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DNA interstrand crosslinks (ICLs) pose a major obstacle for DNA replication and transcription if left unrepaired. The cellular response to ICLs requires the coordination of various DNA repair mechanisms. Homologous recombination (HR) intermediates generated in response to ICLs, require efficient and timely conversion by structure-selective endonucleases. Our knowledge on the precise coordination of this process remains incomplete. Here, we designed complementary genetic screens to map the machinery involved in the response to ICLs and identified FIRRM/C1orf112 as an indispensable factor in maintaining genome stability. FIRRM deficiency leads to hypersensitivity to ICL-inducing compounds, accumulation of DNA damage during S-G 2 phase of the cell cycle, and chromosomal aberrations, and elicits a unique mutational signature previously observed in HR-deficient tumors. In addition, FIRRM is recruited to ICLs, controls MUS81 chromatin loading, and thereby affects resolution of HR intermediates. FIRRM deficiency in mice causes early embryonic lethality and accelerates tumor formation. Thus, FIRRM plays a critical role in the response to ICLs encountered during DNA replication.

Supplementary Methods - PDF 55K, A detailed protocol for the classification of BRCA1 sequence variants using R26CreERT2/RMCE;Brca1SCo/{delta} ES cells

Supplementary Tables 1-4 - PDF 99K, Information on BRCA1 mutants used to validate the functional complementation assays (Table S1), BRCA1 VUS showing low protein levels (Table S2), the results of all proliferation and cisplatin sensitivity assays (Table S3), and predictions of functional consequences from literature and in silico analyses together with a summary of the results of the functional complementation assays for all BRCA1 variants analyzed (Table S4)

Supplementary Figures 1-4 - PDF 213K, This file contains additional data on the generation of R26CreERT2/RMCE;Brca1SCo/{delta} ES cells and analysis of Rosa26 targeted human BRCA1 cDNA expression (Fig. S1), analysis of BRCA1 VUS protein and mRNA expression (Fig. S2 and S3) and growth curves supplementary to Figure 6 (Fig. S4)

Genetic analyses of BRCA1 mutant cell lines and PDX models.

Breast and ovarian cancer patients harboring BRCA1/2 germline mutations have clinically benefitted from therapy with PARP inhibitor (PARPi) or platinum compounds, but acquired resistance limits clinical impact. In this study, we investigated the impact of mutations on BRCA1 isoform expression and therapeutic response. Cancer cell lines and tumors harboring mutations in exon 11 of BRCA1 express a BRCA1-Δ11q splice variant lacking the majority of exon 11. The introduction of frameshift mutations to exon 11 resulted in nonsense-mediated mRNA decay of full-length, but not the BRCA1-Δ11q isoform. CRISPR/Cas9 gene editing as well as overexpression experiments revealed that the BRCA1-Δ11q protein was capable of promoting partial PARPi and cisplatin resistance relative to full-length BRCA1, both in vitro and in vivo. Furthermore, spliceosome inhibitors reduced BRCA1-Δ11q levels and sensitized cells carrying exon 11 mutations to PARPi treatment. Taken together, our results provided evidence that cancer cells employ a strategy to remove deleterious germline BRCA1 mutations through alternative mRNA splicing, giving rise to isoforms that retain residual activity and contribute to therapeutic resistance. Cancer Res; 76(9); 2778–90. ©2016 AACR.

Purpose:Because BRCA1 is a high-risk breast/ovarian cancer susceptibility gene, BRCA1 sequence variants of uncertain clinical significance (VUS) complicate genetic counseling. As most VUS are rare, reliable classification based on clinical and genetic data is often impossible. However, all pathogenic BRCA1 variants analyzed result in defective homologous recombination DNA repair (HRR). Thus, BRCA1 VUS may be categorized based on their functional impact on this pathway.Experimental Design:Two hundred thirty-eight BRCA1 VUS—comprising most BRCA1 VUS known in the Netherlands and Belgium—were tested for their ability to complement Brca1-deficient mouse embryonic stem cells in HRR, using cisplatin and olaparib sensitivity assays and a direct repeat GFP (DR-GFP) HRR assay. Assays were validated using 25 known benign and 25 known pathogenic BRCA1 variants. For assessment of pathogenicity by a multifactorial likelihood analysis method, we collected clinical and genetic data for functionally deleterious VUS and VUS occurring in three or more families.Results:All three assays showed 100% sensitivity and specificity (95% confidence interval, 83%–100%). Out of 238 VUS, 45 showed functional defects, 26 of which were deleterious in all three assays. For 12 of these 26 variants, we could calculate the probability of pathogenicity using clinical and genetic data, resulting in the identification of 7 (likely) pathogenic variants.Conclusions:We have functionally categorized 238 BRCA1 VUS using three different HRR-related assays. Classification based on clinical and genetic data alone for a subset of these variants confirmed the high sensitivity and specificity of our functional assays.

Legend for Supplementary Figures S1-S6.

RNA-seq analyses of global exon splicing events (S1); Cell line therapy sensitivities (S2); Hazard ratios for survival of patients with serous ovarian carcinomas associated with BRCA1 mutation, age at diagnosis and stage (S3); Characteristics of studies included in survival analysis from Fig. 5A (S4).

Description of additional methods and procedures used in the study. Also includes Supplementary References.

