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Codon specific readthrough as a mechanism of BRCA2 restoration in acquired PARP inhibitor and chemotherapy resistance

Nucleic Acids Res. 2025 Oct 14;53(19):gkaf990. doi: 10.1093/nar/gkaf990.

ABSTRACT

BRCA2 mutations contribute to the pathogenesis and treatment sensitivity of a subset of ovarian, breast, prostate, and pancreatic cancers. When these cancers become therapy resistant, secondary mutations that restore the BRCA2 open reading frame are found in half the cases, but other causes of resistance remain incompletely understood. Here, we identified translational readthrough of a premature termination codon (PTC) as a cause of resistance to poly(ADP-ribose) polymerase inhibitors (PARPis) and cisplatin in cells derived from the BRCA2-mutated ovarian cancer line PEO1 by PARPi selection. Despite persistence of the signature 4965C > G (p.Y1655X) BRCA2 mutation, low-level expression of full-length BRCA2 protein was detectable in these cells by immunoblotting and tandem mass spectrometry. Either BRCA2 knockdown or gene interruption 5' or 3' to the PTC restored treatment sensitivity, implicating BRCA2 in the resistance. Reporter assays demonstrated UAG-selective readthrough in the resistant clones but not parental cells. Moreover, custom searching of global proteomic data indicated readthrough of stop codons, particularly UAGs, in additional proteins in the resistant clones. Finally, multi-omic analysis identified multiple changes in the nonsense-mediated decay and termination machineries that favor readthrough. Accordingly, the present results identify PTC readthrough as a potential mechanism of drug resistance in cells with BRCA2 nonsense mutations.

PMID:41099700 | PMC:PMC12526053 | DOI:10.1093/nar/gkaf990

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HT SpaceM: A high-throughput and reproducible method for small-molecule single-cell metabolomics

Single-cell metabolomics (SCM) can probe metabolic heterogeneity but is hindered by low sensitivity for small molecules, limited scalability, and the lack of standardized frameworks for data analysis. HT SpaceM is a high-throughput MALDI-imaging-based SCM method to robustly detect small-molecule metabolites in single cells. Applied to over 140,000 cells across diverse conditions and cancer cell lines, HT SpaceM enabled reproducible metabolic profiling of over 100 small-molecule metabolites, identification of subpopulation-specific markers, and detection of heterogeneity and pathways coordination, thus facilitating scalable and reproducible SCM.
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Framework for the Development and Delivery of Digital Peer Support Programs: Qualitative Study on in-Person and Digital Delivery for People With Cardiovascular Disease

Background: Peer support (sharing experiences/support with others with the same condition) improves health outcomes among people with cardiovascular disease (CVD), including self-management behaviours and self-efficacy. However, current peer support interventions are diverse. Evidence is lacking on peer support attenders perceptions of benefits and the elements that are considered priorities, especially for digital interventions. Objective: The study objectives were to 1) describe perceived benefits and recommendations for CVD peer support programs from people attending in-person peer support, 2) identify priorities for digital peer support from consumers and clinicians testing a peer support app prototype, and 3) develop a framework to inform future peer support intervention development. Methods: Qualitative methodology was used across two components to address the objectives of this study. In Component 1, semi-structured focus groups were conducted with attenders of established in-person CVD peer support groups, exploring the perceived benefits of peer support and recommendations for future programs. In Component 2, semi-structured interactive workshops with consumers with CVD and semi-structured online interviews with CVD clinicians/researchers were undertaken seeking feedback and recommendations for digital peer support using an exploratory digital CVD peer support application prototype. Data were recorded digitally, transcribed verbatim, and analysed thematically. Findings from both components were iteratively synthesised to inform a digital peer support development framework. Results: In Component 1, 22 participants (age range 29-84 years, male 45%) took part in focus groups. The overarching theme was that peer support provides benefits through sharing experiences. Five themes were refined and defined; (i) peer support provides a way of coping, (ii) peers learn from each other, (iii) peers understand what each other are going through, (iv) the peer community uplifts mood and build confidence, and (v) awareness, flexibility and resources are important for engagement. In Component 2, five participants (age range 55-74 years, male 60%) attended two workshops and eight clinicians/researchers (age range 30-65 years, male 10%) were interviewed. Three themes were refined and defined: (i) autonomy is essential to promote engagement, (ii) safeguarding is important to both users and clinicians, and (iii) interfaces that are simple, easy to use and visually attractive enable use. Priorities identified from both components included greater peer support awareness and uptake, flexibility with timing and family participation, healthcare professional involvement, provision of resources, autonomous features enabling choice, checklists and clinician moderation for safeguarding, and simple to use interfaces. Conclusions: Participants in peer support programs derive benefit from sharing their experience of living with CVD which enable coping, learning, feeling understood and a sense of community. Priorities were synthesised to create a framework for digital peer support development for future peer support with recommendations to focus on six key areas: uptake, flexibility, resources, autonomy, safeguarding and interface.
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