Targeting APC loss using synthetic lethality in Colorectal Cancer

PhDMutations in the tumour suppressor gene Adenomatous polyposis coli (APC) are found in 80 % of sporadic colorectal cancer (CRC) tumours and are also responsible for the inherited form of CRC, Familial adenomatous polyposis (FAP). In order to identify novel therapeutic targets for the treatment of APC mutated CRC, we have generated an in vitro model of APC mutant CRC using CRISPR-cas9 gene editing. Using the APC wildtype colorectal carcinoma cell line RKO, we targeted the cells with guide RNA (gRNA) targeting exon 2 or exon 15 (encodes 80 % of APC) of the APC gene. We generated isogenic cell lines which differed in the expression of APC, the controls were APC wildtype and the APC mutant (APC Lys736fs) cell lines expressed a truncated ~80 kDa APC protein. We used these cell lines to perform an siRNA screen against 720 kinases and kinase-related genes. We selected seven genes to investigate further, unfortunately none of the potential hits validated. Additionally, we performed an FDA-approved compound screen targeting over 1000 compounds. From this, we identified a group of HMG-CoA reductase (HMGCR) inhibitors known as statins, which selectively cause a greater loss in cell viability in the APC mutated cell lines, compared to the APC wildtype cells. Mechanistically, our data suggests this synthetic lethal relationship is due to a greater decrease in the anti-apoptotic protein survivin. We propose this is due to statins altering the localisation of Rac1, reducing Pak1 activation and reducing the level of Wnt signalling. This results in the reduction of the Wnt target gene survivin. We have successfully identified an FDA-approved family of compounds, which show synthetic lethality with the APC mutation in our in vitro model.Bowel and Cancer Research, and the Rosetree trus

Are current approaches for measuring access to clean water and sanitation inclusive of people with disabilities? Comparison of individual- and household-level access between people with and without disabilities in the Tanahun district of Nepal.

BACKGROUND: The critical importance of safe and affordable access to water, sanitation and hygiene (WASH) is highlighted in Goal 6 of the Sustainable Development Goals, which seeks to achieve universal and equitable access for all by 2030. However, people with disabilities-who comprise 15% of the global population-frequently face difficulties meeting their WASH needs. Unmet WASH needs amongst people with disabilities may not be captured through current approaches to tracking progress towards Goal 6, which focus on household- rather than individual-level access. OBJECTIVE: To evaluate access to safe water, sanitation and hygiene (WASH), at the individual- and household-level, amongst people with disabilities in the Tanahun district of Nepal. METHODS: A population-based survey of disability was conducted from August-October 2016 to evaluate access to improved water and sanitation facilities between households with members with disabilities (n = 198) and those without (n = 1,265) in the Tanahun district of Nepal. A nested case-control then compared individual-level access between cases aged 15 and above with disabilities (n = 192) and age-sex-location matched controls without disabilities (n = 189), using the newly developed 21-item "Quality of WASH Access" questionnaire. Multivariate regression was used to compare household- and individual-level indicators between people and households with and without disabilities. In-depth interviews with 18 people with disabilities and their caregivers was conducted to assess the acceptability and appropriateness of the "Quality of WASH Access" questionnaire. FINDINGS: There were no significant differences between households with and without members with disabilities in access to an improved sanitation facility or water source. However, at the individual-level, people with disabilities experienced significantly greater difficulties accessing water, sanitation and hygiene compared to people without disabilities (p<0.001 for all three scores). Amongst people with disabilities, water difficulty scores were associated with having a physical impairment and greater disability severity; sanitation difficulty scores were associated with lower socioeconomic status and physical or self-care limitations; and hygiene difficulty scores were positively associated with self-care limitations and lower socioeconomic status, and inversely associated with hearing impairments. Qualitative research found the "Quality of WASH Access" questionnaire was well understood by participants and captured many of the challenges they faced. Additional challenges not covered by the tool included: (1) time spent on WASH, (2) consistency of access, (3) sufficiency of access, and (4) dignity of access. CONCLUSION: People with disabilities face substantial challenges to meeting their WASH needs, particularly in using services autonomously, consistently, hygienically, with dignity and privacy, and without pain or fear of abuse. These challenges are not captured through household-level data, and so individual-level WASH access are needed to monitor progress towards universal WASH access. The Quality of WASH Access questionnaire may provide a useful data collection tool

Does disability increase the risk of poverty ‘in all its forms’? Comparing monetary and multidimensional poverty in Vietnam and Nepal

