PAM-flexible Engineered FnCas9 variants for robust and ultra-precise genome editing and diagnostics.

The clinical success of CRISPR therapies hinges on the safety and efficacy of Cas proteins. The Cas9 from Francisella novicida (FnCas9) is highly precise, with a negligible affinity for mismatched substrates, but its low cellular targeting efficiency limits therapeutic use. Here, we rationally engineer the protein to develop enhanced FnCas9 (enFnCas9) variants and broaden their accessibility across human genomic sites by ~3.5-fold. The enFnCas9 proteins with single mismatch specificity expanded the target range of FnCas9-based CRISPR diagnostics to detect the pathogenic DNA signatures. They outperform Streptococcus pyogenes Cas9 (SpCas9) and its engineered derivatives in on-target editing efficiency, knock-in rates, and off-target specificity. enFnCas9 can be combined with extended gRNAs for robust base editing at sites which are inaccessible to PAM-constrained canonical base editors. Finally, we demonstrate an RPE65 mutation correction in a Leber congenital amaurosis 2 (LCA2) patient-specific iPSC line using enFnCas9 adenine base editor, highlighting its therapeutic utility.

Depression predicts emergency care use in people with chronic obstructive pulmonary disease: a large cohort study in primary care.

Background: Depression is common in people with chronic obstructive pulmonary disease (COPD) and has been associated with a variety of poor outcomes. A large proportion of health care costs in the UK are spent on emergency care. This study examined the prospective relationship between depression and use of emergency care in patients with COPD managed in primary care. Methods: This was a twelve-month, prospective longitudinal study of 355 patients with COPD in six primary care practices in the UK. Baseline measures included demographic characteristics, depression and anxiety, severity of COPD, presence or absence of other chronic diseases, and prior use of emergency care. Outcome measures were (a) number of emergency department (ED) visits; or (b) an emergency hospital admission in the follow-up year. Results: Older age, number of comorbid physical health conditions, severity of COPD, prior use of emergency care, and depression were all independently associated with both ED attendance and an emergency hospital admission in the follow-up year. Subthreshold depression (HADS depression score 4-7) was associated with a 2.8 times increased odds of emergency hospital admission, and HADS depression >8 was associated with 4.8 times increased odds. Conclusion: Depression is a predictor of emergency care in COPD, independent of severity of disease or physical comorbidity. Even mild (subthreshold) symptoms of depression more than double the risk of using emergency care, suggesting there is a strong case to develop and deploy integrated preventive strategies in primary care that can promote mental health in people with COPD.

Acculturation and the prevalence of pain amongst South Asian minority ethnic groups in the UK.

BACKGROUND: Musculoskeletal pain is reported more commonly by South Asians in the UK than by white Europeans. This may result from a variety of factors, including cultural differences, and thus we investigated the extent to which differences in the prevalence of pain within the South Asian population could be explained by differences in acculturation (the extent to which immigrants take on the culture of their host population). METHODS: Nine hundred and thirty-three Europeans and 1914 South Asian (1165 Indian, 401 Pakistani and 348 Bangladeshi) subjects were recruited from the age-sex registers of 13 general practices in areas with high densities of South Asian populations (Bolton, Oldham, Ashton-under-Lyne and Birmingham). A 28-item acculturation scale was developed, based, on aspects including use of language, clothing style, and use of own-culture media. Principle component analysis generated a score (range 0-100), which was validated against constructs expected to relate to acculturation, such as years of full time education and time spent in the UK. The presence of widespread pain was assessed by the answer to the question 'Have you suffered from pain all over the body in the past month?' RESULTS: Widespread pain was more common in all three South Asian ethnic groups than in the white Europeans [odds ratio (OR) = 3.7, 95% confidence interval (CI) 2.9-4.9], with this increase ranging from 2.7 to 5.8 in the different South Asian subgroups. There was a similar increase in consultation rates for pain. Within the South Asians, pooling all three groups, there was a strong negative association between acculturation score and widespread pain, which remained after adjusting for age and sex: [OR (95% CI) per standard deviation decrease in acculturation score -1.2 (1.0-1.3)]. Adjusting for acculturation accounted for some, but not all, of the differences between the ethnic groups in the prevalence of widespread pain: OR 2.0 (95% CI 1.4-3.0). CONCLUSIONS: Widespread pain is more commonly reported in South Asians though there are interesting differences within the South Asian community. Lower acculturation has a strong influence on the reporting of pain, but cannot explain all of the difference between South Asian and European populations.

An excess of widespread pain among South Asians: are low levels of vitamin D implicated?

BACKGROUND: Anecdotal reports from rheumatologists in the United Kingdom suggest that patients from South Asian backgrounds are more likely to report widespread body pain. OBJECTIVE: To confirm the presence of an excess of widespread pain in South Asians, and to evaluate the relationship of their symptoms with levels of 25-OH vitamin D. METHODS: Two population studies involving over 3135 subjects were carried out in the North West and Midlands areas of England. RESULTS: The first study confirmed an excess of widespread pain among South Asians (OR = 1.6, 95% CI 1.3 to 2.1). The second smaller study conducted only among young women also showed a similar excess of widespread pain among South Asians (OR = 1.8, 95% CI 0.7 to 4.7) and found that low levels of 25-OH vitamin D (<10 ng/ml) were more common among those with widespread pain (OR = 3.5, 95% CI 0.4 to 31.0). CONCLUSIONS: Owing to the small numbers, the relationship between 25-OH vitamin D and widespread pain must be considered preliminary and requires further investigation. However, it may be one potentially treatable cause of widespread pain.