Supplementary Data from Functional Categorization of BRCA1 Variants of Uncertain Clinical Significance in Homologous Recombination Repair Complementation Assays

Supplementary Table from Functional Categorization of BRCA1 Variants of Uncertain Clinical Significance in Homologous Recombination Repair Complementation Assays

SUMMARYBRCA1 and BRCA2 both function in DNA double-strand break repair by homologous recombination (HR). Due to their HR-defect, BRCA1/2-deficient cancers are sensitive to poly(ADP-ribose) polymerase inhibitors (PARPi) but they eventually acquire resistance. Preclinical studies yielded several PARPi resistance mechanisms that do not involve BRCA1/2 reactivation, but their relevance in the clinic remains elusive. To investigate which BRCA1/2-independent mechanisms drive spontaneous resistance in vivo, we combined molecular profiling with functional analysis of the HR status of matched PARPi-naïve and PARPi-resistant mouse mammary tumors harboring large intragenic deletions that prevent functional restoration of BRCA1/2. We observed restoration of HR in 64% of PARPi-resistant BRCA1-deficient tumors but none in the PARPi-resistant BRCA2-deficient tumors. Moreover, we found that 53BP1 loss is the prevalent resistance mechanism in HR-proficient BRCA1-deficient tumors, whereas resistance in BRCA2-deficient tumors is mainly induced by the loss of PARG. Our combined multi-omics analysis catalogued additional genes and pathways potentially involved in modulating PARPi response.

Globally, South Africa has one of the highest rates of gender based violence (GBV). While little is known about the levels of GBV against women with disabilities in the country, less is known about...

Cancer-associated systemic inflammation is strongly linked to poor disease outcome in patients with cancer1,2. For most human epithelial tumour types, high systemic neutrophil-to-lymphocyte ratios are associated with poor overall survival3, and experimental studies have demonstrated a causal relationship between neutrophils and metastasis4,5. However, the cancer-cell-intrinsic mechanisms that dictate the substantial heterogeneity in systemic neutrophilic inflammation between tumour-bearing hosts are largely unresolved. Here, using a panel of 16 distinct genetically engineered mouse models for breast cancer, we uncover a role for cancer-cell-intrinsic p53 as a key regulator of pro-metastatic neutrophils. Mechanistically, loss of p53 in cancer cells induced the secretion of WNT ligands that stimulate tumour-associated macrophages to produce IL-1β, thus driving systemic inflammation. Pharmacological and genetic blockade of WNT secretion in p53-null cancer cells reverses macrophage production of IL-1β and subsequent neutrophilic inflammation, resulting in reduced metastasis formation. Collectively, we demonstrate a mechanistic link between the loss of p53 in cancer cells, secretion of WNT ligands and systemic neutrophilia that potentiates metastatic progression. These insights illustrate the importance of the genetic makeup of breast tumours in dictating pro-metastatic systemic inflammation, and set the stage for personalized immune intervention strategies for patients with cancer.

Background: Children with disabilities are more likely to experience violence or injury at school and at home, but there is little evidence from Central Asia. Objective: To describe the prevalence of disability and associations with peer violence perpetration and victimization, depression, corporal punishment, school performance and school attendance, among middle school children in Pakistan and Afghanistan. Method: This is a secondary analysis of data gathered in the course of evaluations of interventions to prevent peer violence conducted in Pakistan and Afghanistan as part of the ‘What Works to Prevent Violence against Women and Girls Global Programme’. In Pakistan, the research was conducted in 40 schools, and disability was assessed at midline in 1516 interviews with Grade 7s. In Afghanistan, the data were from the baseline study conducted in 11 schools with 770 children. Generalized Linear Mixed Modeling was used to assess associations with disability. Results: In Afghanistan, the prevalence of disability was much higher for girls (22.1%) than boys (12.9%), while in Pakistan 6.0% of boys and girls reported a disability. Peer violence victimization was strongly associated with disability in Afghanistan and marginally associated in Pakistan. In Pakistan, perpetration of peer violence was associated with disability. In both countries, disability was significantly associated with higher depression scores. Food insecurity was strongly associated with disability in Afghanistan. Conclusion: Disability is highly prevalent in Afghanistan and Pakistan schools and this is associated with a greater risk of experiencing and perpetrating peer violence. It is important to ensure that all children can benefit from school-based prevention interventions.

IntroductionWomen with disabilities experience higher rates of intimate partner violence (IPV) than women without disabilities. There remains limited evidence about whether IPV prevention interventions for the general population have benefits for women with disabilities that compare to those for women without disabilities. Using data from IPV prevention randomised controlled trials in diverse locations (Rwanda, South Africa and Afghanistan), we assess whether outcomes differed by disability status.MethodsWe assessed disability at baseline in three IPV prevention trials. We performed post-hoc analysis of intervention impacts at endline (22 or 24 months post-baseline) stratified by disability status at study baseline and tested an interaction term for disability at baseline by intervention arm for three sets of outcomes: (1) past year experiences of physical, sexual and severe IPV; (2) economic and livelihood outcomes; and (3) health, mental health and substance use outcomes.ResultsAt baseline between 17.7% and 26.2% of women reported being disabled. For IPV prevention, in seven out of eight tests across three studies, women with and without disabilities had similar outcomes. For economic, health and substance use outcomes, there was more variation, with women with disabilities reporting both better and worse outcomes than women without disabilities; however there was no clear pattern in these differential results.ConclusionIPV prevention programmes targeting general populations can prevent IPV among women with disabilities participants with benefits that mirror those for women without disabilities. Benefits for participants with and without disabilities on secondary programme outcomes related to economic empowerment and health may be more varied and should be explicitly monitored.

SUMMARYInhibitors of poly(ADP-ribose) (PAR) polymerase (PARPi) have entered the clinic for the treatment of homologous recombination (HR)-deficient cancers. Despite the success of this approach, preclinical and clinical research with PARPi has revealed multiple resistance mechanisms, highlighting the need for identification of novel functional biomarkers and combination treatment strategies. Functional genetic screens performed in cells and organoids that acquired resistance to PARPi by loss of 53BP1 identified loss of LIG3 as an enhancer of PARPi toxicity in BRCA1-deficient cells. Enhancement of PARPi toxicity by LIG3 depletion is dependent on BRCA1 deficiency but independent of the loss of 53BP1 pathway. Mechanistically, we show that LIG3 loss promotes formation of MRE11-mediated post-replicative ssDNA gaps in BRCA1-deficient and BRCA1/53BP1 double-deficient cells exposed to PARPi, leading to an accumulation of chromosomal abnormalities. LIG3 depletion also enhances efficacy of PARPi against BRCA1-deficient mammary tumors in mice, suggesting LIG3 as a potential therapeutic target.

