Epigenetic variation in the mu-opioid receptor gene in infants with neonatal abstinence syndrome

Abstract

Objective: Neonatal abstinence syndrome (NAS) from in utero opioid exposure is highly variable with genetic factors appearing to play an important role. Epigenetic changes in cytosine:guanine (CpG) dinucleotide methylation can occur after drug exposure and may help to explain NAS variability. We correlated DNA methylation levels in the mu-opioid receptor (OPRM1) promoter in opioid-exposed infants with NAS outcomes.

Study design: DNA samples from cord blood or saliva were analyzed for 86 infants who were being treated for NAS according to institutional protocol. Methylation levels at 16 OPRM1 CpG sites were determined and correlated with NAS outcome measures, including need for treatment, treatment with ≥ 2 medications, and length of hospital stay. We adjusted for covariates and multiple genetic testing.

Results: Sixty-five percent of infants required treatment for NAS, and 24% required ≥ 2 medications. Hypermethylation of the OPRM1 promoter was measured at the -10 CpG in treated vs nontreated infants (adjusted difference δ = 3.2% [95% CI, 0.3-6.0%], P = .03; nonsignificant after multiple testing correction). There was hypermethylation at the -14 (δ = 4.9% [95% CI, 1.8%-8.1%], P = .003), -10 (δ = 5.0% [95% CI, 2.3-7.7%], P = .0005), and +84 (δ = 3.5% [95% CI, 0.6-6.4], P = .02) CpG sites in infants requiring ≥ 2 medications, which remained significant for -14 and -10 after multiple testing correction.

Conclusions: Increased methylation within the OPRM1 promoter is associated with worse NAS outcomes, consistent with gene silencing.

Figure 1. OPRM1 Promoter Region

The OPRM1 gene promoter region is shown with the two CpG islands boxed in and CpG dinucleotides indicated as ∣. The major transcription start site is indicated by the arrow, located −253 upstream of the ATG translation start site. The sequence of the amplified CpG island is also shown with the 16 CpG sites analyzed for cytosine methylation (bold) and their position relative to the ATG translation start site (underlined) indicated. Three putative Sp1 transcription factor binding sites are boxed.

Figure 2

Percent methylation of 16 CpG sites within the OPRM1 promoter in opioid-exposed infants who were treated (n=65) versus non-treated (n=21) for NAS. CpG sites in Sp1 transcription factor binding sites are indicated. * p<0.05 in bivariable analyses.

Figure 3

Percent methylation of 16 CpG sites within the OPRM1 promoter in opioid-exposed infants treated with ≥2 medications (n=21) versus <2 medications (n=65) for NAS. CpG sites in Sp1 transcription factor binding sites are indicated. * p<0.05 in bivariable analyses.