BCOR Internal Tandem Duplication Associated Uterine Sarcoma: Expanding the Clinicopathologic Spectrum.

The diagnosis of high-grade endometrial stromal sarcoma has become more refined following molecular characterization of these tumors. Recently BCOR internal tandem duplications (ITD) have been identified in a small number of high-grade endometrial stromal sarcoma. Here we present an additional case of this rare entity in a young woman in her late teens. She presented with menorrhagia and underwent resection of 2 uterine lesions. The tumor was a spindle cell neoplasm composed of long fascicles with low to moderate cellularity, mild to moderate cytologic atypia, and up to 2 mitotic figures per 10 high power fields. Necrosis was not identified. Immunohistochemical stains showed the tumor to be positive for cyclin D1 in >50% of tumor cells, focally positive for CD10, and negative for SMA, desmin, h-caldesmon, and ALK1. BCOR ITD was confirmed by polymerase chain reaction with subsequent Sanger sequencing. Clues to the diagnosis of BCOR ITD uterine sarcoma include young patient age, uniform nuclear features, and diffuse positivity for cyclin D1. These features should prompt further molecular interrogation for definitive diagnosis, which is important for prognostication.

Tumor BRCA Testing in High Grade Serous Carcinoma: Mutation Rates and Optimal Tissue Requirements.

BACKGROUND: Approximately 25% of women diagnosed with tubo-ovarian high-grade serous carcinoma have germline deleterious mutations in BRCA1 or BRCA2, characteristic of hereditary breast and ovarian cancer syndrome, while somatic mutations have been detected in 3-7%. We set out to determine the BRCA mutation rates and optimal tissue requirements for tumor BRCA testing in patients diagnosed with tubo-ovarian high-grade serous carcinoma. METHODS: Sequencing was performed using a multiplexed polymerase chain reaction-based approach on 291 tissue samples, with a minimum sequencing depth of 500X and an allele frequency of >5%. RESULTS: There were 253 surgical samples (87%), 35 biopsies (12%) and 3 cytology cell blocks (1%). The initial failure rate was 9% (25/291), including 9 cases (3%) with insufficient tumor, and 16 (6%) with non-amplifiable DNA. Sequencing was successful in 78% (228/291) and deemed indeterminate due to failed exons or variants below the limit of detection in 13% (38/291). Repeat testing was successful in 67% (28/42) of retested samples, with an overall success rate of 86% (251/291). Clinically significant (pathogenic, likely pathogenic) variants were identified in 17% (48/276) of complete and indeterminate cases. Successful sequencing was dependent on sample type, tumor cellularity and size (p ≤ 0.001) but not on neoadjuvant chemotherapy or age of blocks (p > 0.05). CONCLUSIONS: Our study shows a 17% tumor BRCA mutation rate, with an overall success rate of 86%. Biopsy and cytology samples and post-chemotherapy specimens can be used for tumor BRCA testing, and optimal tumors measure ≥5 mm in size with at least 20% cellularity.

Long-range PCR and next-generation sequencing of BRCA1 and BRCA2 in breast cancer.

Individuals and families carrying mutations in BRCA1 and BRCA2 (BRCA1/2) have a markedly elevated risk of developing breast and ovarian cancers. The first-generation of BRCA1/2 mutation analysis targeted only the coding exons and has implicated protein-truncating mutations (indel, nonsense) in BRCA1/2 inactivation. Recently, heritable breast cancers have also been attributed to other exonic mutations (missense, silent) and mutations in introns and untranslated regions. However, analysis of these alterations has been prohibitively laborious and cost intensive, and the proportion of cases carrying mutations in unscreened regions of BRCA1/2 and other predisposition genes is unknown. We have developed and validated a next-generation sequencing (NGS) approach for BRCA1/2 mutation analysis by applying long-range PCR and deep sequencing. Genomic DNA from familial breast cancer patients (N = 12) were screened and NGS successfully identified all 19 distinct (51 total) BRCA1 and 35 distinct (63 total) BRCA2 sequence alterations detectable by the Sanger sequencing, with no false-negative or positive results. In addition, we report the robust detection of variants from introns and untranslated regions. These results illustrate that NGS can provide comprehensive genetic information more quickly, accurately, and at a lower cost than conventional approaches, and we propose NGS to be a more effective method for BRCA1/2 mutational analysis. Advances in NGS will play an important role in enabling molecular diagnostics and personalized treatment of breast and ovarian cancers.