Purpose: Because BRCA1 is a high-risk breast/ovarian cancer susceptibility gene, BRCA1 sequence variants of uncertain clinical significance (VUS) complicate genetic counseling. As most VUS are rare, reliable classification based on clinical and genetic data is often impossible. However, all pathogenic BRCA1 variants analyzed result in defective homologous recombination DNA repair (HRR). Thus, BRCA1 VUS may be categorized based on their functional impact on this pathway. Experimental Design: Two hundred thirty-eight BRCA1 VUS—comprising most BRCA1 VUS known in the Netherlands and Belgium—were tested for their ability to complement Brca1-deficient mouse embryonic stem cells in HRR, using cisplatin and olaparib sensitivity assays and a direct repeat GFP (DR-GFP) HRR assay. Assays were validated using 25 known benign and 25 known pathogenic BRCA1 variants. For assessment of pathogenicity by a multifactorial likelihood analysis method, we collected clinical and genetic data for functionally deleterious VUS and VUS occurring in three or more families. Results: All three assays showed 100% sensitivity and specificity (95% confidence interval, 83%–100%). Out of 238 VUS, 45 showed functional defects, 26 of which were deleterious in all three assays. For 12 of these 26 variants, we could calculate the probability of pathogenicity using clinical and genetic data, resulting in the identification of 7 (likely) pathogenic variants. Conclusions: We have functionally categorized 238 BRCA1 VUS using three different HRR-related assays. Classification based on clinical and genetic data alone for a subset of these variants confirmed the high sensitivity and specificity of our functional assays.

BecauseTwo hundred thirty-eightAll three assays showed 100% sensitivity and specificity (95% confidence interval, 83%-100%). Out of 238 VUS, 45 showed functional defects, 26 of which were deleterious in all three assays. For 13 of these 26 variants, we could calculate the probability of pathogenicity using clinical and genetic data, resulting in the identification of 7 (likely) pathogenic variants.We have functionally categorized 238

Inhibitors of poly(ADP-ribose) (PAR) polymerase (PARPi) have entered the clinic for the treatment of homologous recombination (HR)-deficient cancers. Despite the success of this approach, preclinical and clinical research with PARPi has revealed multiple resistance mechanisms, highlighting the need for identification of novel functional biomarkers and combination treatment strategies. Functional genetic screens performed in cells and organoids that acquired resistance to PARPi by loss of 53BP1, identified loss of LIG3 as an enhancer of PARPi toxicity in BRCA1-deficient cells. Enhancement of PARPi toxicity by LIG3 depletion is dependent on BRCA1 deficiency but independent of the loss of 53BP1 pathway. Mechanistically, we show that LIG3 is required for PARPi-induced fork acceleration in BRCA1-deficient cells and that LIG3 loss increases fork asymmetry. Furthermore, LIG3 depletion in BRCA1-deficient cells results in an increase in ssDNA gaps behind the replication forks, resulting in accumulation of chromosomal abnormalities. We also report that high expression of LIG3 in patients with invasive breast cancer correlates in with poorer overall survival, rendering LIG3 as a potential therapeutic target for enhancing PARPi sensitivity.

Effective treatment of invasive lobular carcinoma (ILC) of the breast is hampered by late detection, invasive growth, distant metastasis, and poor response to chemotherapy. Phosphoinositide 3-kinase (PI3K) signaling, one of the major druggable oncogenic signaling networks, is frequently activated in ILC. We investigated treatment response and resistance to AZD8055, an inhibitor of mammalian target of rapamycin (mTOR), in the K14-cre;Cdh1Flox/Flox;Trp53Flox/Flox (KEP) mouse model of metastatic ILC. Inhibition of mTOR signaling blocked the growth of primary KEP tumors as well as the progression of metastatic disease. However, primary tumors and distant metastases eventually acquired resistance after long-term AZD8055 treatment, despite continued effective suppression of mTOR signaling in cancer cells. Interestingly, therapeutic responses were associated with increased expression of genes related to antigen presentation. Consistent with this observation, increased numbers of tumor-infiltrating major histocompatibility complex class II-positive (MHCII+) immune cells were observed in treatment-responsive KEP tumors. Acquisition of treatment resistance was associated with loss of MHCII+ cells and reduced expression of genes related to the adaptive immune system. The therapeutic efficacy of mTOR inhibition was reduced in Rag1−/- mice lacking mature T and B lymphocytes, compared to immunocompetent mice. Furthermore, therapy responsiveness could be partially rescued by transplanting AZD8055-resistant KEP tumors into treatment-naïve immunocompetent hosts. Collectively, these data indicate that the PI3K signaling pathway is an attractive therapeutic target in invasive lobular carcinoma, and that part of the therapeutic effect of mTOR inhibition is mediated by the adaptive immune system.