To meet the Sustainable Development Goals target of ending poverty “in all its forms”, it is critical to monitor progress towards poverty alleviation, including amongst people with disabilities. This research used data from a population-based nested case control studies (n=667) and compares monetary and multidimensional poverty levels amongst people with and without disabilities in the districts of Cam Le, Vietnam and Tanahun, Nepal. Overall, there were no significant differences in incidence of monetary poverty between people with and without disabilities. However, approximately half of people with disabilities were multidimensionally poor in both settings, twice as frequent as compared to people without disabilities. Amongst people with disabilities, multidimensional poverty was associated with having a functional limitation affecting cognition and self-care, disability severity and younger age. The high incidence of multidimensional poverty amongst people with disabilities even in the absence of monetary poverty indicates a need for social protection and other interventions

Disability among Older People: Analysis of Data from Disability Surveys in Six Low- and Middle-Income Countries

This analysis of surveys from six low- and middle-income countries (LMICs) aimed to (i) estimate the prevalence of disability among older adults and (ii) compare experiences and participation in key life areas among older people with and without disabilities which may show vulnerability during the COVID-19 pandemic. Data were analysed from district-level or national surveys in Cameroon, Guatemala, Haiti, India, Nepal and the Maldives, which across the six databases totalled 3499 participants aged 60 years and above including 691 people with disabilities. Disability was common among adults 60+, ranging from 9.7% (8.0–11.8) in Nepal to 39.2% in India (95% CI 34.1–44.5%). Mobility was the most commonly reported functional difficulty. In each setting, older people with disabilities were significantly less likely to be working and reported greater participation restrictions and environmental barriers in key life areas compared to people in the same age categories without disabilities (p < 0.05). Disability is common in this population, and older people with disabilities may have greater difficulties participating in COVID-19 responses and have high economic vulnerabilities. It is imperative to prioritise the needs of older people with disabilities in the COVID-19 pandemic, including ensuring accessibility of both health services and the community in genera

Interrogating and Reflecting on Disability Prevalence Data Collected Using the Washington Group Tools: Results from Population-Based Surveys in Cameroon, Guatemala, India, Maldives, Nepal, Turkey and Vanuatu.

The Washington Group (WG) tools capture self-reported functional limitations, ranging from 6 domains in the Short Set (SS) to 11 in the Extended Set (ESF). Prevalence estimates can vary considerably on account of differences between modules and the different applications of them. We compare prevalence estimates by WG module, threshold, application and domain to explore these nuances and consider whether alternative combinations of questions may be valuable in reduced sets. We conducted secondary analyses of seven population-based surveys (analyses restricted to adults 18+) in Low- and Middle-Income Countries that used the WG tools. The prevalence estimates using the SS standard threshold (a lot of difficulty or higher in one or more domain) varied between 3.2% (95% Confidence Interval 2.9-3.6) in Vanuatu to 14.1% (12.2-16.2) in Turkey. The prevalence was higher using the ESF than the SS, and much higher (5 to 10-fold) using a wider threshold of "some" or greater difficulty. Two of the SS domains (communication, self-care) identified few additional individuals with functional limitations. An alternative SS replacing these domains with the psychosocial domains of anxiety and depression would identify more participants with functional limitations for the same number of items. The WG tools are valuable for collecting harmonised population data on disability. It is important that the impact on prevalence of use of different modules, thresholds and applications is recognised. An alternative SS may capture a greater proportion of people with functional domains without increasing the number of items

Diagnostic Test Accuracy of a 2-Transcript Host RNA Signature for Discriminating Bacterial vs Viral Infection in Febrile Children.

