Immunoepigenetic targeting of MHC regulators in FAP

Major histocompatibility complex class I and class II (MHC-I/II) members are critical for antigen presentation and adaptive immune responses but are downregulated in sporadic and genetic cases of duodenal and colorectal cancer (CRC), such as Familial Adenomatous Polyposis (FAP). There is an urgent need for novel approaches to reverse MHC-I/II silencing and overcome tumor immune evasion, starting at the adenoma stage. Based on the prior Critique, standard-of-care sulindac and erlotinib (SUL, ERL), and a new histone methyl `reader' inhibitor MAK683 that shows clinical promise, will be repurposed for immune-based interception of FAP. From the rigor of prior research and extensive preliminary data, the revised CENTRAL HYPOTHESIS is that low-dose ERL+SUL combinations along with MAK683 upregulate MHC-I/II components to enhance immune surveillance via CD8 and CD4 T cell recruitment into preneoplastic and later stages of FAP. Low-dose, non-cytotoxic test agents and their combinations augment MHC complexes at the cell surface to re-engage the host immune system for anticancer efficacy, and will be translatable to FAP patients via oral administration with a good safety margin. ✓Aim 1 MECHANISMS Aim 1A Perform dose-response and time-course studies with ERL, SUL, MAK683 and selected combinations for MHC-I/II upregulation in human and murine CRC, duodenal cancer, and adenoma cells compared with normal cells. Aim 1B Corroborate for ERL, SUL and MAK683 the increased cell surface occupancy of MHC-I/II complexes, coupled to enhanced cancer cell killing in CD8+ and CD4+ T cell co-cultures. Aim 1C Overexpress, silence, and pharmacologically inhibit mechanistic targets (e.g., EED, NLRC5, CIITA) to identify key roles in MHC-I/II regulation and immune evasion in adenoma and later stages relevant to FAP. ✓Aim 2 PRECLINICAL Aim 2A In the polyposis in rat colon (Pirc) model, perform biomarker analyses with MAK683 via i.g. drug administration and corroborate histone methylation changes and immune target modulation in colon and duodenal polyps for up to 14 days. Aim 2B Test the antitumor efficacy and safety of MAK683 alone and combined with ERL or SUL in the Pirc model. Mechanistic targets will be validated via RT-qPCR, immunoblot and multiplex CyTOF as predictors of antitumor efficacy. Aim 2C Test selected drug combinations in ApcMin/+, ApcMin/+Nlrc5–/– and ApcMin/+Ciita–/– mice for insights into Nlrc5- and Ciita-dependent/independent mechanisms. ✓Aim 3 TRANSLATIONAL Aim 3A In colon and duodenal biopsies from FAP patients, corroborate the predicted interrelationships between MHC-I/II members (B2M, CD74), coactivators (NLRC5, CIITA), mechanistic targets (EED) and epigenetic readouts (histone H3K27me3) in epithelial and immune cell components, via CyTOF. Aim 3B In FAP patient organoids, define MAK683, ERL and SUL doses that increase MHC-I/II-dependent gene expression, and examine autologous effector functions and the capacity to kill tumor organoids in T cell cocultures. Aim 3C In organoids from Aim 3B, test the hypothesis that MAK683, ERL and SUL increase MHC-I/IIbound peptides using MS-based peptidomic approaches to aid in precision immunoepigenetic clinical strategies.