Epigenetic Biomarkers of Prenatal Tobacco Smoke Exposure Are Associated with Gene Deletions in Childhood Acute Lymphoblastic Leukemia

Abstract

Background: Parental smoking is implicated in the etiology of acute lymphoblastic leukemia (ALL), the most common childhood cancer. We recently reported an association between an epigenetic biomarker of early-life tobacco smoke exposure at the AHRR gene and increased frequency of somatic gene deletions among ALL cases.

Methods: Here, we further assess this association using two epigenetic biomarkers for maternal smoking during pregnancy-DNA methylation at AHRR CpG cg05575921 and a recently established polyepigenetic smoking score-in an expanded set of 482 B-cell ALL (B-ALL) cases in the California Childhood Leukemia Study with available Illumina 450K or MethylationEPIC array data. Multivariable Poisson regression models were used to test the associations between the epigenetic biomarkers and gene deletion numbers.

Results: We found an association between DNA methylation at AHRR CpG cg05575921 and deletion number among 284 childhood B-ALL cases with MethylationEPIC array data, with a ratio of means (RM) of 1.31 [95% confidence interval (CI), 1.02-1.69] for each 0.1 β value reduction in DNA methylation, an effect size similar to our previous report in an independent set of 198 B-ALL cases with 450K array data [meta-analysis summary RM (sRM) = 1.32; 95% CI, 1.10-1.57]. The polyepigenetic smoking score was positively associated with gene deletion frequency among all 482 B-ALL cases (sRM = 1.31 for each 4-unit increase in score; 95% CI, 1.09-1.57).

Conclusions: We provide further evidence that prenatal tobacco-smoke exposure may influence the generation of somatic copy-number deletions in childhood B-ALL.

Impact: Analyses of deletion breakpoint sequences are required to further understand the mutagenic effects of tobacco smoke in childhood ALL.

Figure 1.. Sample flowchart.

Left box: ALL cases included in the previous analysis for the association between early-life tobacco smoke and gene deletion frequencies (n=559) , in which 361 cases were analyzed only with interview data and 198 B-ALL cases had Illumina 450K genome-wide DNA methylation array data available and were thus included in the analyses of DNA methylation at the AHRR CpG cg05575921. Right box: samples included in the current study, including 198 B-ALL cases that were analyzed previously and 284 B-ALL cases now with available EPIC array DNA methylation data and MLPA gene deletion frequency data, of which 178 overlapped with the 361 cases that were analyzed previously only with interview data. In total, 482 B-ALL cases are included in our case-only analysis of prenatal tobacco smoke exposure and gene deletion frequency. *11 out of 106 B-ALL cases have interview data now available.

Figure 2.. Linear regression results for the associations of parental self-reported tobacco smoke exposures with DNA methylation at the AHRR CpG cg05575921 and with the polyepigenetic smoking score.

Paternal and maternal ever smoking were defined as having smoked at least 100 cigarettes, pipes, or cigars before the child’s diagnosis. Additional dichotomous variables only accounted for whether the mother or father smoked at all during the time period described. Child postnatal passive smoking was measured by child secondhand smoking from either parent, or from other persons aside from parents who smoked indoors, in order to show the presence of a regular smoker (e.g. the mother, father, or other individual) in the household up to the child’s third birthday or ALL diagnosis (whichever came first). Cumulative tobacco exposures were calculated from four binary exposures: paternal smoking during preconception, maternal smoking during preconception, maternal smoking during pregnancy, and child’s postnatal passive smoking. Parental continuous smoking exposures were measured by number of cigarettes, pipes, or cigars per day (CPD) in 5-unit increments. All linear regression models were adjusted for cell type heterogeneity and genetic ancestry. AHRR CpG cg05575921 beta value was multiplied by −10. Linear regression models were fitted for each smoking exposure variable in the 450K dataset and EPIC dataset. Panels show results from linear regression models for the outcome variable DNA methylation at the AHRR CpG cg05575921 (left) or for the outcome variable polyepigenetic smoking score (right). Centers of points and horizontal bars indicate point estimates and 95% confidence intervals. (A) Results from linear regression models adjusted for cell type heterogeneity and genetic ancestry only. (B) Independent effects from linear regression models additionally mutually adjusted for paternal and maternal smoking variables. (C) The joint exposures of prenatal and postnatal tobacco smoking from linear regression models for newly derived variables of paternal smoke preconception plus maternal prenatal smoking, maternal prenatal smoking plus child postnatal passive smoking, and paternal smoke preconception plus child postnatal passive smoking. Reference group: cases who were unexposed to both exposures that make up the joint effect.

Figure 3.. Forest plots showing meta-analysis results of the association between epigenetic biomarkers of prenatal tobacco smoke exposure and gene deletion frequency in B-ALL cases.

The panels include Poisson regression results for the association between deletion numbers and DNA methylation at the AHRR CpG cg05575921 (top), the polyepigenetic smoking score (middle), and the polyepigenetic smoking score excluding AHRR CpG cg05575921 (bottom). Ratio of means (RM) were calculated for every 0.1 beta value decrease of cg05575921 and every 4-unit increase of polyepigenetic smoking score. All Poisson regression models were adjusted for cell type heterogeneity and genetic ancestry. Models with exposure variable DNA methylation at the AHRR CpG cg05575921 were additionally adjusted for methyl-QTL SNP genotypes (rs148405299 in the 450K dataset and rs77111113 in the EPIC dataset). Centers of squares and horizontal bars through each indicate point estimates and 95% confidence intervals (CI) of individual set RM. Area of squares indicate relative weights of individual set. Vertical apices of diamonds and horizontal bars through each indicate summary RM and 95% CI. Relative weights (%) (proportional to the reciprocal of the sampling variance of the individual set) of two sets, RM, sRM, and 95% CI are summarized in the right panel.