IntroductionIntimate partner violence (IPV) is a serious public health and human rights violation which impacts approximately one in three women worldwide. Some existing evidence suggests that women with disabilities are at higher risk of IPV, but is largely limited in geographical scope to the Global North, and comparison across settings has been hampered by inconsistent measurement of both IPV and disability.MethodsPooled analysis of baseline data from 8549 adult women participating in seven IPV prevention studies in five countries across Africa and Asia that used collaborative, comparative measurement strategies to assess both disability and IPV.ResultsAfter adjusting for age, women with disabilities were more likely to experience past 12-month physical IPV (adjusted OR (aOR)=1.79; 95% CI 1.49 to 2.17), sexual IPV (aOR=1.98; 95% CI 1.36 to 2.89), emotional IPV (aOR=1.84; 95% CI 1.49 to 2.27) and economic IPV (aOR=1.66; 95% CI 1.45 to 1.89), with an overall association between disability and past 12-month physical/sexual IPV of aOR=1.93 (95% CI 1.52 to 2.46). Compared to women without disability, women with moderate and severe disability showed a trend of increasing risk of IPV in the past 12 months for each of physical, sexual, emotional and economic IPV. Overall, both women with moderate disability (aOR=1.86, 95% CI 1.57 to 2.21) and women with severe disability (aOR=2.63; 95% CI 1.95 to 3.55) were significantly more likely to experience any form of IPV when compared with women without disability.ConclusionWomen with disabilities are at increased risk of past-year IPV compared to women without disabilities across a range of settings in the Global South, and the risk of IPV increases with increasing severity of disability. IPV prevention and response efforts in these settings must find ways to include and address the needs of women with disabilities, including increased outreach and improved accessibility of programmes.

Women with disabilities are more vulnerable to violence, including intimate partner violence (IPV), yet the majority of emerging IPV prevention programs fail to explicitly consider the needs of participants with disabilities. Women and men living with disabilities engaged with IPV prevention programs in four countries were interviewed to explore how disability shaped their experiences of gender, violence, IPV, and whether the programs met their disability related needs. In-depth interviews were conducted with 16 women and 15 men living with disabilities in Ghana, Rwanda, Tajikistan and South Africa. The data were analysed thematically and compared across the settings. Participants described experiencing disability-related stigma, discrimination, exclusion, and for women, increased vulnerability to IPV. Barriers to full participation in programs included limited accessibility, and lack of disability-specific materials, recruitment or outreach. Enablers of inclusion included recruitment and monitoring strategies aimed at people with disabilities, partnering with a local disabled people’s organization, training staff in disability inclusion, and raising awareness of disability rights. The data encouragingly suggests that inclusion of women and men with disabilities in IPV prevention programs designed for the general population has beneficial outcomes. Inclusion can prevent violence, promote their wellbeing, support economic empowerment, and challenge disability-related stigma and discrimination.

Globally, women with disabilities experience heightened and unique forms of violence compared to men with disabilities and women without disabilities. Yet formalised guidelines for their inclusion in gender-based violence (GBV) research is lacking. This paper draws on ethical guidelines for researching violence against women, and studies on the ethicality of including people with disabilities in research, to advocate for women with disabilities' inclusion and safety in GBV research. Reflecting on lessons from a qualitative study on violence against women living with disabilities in South Africa, the paper considers what could be of value for GBV researchers and ethics review committees in low-middle income countries (LMICs). It aims to stimulate debate around the integration of reasonable accommodation, accessibility, and equal participation of women with disabilities in planning and conducting ethical GBV research. The paper recommends that considerations are practically applied and tested in other LMICs, and thereafter critiqued in consultation with a range of stakeholders and women with disabilities, to enhance best practice and form a basis for developing guidelines for undertaking ethical and inclusive GBV research in LMICs.

Notions of womanhood inculcate naturalised ideologies of femininity, sexuality, motherhood and caregiving. The paper asks how disability stigma intersects with womanhood to characterise intimate partnerships in South Africa. In-depth interviews with 30 women with a range of disabilities were conducted in informal settlements in Cape Town. Findings suggest that disability stigma may hamper attainment of normative womanhood and sexual relationships for women with disabilities in South Africa. Limited opportunities to meet potential partners, hegemonic gender expectations and restricted sexual and physical contact shape their intimate partnerships. However, women with disabilities also challenge ableist constructs of normalcy and discredit negative images of disabled womanhood. Because of this, theoretical models of intimate partner violence should consider the influence of disability on constructions of sexuality and norms in intimate partnerships. Building on women with disabilities' stigma-avoidance strategies will help facilitate better relationship outcomes. Social norms interventions with broader society, communities, women with disabilities and their partners, family and carers can help destabilise assumptions that women with disabilities are unable to have long-lasting and fulfilling sexual and intimate partnerships. Moreover, accessible and relevant sexuality education and information on relationships, intimate partner violence, maternal and sexual and reproductive health care can ensure healthy and safe intimate partnerships for women with disabilities.