IMPORTANCE: Because clinical features do not reliably distinguish bacterial from viral infection, many children worldwide receive unnecessary antibiotic treatment, while bacterial infection is missed in others. OBJECTIVE: To identify a blood RNA expression signature that distinguishes bacterial from viral infection in febrile children. DESIGN, SETTING, AND PARTICIPANTS: Febrile children presenting to participating hospitals in the United Kingdom, Spain, the Netherlands, and the United States between 2009-2013 were prospectively recruited, comprising a discovery group and validation group. Each group was classified after microbiological investigation as having definite bacterial infection, definite viral infection, or indeterminate infection. RNA expression signatures distinguishing definite bacterial from viral infection were identified in the discovery group and diagnostic performance assessed in the validation group. Additional validation was undertaken in separate studies of children with meningococcal disease (n = 24) and inflammatory diseases (n = 48) and on published gene expression datasets. EXPOSURES: A 2-transcript RNA expression signature distinguishing bacterial infection from viral infection was evaluated against clinical and microbiological diagnosis. MAIN OUTCOMES AND MEASURES: Definite bacterial and viral infection was confirmed by culture or molecular detection of the pathogens. Performance of the RNA signature was evaluated in the definite bacterial and viral group and in the indeterminate infection group. RESULTS: The discovery group of 240 children (median age, 19 months; 62% male) included 52 with definite bacterial infection, of whom 36 (69%) required intensive care, and 92 with definite viral infection, of whom 32 (35%) required intensive care. Ninety-six children had indeterminate infection. Analysis of RNA expression data identified a 38-transcript signature distinguishing bacterial from viral infection. A smaller (2-transcript) signature (FAM89A and IFI44L) was identified by removing highly correlated transcripts. When this 2-transcript signature was implemented as a disease risk score in the validation group (130 children, with 23 definite bacterial, 28 definite viral, and 79 indeterminate infections; median age, 17 months; 57% male), all 23 patients with microbiologically confirmed definite bacterial infection were classified as bacterial (sensitivity, 100% [95% CI, 100%-100%]) and 27 of 28 patients with definite viral infection were classified as viral (specificity, 96.4% [95% CI, 89.3%-100%]). When applied to additional validation datasets from patients with meningococcal and inflammatory diseases, bacterial infection was identified with a sensitivity of 91.7% (95% CI, 79.2%-100%) and 90.0% (95% CI, 70.0%-100%), respectively, and with specificity of 96.0% (95% CI, 88.0%-100%) and 95.8% (95% CI, 89.6%-100%). Of the children in the indeterminate groups, 46.3% (63/136) were classified as having bacterial infection, although 94.9% (129/136) received antibiotic treatment. CONCLUSIONS AND RELEVANCE: This study provides preliminary data regarding test accuracy of a 2-transcript host RNA signature discriminating bacterial from viral infection in febrile children. Further studies are needed in diverse groups of patients to assess accuracy and clinical utility of this test in different clinical settings

Identification of novel locus associated with coronary artery aneurysms and validation of loci for susceptibility to Kawasaki disease

Kawasaki disease (KD) is a paediatric vasculitis associated with coronary artery aneurysms (CAA). Genetic variants influencing susceptibility to KD have been previously identified, but no risk alleles have been validated that influence CAA formation. We conducted a genome-wide association study (GWAS) for CAA in KD patients of European descent with 200 cases and 276 controls. A second GWAS for susceptibility pooled KD cases with healthy paediatric controls from vaccine trials in the UK (n = 1609). Logistic regression mixed models were used for both GWASs. The susceptibility GWAS was meta-analysed with 400 KD cases and 6101 controls from a previous European GWAS, these results were further meta-analysed with Japanese GWASs at two putative loci. The CAA GWAS identified an intergenic region of chromosome 20q13 with multiple SNVs showing genome-wide significance. The risk allele of the most associated SNV (rs6017006) was present in 13% of cases and 4% of controls; in East Asian 1000 Genomes data, the allele was absent or rare. Susceptibility GWAS with meta-analysis with previously published European data identified two previously associated loci (ITPKC and FCGR2A). Further meta-analysis with Japanese GWAS summary data from the CASP3 and FAM167A genomic regions validated these loci in Europeans showing consistent effects of the top SNVs in both populations. We identified a novel locus for CAA in KD patients of European descent. The results suggest that different genes determine susceptibility to KD and development of CAA and future work should focus on the function of the intergenic region on chromosome 20q13

Diagnosis of Kawasaki Disease Using a Minimal Whole-Blood Gene Expression Signature.

