A Flexible, Pooled CRISPR Library for Drug Development Screens.

Functional genomic screening with CRISPR has provided a powerful and precise new way to interrogate the phenotypic consequences of gene manipulation in high-throughput, unbiased analyses. However, some experimental paradigms prove especially challenging and require carefully and appropriately adapted screening approaches. In particular, negative selection (or sensitivity) screening, often the most experimentally desirable modality of screening, has remained a challenge in drug discovery. Here we assess whether our new, modular genome-wide pooled CRISPR library can improve negative selection CRISPR screening and add utility throughout the drug development pipeline. Our pooled library is split into three parts, allowing it to be scaled to accommodate the experimental challenges encountered during drug development, such as target identification using unlimited cell numbers compared with target identification studies for cell populations where cell numbers are limiting. To test our new library, we chose to look for drug-gene interactions using a well-described small molecule inhibitor targeting poly(ADP-ribose) polymerase 1 (PARP1), and in particular to identify genes which sensitise cells to this drug. We simulate hit identification and performance using each library partition and support these findings through orthogonal drug combination cell panel screening. We also compare our data with a recently published CRISPR sensitivity dataset obtained using the same PARP1 inhibitor. Overall, our data indicate that generating a comprehensive CRISPR knockout screening library where the number of guides can be scaled to suit the biological question being addressed allows a library to have multiple uses throughout the drug development pipeline, and that initial validation of hits can be achieved through high-throughput cell panels screens where clinical grade chemical or biological matter exist.

Brain complications of diabetes mellitus: a cross-sectional study of awareness among individuals with diabetes and the general population in Ireland.

AIMS: To identify awareness of potential brain complications of diabetes among individuals with diabetes and the public. METHODS: For this observational, cross-sectional survey study, we recruited consecutive adult attendees of a specialist diabetes clinic and two primary care practices. Primary care attendees represented members of the general population of Ireland. An interviewer-administered questionnaire was used to gather data on respondents' awareness of brain complications of diabetes and modifiable risk factors for dementia. Multivariable logistic regression was undertaken to identify variables independently associated with awareness. RESULTS: Respondents included a total of 502 adults: 250 in the diabetes group (37% women, mean age 63 ± 14 years, 88% with Type 2 diabetes) and 252 in the general population group (51% women, mean age 47 ± 17 years, 7% with Type 2 diabetes). The diabetes group had significantly greater awareness of diabetes complications, except for depression, compared with the general population group. In the group as a whole, respondent awareness of dementia (35%) and memory problems (47%) as potential complications of diabetes was poor compared with awareness of kidney (84%) and eye damage (84%). Respondents were 1.5 times more likely to identify that individuals can modify their risk of developing Type 2 diabetes than their risk of dementia. CONCLUSIONS: This study shows that there is poor awareness of brain complications of diabetes among individuals with diabetes and the general population in Ireland. The results suggest a need for expansion of public awareness campaigns and diabetes education programmes to promote awareness of the brain complications of diabetes and of the modifiable risk factors for dementia, as part of a life-course approach to dementia prevention.

Bikini versus traditional incision direct anterior approach: is there any difference in soft tissue damage?

INTRODUCTION: The direct anterior approach for total hip replacements has reported advantages of improved early function and muscle preservation. In an effort to improve healing and cosmesis, a change in the orientation of the incision has been proposed. Traditionally, the skin incision is in-line with the tensor fasciae latae muscle belly. The bikini incision is orthogonal to this orientation. The hypothesis was that muscle damage would be increased by using the bikini incision. METHODS: A traditional or bikini incision was performed on 18 cadaveric hips. On each of the 9 specimens, the traditional incision was performed on 1 side, and a bikini incision on the contralateral hip, with an even distribution of right or left side. Blinded anatomists performed the hip dissections, and assessed for muscle damage as well as for damage to the lateral femoral cutaneous nerve. RESULTS: No difference in muscle damage was identified in the tensor fasciae latae between muscle groups. Muscle damage was very minimal to the gluteus medius and minimus. Damage to the lateral femoral cutaneous nerve occurred equally for both the bikini and traditional skin incisions. CONCLUSIONS: The bikini incision for the direct anterior approach to the hip can be performed safely, with no increase in muscle damage or damage to the lateral femoral cutaneous nerve compared to the traditional incision.

Self-Harm, Methadone Use and Drug-Related Deaths amongst Those Registered As Being of No Fixed Abode or Homeless in Ireland.

