SAT177 Molecular Analysis Of AMBRA1 As A Candidate Tumor Suppressor In Sporadic Parathyroid Adenomas

Disclosure: S. Chinwo: None. J. Costa: None. A. Arnold: None. J. Bellizzi: None. Primary hyperparathyroidism is a common endocrine disorder that is most often caused by a sporadic single-gland parathyroid adenoma. Currently, the only known and experimentally validated oncoprotein for parathyroid adenomas is cyclin D1. The overexpression of cyclin D1 protein has been reported in 20-40% of cases and about 8 percent of cases harbor a DNA rearrangement involving the cyclin D1 (CCND1) locus. Otherwise, the molecular mechanisms underlying this cyclin D1 overexpression have yet to be identified. Here, we explored a potential parathyroid tumorigenic mechanism that could increase cyclin D1 stability through a defect in the AMBRA1 E3 ligase adaptor, a molecule responsible for its degradation. Evidence that AMBRA1 may be a tumor suppressor gene includes the accumulation of cyclin D with hyperproliferation in AMBRA1-deficient cells, the context-dependent growth of tumors in Ambra1-deficient mice, and the finding that low levels of AMBRA1 correlated with poor clinical outcomes in The Cancer Genome Atlas. Therefore, we proceeded to examine AMBRA1 for evidence of tumor suppressor-type inactivation within a cohort of 98 typical parathyroid adenomas. Genomic DNA samples from these tumors were subjected to PCR-amplification of all 18 coding exons of AMBRA1, followed by Sanger sequencing. Sequences were analyzed by comparison to the normal reference sequence: ENST00000683756.1, seeking to assess any observed intragenic or splice-site variants as potential or likely loss-of-function mutations. Thus far, with 97 percent of the coding region for this 1298-amino acid protein fully sequenced, we have identified an inactivating mutation in 1 of the 98 tumors (1%): c.126G>A in exon 2, resulting in an early stop codon p.Trp42*. Sequencing the patient’s matched germline DNA confirmed the mutation was somatic and showed loss of heterozygosity. We also identified likely-nonpathogenic variants in 5 parathyroid adenomas: missense variant c.3385G>T (p.Ala1129Ser), 3’ UTR variants c.*576A>T and c.*491T>C and synonymous variants c.579C>T and c.2776C>T, each in a single tumor. 4 of 5 were found as germline variants in the normal population and the one missense scored benign by in silico criteria. To conclude, our observations suggest that AMBRA1 may function as a classical tumor suppressor gene in sporadic parathyroid adenomas at very low frequency. Further study may include protein expression analysis of AMBRA1 to investigate influences on gene expression. To further elucidate the mechanisms behind cyclin D1 overexpression in parathyroid adenomas, other molecules that participate in its degradation should be explored as potential tumor suppressors. Presentation: Saturday, June 17, 2023