Regulatory polymorphisms of MSH6, MSH2, FBXO11, and PPP1R21 genes affect survival of patients with immunotherapy-treated lung cancer

Abstract

Background: Immune checkpoint inhibitors (ICI) improved survival of patients with non-small cell lung cancer (NSCLC), yet many patients do not respond to treatment. The identification of markers for ICI response remains an unmet clinical need. This study hypothesizes that host genetics influences the response to ICI, contributing to the variability in efficacy among individuals.

Methods: We conducted a genome-wide association study (GWAS) in patients with NSCLC on ICI monotherapy with nivolumab, pembrolizumab, or atezolizumab, to identify germline variants associated with objective response rate (ORR), progression-free survival (PFS), and overall survival (OS) at 24 months after the start of ICI therapy. Genomic DNA was genotyped using Axiom Precision Medicine Research Arrays. Raw data were processed with Axiom Analysis Suite, and quality checked with PLINK software. Imputation to the whole genome was done on the Michigan Imputation Server. Association analyses were performed for ORR (logistic regression with PLINK2 software) and survival (Cox proportional hazards model, with GenAbel package in R environment), with appropriate covariates. Variants were annotated for functional significance using SNPnexus and FUMA. Post-GWAS analyses, including colocalization, were performed to explore the function of the identified variants. Their possible role as expression quantitative trait loci was investigated in different databases (GTEx, eQTLGen, TCGA).

Results: No genome-wide significant associations were found for ORR or PFS, while a locus on chromosome 2 (lead variant: rs111648355) showed near genome-wide significance (p value=6.3×10⁻⁸) for OS. Patients with minor alleles of these variants exhibited significantly worse OS (HR=5.1, 95% CI: 2.9 to 9.2). Functional annotation linked these variants to regulatory effects on genes including MSH2, MSH6, PPP1R21, FBXO11, and STON1. These genes play a role in mismatch repair, endosomal trafficking, or major histocompatibility complex class II regulation, and might influence the response to immunotherapy.

Conclusions: This study identifies an association between a genomic locus on chromosome 2 and OS in patients with NSCLC treated with ICI. Although these results need validation in larger cohorts and functional studies to elucidate the underlying mechanisms, they highlight the potential of germline variants as predictive biomarkers of response to ICI.

Keywords: Genetic; Genome; Immune Checkpoint Inhibitors; Non-Small Cell Lung Cancer.

Figure 1. Variables associated with PFS and OS in univariable Cox models. Kaplan-Meier (KM) survival curves (at 24 months of follow-up) of patients with lung adenocarcinoma treated with ICI, according to their ECOG performance status (a, d), programmed death-ligand 1 expression level in their tumors (b, e), and taken ICI (c, f). PFS and OS KM are reported in the left and right panels, respectively. Crosses denote censored samples. Below each plot are indicated the numbers of patients at risk in each group. Log-rank p values are shown. ECOG, Eastern Cooperative Oncology Group; OS, overall survival; PFS, progression-free survival.

Figure 2. A nearly significant locus on chromosome 2 is associated with overall survival of patients with immune checkpoint inhibitor-treated lung adenocarcinoma. Manhattan plot of the results of the genome-wide association for overall survival, at 24 months of follow-up, with age, sex, drug, and ECOG as covariates (n=150, due to missing ECOG data for two patients). Imputed variants are plotted along the x-axis according to their genomic position (GChr37, hg19 release). On the y-axis is reported the level of association (− log₁₀(p values)) with survival probability of each single nucleotide polymorphism. The horizontal red and blue lines represent the threshold of genome-wide significance (p< 5.0×10⁻⁸) and a suggestive threshold at p<1.0×10⁻⁵, respectively. In the upper-right corner the QQ plot of observed and expected p values is shown. Genomic inflation factor (λ) is reported. ECOG, Eastern Cooperative Oncology Group.

Figure 3. Regional plot of the locus on chromosome 2 identified in the genome-wide association. Plot spans the region from 47.15 Mbp to 49.25 Mbp (GRCh37/hg19), containing the mapped genes and the analyzed imputed variants. SNPs are plotted on the x‐axis according to their position on chromosome 2. Dot color represents the level of linkage disequilibrium, expressed as R² between each SNP and the lead variant (rs111648355, purple dot). SNP, single nucleotide polymorphism.

Figure 4. The evidence for shared variants between OS and gene expression is moderate. Colocalization plots for the (a) PPP1R21, (b) MSH2, (c) MSH6, and (d) STON1 genes using the whole-blood tissue expression data from the eQTLGen database (numbers of SNPs included in the analyses were 190); (e) colocalization plot for FBOX11, with expression data from, including 180 SNPs. Dots represent variants along the portion of chromosome 2 spanning from 48.0 to 48.2 Mb and are colored based on linkage disequilibrium (R²) with the lead variant of our genome-wide association for OS, rs111648355 (purple dot). Dot coordinates are defined, on the x-axis, by the p values (−log₁₀-transformed) of association with OS, whereas, on the y-axis, those with the gene expression levels. OS, overall survival; SNP, single nucleotide polymorphism.