This work aims to contribute to the evidence base regarding the health of those who experience homelessness in Ireland by collating data on methadone use, drug-related deaths and emergency department presentations due to self-harm. Data from the Central Methadone Treatment List (CTL), National Self-Harm Registry Ireland and the National Drug-Related Deaths Index were analysed. The percentage on the CTL registered as being of no fixed abode (NFA) or homeless increased from 2% to 7% from 2011-2014. The absolute number of presentations with deliberate self-harm from those of NFA increased by 49% from 2007-2014. The number of drug-related deaths amongst those of NFA or homeless and who died in Dublin fluctuated from 2004-13 with an overall upward trend. There is an urgent need to adequately resource and coordinate those services which aim to address factors (social and health inequalities, mental ill-health and addiction) which lead people into - and prevent them exiting from - homelessness.

Ebola, Zika and the International Health Regulations - implications for Port Health Preparedness.

BACKGROUND: The outbreak of Ebola Virus Disease in West Africa in 2014-2015 was unprecedented in terms of its scale and consequence.  This, together with the emergence of Zika virus as a Public Health Emergency of International Concern in 2016, has again highlighted the potential for disease to spread across international borders and provided an impetus for countries to review their Port Health preparedness. This report reviews the legislative framework and actions taken under this framework in advancing and improving Port Health preparedness in Ireland, in response to the declaration of the Public Health Emergency of International Concern for Ebola Virus Disease in August 2014. FINDINGS: Infectious disease Shipping and Aircraft Regulations were brought into force in Ireland in 2008 and 2009, respectively. Preparatory actions taken under these and the International Health Regulations necessitated significant levels of cross disciplinary working with other organisations, both within and beyond traditional healthcare settings. Information packs on Ebola Virus Disease were prepared and distributed to airports, airlines, port authorities and shipping agents, and practical exercises were held at relevant sites. Agreements were put in place for contact tracing of passenger and crew on affected conveyances and protocols were established for the management of Medical Declarations of Health from ships coming from West Africa. CONCLUSIONS: The outbreak of Ebola Virus Disease in West Africa resulted in significant strengthening of Ireland's Port Health preparedness, while also highlighting the extent to which preparedness requires ongoing and sustained commitment from all stakeholders, both nationally and internationally, in ensuring that countries are ready when the next threat presents at their borders.

Comparative cardiovascular safety of glucagon-like peptide-1 receptor agonists versus other antidiabetic drugs in routine care: a cohort study.

AIMS: To evaluate the comparative cardiovascular disease (CVD) safety of glucagon-like peptide-1 receptor agonists (GLP-1 RAs) in head-to-head comparisons with dipeptidyl peptidase-4 (DPP-4) inhibitors, sulphonylureas or insulin, when added to metformin, as used in 'real-world' patients with type 2 diabetes mellitus (T2DM). METHODS: Within a large US commercial health plan database linked to laboratory test results, we identified three pairwise 1 : 1 propensity-score-matched cohorts of patients with T2DM aged ≥18 years treated with metformin who initiated a GLP-1 RA or a comparator, i.e. DPP-4 inhibitor (n = 35 534), second-generation sulphonylureas (n = 28 138) or insulin (n = 47 068), between 2005 and 2013. We examined the association between drug initiation and a composite CVD endpoint, comprising hospitalizations for acute myocardial infarction, unstable angina, stroke or coronary revascularization. RESULTS: During the course of 1 year, there were 13.9 and 13.7 CVD events per 1000 person-years among propensity-score-matched initiators of GLP-1 RAs versus DPP-4 inhibitors [hazard ratio (HR) 1.02; 95% confidence interval (CI) 0.84-1.24]; and 12.1 versus 14.0 events among initiators of GLP-1 RAs versus sulphonylureas (HR 0.86; 95% CI 0.69-1.08). The effect estimates for GLP-1 RAs versus insulin were sensitive to the adjustment for glycated haemoglobin, after which the HR was 1.01 (95% CI 0.73-1.41). Results were robust across several sensitivity analyses, including an as-treated analysis considering up to 8.7 years of follow-up. CONCLUSIONS: This large study, performing head-to-head comparisons of GLP-1 RAs with other antidiabetic agents in real-world patients, provides estimates of relative safety precise enough to exclude large differences in CVD risk and adds further understanding to results from recent clinical trials.

A cost analysis of single-stage bilateral versus two-stage direct anterior total hip arthroplasty.

