Sixty-four Polish families with a history of breast and/or ovarian cancer were screened for mutations in the BRCA1/2 genes using a combination of denaturing high performance liquid chromatography (DHPLC) and sequencing. Two thirds (43/64; 67%) of the families were found to carry deleterious mutations, of which the most frequent were BRCA1 5382insC (n=22/43; 51%) and Cys61Gly (n=9/43; 20%). Two other recurrent mutations were BRCA1 185delAG (n=3) and 3819del5 (n=4), together accounting for 16% of the 43 mutation-positive cases. We also found three novel mutations (BRCA1 2991del5, BRCA2 6238ins2del21 and 8876delC) which combined with findings from our earlier study of 60 Northern Polish families. Moreover, screening of 43 BRCA1/2 negative families for the presence of large rearrangements by multiplex ligation-dependent probe amplification (MLPA) resulted in the finding of two additional BRCA1 mutations: a deletion of exons 1A, 1B and 2, and a deletion of exons 17-19, both present in single families. We conclude that the Polish population has a diverse mutation spectrum influenced by strong founder effects. However, families with strong breast/ovarian cancer history who are negative for these common mutations should be offered a complete BRCA gene screening, including MLPA analysis.

Background: Liquid biopsy is a minimally invasive collection of a patient body fluid sample. In oncology, they offer several advantages compared to traditional tissue biopsies. However, the potential of this method in endometrial cancer (EC) remains poorly explored. We studied the utility of tumor educated platelets (TEPs) and circulating tumor DNA (ctDNA) for preoperative EC diagnosis, including histology determination. Methods: TEPs from 295 subjects (53 EC patients, 38 patients with benign gynecologic conditions, and 204 healthy women) were RNA-sequenced. DNA sequencing data were obtained for 519 primary tumor tissues and 16 plasma samples. Artificial intelligence was applied to sample classification. Results: Platelet-dedicated classifier yielded AUC of 97.5% in the test set when discriminating between healthy subjects and cancer patients. However, the discrimination between endometrial cancer and benign gynecologic conditions was more challenging, with AUC of 84.1%. ctDNA-dedicated classifier discriminated primary tumor tissue samples with AUC of 96% and ctDNA blood samples with AUC of 69.8%. Conclusions: Liquid biopsies show potential in EC diagnosis. Both TEPs and ctDNA profiles coupled with artificial intelligence constitute a source of useful information. Further work involving more cases is warranted.

BARD1 has tumor suppressor functions by binding to BRCA1 and to p53 via RING finger and ankyrin repeats, respectively. The BARD1-BRCA1 dimer has E3 ligase activity, and BARD1-p53 interaction induces apoptosis. The C-terminus of BARD1 is binding to many proteins that are targets of the BRCA1-BARD1 ubiquitin ligase, and expression of N-terminally truncated RING-less BARD1 isoforms was correlated with poor prognosis in various cancers, consistent with their oncogenic functions by antagonistic binding to target proteins. Expression and interaction assays showed that the BARD1 BRCT domains were critically involved in telomere maintenance and interacted with the telomere binding protein TRF2. Overexpression of BARD1 or BRCA1 lead to TRF2 degradation, suggesting that TRF2 presents a novel target of the BARD1-BRCA1 E3 ubiquitination ligase. Repression of BARD1 or BRCA1 by siRNA leads to genetic instability, as shown before, and telomere alterations. We observed distinct telomeric alterations in blood cells of patients with a BARD1 germline mutation that causes a translation stop and translation of a truncated protein lacking the BRCT domains. Cells of carriers of a BRCA1 mutation that disrupts the BARD1-BRCA1 interaction show similar telomere alterations. These in vitro and in vivo observations suggest that BARD1 and BRCA1 play distinct roles in maintaining telomere integrity in a dosage dependent manner. BARD1δ, an isoform lacking RING and ankyrin motifs, but retaining the BRCT domains, is abundantly expressed in all cancers investigated so far. Overexpression of BARD1δ in vitro blocked cell cycle progression and induced chromosomal and telomere abnormalities, suggesting that BARD1δ also affected telomere integrity. We conclude that the correct dose of BARD1 and/ or its BRCT domains is critically important for maintaining telomere integrity. Expression of BARD1δ in cancer, or reduced expression of BARD1 in mutation carriers causes chromosomal instability and promotes carcinogenesis and tumor progression. Citation Format: Irmgard Irminger-Finger, Maxim Pilyugin, Magdalena Ratajska. The BARD1 BRCT domains are essential for maintenance of telomere integrity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2393. doi:10.1158/1538-7445.AM2014-2393

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