Beyond Intended Mechanisms: Immune Interactions of Trop2-Directed Antibody-Drug Conjugates and a HER2-Directed Self-Replicating RNA Vaccine in Breast Cancer

Abstract

Breast cancer immunotherapeutics are generally developed and evaluated according to their intended mechanisms of action, yet their interactions with the tumor-immune microenvironment are often broader and less well understood. This dissertation addresses the problem that unintended or overlooked immune interactions may substantially influence therapeutic efficacy, but remain insufficiently characterized for both antibody-drug conjugates (ADCs) and self-replicating RNA cancer vaccines. To investigate this issue, I studied two breast cancer immunotherapy platforms: Trop2-directed ADCs and a Venezuelan equine encephalitis virus (VEEV)-based viral replicon particle self-replicating RNA (VRP-srRNA) vaccine targeting HER2.A combination of in vitro, in vivo, and translational approaches was used. For studies of Trop2-directed ADCs, human Trop2 knock-in and knockout breast tumor cell lines and mouse tumor models were used to evaluate antigen-dependent and antigen-independent antitumor effects of Sacituzumab Govitecan (SG) and Datopotamab Deruxtecan (Dato-DXd). Additional functional assays were performed to assess Fcγ receptor activation, macrophage-mediated antibody-dependent cellular phagocytosis, and treatment-induced adaptive antitumor immunity. For studies of the VRP-srRNA HER2 vaccine, patient samples from a phase I clinical trial and complementary mouse vaccination models were analyzed to characterize vector-directed humoral and cellular immune responses and to determine how repeated homologous vaccination influenced subsequent boosting. These studies demonstrate that the activity of breast cancer immunotherapeutics is shaped not only by their intended targets, but also by unintended interactions with antigen-negative tumor cells, innate immune cells, and vaccine vector components. Trop2-directed ADCs exhibited context-dependent antigen-independent effects while retaining Fc-mediated immune function, and VRP-srRNA vaccination induced anti-vector immunity that limited the benefit of repeated homologous boosting on a short dosing schedule. Together, these findings show that bidirectional interactions between immunotherapeutic agents and the immune system are important determinants of therapeutic outcome and should be more deliberately considered in the design and optimization of breast cancer therapies.