PURPOSE: The potential cost savings of single-stage bilateral total hip arthroplasty (THA) are unclear, and the risks associated with it are not well defined. We sought to compare the costs and perioperative complications of single-stage bilateral THA via the direct anterior approach (DAA) to a two-stage bilateral protocol. METHODS: We retrospectively reviewed patients who underwent a single- stage bilateral DAA THA and compared them to a two-stage THA group. We conducted a cost analysis from both the hospital perspective and the Ministry of Health (MOH) perspective. RESULTS: 24 patients were included in this study. The 2 groups were similar in age (58.9 vs 63.9 yrs), height (169.2 vs 170.9 cm), weight (80.2 vs 78.6 kg), BMI (27.9 vs 26.3 kg/m2), ASA score (2.2 vs 2.2), and CCI score (2.3 vs 2.9). The mean cost per patient from the hospital perspective for the single-stage group was $10,728.13 (SD = 621.46) compared to $12,670.63 (SD = 519.72) for the two-stage group (Mean Difference = $1,942.50, 95% CI = $1,457.49 to $2,427.51, p<0.001). Similarly, from the MOH perspective, the cost for the single-stage group was $12,552.34 (SD = 644.93) compared to $14,740.58 (SD = 598.07) for the two-stage group (Mean Difference = $2,188.24, 95% CI = $1,661.67 to $2,714.81, p<0.001). There were no significant differences in complication rate between groups. The largest percent of total cost savings from a hospital perspective was attributed to cost of operating room staff and OR set-up (55%). CONCLUSIONS: Our results suggest that single-stage bilateral DAA THA results in significant cost savings compared to two-stage DAA THA.

Is grand-parental smoking associated with adolescent obesity? A three-generational study.

BACKGROUND/OBJECTIVES: Data from previous studies consistently suggest that maternal smoking is positively associated with obesity later in life. Whether this association persists across generations is unknown. We examined whether grand-parental smoking was positively associated with overweight status in adolescence. SUBJECT/METHODS: Participants were grandmother-mother-child triads in the Nurses' Health Study II (NHS II), the Nurses Mothers' Cohort Study and the Growing up Today Study (GUTS). Grandmothers provided information on their and their partner's smoking during pregnancy with the child's mother. Information on child's weight and height at ages 12 (N=3094) and 17 (N=3433) was obtained from annual or biennial GUTS questionnaires. We used logistic regression to estimate the odds ratios (ORs) of being overweight or obese, relative to normal weight. RESULTS: Grand-maternal smoking during pregnancy was not associated with overweight status in adolescence. After adjusting for covariates, the OR of being overweight or obese relative to normal weight at age 12 years in girls whose grandmothers smoked 15+ cigarettes daily during pregnancy was 1.21 (95% confidence interval (CI) 0.74-1.98; P(trend)=0.31) and 1.07 (0.65-1.77; P(trend)=0.41) in boys. Grand-paternal smoking during pregnancy was associated with being overweight or obese at age 12 in girls only, but not at age 17 for either sex: the OR for being overweight or obese at age 12 was 1.38 (95% CI 1.01-1.89; P(trend)=0.03) in girls and 1.31 (95% CI 0.97-1.76; P(trend)=0.07) in boys. Among children of non-smoking mothers, the OR for granddaughter obesity for grand-paternal smoking was attenuated and no longer significant (OR 1.28 (95% CI 0.87-1.89; P(trend)=0.18)). CONCLUSIONS: Our findings suggest that the association between maternal smoking and offspring obesity may not persist beyond the first generation. However, grand-paternal smoking may affect the overweight status of the granddaughter, likely through the association between grand-paternal smoking and maternal smoking.

Propensity scores for confounder adjustment when assessing the effects of medical interventions using nonexperimental study designs.

Treatment effects, especially when comparing two or more therapeutic alternatives as in comparative effectiveness research, are likely to be heterogeneous across age, gender, co-morbidities and co-medications. Propensity scores (PSs), an alternative to multivariable outcome models to control for measured confounding, have specific advantages in the presence of heterogeneous treatment effects. Implementing PSs using matching or weighting allows us to estimate different overall treatment effects in differently defined populations. Heterogeneous treatment effects can also be due to unmeasured confounding concentrated in those treated contrary to prediction. Sensitivity analyses based on PSs can help to assess such unmeasured confounding. PSs should be considered a primary or secondary analytic strategy in nonexperimental medical research, including pharmacoepidemiology and nonexperimental comparative effectiveness research.

Siblings caring for and about pediatric palliative care patients.

