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p50, the mature product of NFKB1, is constitutively produced from its precursor, p105. Here, we identify BARD1 as a p50-interacting factor. p50 directly associates with the BARD1 BRCT domains via a C-terminal phospho-serine motif. This interaction is induced by ATR and results in mono-ubiquitination of p50 by the BARD1/BRCA1 complex. During the cell cycle, p50 is mono-ubiquitinated in S phase and loss of this post-translational modification increases S phase progression and chromosomal breakage. Genome-wide studies reveal a substantial decrease in p50 chromatin enrichment in S phase and Cycln E is identified as a factor regulated by p50 during the G1 to S transition. Functionally, interaction with BARD1 promotes p50 protein stability and consistent with this, in human cancer specimens, low nuclear BARD1 protein strongly correlates with low nuclear p50. These data indicate that p50 mono-ubiquitination by BARD1/BRCA1 during the cell cycle regulates S phase progression to maintain genome integrity., p50 is a constitutively produced NF-κB subunit that modulates the response to DNA damage. Here, the authors show that activation of ATR during S phase induces p50 interaction with BARD1 resulting in p50 mono-ubiquitination, facilitating cell cycle progression and promoting chromosome integrity.

Implementing cancer precision medicine in the clinic requires assessing the therapeutic relevance of genomic alterations. A main challenge is the systematic interpretation of whole-exome sequencing (WES) data for clinical care. One hundred sixty-five adults with metastatic colorectal and lung adenocarcinomas were prospectively enrolled in the CanSeq study. WES was performed on DNA extracted from formalin-fixed paraffin-embedded tumor biopsy samples and matched blood samples. Somatic and germ-line alterations were ranked according to therapeutic or clinical relevance. Results were interpreted using an integrated somatic and germ-line framework and returned in accordance with patient preferences. At the time of this analysis, WES had been performed and results returned to the clinical team for 165 participants. Of 768 curated somatic alterations, only 31% were associated with clinical evidence and 69% with preclinical or inferential evidence. Of 806 curated germ-line variants, 5% were clinically relevant and 56% were classified as variants of unknown significance. The variant review and decision-making processes were effective when the process was changed from that of a Molecular Tumor Board to a protocol-based approach. The development of novel interpretive and decision-support tools that draw from scientific and clinical evidence will be crucial for the success of cancer precision medicine in WES studies. Genet Med advance online publication 26 January 2017

Inherited mutations in DNA-repair genes such as BRCA2 are associated with increased risks of lethal prostate cancer. Although the prevalence of germline mutations in DNA-repair genes among men with localized prostate cancer who are unselected for family predisposition is insufficient to warrant routine testing, the frequency of such mutations in patients with metastatic prostate cancer has not been established.We recruited 692 men with documented metastatic prostate cancer who were unselected for family history of cancer or age at diagnosis. We isolated germline DNA and used multiplex sequencing assays to assess mutations in 20 DNA-repair genes associated with autosomal dominant cancer-predisposition syndromes.A total of 84 germline DNA-repair gene mutations that were presumed to be deleterious were identified in 82 men (11.8%); mutations were found in 16 genes, including BRCA2 (37 men [5.3%]), ATM (11 [1.6%]), CHEK2 (10 [1.9% of 534 men with data]), BRCA1 (6 [0.9%]), RAD51D (3 [0.4%]), and PALB2 (3 [0.4%]). Mutation frequencies did not differ according to whether a family history of prostate cancer was present or according to age at diagnosis. Overall, the frequency of germline mutations in DNA-repair genes among men with metastatic prostate cancer significantly exceeded the prevalence of 4.6% among 499 men with localized prostate cancer (P0.001), including men with high-risk disease, and the prevalence of 2.7% in the Exome Aggregation Consortium, which includes 53,105 persons without a known cancer diagnosis (P0.001).In our multicenter study, the incidence of germline mutations in genes mediating DNA-repair processes among men with metastatic prostate cancer was 11.8%, which was significantly higher than the incidence among men with localized prostate cancer. The frequencies of germline mutations in DNA-repair genes among men with metastatic disease did not differ significantly according to age at diagnosis or family history of prostate cancer. (Funded by Stand Up To Cancer and others.).

