A statin-dependent QTL for GATM expression is associated with statin-induced myopathy

Abstract

Statins are prescribed widely to lower plasma low-density lipoprotein (LDL) concentrations and cardiovascular disease risk and have been shown to have beneficial effects in a broad range of patients. However, statins are associated with an increased risk, albeit small, of clinical myopathy and type 2 diabetes. Despite evidence for substantial genetic influence on LDL concentrations, pharmacogenomic trials have failed to identify genetic variations with large effects on either statin efficacy or toxicity, and have produced little information regarding mechanisms that modulate statin response. Here we identify a downstream target of statin treatment by screening for the effects of in vitro statin exposure on genetic associations with gene expression levels in lymphoblastoid cell lines derived from 480 participants of a clinical trial of simvastatin treatment. This analysis identified six expression quantitative trait loci (eQTLs) that interacted with simvastatin exposure, including rs9806699, a cis-eQTL for the gene glycine amidinotransferase (GATM) that encodes the rate-limiting enzyme in creatine synthesis. We found this locus to be associated with incidence of statin-induced myotoxicity in two separate populations (meta-analysis odds ratio = 0.60). Furthermore, we found that GATM knockdown in hepatocyte-derived cell lines attenuated transcriptional response to sterol depletion, demonstrating that GATM may act as a functional link between statin-mediated lowering of cholesterol and susceptibility to statin-induced myopathy.

Figure 1. Simvastatin treatment alters transcript expression in LCLs

Log change in expression following simvastatin- and control-exposed lymphoblastoid cell lines (n=480) displayed as a function of log sum of expression traits. Grey: genes for which expression was significantly changed in response to simvastatin exposure (N=5509/10105, 0.12±0.08 mean absolute log₂ change±SD, q<0.0001); Black: genes for which expression was not significantly changed (N=4686). Red: genes in the cholesterol biosynthesis pathway, all of which exhibited significant changes in expression.

Figure 2. Treatment-specific QTL associated with GATM expression

(a) Association of rs9806699 with quantile normalized GATM expression levels following (i) control exposure (not significant); (ii) simvastatin exposure (log₁₀BF=5.1, effect size = -0.43). (iii), fold change (log₁₀BF=5.7, effect size = -0.40); (iv), control versus simvastatin-exposed GATM expression (black: GG, N=225; red: GA, N=207; green: AA, N=48). Box height and whiskers described in Supplemental Methods. (b) SNPs associated with GATM expression (log₁₀BF, left y-axis); SNPs associated with statin-induced myopathy (red), significance threshold (dotted line) recombination rates (blue, right y-axis); Bottom panel: transcribed genes (green), DNAse I hypersensitive (DHS) sites and transcription factor binding sites (TFBS; black), predicted chromosomal enhancers (orange) and promoters (red) as identified in hepatocyte (HepG2), lymphoblastoid (GM12878) and myocyte (HSMM) cell lines.

Figure 3. GATM knockdown attenuated sterol-mediated induction in expression of SREBP-responsive genes

(a) Changes in transcript concentrations following sterol depletion via 24-hr exposure to lipoprotein deficient serum (LPDS)-containing media vs. standard FBS-containing media in hepatocyte-derived HepG2 (left, N=12) and Huh7 (right, N=12) cell lines. Asterisk indicates P<0.05 for the comparison of GATM versus NTC siRNA treated cells. (b) Fold changes in media accumulation of apolipoprotein B (ApoB) and apolipoprotein AI (ApoAI) following gene knockdown with GATM versus (NTC) siRNA in HepG2 cells (left, N=6-10) or Huh7 (right, N=4-6) cells under standard culture conditions. Error bars represent SEM.