BACKGROUND: The experiences of young people who have siblings with life-limiting illnesses are not well understood. AIM: The study proposed to identify the concerns of siblings of pediatric palliative care (PPC) patients. DESIGN AND MEASUREMENT: Semistructured interviews were administered to participants and analyzed using qualitative inductive thematic analysis. SETTING AND PARTICIPANTS: Study subjects were 18 siblings of PPC patients aged 9 to 22 living in the Auckland area. RESULTS: The siblings of PPC patients held concerns about their siblings' impending death and desires to be involved in their lives and care. CONCLUSIONS: Siblings may benefit from opportunities to be involved in conversations about mortality and the care of their ill sibling. They are able to express their concerns and help provide care to PPC patients.

Patterns of internet use by parents of children attending a pediatric surgical service.

PURPOSE: The internet has revolutionised the way we search for information. We determined the level of internet use by parents of children attending general surgical services and identified trends in online information-seeking behaviour. METHODS: A questionnaire based on the work by Boston and Tassone was distributed to parents attending both the day surgical units and surgical outpatients department in a paediatric tertiary referral centre. RESULTS: There were 214 (82.3 %) questionnaires returned, with 82 (38.3 %) of respondents having searched the internet regarding their child's surgical issue. Access to a smartphone, a university education and private health insurance were factors that positively influenced online searching (p < 0.005). Of those respondents who searched the internet, 42 (51 %) felt that information they found online was understandable, while only 14 (17 %) admitted to online sourced information influencing the treatment decisions they had made for their children. When asked to rank information sources on Likert-type scales in terms of importance; parents ranked the surgeon as most important (mean = 4.73), whilst the internet ranked lowest (mean = 3.02). CONCLUSION: We demonstrated significant use of the internet amongst those attending paediatric general surgical services. Clinician sourced information remains important, however we should engage with patients to utilise this vast resource effectively.

Long-term outcomes for Kienböck's disease.

BACKGROUND: The precise etiology of Kienböck's disease is unclear. Controversy exists regarding the appropriate treatment modality. The present study sought to investigate and compare surgical and nonsurgical treatment outcomes of patients suffering from Kienböck's disease in the province of Newfoundland and Labrador (NL), Canada. METHODS: The present study was a retrospective analysis of 66 patients. The primary outcome was the Disabilities of the Arm, Shoulder, and Hand (DASH) score. Student's t test was used to assess differences in outcomes between treatment groups. One-way ANOVA was used to assess differences in primary outcome in time since first assessed in an effort to examine progression over time. Pearson correlation was used to assess for correlation between primary outcome and age at diagnosis. RESULTS: The average age was 38.6 ± 11.4 (18-70) years; Four patients were excluded due to inaccessible imaging. Of the remaining patients, 44 were treated conservatively, while 18 were treated surgically. The DASH scores for the surgical group were 23.7 ± 24.5 (0.9-82.8) and nonsurgical group were 20.0 ± 20.1 (1.7-81). As expected, the surgical group was mainly comprised of late-stage Kienböck's. When both groups were compared, there was no significant difference in the DASH scores. There were no difference in DASH scores within groups according to time since first diagnosed (<5 years; between 5 and 10 years; and >10 years). A positive correlation was found between age at diagnosis and DASH score (r = 0.42, p = 0.007), despite treatment modality. This finding remained significant after accounting for confounding factors (p = 0.029). CONCLUSION: The DASH score for the surgical group was 23.7 ± 24.5 (0.9-82.8) and nonsurgical group was 20.0 ± 20.1 (1.7-81). No significant difference in DASH scores was found between surgically and nonsurgically treated patients. A positive association was found between the age at diagnosis of Kienböck's and DASH score, which suggests that patients diagnosed and treated later in life tend not to do as well.

Caregivers' estimations of their children's perceptions of death as a biological concept.

Communication about death is often a sensitive topic in families with children. The present studied compared answers of 141 school children aged 5-7 to questions about death, and their caregivers' predictions. Children were interviewed, and caregivers answered on paper, questions on inevitability, applicability, irreversibility, cessation, causation, and personal mortality. For causation, cessation, and irreversibility, children were significantly more correct than caregivers expected, and girls were more correct on applicability. Communicating with children about death may not always be as caregivers expect.

Division of tongue tie: review of practice through a tertiary paediatric otorhinolaryngology service.

