Analysis of a missense SNP in the aryl hydrocarbon receptor repressor gene that may disproportionately increase lung cancer risk in Black Americans - This is a project to study to a single nucleotide polymorphism (SNP) in the aryl hydrocarbon receptor repressor (AHRR) gene that may influence the lung cancer risk of Black/African American (B/AA) subjects. AHRR is a negative regulator of detoxification responses involved in the metabolism and elimination of tobacco smoke carcinogens. B/AA men have the highest rate of lung cancer death compared to all other groups; they show a 12% higher lung cancer incidence rate and a 15% higher lung cancer death rate than White/European (W/E) men. Elevated AHRR expression has been implicated in lung cancer risk; thus, a SNP that improves AHRR function would likely increase lung cancer risk. We have identified a missense SNP, rs35008248, in AHRR exon 4 that, while rare in W/E subjects, is present one third of B/AA subjects. The SNP was not included in most previously used arrays and has only recently begun to be studied. Preliminary data from GWAS explorer shows no significant associations in W/E subjects but suggests an association with smoking in B/AA males (p=3.87E- 3) and with smoking-related cancers in B/AA subjects (p=0.04927). As B/AA individuals comprise only a small fraction of the subjects (<4%), power to determine the role of SNPs in this group has been limited. AHRR is a negative regulator of the aryl hydrocarbon receptor (AHR). Upon tobacco smoke exposure, AHR binds to polycyclic aromatic hydrocarbon, enters the nucleus, and dimerizes with ARNT. The AHR/ARNT dimer binds to xenobiotic response elements (XREs), switching on genes involved in the inactivation/secretion of xenobiotic compounds. In a negative regulatory loop, AHR/ARNT turns on AHRR, which also binds to ARNT. The transcriptionally inactive AHRR/ARNT complex competes with AHR/ARNT for XRE sites, dampening the detoxification response. The reference allele of rs35008248 (T) encodes a leucine at position 114, while the alternate allele (C) encodes a turn-inducing proline. Combined annotation-dependent depletion (CADD) suggests the SNP might affect AHRR function. In the co-crystal structure of the AHRR/ARNT heterodimer, leucine 114 borders a loop entwining ARNT. We hypothesize that a proline at position 114 constrains AHRR loop conformation, thereby stabilizing AHRR/ARNT interaction, increasing the repressive effect of AHRR/ARNT heterodimers, interfering with the detoxification response, and increasing lung cancer risk of B/AA smokers, 34% of whom carry the minor allele. Here we propose to test for associations between genetic variation at AHRR with lung cancer outcomes using existing large-scale genetic data in B/AA individuals (Aim 1), to use CRISPR-based genome editing to engineer the minor allele into the genome of our unique immortalized human alveolar epithelial cell lines and assess its effects on the response to tobacco smoke in vitro (Aim 2), and to test whether the amino acid at position 114 affects the kinetics of the interaction between AHRR and ARNT (Aim 3).Together, the specific aims of this pilot project will provide insight into the role of a potential functional SNP in AHRR that is common in B/AA subjects and that could help explain their increased lung cancer risk.
