Vivekanandhan, Sneha

Abstract In this study, we tested a novel approach of “repurposing” a biomarker typically associated with breast cancer for use in melanoma. HER2/neu is a well characterized biomarker in breast cancer for which effective anti-HER2/neu therapies are readily available. We constructed a lentivirus encoding c-erb-B2 (the animal homolog to HER2/neu). This was used to transfect B16 melanoma in vitro for use in an orthotopic preclinical mouse model, which resulted in expression of c-erb-B2 as a neoantigen target for anti-c-erb-B2 monoclonal antibody (7.16.4). The c-erb-B2-expressing melanoma was designated B16/neu. 7.16.4 produced statistically significant in vivo anti-tumor responses against B16/neu. This effect was mediated by NK-cell antibody-dependent cell-mediated cytotoxicity. To further model human melanoma (which expresses <5% HER2/neu), our c-erb-B2 encoding lentivirus was used to inoculate naïve (wild-type) B16 tumors in vivo, resulting in successful c-erb-B2 expression. When combined with 7.16.4, anti-tumor responses were again demonstrated where approximately 40% of mice treated with c-erb-B2 lentivirus and 7.16.4 achieved complete clinical response and long-term survival. For the first time, we demonstrated a novel strategy to repurpose c-erb-B2 as a neoantigen target for melanoma. Our findings are particularly significant in the contemporary setting where newer anti-HER2/neu antibody-drug candidates have shown increased efficacy.

One of the most impactful biologics for the treatment of breast cancer is the humanized monoclonal antibody, trastuzumab, which specifically recognizes the HER2/neu (HER2) protein encoded by the ERBB2 gene. Useful for both advanced and early breast cancers, trastuzumab has multiple mechanisms of action. Classical mechanisms attributed to trastuzumab action include cell cycle arrest, induction of apoptosis, and antibody-dependent cell-mediated cytotoxicity (ADCC). Recent studies have identified the role of the adaptive immune system in the clinical actions of trastuzumab. Despite the multiple mechanisms of action, many patients demonstrate resistance, primary or adaptive. Newly identified molecular and cellular mechanisms of trastuzumab resistance include induction of immune suppression, vascular mimicry, generation of breast cancer stem cells, deregulation of long non-coding RNAs, and metabolic escape. These newly identified mechanisms of resistance are discussed in detail in this review, particularly considering how they may lead to the development of well-rationalized, patient-tailored combinations that improve patient survival.

PLEXIND1 is upregulated in several cancers, including pancreatic ductal adenocarcinoma (PDAC). It is an established mediator of semaphorin signaling, and neuropilins are its known coreceptors. Herein, we report data to support the proposal that PLEXIND1 acts as a transforming growth factor beta (TGFβ) coreceptor, modulating cell growth through SMAD3 signaling. Our findings demonstrate that PLEXIND1 plays a pro-tumorigenic role in PDAC cells with oncogenic KRAS (KRASmut). We show in KRASmut PDAC cell lines (PANC-1, AsPC-1,4535) PLEXIND1 downregulation results in decreased cell viability (in vitro) and reduced tumor growth (in vivo). Conversely, PLEXIND1 acts as a tumor suppressor in the PDAC cell line (BxPC-3) with wild-type KRAS (KRASwt), as its reduced expression results in higher cell viability (in-vitro) and tumor growth (in vivo). Additionally, we demonstrate that PLEXIND1-mediated interactions can be selectively disrupted using a peptide based on its C-terminal sequence (a PDZ domain-binding motif), an outcome that may possess significant therapeutic implications. To our knowledge, this is the first report showing that (1) PLEXIND1 acts as a TGFβ coreceptor and mediates SMAD3 signaling, and (2) differential roles of PLEXIND1 in PDAC cell lines correlate with KRASmut and KRASwt status.

Abstract The increase in fatty acid metabolism is a hallmark of cancer. Stearoyl CoA desaturase (SCD) is an endoplasmic reticulum (ER) enzyme that catalyzes D9 cis désaturation of fatty acyl-CoA substrates. One of its isoforms, SCD1, is overexpressed in several cancers including breast cancer. The higher expression of SCD1 in breast cancer patients correlates with significantly shorter relapse-free and overall survival rates, indicating that SCD1 may be a therapeutic target. We have previously developed and published a novel SCD1 inhibitor, SSI4. Metabolic reprogramming facilitates tumor growth by promoting immune suppression in tumors. Fundamental metabolic processes like fatty acid (FA) metabolism are involved in T cell activation and differentiation and their modulation can differentially affect the development of T helper cell lineages. FA synthesis and oxidation are essential for the development and functioning of CD8+ T memory cells and are contributing factors that influence CD4+ T effector and Treg cell development. These associations between metabolism and immune cells lead us to our hypothesis that aberrant de novo lipogenesis is linked to immune suppression. Thus, SCD1 inhibitors should increase anti-tumor immunity. We evaluated the effect of SSI4-mediated SCD1 inhibition in two mouse triple negative breast cancer cell lines. The data showed that SSI4 stimulates calreticulin (CRT) translocation to cell membrane. CRT is an ER resident protein that translocates to the cell surface under ER stress conditions. Cell surface expression of CRT is known to induce immune responses including phagocytosis, translocation of cancer antigen to the cell surface, and apoptosis. We are currently studying the effect of CRT translocation in our model. A major focus of our project is to assess the efficacy of combination treatment of SSI4 and an immune checkpoint blockade. To this end, this combination therapy synergistically blocked E0771 tumor growth in vivo. Further, to understand the immune responses generated, we performed immunohistochemical analysis of the tumors (day 20 post-treatment) indicating that the combination therapy resulted in higher membrane localized CRT, supporting our rationale that CRT translocation is involved in mediating the anti-tumor responses. Flow cytometry analysis indicated that the combination treatment increased immune cell infiltration. Specifically, SSI4 treatment induced infiltration of double negative T cells, natural killer cells and macrophages, suggesting that it promoted a pro-inflammatory immune response. SSI4 treatment down regulated immune suppressive cells including Tregs, dendritic cells and mesenchymal derived stem cells. The response of cancers with low and intermediate tumor mutational burden such as breast cancer to immune checkpoint inhibition is limited. Our data indicates that combination treatment of SSI4 and anti PD1 is a promising therapeutic strategy. Citation Format: Sneha Vivekanandhan, Justyna Trynda, Laura A. Marlow, Barath shreeder, James L. Miller, Adam M. Kase, Winston Tan, Keith L. Knutson, John A. Copland. SCD-1 blockade sensitizes triple negative breast cancer to immune checkpoint inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1484.