OBJECTIVES: Recent NICE guidance declared that evidence regarding surgical division of tongue tie was adequate to support the use of the procedure, provided that normal arrangements are in place for consent, audit and clinical governance. This work aimed to carry out a retrospective review of those patients who have previously undergone tongue tie division through a tertiary paediatric otorhinolaryngology service. We further aimed to identify the referral patterns, indications for, and outcomes following, division. METHODS: A retrospective chart review of all patients attending for division of tongue-tie between January 2010 and June 2011, inclusive. RESULTS: A total of 48 patients underwent division of tongue-tie over the study period. The median age of patients at the time of operation was 16 months (range 3-192). Delayed speech or articulation problems (27%), and parental concern (23%) were the most common reasons for referral. Overt difficulty with feeding was responsible for referral in just 4 cases. All cases were performed under general anaesthetic as day-case procedures; 7 patients had a second procedure performed at the same time. No complications, either early or late, were recorded across this cohort. CONCLUSIONS: Tongue-tie division is a safe procedure with minimal associated morbidity. Whilst the primary indication cited in the literature is that of feeding difficulty, it appears that the majority of division is performed for other reasons.

Active safety monitoring of newly marketed medications in a distributed data network: application of a semi-automated monitoring system.

We developed a semi-automated active monitoring system that uses sequential matched-cohort analyses to assess drug safety across a distributed network of longitudinal electronic health-care data. In a retrospective analysis, we show that the system would have identified cerivastatin-induced rhabdomyolysis. In this study, we evaluated whether the system would generate alerts for three drug-outcome pairs: rosuvastatin and rhabdomyolysis (known null association), rosuvastatin and diabetes mellitus, and telithromycin and hepatotoxicity (two examples for which alerting would be questionable). Over >5 years of monitoring, rate differences (RDs) in comparisons of rosuvastatin with atorvastatin were -0.1 cases of rhabdomyolysis per 1,000 person-years (95% confidence interval (CI): -0.4, 0.1) and -2.2 diabetes cases per 1,000 person-years (95% CI: -6.0, 1.6). The RD for hepatotoxicity comparing telithromycin with azithromycin was 0.3 cases per 1,000 person-years (95% CI: -0.5, 1.0). In a setting in which false positivity is a major concern, the system did not generate alerts for the three drug-outcome pairs.

Comparison of associations of adherence to a Dietary Approaches to Stop Hypertension (DASH)-style diet with risks of cardiovascular disease and venous thromboembolism.

BACKGROUND: Venous thromboembolism (VTE) and cardiovascular disease (CVD) share some risk factors, including obesity, but it is unclear how dietary patterns associated with reduced risk of CVD relate to risk of VTE. OBJECTIVE: To compare the relationships of adherence to a Dietary Approaches to Stop Hypertension (DASH)-style diet with the risks of CVD and VTE. PATIENTS/METHODS: We confirmed by medical record review 1094 incident cases of CVD and 675 incident VTEs during a mean follow-up of 14.6 years in 34 827 initially healthy participants in the Women's Health Study who completed at baseline a 133-item food frequency questionnaire scored for adherence to a DASH diet. We compared estimated associations of dietary patterns with CVD and VTE from proportional hazards models in a competing risk framework. RESULTS: Initial analyses adjusted for age, energy intake and randomized treatments showed 36-41% reduced hazards of CVD among women in the top two quintiles of DASH score relative to those in the bottom quintile (P(trend) < 0.001). In multivariate analysis, women in the top two quintiles had 12-23% reduced hazards of CVD relative to women in the bottom quintile (P(trend) = 0.04). Analyses restricted to coronary events showed more variable 10-33% reduced hazards in the top two quintiles (P(trend) = 0.09). In contrast, higher DASH scores were unrelated to risk of VTE, with a 1% reduced hazard for the top vs. bottom quintile (P(trend) = 0.95). CONCLUSION: An apparently strong association of adherence to the DASH diet with incidence of CVD was attenuated upon control for confounding variables. Adherence to the DASH diet was not associated with risk of VTE in women.

Assessing the comparative effectiveness of newly marketed medications: methodological challenges and implications for drug development.

Comparative-effectiveness research (CER) aims to produce actionable evidence regarding the effectiveness and safety of medical products and interventions as they are used outside of controlled research settings. Although CER evidence regarding medications is particularly needed shortly after market approval, key methodological challenges include (i) potential bias due to channeling of patients to the newly marketed medication because of various patient-, physician-, and system-related factors; (ii) rapid changes in the characteristics of the user population during the early phase of marketing; and (iii) lack of timely data and the often small number of users in the first few months of marketing. We propose a mix of approaches to generate comparative-effectiveness data in the early marketing period, including sequential cohort monitoring with secondary health-care data and propensity score (PS) balancing, as well as extended follow-up of phase III and phase IV trials, indirect comparisons of placebo-controlled trials, and modeling and simulation of virtual trials.