Exome-wide association analysis identifies novel risk loci for alcohol-associated hepatitis

Abstract

Background and aims: Alcohol-associated hepatitis (AH) is a clinically severe, acute disease that afflicts only a fraction of patients with alcohol use disorder. Genomic studies of alcohol-associated cirrhosis (AC) have identified several genes of large effect, but the genetic and environmental factors that lead to AH and AC, and their degree of genetic overlap, remain largely unknown. This study aims to identify genes and genetic variations that contribute to the development of AH.

Approach and results: Exome-sequencing of patients with AH (N=784) and heavy drinking controls (N=951) identified an exome-wide significant association for AH at patalin-like phospholipase domain containing 3, as previously observed for AC in genome-wide association study, although with a much lower effect size. Single nucleotide polymorphisms (SNPs) of large effect size at inducible T cell costimulatory ligand ( ICOSLG ) (Chr 21) and TOX4/RAB2B (Chr 14) were also exome-wide significant. ICOSLG encodes a co-stimulatory signal for T-cell proliferation and cytokine secretion and induces B-cell proliferation and differentiation. TOX high mobility group box family member 4 ( TOX4 ) was previously implicated in diabetes and immune system function. Other genes previously implicated in AC did not strongly contribute to AH, and the only prominently implicated (but not exome-wide significant) gene overlapping with alcohol use disorder was alcohol dehydrogenase 1B ( ADH1B ). Polygenic signals for AH were observed in both common and rare variant analysis and identified genes with roles associated with inflammation.

Conclusions: This study has identified 2 new genes of high effect size with a previously unknown contribution to alcohol-associated liver disease and highlights both the overlap in etiology between liver diseases and the unique origins of AH.

Graphical abstract

FIGURE 1

Exome wide association to alcohol-associated hepatitis of 160,074 common SNVs from exome sequencing. (A) Manhattan plot of −log10 raw p-values. The red line represents the exome-wide significance threshold of p=7×10⁻⁷. The blue line represents suggestive significance at p=10⁻⁵; (B) Quantile-quantile (QQ)-plot of randomly expected versus observed p-values, with 0.95 CI. λ=1.040.

FIGURE 2

Enrichment in alcohol-associated hepatitis (AH) of exome-wide significant and subsignificant genes detecting potential types of polygenic contributions to AH. In each GSEA graph, the top p-values from 18,705 genes are rank ordered, and cumulative enrichment in AH for the nonreference (alternative) allele against rank is plotted. Both positive (pink) and negative (green) enrichment values indicate association of the pathway to AH, positive values reflecting greater risk effects of the nonreference alleles, and negative values reflecting protection conferred by the nonreference alleles. p-values were estimated empirically via permutation analyses with 10⁴ trials, and if p-value < 10^-4, a unidirectional enrichment score of that magnitude was not observed in any of the 10⁴ trials. (A) p-values of 227 liver disease-related genes precompiled from PheWeb and genome-wide association study Catalog (v1.0.3) and against all rank-ordered 18,705 genes, showing polygenic enrichment of AH for liver disease genes (NES=1.73, p<10⁻⁴), with symmetrical and thus approximately equivalent contributions of reference and alternative alleles. (B) Enrichment (NES=2.28, p<10⁻⁴) of “vitamin D sensitive calcium signaling in depression,” a 36 gene Wikipathway (WP4698). This enrichment is unidirectional, being driven by the risk effects of alternative alleles. (C) Enrichment (NES=2.23, p<10⁻⁴) of “pyrimidine metabolism and related diseases,” a 20 gene Wikipathway (WP4225). This enrichment is also unidirectional, being driven by the risk effects of alternative alleles. (D) “SREBP signaling,” a 69 Wikipathway (WP1982) was significantly enriched at the bottom of ranked genes (NES=−2.14, p<0.0001). (E) Enrichment (NES=1.97, p=0.0042) of “glycerolipids and glycerophospholipids,” a 22-gene Wikipathway (WP4722). This enrichment is also unidirectional, being driven by risk effects of alternative alleles. (F) Moderate significant enrichment of “Inflammation” genes from The Human Phenotype Ontology (HP:0004386, HP:0012115, HP:0030151, HP:0033196, HP:0200123) in AH (NES=−1.53, p=0.006). The bottom rank metric is shared by each panel. (G) Modest enrichment in AH of “alcohol drinking” (288 genes) implicated by genome-wide association study Catalog (EFO_0004329, NES=1.494, p=0.013). (H) 184 “alcohol dependence” genes from genome-wide association study Catalog (MONDO_0007079) was not enriched either (NES=1.245, p=0.136). Abbreviation: NES, Normalized Enrichment Score.