predefined pathways related to endocytosis, receptor recycling, lysosomal trafficking, drug efflux, and DNA repair. Differential expression analysis revealed hundreds of significantly modulated genes. HER2 expression was assessed by immunohistochemistry, and RNA-Seq expression levels and mutational status of lysosomal cathepsins (CTSD, CTSB, CTSL) were examined in detail, given their potential involvement in the observed resistance mechanisms. In addition, resistant models were tested with HER2-targeting agents and a topoisomerase I inhibitor to evaluate the respective contributions of the two pharmacological targets of Enhertu® in the resistance mechanisms. Overall, resistance to Enhertu® in TNBC PDX models involves multifactorial adaptations, including signaling pathway reprogramming, altered drug efflux, and modulation of DNA repair. This collection of resistant models represents a valuable preclinical resource to investigate resistance mechanisms and to support the development of novel therapeutic strategies.