Background Being diagnosed with human immunodeficiency virus (HIV), and labelled with a chronic, life-threatening, and often stigmatizing disease, can impact on a person's well-being. Psychosocial group interventions aim to improve life-functioning and coping as individuals adjust to the diagnosis. Objectives To examine the effectiveness of psychosocial group interventions for improving the psychological well-being of adults living with HIV/AIDS. Search methods We searched the following electronic databases up to 14 March 2016: the Cochrane Central Register of Controlled Trials (CENTRAL) published in the Cochrane Library (Issue 2, 2016), PubMed (MEDLINE) (1996 to 14 March 2016), Embase (1996 to 14 March 2016), and Clinical Trials.gov. Selection criteria Randomized controlled trials (RCTs) or quasi-RCTs that compared psychosocial group interventions with versus control (standard care or brief educational interventions), with at least three months follow-up post-intervention. We included trials that reported measures of depression, anxiety, stress, or coping using standardized scales. Data collection and analysis Two review authors independently screened abstracts, applied the inclusion criteria, and extracted data. We compared continuous outcomes using mean differences (MD) with 95% confidence intervals (95% CIs), and pooled data using a random-effects model. When the included trials used different measurement scales, we pooled data using standardized mean difference (SMD) values. We reported trials that we could not include in the meta analysis narratively in the text. We assessed the certainty of the evidence using the GRADE approach. Main results We included 16 trials (19 articles) that enrolled 2520 adults living with HIV. All the interventions were multifaceted and included a mix of psychotherapy, relaxation, group support, and education. The included trials were conducted in the USA (12 trials), Canada (one trial), Switzerland (one trial), Uganda (one trial), and South Africa (one trial), and published between 1996 and 2016. Ten trials recruited men and women, four trials recruited homosexual men, and two trials recruited women only. Interventions were conducted with groups of four to 15 people, for 90 to 135 minutes, every week for up to 12 weeks. All interventions were conducted face-to-face except two, which were delivered by telephone. All were delivered by graduate or postgraduate trained health, psychology, or social care professionals except one that used a lay community health worker and two that used trained mindfulness practitioners. Group-based psychosocial interventions based on cognitive behavioural therapy (CBT) may have a small effect on measures of depression, and this effect may last for up to 15 months after participation in the group sessions (SMD −0.26, 95% CI −0.42 to −0.10; 1139 participants, 10 trials, low certainty evidence). Most trials used the Beck Depression Inventory (BDI), which has a maximum score of 63, and the mean score in the intervention groups was around 1.4 points lower at the end of follow-up. This small benefit was consistent across five trials where participants had a mean depression score in the normal range at baseline, but trials where the mean score was in the depression range at baseline effects were less consistent. Fewer trials reported measures of anxiety, where there may be little or no effect (four trials, 471 participants, low certainty evidence), stress, where there may be little or no effect (five trials, 507 participants, low certainty evidence), and coping (five trials, 697 participants, low certainty evidence). Group-based interventions based on mindfulness have not demonstrated effects on measures of depression (SMD −0.23, 95% CI −0.49 to 0.03; 233 participants, 2 trials, very low certainty evidence), anxiety (SMD −0.16, 95% CI −0.47 to 0.15; 62 participants, 2 trials, very low certainty evidence), or stress (MD −2.02, 95% CI −4.23 to 0.19; 137 participants, 2 trials, very low certainty evidence). No mindfulness based interventions included in the studies had any valid measurements of coping. Authors' conclusions Group-based psychosocial interventions may have a small effect on measures of depression, but the clinical importance of this is unclear. More high quality evidence is needed to assess whether group psychosocial intervention improve psychological well-being in HIV positive adults. Does group therapy improve well-being in people living with HIV? Cochrane researchers conducted a review of the effects of group therapy for people living with human immunodeficiency virus (HIV). After searching for relevant trials up to 14 March 2016, they included 16 trials reported in 19 articles that enrolled 2520 adults living with HIV. The included trials were conducted in the USA (12 trials), Canada (one trial), Switzerland (one trial), Uganda (one trial), and South Africa (one trial), and published between 1996 and 2016. Ten trials recruited men and women, four trials recruited homosexual men, and two trials recruited women only. What is group therapy and how might if benefit people with HIV? Group therapy aims to improve the well-being of individuals by delivering psychological therapy in a group format, which can encourage the development of peer support and social networks. Group therapy often also incorporates training in relaxation techniques and coping skills, and education on the illness and its management. Human immunodeficiency virus (HIV) causes a chronic, life threatening, and often stigmatising disease, which can impact on a person's well-being. Group therapy could help people living with HIV to adapt to knowing they have HIV, or recover from depression, anxiety, and stress. What the research says Group-based therapy based on cognitive behavioural therapy may have a small effect on measures of depression, and this effect may last for up to 15 months after participation in the group sessions (low certainty evidence). This effect was apparent in groups who did not appear to be depressed on clinical scoring systems before the therapy started. The research also showed there may be little or no effect on measures of anxiety, stress, and coping (low certainty evidence). Group-based interventions based on mindfulness have been studied in two small trials, and have not demonstrated effects on measures of depression, anxiety or stress (all very low certainty evidence). No mindfulness based interventions included in the studies had any valid measurements of coping. Overall, the review suggests that existing interventions have little to no effect in increasing psychological adjustment to living with HIV. More good quality studies are required to inform good practice and evidence.

Transactional sex is a common feature of sexual relationships in South Africa but has severe health implications for those who engage in it. This paper presents perspectives on transactional sex based on interviews and focus group discussions with young people in Gauteng and Limpopo, South Africa. The discussions were part of an evaluation of a peer education programme promoting HIV prevention called Vhutshilo , aimed at 14–16 year olds. The session on transactional sex entitled ‘Something for Something’ evoked strong responses from youth. Youth recognised transactional sex as a common phenomenon in their communities and associated it with many risks. However, when comparing young people’s qualitative responses to the impact of the session as measured by the quantitative impact survey, no significant differences were discernible between youth in the programme and those in a control group who were not exposed to the session. Further analysis showed that the content of the session was limited to the negative consequences and health risks of transactional sex and focused mostly on the adverse contexts in South Africa that force youth into such relationships. The session did little to situate transactional relationships within the everyday realities of sexual decision making and youth values of peer status and consumerism. We argue that the session’s findings reveal a narrow understanding of the dynamic contexts under which transactional sex occurs and fails to take into account the resilience of youth to make choices of whether or not to engage in such relationships, and how they can engage in these types of relationships safely. We conclude that HIV prevention curriculums need to leverage youth resilience and protective skills within the confines of difficult economic and social circumstances to allow them to successfully navigate safer sexual relationships. Keywords: transactional sex, HIV/AIDS, sexual health, youth, risk behaviour, peer education, resilience, agency African Journal of AIDS Research 2014, 13(1): 53–63

