Identification and characterization of immunogenic neoantigens in biliary cancer (BC) and pancreatic cancer (PC).
Authors: Battaglin Francesca , Elliott Andrew , Xiu Joanne , Algaze Sandra , Wang Jingyuan , Jayachandran Priya , Soni Shivani , Ashouri Karam , Wong Alexandra , Mittal Pooja , Lo Jae Ho , Torres-Gonzalez Lesly , Zhang Wu , Weinberg Benjamin Adam , Goel Sanjay , Lou Emil , El-Khoueiry Anthony B. , Lenz Heinz-Josef 
Published: January 2024

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Background: Recognition of tumor neoantigens by autologous T cells activates immune surveillance and has been reported to promote sensitivity to immune checkpoint inhibitors (ICI) in mismatch repair deficient (MMRd)/microsatellite instability high (MSI-H) tumors. Neoantigen-targeted reactivity has also been reported in microsatellite stable (MSS) tumors. Neoantigens are emerging targets for novel immunotherapy strategies, including tumor vaccines, in BC and PC. We aimed to comprehensively assess the spectrum of immunogenic neoantigens in BC and PC. Methods: 3728 tumor specimens (1389 BC; 2339 PC) tested at Caris Life Sciences (Phoenix, AZ) with NextGen Sequencing on DNA (720-gene panel) and RNA (whole transcriptome) were analyzed. MSI status was determined by immunohistochemistry of MMR protein and/or NGS. Immune epitope prediction was performed on translated peptide sequences harboring detected mutations using the NetMHCpan v4.0 method in the Immune Epitope Database, with HLA genotyping performed using arcasHLA. Immune/stromal cell abundance in the tumor microenvironment (TME) was quantified using the MCP Counter method. Gene expression profiles were analyzed for a transcriptional signature predictive of response to immunotherapy (T cell-inflamed signature, TIS). Results: MMRd/MSI-H rate was 1.8% in BC and 1.4% in PC. 117219 unique peptide:allele interactions with predicted binding-level affinity for patient-specific HLA alleles were identified (48781 in BC; 71182 in PC). MMRd/MSI-H tumors had higher neoantigen load at all affinity levels compared to MSS. Only 4 recurrent neoantigens with binding affinity (observed in >10 samples) were identified in BC and derived from mutations in KRAS (3/4) and IDH1 (1/4), while 78 were found in PC, mostly associated with mutations in KRAS (53/78) and TP53 (16/78). The frequency of individual neoantigens was particularly low in MSS BC (< 2%). Across both cancer types, TIS scores positively correlated with the abundance of immune cell populations in the TME, notably cytotoxic lymphocytes (r > 0.40). Recurrent neoantigens associated with highest average TIS scores resulted from mutation of KRAS (G12D/V, 1.8%/1.5% of samples, respectively) in BC yet mean TIS scores were low (~60th percentile overall). Similarly, TP53 (Y220C/R273C, 0.6%/0.7%, respectively) and CDKN2A (multiple variants, 0.5%) were associated with the highest yet relatively low mean TIS scores in MSS PC (~73th percentile) compared to mean TIS scores associated with MSH3 (K383fs, 12%) and KRAS (G12D, 12%) in MMRd/MSI-H PC (~90th percentile). Conclusions: This is the largest study to investigate the landscape of immunogenic neoantigens in BC and PC. The frequency of high-level binding affinity neoantigens was relatively low and associated with relatively lower TIS scores in MSS tumors, which may contribute to the immunogenic cold TME characterizing these tumor types.