T. D. Murtha1, R. Korah1, T. Carling1 1Yale University School Of Medicine,New Haven, CT, USA
Introduction: Recent comprehensive genetic analyses have demonstrated a high prevalence of gene fusions–suggestive of defective DNA break repair–in papillary thyroid carcinoma (PTC). Polymerase theta (POLQ) is an A-family mammalian polymerase that is unique in mediating replication-independent DNA repairs, including single and double strand DNA breaks. Differential expression of POLQ has been shown to correlate with poor outcomes in breast, ovarian, and lung cancer. Because of the high concentration of hydrogen peroxide in thyroid follicles and the consequent oxidation-induced single and double strand breaks in DNA, we hypothesized that genetic polymorphism of the DNA repair enzyme POLQ may play a role in predisposing follicular cells to de novo mutations and potentially tumorigenesis. To identify novel mutations and enrichment of SNPs, we performed classical Sanger sequencing of the exonic regions of POLQ in a large cohort of sporadic PTCs.
Methods: Genomic DNA was isolated from 51 histologically confirmed PTC specimens and matched adjacent non-tumor thyroid tissue. Genetic sequencing via capillary electrophoresis (Sanger method) was performed to detect novel variants and SNPs for all 30 exons and exon/intron splice sites of the POLQ gene. Discovered nonsynonymous and missense SNPs, and the corresponding amino acid substitutions, were analyzed using multiple bioinformatic algorithms to determine if alterations were deleterious to protein integrity or function. Novel variants were annotated and SNP frequency was compared to The Exome Aggregation Consortium (ExAC) database.
Results: Fourteen germline variants were confirmed in tumor and corresponding matched adjacent non-tumor thyroid tissue. Eight of those germline variants (57%) were determined to be disease-causing by mutation prediction algorithms. In the C-terminal polymerase domain (amino acids 2060–2590), there was an increased frequency of germline SNPs compared to the comprehensive population SNP database ExAC. The recurrent SNP rs532411, previously found to be significantly associated with breast cancer, was found in 22% (11/51) of PTC specimens while its frequency in the population is 6.5% (P < .0001, chi-squared test). We also identified 4 novel variants not previously reported.
Conclusion: The unique contribution of POLQ in DNA repair via non-homologous end-joining of double stand breaks makes it a critical regulator of genomic integrity in the hydrogen peroxide-rich thyroid gland. Sequencing demonstrates enrichment of multiple SNPs in the polymerase region of POLQ, potentially increasing the probability of carcinogenic point mutations and gene fusions. While expanded genome-wide association studies (GWAS) are essential for confirming the relationship, the current study offers preliminary information suggesting a role for POLQ SNPs in genetically predisposing individuals to PTC.