Breast and ovarian cancer patients harboring BRCA1/2 germline mutations have clinically benefitted from therapy with PARP inhibitor (PARPi) or platinum compounds, but acquired resistance limits clinical impact. In this study, we investigated the impact of mutations on BRCA1 isoform expression and therapeutic response. Cancer cell lines and tumors harboring mutations in exon 11 of BRCA1 express a BRCA1-Δ11q splice variant lacking the majority of exon 11. The introduction of frameshift mutations to exon 11 resulted in nonsense-mediated mRNA decay of full-length, but not the BRCA1-Δ11q isoform. CRISPR/Cas9 gene editing as well as overexpression experiments revealed that the BRCA1-Δ11q protein was capable of promoting partial PARPi and cisplatin resistance relative to full-length BRCA1, both in vitro and in vivo. Furthermore, spliceosome inhibitors reduced BRCA1-Δ11q levels and sensitized cells carrying exon 11 mutations to PARPi treatment. Taken together, our results provided evidence that cancer cells employ a strategy to remove deleterious germline BRCA1 mutations through alternative mRNA splicing, giving rise to isoforms that retain residual activity and contribute to therapeutic resistance. Cancer Res; 76(9); 2778–90. ©2016 AACR.

South Africa has unprecedented levels of violence and many South African women are exposed to violence during their lifetime. This article explores how gender and disability intersect in women’s experiences of violence during their lifetime. Repeat in-depth qualitative interviews with 30 physically disabled women in Cape Town reveal that women with physical disabilities are exposed to various forms of violence, and shows how their impairments shape their violence experiences. The most common forms of violence women with disabilities experience are psychological violence, financial abuse, neglect, and deprivation, with disability stigma playing a central role and contributing to how women with disabilities are exploited and dehumanized. Constructions of women as asexual shape their sexual relationships and experiences of sexual violence. This article identifies that women with disabilities are more at risk and experience additional layers of violence than women without disabilities. These additional risks and layers of violence need to be recognized and inform interventions to prevent and respond to violence against women with disabilities in the country. Prevention of violence against women with physical disabilities in South Africa needs to address the role of disability stigma that shapes the types of violence they experience, change gender norms, and create accessible and safe environments and economic empowerment opportunities.

Introduction: In the context of poverty and HIV and AIDS, peer education is thought to be capable of providing vulnerable youth with psychosocial support as well as information and decision-making skills otherwise limited by scarce social and material resources. As a preventative education intervention method, peer education is a strategy aimed at norms and peer group influences that affect health behaviours and attitudes. However, too few evaluations of peer-led programmes are available,and they frequently fail to reflect real differences between those who have been recipients of peer education and those who have not. This article reports on an evaluation of a pilot peer-led intervention, entitled Vhutshilo, implemented on principles agreed upon through a collaborative effort in South Africa by the Harvard School of Public Health and the Centre for theSupport of Peer Education (the Rutanang collaboration). Vhutshilo targeted vulnerable adolescents aged 14–16 years living in some of South Africa’s under-resourced communities. Methodology: The research design was a mixed-method (qualitative and quantitative), longitudinal, quasi-experimental evaluation. Tools used included a quantitative survey questionnaire (n ¼ 183) and semi-structured interviews (n ¼ 32) with beneficiaries of peer education. Surveys were administered twice forbeneficiaries of peer education (n ¼ 73), immediately after completion of the programme (post-test) and 4 months later (delayed post-test), and once for control group members (n ¼ 110). The three main methodological limitations in this study were the use of a once-off control group assessment as the baseline for comparison, without a pre-test, due to timing and resource constraints; a small sample size (n ¼ 183), which reduced the statistical power of the evaluation; and the unavailability of existing tested survey questions to measure the impact of peer education and its role in behaviour change. Findings: This article reports on the difficulties of designing a comprehensive evaluation within time and financial constraints, critically evaluates survey design with multi-item indicators, and discusses six statistically significant changes observed in Vhutshilo participants out of a 92-point survey. Youth struggling with poor quality education and living in economically fraught contexts with little social support, nonetheless, showed evidence of having greater knowledge of support networks and an expanded emotional repertoire by the end of the Vhutshilo programme, and 4 months later. At both individual and group level, many with low socio-economic status showed great improvement with regard to programme indicator scores. Conclusion: For the poorest adolescents, especially those living in the rural parts of South Africa, peer education has the potential to change future orientation, attitudes and knowledge regarding HIV and AIDS, including an intolerance for multiple concurrent partnerships. When well organised and properly supported, peer education programmes (and the Vhutshilo curriculum, in particular) provide vulnerable youth with opportunities to develop psychosocial skills and informational resources that contribute to the changing of norms, attitudes and behaviours. However, the article also flags the need for effective peer education evaluations that take into account limited financial resources and that possess tested indicators of programme effectiveness.

We have studied in vivo responses of “spontaneous” Brca1 - and p53 -deficient mammary tumors arising in conditional mouse mutants to treatment with doxorubicin, docetaxel, or cisplatin. Like human tumors, the response of individual mouse tumors varies, but eventually they all become resistant to the maximum tolerable dose of doxorubicin or docetaxel. The tumors also respond well to cisplatin but do not become resistant, even after multiple treatments in which tumors appear to regrow from a small fraction of surviving cells. Classical biochemical resistance mechanisms, such as up-regulated drug transporters, appear to be responsible for doxorubicin resistance, rather than alterations in drug-damage effector pathways. Our results underline the promise of these mouse tumors for the study of tumor-initiating cells and of drug therapy of human cancer.

