Impact of regulatory genetics in the susceptibility to childhood leukemia

^1,2Daniel Sinnett, ¹Hélène Bélanger, ¹Joëlle Dionne, ¹Mathieu Larivière, ¹Jasmine Healy
¹Division of Hematology-Oncology, CHU Sainte-Justine Research Center, Montreal, QC, Canada, ²Department of Pediatrics, University of Montreal, Montreal, QC, Canada

Acute lymphoblastic leukemia (ALL) is the most frequent form of pediatric cancer. It is a complex genetic disease in which the effect of a series of low penetrance genes is modulated by external factors, underlining the important role of genetic variation in determining inter-individual variability in the susceptibility to childhood leukemia. We have shown that genes involved in effective carcinogen metabolism, maintenance of DNA integrity and oxidative stress response, may collectively contribute to the development of childhood ALL, illustrating the need for thorough genetic analyses to further define the genetic determinants of this complex disorder. Until recently, association studies have mainly focused on DNA variants in coding regions while those in regulatory sequences have been studied less intensively, despite the fact that these variants in promoter regions (rSNP) could alter transcription factor binding sites (TFBS) and lead to differences in expression that would modify the risk of disease. Moreover, due to their highly regulated nature, important biological processes such as the cell cycle could be particularly sensitive to variations in gene expression levels. Thus we hypothesized that the susceptibility (or resistance) to childhood ALL is influenced by functional cis-acting regulatory DNA variants in G1/S cell cycle checkpoint genes. The targeted regulatory regions (defined as 2kb upstream of the transcription start site) were screened by dHPLC for the presence of rSNPs in a panel of 40 individuals from different ethnic backgrounds. We identified 127 rSNPs in 16 cell cycle checkpoint genes. Their functional impact was assessed by combining in silico analysis and in vitro functional assays: 90 rSNPs had a predicted impact on putative TFBSs; certain rSNPs showed differential DNA-protein binding; and several regulatory haplotypes (rHAP) significantly influenced transcriptional activity in an allele-specific manner. To evaluate the impact of these rSNPs/rHAPs on the risk to childhood ALL, we performed case-control and family-based association studies targeting cyclin-dependent kinase inhibitor genes. Using both study designs, we showed evidence of association between variants in CDKN2A, CDKN2B and CDKN1B and an increased risk of ALL. These findings suggest that variable expression levels of cell cycle inhibitor genes due to regulatory polymorphisms could indeed influence an individual's risk of cancer and contribute at least to childhood leukemogenesis.