Abstract In our previous studies, we have identified that HER2 monoclonal antibodies (mAb) function to stimulate antibody dependent cellular phagocytosis (ADCP) through innate pathways, which are critical to establish effective anti-tumor responses (Tsao et al., JCI-Insight, 2019). In our ongoing funded studies (R01 CA238217- 01A1), we are exploring if HER2-specific ADCP eliciting antibodies could be enhanced by innate ICB (targeting CD47) and adaptive ICBs (targeting CTLA4/PD1). These studies utilize an endogenous immune-tolerant model of HER2+ BC (Turpin et al., 2016) driven by an oncogenic human isoform of HER2, HER2Δ16. While this model is immune-tolerant to HER2 epitopes and generates HER2Δ16-driven tumors in an appropriate mammary microenvironment, it does not recapitulate the tumor heterogeneity seen clinically in HER2+ BCs, or permit the development of major resistance mechanisms to HER2 mAb therapies, which often occur in advanced HER2+ BC patients. To address these shortcomings and further our understanding of HER2 mAb therapies in HER2+ BC, we propose to utilize a novel HER2BOW mouse developed through the IMAT program (5R33CA191198;Lubkov et al., Nat. Cell Bio., in review). This model utilizes the stochastic activation of several different variants of HER2 (HER2-WT, HER2Δ16, and HER2-p95) within the mammary compartment to evolve heterogeneous HER2+ BCs with disparate pathologies. Critically, these different pathologies stem from the expression of these alternate HER2 isoforms, which may explain HER2+ tumor heterogeneity observed clinically. Moreover, this model contains HER2-p95, which lacks the extracellular binding domain for HER2 mAbs to function as a driver of resistance. However, HER2-specific mAbs are able to prime T cell responses against HER2 intracellular epitopes within the heterogeneous context of clinical HER2+ BCs, thus its importance in mediating resistance in a heterogeneous context is unclear. Moreover, it is unknown if strategies to enhance HER2 mAb stimulated immunity could overcome HER2-p95 mediated resistance thorough enhanced HER2 immune cross-priming. We hypothesize that HER2-p95 expression confers resistance against HER2 mAb therapies within heterogeneous HER2+ BCs, but that dual (innate and adaptive) ICB-enhanced HER2 mAb therapies could overcome this resistance through enhance HER2 epitope cross-priming. Guided by our preliminary data, this hypothesis will be tested by utilizing the HER2BOW model that can be interrogated with the following combinations that comprise our specific aims: 1) HER2+CD47 mAbs and 2) HER2/CD47 mAbs + CTLA4/PD1 ICB combinations. These studies will determine if responsiveness is due to a lack of local immunity related to substantial intratumoral heterogeneity, or due to the rapid development of resistance from alternate HER2 oncogenic isoforms. Critically, they will also validate the therapeutic utility of the HER2BOW model and the therapeutic potential of HER2 mAb+ICB combinations acros...