The human and murine genes for MRP9 (multidrug resistance-associated protein 9; ABCC12) yield many alternatively spliced RNAs. Using a panel of monoclonal antibodies, we detected full-length Mrp9 only in testicular germ cells and mouse sperm; we obtained no evidence for the existence of the truncated 100 kDa MRP9 protein reported previously. In contrast with other MRPs, neither murine Mrp9 nor the human MRP9 produced in MRP9-transfected HEK-293 cells (human embryonic kidney cells) appears to contain N-linked carbohydrates. In mouse and boar sperm, Mrp9 localizes to the midpiece, a structure containing all sperm mitochondria. However, immunolocalization microscopy and cell fractionation studies with transfected HEK-293 cells and mouse testis show that MRP9/Mrp9 does not localize to mitochondria. In HEK-293 cells, it is predominantly localized in the endoplasmic reticulum. We have been unable to demonstrate transport by MRP9 of substrates transported by other MRPs, such as drug conjugates and other organic anions.

cGMP secretion from cells can be mediated by ATP-binding cassette (ABC) transporters ABCC4, ABCC5, and ABCC11. Indirect evidence suggests that ABCC4 and ABCC5 contribute to cGMP transport by erythrocytes. We have re-investigated the issue using erythrocytes from wild-type and transporter knockout mice. Murine wild-type erythrocyte vesicles transported cGMP with an apparent Km that was 100-fold higher than their human counterparts, the apparent Vmax being similar. Whereas cGMP transport into human vesicles was efficiently inhibited by the ABCC4-specific substrate prostaglandin E1, cGMP transport into mouse vesicles was inhibited equally by Abcg2 and Abcc4 inhibitors/substrates. Similarly, cGMP transport into vesicles from Abcc4–/– and Abcg2–/– mice was 42% and 51% of that into wild-type mouse vesicles, respectively, whereas cGMP transport into vesicles from Abcc4–/–/Abcg2–/– mice was near background. The knockout mice were used to show that Abcg2-mediated cGMP transport occurred with lower affinity but higher Vmax than Abcc4-mediated transport. Involvement of Abcg2 in cGMP transport by Abcc4–/– erythrocyte vesicles was supported by higher transport at pH 5.5 than at pH 7.4, a characteristic of Abcg2-mediated transport. The relative contribution of ABCC4/Abcc4 and ABCG2/Abcg2 in cGMP transport was confirmed with a new inhibitor of ABCC4 transport, the protease inhibitor 4-(2-aminoethyl)benzenesulfonyl fluoride.

Prostaglandins are involved in a wide variety of physiological and pathophysiological processes, but the mechanism of prostaglandin release from cells is not completely understood. Although poorly membrane permeable, prostaglandins are believed to exit cells by passive diffusion. We have investigated the interaction between prostaglandins and members of the ATP-binding cassette (ABC) transporter ABCC [multidrug resistance protein (MRP)] family of membrane export pumps. In inside-out membrane vesicles derived from insect cells or HEK293 cells, MRP4 catalyzed the time- and ATP-dependent uptake of prostaglandin E 1 (PGE 1 ) and PGE 2 . In contrast, MRP1, MRP2, MRP3, and MRP5 did not transport PGE 1 or PGE 2 . The MRP4-mediated transport of PGE 1 and PGE 2 displayed saturation kinetics, with K m values of 2.1 and 3.4 μM, respectively. Further studies showed that PGF 1 α , PGF 2 α , PGA 1 , and thromboxane B 2 were high-affinity inhibitors (and therefore presumably substrates) of MRP4. Furthermore, several nonsteroidal antiinflammatory drugs were potent inhibitors of MRP4 at concentrations that did not inhibit MRP1. In cells expressing the prostaglandin transporter PGT, the steady-state accumulation of PGE 1 and PGE 2 was reduced proportional to MRP4 expression. Inhibition of MRP4 by an MRP4-specific RNA interference construct or by indomethacin reversed this accumulation deficit. Together, these data suggest that MRP4 can release prostaglandins from cells, and that, in addition to inhibiting prostaglandin synthesis, some nonsteroidal antiinflammatory drugs might also act by inhibiting this release.

Human multidrug-resistance protein (MRP) 4 transports cyclic nucleotides and when overproduced in mammalian cells mediates resistance to some nucleoside analogues. Recently, it has been shown that Mrp4 is induced in the livers of Fxr ((-/-)) mice, which have increased levels of serum bile acids. Since MRP4, like MRP1-3, also mediates transport of a model steroid conjugate substrate, oestradiol 17-beta-D-glucuronide (E(2)17betaG), we tested whether MRP4 may be involved in the transport of steroid and bile acid conjugates. Bile salts, especially sulphated derivatives, and cholestatic oestrogens inhibited the MRP4-mediated transport of E(2)17betaG. Inhibition by oestradiol 3,17-disulphate and taurolithocholate 3-sulphate was competitive, suggesting that these compounds are MRP4 substrates. Furthermore, we found that MRP4 transports dehydroepiandrosterone 3-sulphate (DHEAS), the most abundant circulating steroid in humans, which is made in the adrenal gland. The ATP-dependent transport of DHEAS by MRP4 showed saturable kinetics with K (m) and V (max) values of 2 microM and 45 pmol/mg per min, respectively (at 27 degrees C). We further studied the possible involvement of other members of the MRP family of transporters in the transport of DHEAS. We found that MRP1 transports DHEAS in a glutathione-dependent manner and exhibits K (m) and V (max) values of 5 microM and 73 pmol/mg per min, respectively (at 27 degrees C). No transport of DHEAS was observed in membrane vesicles containing MRP2 or MRP3. Our findings suggest a physiological role for MRP1 and MRP4 in DHEAS transport and an involvement of MRP4 in transport of conjugated steroids and bile acids.