Importance: To date, there is no diagnostic test for Kawasaki disease (KD). Diagnosis is based on clinical features shared with other febrile conditions, frequently resulting in delayed or missed treatment and an increased risk of coronary artery aneurysms. Objective: To identify a whole-blood gene expression signature that distinguishes children with KD in the first week of illness from other febrile conditions. Design, Setting, and Participants: The case-control study comprised a discovery group that included a training and test set and a validation group of children with KD or comparator febrile illness. The setting was pediatric centers in the United Kingdom, Spain, the Netherlands, and the United States. The training and test discovery group comprised 404 children with infectious and inflammatory conditions (78 KD, 84 other inflammatory diseases, and 242 bacterial or viral infections) and 55 healthy controls. The independent validation group comprised 102 patients with KD, including 72 in the first 7 days of illness, and 130 febrile controls. The study dates were March 1, 2009, to November 14, 2013, and data analysis took place from January 1, 2015, to December 31, 2017. Main Outcomes and Measures: Whole-blood gene expression was evaluated using microarrays, and minimal transcript sets distinguishing KD were identified using a novel variable selection method (parallel regularized regression model search). The ability of transcript signatures (implemented as disease risk scores) to discriminate KD cases from controls was assessed by area under the curve (AUC), sensitivity, and specificity at the optimal cut point according to the Youden index. Results: Among 404 patients in the discovery set, there were 78 with KD (median age, 27 months; 55.1% male) and 326 febrile controls (median age, 37 months; 56.4% male). Among 202 patients in the validation set, there were 72 with KD (median age, 34 months; 62.5% male) and 130 febrile controls (median age, 17 months; 56.9% male). A 13-transcript signature identified in the discovery training set distinguished KD from other infectious and inflammatory conditions in the discovery test set, with AUC of 96.2% (95% CI, 92.5%-99.9%), sensitivity of 81.7% (95% CI, 60.0%-94.8%), and specificity of 92.1% (95% CI, 84.0%-97.0%). In the validation set, the signature distinguished KD from febrile controls, with AUC of 94.6% (95% CI, 91.3%-98.0%), sensitivity of 85.9% (95% CI, 76.8%-92.6%), and specificity of 89.1% (95% CI, 83.0%-93.7%). The signature was applied to clinically defined categories of definite, highly probable, and possible KD, resulting in AUCs of 98.1% (95% CI, 94.5%-100%), 96.3% (95% CI, 93.3%-99.4%), and 70.0% (95% CI, 53.4%-86.6%), respectively, mirroring certainty of clinical diagnosis. Conclusions and Relevance: In this study, a 13-transcript blood gene expression signature distinguished KD from other febrile conditions. Diagnostic accuracy increased with certainty of clinical diagnosis. A test incorporating the 13-transcript disease risk score may enable earlier diagnosis and treatment of KD and reduce inappropriate treatment in those with other diagnoses

Identification of novel locus associated with coronary artery aneurysms and validation of loci for susceptibility to Kawasaki disease.

Kawasaki disease (KD) is a paediatric vasculitis associated with coronary artery aneurysms (CAA). Genetic variants influencing susceptibility to KD have been previously identified, but no risk alleles have been validated that influence CAA formation. We conducted a genome-wide association study (GWAS) for CAA in KD patients of European descent with 200 cases and 276 controls. A second GWAS for susceptibility pooled KD cases with healthy paediatric controls from vaccine trials in the UK (n = 1609). Logistic regression mixed models were used for both GWASs. The susceptibility GWAS was meta-analysed with 400 KD cases and 6101 controls from a previous European GWAS, these results were further meta-analysed with Japanese GWASs at two putative loci. The CAA GWAS identified an intergenic region of chromosome 20q13 with multiple SNVs showing genome-wide significance. The risk allele of the most associated SNV (rs6017006) was present in 13% of cases and 4% of controls; in East Asian 1000 Genomes data, the allele was absent or rare. Susceptibility GWAS with meta-analysis with previously published European data identified two previously associated loci (ITPKC and FCGR2A). Further meta-analysis with Japanese GWAS summary data from the CASP3 and FAM167A genomic regions validated these loci in Europeans showing consistent effects of the top SNVs in both populations. We identified a novel locus for CAA in KD patients of European descent. The results suggest that different genes determine susceptibility to KD and development of CAA and future work should focus on the function of the intergenic region on chromosome 20q13

Access to Social Protection Among People with Disabilities:Mixed Methods Research from Tanahun, Nepal

While people with disabilities are often targeted as key beneficiaries of social protection, there is little evidence available on their participation in existing programmes. This study uses mixed methods to explore access to disability-targeted and non-targeted social protection programmes in Nepal, particularly the District of Tanahun. In total, 31% of people with disabilities had Disability Identification Cards, which entitles them to a range of different social protection benefits depending on the card level, including disability-targeted social assistance (received by 13% of people with disabilities). Overall, 37% of people with disabilities received social assistance, which was higher than for people without disabilities (21%). The most commonly accessed form of social assistance was the Old Age Allowance, which had universally high coverage amongst both people with and without disabilities. Uptake of disability-targeted social protection entitlements other than social assistance (e.g. scholarships, discounted transportation and health services) was generally low. Factors impacting upon access included the geographic and financial accessibility of the application process, procedures for determining eligibility and compliance of service providers