A research-based gene panel to investigate breast, ovarian and prostate cancer genetic risk

Abstract

There is a need to investigate and better understand the inherited risk of cancer to ensure that clinical applications provide more accurate assessments and management strategies. Developing research-based next-generation sequencing gene panels that not only target (present-day) clinically actionable susceptibility genes but also genes that currently lack sufficient evidence for risk as well as candidate genes, such as those in DNA repair pathways, can help aid this effort. Therefore, gene panel B.O.P. ( B reast, O varian, and P rostate) was developed to evaluate the genetic risk of breast, ovarian and/or prostate cancer, and this manuscript serves as an introduction to B.O.P. and highlights its initial analytical validity assessment. B.O.P targets 87 genes that have been suggested, predicted, or clinically proven to be associated with breast, ovarian, and/or prostate cancer risk using Agilent Technologies Haloplex probes. The probes were designed for 100 base pair reads on an Illumina platform and target both coding and non-coding exons as well as 10 intronic base pairs flanking the intron-exon boundaries. The initial B.O.P screening involved 43 individuals from the Alabama Hereditary Cancer Cohort, and an average sequencing depth of 809X was obtained. Upon variant filtering and validation, true positives had an average sequencing depth of 659X and allele balance of 0.51. The average false positive sequencing depth was 34X and allele balance was 0.33. Although low sequencing depth was not always indicative of a false positive, high sequencing depths (>100X) signified a true positive. Furthermore, sensitivity and specificity of BRCA1/2 were calculated to be 100% and 92.3%, respectively. Overall, this screening enabled the establishment of criteria to alleviate future validation efforts and strongly supports the use of B.O.P. to further elucidate hereditary cancer susceptibility. Ultimately, continued B.O.P. screening will provide insights toward the genetic risk of and overlap between breast, ovarian, and/or prostate cancer.

Rare and Potentially Pathogenic Variants in Hydroxycarboxylic Acid Receptor Genes Identified in Breast Cancer Cases.

“…This individual did not have a family history of BC but had a rst-and second-degree relative diagnosed with other cancer types. Lastly, in the absence of clinical genetic screening, previous research-based screening did not reveal any clinically relevant mutations (12).…”

“…Lastly, an extremely rare HCAR3 frameshift mutation (Table 1) was identi ed in a female diagnosed with ER+ PR+ and HER2-in ltrating ductal carcinoma at 62 years of age. She had previous negative clinical genetic screening, which was con rmed through our research-based screening (12). The mutation, c.1117delC;p.Gln373Lysfs*82, greatly extends the C-terminus of HCAR3 and changes the secondary and tertiary protein structure (Figure 2).…”

“…The variant was detected in a female with an in ltrating lobular carcinoma (estrogen receptor-positive (ER+) and human epidermal growth factor receptor 2-negative (HER2-)) diagnosed at 35 years of age and a family history of BC and other cancers in rst-, second-and third-degree relatives. This individual did not have clinical genetic screening but screened negative for clinically signi cant mutations through our research-based multi-gene panel screening (12). Regarding HCAR1 c.721C>T;p.Leu241Phe, a statistically signi cant difference was not identi ed between cases and controls, and two of the ve prediction software suggested it was pathogenic ( Table 1).…”

“…That being said, with such slight differences between the two proteins, perhaps each alteration is key to protein function. On another note, this variant was detected in an early onset BC case also determined to harbor a clinically signi cant frameshift mutation in NBN (12,30). The interaction of these two variants and their combined ability to promote BC is unknown, but, intriguingly, expression of both HCAR3 and NBN have been reported to be dysregulated in oocytes of older women, when investigating why aneuploidy pregnancies occur in women of older ages.…”

“…Sequences were analyzed using Mutation Surveyor (Soft Genetics). Similar to the screening/validation process previously described (12), a systematic approach was followed for variant detection and validation. This involved screening genomic DNA from "Tube A" for the initial PCR of each study participant and, upon variant detection, validating variants by performing another initial PCR but using "Tube B."…”