Developing interventions that address psychosocial wellbeing of people living with HIV is critical to ensure strong linkages to and retention in HIV care. This paper describes the development of Sondela, an HIV adjustment and coping intervention for heterosexual men and women living with HIV, and its relevance and feasibility in the South African context. Sondela is a six three-hour, small group-based, participatory workshop series. We followed an iterative, multi-phased process of curriculum development that involved research, theoretical frameworks and piloting. A systematic review highlighted the absence of psychosocial interventions targeting heterosexual HIV positive populations living in high HIV prevalence and resource-poor settings. Formative studies demonstrated risk and social factors associated with adjustment and coping with HIV, emphasising the need for interventions that acknowledge gendered experiences. Our pilot of Sondela demonstrated high levels of relevance and feasibility. Men appreciated the workshop "space" to openly talk about their HIV positive status and what is means for their role as partners and fathers and friends. Women valued the skills and approaches because they were relevant to "real life" situations and not just about HIV. Sondela promises to be valuable in supporting health system initiatives and psychosocial support to strengthen linkages to and retention in HIV care, and this suggests a need for rigorous evaluation of Sondela to establish evidence for its effectiveness in a general population.

This is a protocol for a Cochrane Review (Intervention). The objectives are as follows: To examine the effectiveness of psychosocial group interventions for improving psychological health, psychosocial adjustment and QOL in adults living with HIV/AIDS.

Mutations in BRCA1 and BRCA2 account for the majority of hereditary breast and ovarian cancers, and therefore sequence analysis of both genes is routinely conducted in patients with early-onset breast cancer. Besides mutations that clearly abolish protein function or are known to increase cancer risk, a large number of sequence variants of uncertain significance (VUS) have been identified. Although several functional assays for BRCA1 VUSs have been described, thus far it has not been possible to conduct a high-throughput analysis in the context of the full-length protein. We have developed a relatively fast and easy cDNA-based functional assay to classify BRCA1 VUSs based on their ability to functionally complement BRCA1-deficient mouse embryonic stem cells. Using this assay, we have analyzed 74 unclassified BRCA1 missense mutants for which all predicted pathogenic variants are confined to the BRCA1 RING and BRCT domains. Significance: BRCA1 VUSs are frequently found in patients with hereditary breast or ovarian cancer and present a serious problem for clinical geneticists. This article describes the generation, validation, and application of a reliable high-throughput assay for the functional classification of BRCA1 sequence variants of uncertain significance. Cancer Discov; 3(10); 1142–55. ©2013 AACR. This article is highlighted in the In This Issue feature, p. 1083

Heterozygous germline mutations in breast cancer 1 (BRCA1) strongly predispose women to breast cancer. BRCA1 plays an important role in DNA double-strand break (DSB) repair via homologous recombination (HR), which is important for tumor suppression. Although BRCA1-deficient cells are highly sensitive to treatment with DSB-inducing agents through their HR deficiency (HRD), BRCA1-associated tumors display heterogeneous responses to platinum drugs and poly(ADP-ribose) polymerase (PARP) inhibitors in clinical trials. It is unclear whether all pathogenic BRCA1 mutations have similar effects on the response to therapy. Here, we have investigated mammary tumorigenesis and therapy sensitivity in mice carrying the Brca1(185stop) and Brca1(5382stop) alleles, which respectively mimic the 2 most common BRCA1 founder mutations, BRCA1(185delAG) and BRCA1(5382insC). Both the Brca1(185stop) and Brca1(5382stop) mutations predisposed animals to mammary tumors, but Brca1(185stop) tumors responded markedly worse to HRD-targeted therapy than did Brca1(5382stop) tumors. Mice expressing Brca1(185stop) mutations also developed therapy resistance more rapidly than did mice expressing Brca1(5382stop). We determined that both murine Brca1(185stop) tumors and human BRCA1(185delAG) breast cancer cells expressed a really interesting new gene domain-less (RING-less) BRCA1 protein that mediated resistance to HRD-targeted therapies. Together, these results suggest that expression of RING-less BRCA1 may serve as a marker to predict poor response to DSB-inducing therapy in human cancer patients.

Hereditary breast cancers are frequently caused by germline BRCA1 mutations. The BRCA1(C61G) mutation in the BRCA1 RING domain is a common pathogenic missense variant, which reduces BRCA1/BARD1 heterodimerization and abrogates its ubiquitin ligase activity. To investigate the role of BRCA1 RING function in tumor suppression and therapy response, we introduced the Brca1(C61G) mutation in a conditional mouse model for BRCA1-associated breast cancer. In contrast to BRCA1-deficient mammary carcinomas, tumors carrying the Brca1(C61G) mutation responded poorly to platinum drugs and PARP inhibition and rapidly developed resistance while retaining the Brca1(C61G) mutation. These findings point to hypomorphic activity of the BRCA1-C61G protein that, although unable to prevent tumor development, affects response to therapy.

In addressing the psychosocial effects of the HIV and AIDS pandemic among vulnerable children, the issue of bereavement appears inadequately addressed. Amid the global discourse on children orphaned and made vulnerable by HIV and AIDS, this paper explores how cultural contexts and social environments in South Africa shape children’s experience of grief. The argument draws on a number of qualitative studies and uses empirical evidence from an evaluation of a peer-led HIV/AIDS-prevention strategy aimed at providing psychosocial support for 10- to 13-year-old South African children living in resource-poor communities. The paper reveals a central paradox regarding how the intervention’s objective of talking about death and eliciting memories of deceased loved ones with young children is confounded by cultural practices located in notions of silence and the need to protect children. The paper acknowledges the ‘culture of silence’ surrounding death in some African contexts, but concludes that peer-led strategies have the potential to naturally circumvent these cultural taboos, simultaneously creating a much-needed space for young children to cry and talk among themselves, even if remaining silent at home in the presence of adults.Keywords: cultural practices; empowerment; grief; group therapy; orphaned and vulnerable children; peer education; psychosocial aspects; social dynamics; social support programmesAfrican Journal of AIDS Research 2010, 9(1): 41–50