Background: HER2 overexpression and amplification define ~20% of breast cancers (BC) that are currently treated using HER2-specific monoclonal antibodies (mAb), such as trastuzumab. While multiple studies have demonstrated trastuzumab’s efficacy to be largely dependent upon triggering innate and adaptive immune responses, its dominant immune mechanism of therapeutic action (MOA) remains unclear. As trastuzumab’s efficacy is subverted in advanced immunosuppressive cancers, an understanding of its MOA and strategy to boost its immune effect is of critical clinical interest and scientific significance for immunologically enhancing other targeted mAb treatments.

Methods: In this study, we developed a novel fully murine trastuzumab mAb (mTrastuzumab) to determine the MOA for HER2-mAb antitumor efficacy. We utilized these mAbs in multiple models that allowed us to interrogate antibody-dependent cell cytotoxicity and phagocytosis (ADCC and ADCP), as well as complement-dependent cytotoxicity (CDC) in vitro and in vivo. To determine the antitumor efficacy of these HER2 mAbs, we employed orthotopic HER2+ murine cells (transformed by HER2) in immunocompetent, FCGR KO, and immunocompetent settings, as well as utilized a unique transgenic HER2+ BC model driven by an oncogenic isoform of HER2 to best simulate an endogenous mammary tumor immune microenvironment (Turpin et al., 2016).

Results: We found that mTrastuzumab significantly suppressed tumor growth and stimulated the infiltration of tumor-associated macrophages (TAMs), without affecting other immune cell types in the microenvironment. In contrast to other studies, we also found that this antitumor activity did not require adaptive immunity or NK cells, but did require activating Fc-gamma receptor engagement, implicating ADCP as the potential dominant MOA. Consistent with these findings, we further demonstrated that mTrastuzumab activated mFcγR4 and mFcγR3 signaling and elicited significant macrophage-mediated ADCP of HER2+ BC in vitro, but not ADCC or CDC. To test if enhanced ADCP could confer more effective antitumor immunity, we combined mTrastuzumab with blockade of the ADCP checkpoint, CD47 (“don’t eat me signal”), and found that this combination significantly increased mTrastuzumab-mediated ADCP in vitro without altering ADCC. Critically, we also found that this combination significantly enhanced TAM infiltration in vivo, as well as stimulated stronger antitumor efficacy and prolonged survival in highly immunosuppressive tumor microenvironments.

Conclusion: Our study demonstrates the major MOA of clinically relevant HER2 mAbs requires the engagement with TAMs to elicit ADCP of tumor cells, which could be significantly enhanced by CD47 blockade. We conclude CD47 blockade could unleash the full potential of HER2 mAb therapy by abrogating the innate immunosuppressive signals on macrophages to stimulate immunity and enhance antitumor efficacy.

Citation Format: Li-Chung Tsao, Herbert Kim Lyerly, Zachary Hartman. Trastuzumab therapy suppresses HER2+ breast tumor growth through inducing ADCP by tumor-associated macrophages and synergizes with CD47 checkpoint blockade [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr B03.