Constitutive activation of interferon signaling pathways has been reported in colorectal cancer (CRC), leading to a strong CD8+ T cell response through stimulation of NLRC5 expression. Primed CD8+ T cell expansion, however, may be negatively regulated by PD-L1 expression. Additionally, aberrant PD-L1 expression enables cancer cells to escape the immune attack. Our study aimed to select potential regulatory variants in the NLRC5 and PD-L1 genes by using several online in silico tools, such as UCSC browser, HaploReg, Regulome DB, Gtex Portal, microRNA and transcription factor binding site prediction tools and to investigate their influence on CRC risk in a Czech cohort of 1424 CRC patients and 1114 healthy controls. Logistic regression analysis adjusted for age and gender reported a moderate association between rectal cancer risk and two NLRC5 SNPs, rs1684575 T>G (OR: 1.60, 95% CI: 1.13-2.27, recessive model) and rs3751710 (OR: 0.70, 95% CI: 0.51-0.96, dominant model). Given that a combination of genetic variants, rather than a single polymorphism, may explain better the genetic etiology of CRC, we studied the interplay between the variants within NLRC5, PD-L1 and the previously genotyped IFNGR1 and IFNGR2 variants, to evaluate their involvement in the risk of CRC development. Overall we obtained 18 pair-wise interactions within and between the NLRC5 ad PD-L1 genes and 6 more when IFNGR variants were added. Thirteen out of the 24 interactions were below the threshold for the FDR calculated and controlled at an arbitrary level q*<0.10. Furthermore, the interaction IFNGR2 rs1059293 C>T-NLRC5 rs289747 G>A (P<0.0001) remained statistically significant even after Bonferroni correction. Our data suggest that not only a single genetic variant but also an interaction between two or more variants within genes involved in immune regulation may play important roles in the onset of CRC, providing therefore novel biological information, which could eventually improve CRC risk management but also PD-1-based immunotherapy in CRC.

Biomedical Center Faculty of Medicine in Pilsen Charles University Prague Pilsen Czech Republic

Center for Primary Health Care Research Clinical Research Center Lund University Malmö Sweden

Department of Immunology Interfaculty Institute for Cell Biology University of Tübingen Tübingen Germany

Department of Molecular Biology of Cancer Institute of Experimental Medicine of the Czech Academy of Sciences Prague Czech Republic

Department of Surgery 1st Medical Faculty Charles University and Thomayer Hospital Prague Czech Republic

Division of Molecular Genetic Epidemiology German Cancer Research Center Heidelberg Germany

Institute of Biology and Medical Genetics 1st Medical Faculty Charles University Prague Czech Republic

Molecular and Genetic Epidemiology Italian Institute for Genomic Medicine Turin Italy

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