Immunosuppressive impact of the GPCR/MALT1 protease axis in triple-negative breast cancer

PROJECT SUMMARY/ABSTRACT Breast cancer is the most common cancer in women. Triple-negative breast cancer (TNBC) lacks expression of estrogen receptor (ER-), progesterone receptor (PR-) and human epidermal growth factor receptor 2 (HER2-), the most common targetable markers in breast cancer. TNBC demonstrates the worst overall prognosis among

breast cancer subtypes due to lack of well-established targeted therapies. Because TNBC treatment relies heavily on non-specific therapies, a significant need exists to identify innovative ways to harness the tumor immune microenvironment to enable the effective use of immunotherapy. I propose to investigate how the intracellular protein MALT1 promotes immunosuppression in the tumor immune

microenvironment (TIME) in TNBC. MALT1 is the effector protein of the CARMA-BCL10-MALT1 (CBM) signalosome, a protein complex that is stimulated downstream of certain oncogenic G protein-coupled receptors (GPCRs) and induces pro-survival NF-κB transcriptional activation. MALT1 acts via two main biologic functions:

(1) as a molecular scaffold and (2) as a protease. Our laboratory has demonstrated that in TNBC cells, MALT1 protease activity promotes tumor progression and metastasis by inducing epithelial-to-mesenchymal transition (EMT). In many cancers, including TNBC, malignant cells undergoing EMT modulate the surrounding

microenvironment to polarize immune cells toward a cancer-promoting, immunosuppressive phenotype. Based on my preliminary data, I hypothesize that MALT1 protease activity within TNBC cells promotes tumor immune suppression through production of an immunosuppressive secretome. In Aim 1 of this proposal, I will analyze MALT1 regulation of the TNBC TIME by characterizing MALT1 protease-

dependent secretion of immunosuppressive cytokines from TNBC cells. Experimental techniques utilized in this aim will include CRISPR/Cas9 editing, mouse breast cancer models, tumor cell/immune cell co-culture, flow cytometry, single cell RNA sequencing, and Olink® proteomics. In Aim 2 of this proposal, I will evaluate a novel

MALT1 protease activity gene signature for its clinicopathologic utility in predicting TIME status in TNBC. Experimental techniques utilized in this aim will include use of patient-derived organoids, analysis of patient tumor specimens, multiplex multispectral immunofluorescence, and a series of in silico analyses.

In summary, this project evaluates MALT1 protease as a potential driver of tumor progression via its effects on the TNBC TIME. Completion of this proposal will provide me with rigorous research training by a remarkable team of scientists and physician-scientists. This training experience will prepare me for a career as a physician-

scientist who develops new and innovative strategies for the treatment of patients with malignant solid tumors.