Introduction. Minimal residual disease (MRD) is the most powerful prognostic factor in acute lymphoblastic leukemia (ALL). Currently, real-time quantitative PCR (RQ-PCR) is the most widely used molecular method for MRD assessment, rigorously standardized within the EuroMRD consortium. According to the EuroMRD guidelines (Van der Velden et al. Leukemia 2007), a non-negligible fraction of patients with very low MRD levels are classified as positive not-quantifiable (PNQ), a definition that may result problematic in the clinical practice. Digital-droplet-PCR (ddPCR) allows an absolute quantification without the need of a standard curve and has the potential to overcome some limitations of RQ-PCR. High degrees of efficiency, sensitivity and accuracy have been reported for ddPCR compared to RQ-PCR, but no established guidelines for ddPCR MRD analysis and interpretation have so far been defined and its ability to correctly evaluate very low MRD levels is still under investigation. In the present study, we assessed MRD by ddPCR in pediatric ALL cases classified as PNQ and/or negative by RQ-PCR at days +33 and/or +78 of the AIEOP-BFM ALL 2000 trial, to evaluate the potential of ddPCR for low MRD quantification and patients' risk stratification. Patients and Methods. A total of 211 pediatric ALL patients enrolled in the AIEOP-BFM ALL 2000 trial were included in the study. We analyzed 124 B-lineage ALL patients defined as intermediate risk (IR) who had high positive MRD at day +33 and at day +78 were either PNQ (n=45, Slow Early Responders (SER)) or negative (n=79). A case-control design was applied to 36 B- and T-lineage relapsed ALL patients (cases) who at day +33 had PNQ MRD (n=12, IR) or were negative (n=24, standard risk (SR)) and to matched controls (21 and 30 patients who did not present a relapse). ddPCR analysis was performed as previously published (Della Starza et al, BJH 174, 541-9, 2016), by using 1.5 μg and 3.0 μg DNA of the follow-up samples. In the absence of an international consensus, data have been analyzed using two alternative guidelines; results are reported according to Della Starza et al (BJH 2016). Results. Among 45 SER patients, ddPCR performed on 1.5 μg DNA of PNQ samples at day +78 revealed that 13 were quantifiable (Q), 16 PNQ and 16 negative (NEG) . When 3.0 μg of DNA were used (41/45 samples due to material availability), 12 were Q, 19 PNQ and 10 NEG. Event-free survival (EFS) curves are shown in Fig. 1a. Among the 79 patients with high positive MRD at day +33 but who were negative at day +78, ddPCR on 1.5 μg DNA of day +78 identified 5 as Q, 17 PNQ and 57 NEG. When 3.0 μg DNA was used (77/79 samples), 9 patients were Q, 27 PNQ and 41 NEG. EFS curves are reported in Fig. 1b. When ddPCR was applied to 33 PNQ samples at day +33, 2 were Q, 9 PNQ and 22 NEG; when using 3.0 μg of DNA, 1 was Q, 15 were PNQ and 17 NEG. EFS curves are shown in Fig. 1c. Lastly, ddPCR on 1.5 μg of day +33 DNA of 54 SR patients showed 5 PNQ and 49 NEG, whilst by using 3.0 μg on 53 sample, 7 were PNQ and 46 NEG. Conclusions. Our data demonstrate that ddPCR is a very promising tool for the evaluation of MRD in ALL cases with very low or negative RQ-PCR MRD results. In particular, among SER patients most relapses occurred in cases with quantifiable ddPCR MRD at day +78, while patients with negative or PNQ MRD by ddPCR at day +78 had a better outcome. Based on these results, high-risk treatment could be offered only to ddPCR quantifiable cases. Among patients with highly positive MRD at day +33 and negative at day +78, the small number of cases with quantifiable disease by ddPCR at present does not allow to establish the impact of quantification; consistently with SER patients, the outcome was similar for patients with negative or PNQ MRD by ddPCR at day +78. Similarly, among patients with PNQ MRD by RQ-PCR at day +33, a similar outcome was observed for cases negative or PNQ by ddPCR. Lastly, in most SR patients ddPCR confirmed the negative results of RQ-PCR at day +33, associated with an extremely good kinetics of disease reduction, independently of the MRD PCR method. Overall, our data indicate that ddPCR is as sensitive as RQ-PCR and can provide a potentially more accurate prognostic stratification for cases defined as PNQ MRD by RQ-PCR, in view of its ability to quantify without a standard curve. The application of ddPCR in a prospective clinical protocol with international guidelines is needed to define whether it can result in an overall improvement of pediatric ALL patients' stratification and outcome. Disclosures Foà: AMGEN: Other: ADVISORY BOARD; JANSSEN: Other: ADVISORY BOARD, Speakers Bureau; CELTRION: Other: ADVISORY BOARD; INCYTE: Other: ADVISORY BOARD; ABBVIE: Other: ADVISORY BOARD, Speakers Bureau; ROCHE: Other: ADVISORY BOARD, Speakers Bureau; NOVARTIS: Speakers Bureau; CELGENE: Other: ADVISORY BOARD, Speakers Bureau; GILEAD: Speakers Bureau.

5009Background: Inherited mutations in genes that regulate DNA repair processes such as BRCA1 and BRCA2 associate with increased risks of developing lethal prostate cancer (PC), and with response t...