ABSTRACT/SUMMARY Nearly 10% of patients with metastatic colorectal cancer (mCRC) have progression of disease involving the spread of metastatic cancer to the peritoneal cavity. These patients generally have the worst outcomes for all patients with mCRC. A recent phase III multicenter clinical trial has shown that single-agent immune checkpoint inhibitor (ICI) has greater efficacy and better safety profile than standard cytotoxic chemotherapy regimens for patients with mCRC. Importantly, indications to receive ICI therapy are dependent on the measurement of high tumor mutation burden (TMB), neoantigen burden, and microsatellite instability (MSI) status. These direct and indirect measures of genomic instability predict clinical response to ICIs, however, only 5% of patients are currently eligible to receive these therapies. Patients with mCRC with peritoneal spread of disease may undergo major cytoreductive surgery (CRS), during which all gross sites of disease are resected. CRS is combined with heated intraperitoneal chemotherapy (HIPEC) with mitomycin C at 43°C to eradicate microscopic disease. We hypothesize that major surgical inflammation from CRS and the heat and chemotherapy of HIPEC altogether increase genomic instability. Surgical inflammation is known to activate heat shock proteins (Hsps) which may destabilize its regulatory control over mismatch repair (MMR) proteins leading to deficient DNA repair. This condition combined with the DNA damaging effects of mitomycin c and heat may lead to high TMB, increased neoantigens, and altered MSI in occult microscopic mCRC cells. Our overarching hypothesis is that patients undergoing CRS/HIPEC will be eligible to receive and will benefit from ICI therapy. We propose to obtain blood samples and surgical tissues before and after CRS/HIPEC from patients with mCRC. These specimens will be assessed for changes in TMB and neoantigens (Aim 1) and MMR/MSI (Aim 2). From the surgical tissues, we will also create paired patient-derived organoids (PDOs) from which we will investigate Hsps, MMR proteins, and MSI. Our prior studies using PDOs from CRS/HIPEC patients support their use to assess sensitivities to ICI therapies before and after CRS/HIPEC. We propose the following specific aims: Specific Aim 1. To measure alterations in TMB and neoantigens as a result of the heat and DNA damaging effects of mitomycin C in mCRC patients undergoing CRS/HIPEC. Specific Aim 2. To determine whether surgical inflammation combined with heat and DNA damaging effects of mitomycin C from CRS/HIPEC disrupt Hsp-stabilization of MMR proteins to increase MSI.

PROJECT NARRATIVE Our proposal examines whether the major inflammation, heat, and chemotherapy of cytoreductive surgery (CRS) and heated intraperitoneal chemotherapy (HIPEC) increases the genomic instability of patients with metastatic colorectal cancer (mCRC). Our ultimate objective is to identify changes in tumor mutation burden, neoantigens, or microsatellite instability as a result of CRS/HIPEC, so that patients with mCRC are eligible to receive efficacious and safe immune therapies.