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The SARS-CoV-2 main protease (Mpro) plays an essential role in the coronavirus lifecycle by catalysing hydrolysis of the viral polyproteins at specific sites. Mprois the target of drugs, such as nirmatrelvir, though resistant mutants have emerged that threaten drug efficacy. Despite its importance, questions remain on the mechanism of how Mprobinds its substrates. Here, we apply dynamical nonequilibrium molecular dynamics (D-NEMD) simulations to evaluate structural and dynamical responses of Mproto the presence and absence of a substrate. The results highlight communication between the Mprodimer subunits and identify networks, including some far from the active site, that link the active site with a known allosteric inhibition site, or which are associated with nirmatrelvir resistance. They imply that some mutations enable resistance by altering the allosteric behaviour of Mpro. More generally, the results show the utility of the D-NEMD technique for identifying functionally relevant allosteric sites and networks including those relevant to resistance.

Objective: To establish whether there is a significant change in suicidality risk following psychiatric assessment for commencement of isotretinoin and identify factors that underpin any potential risk change. Method: Retrospective cohort study. Suicidality risk was defined as a combination of the following: (i) actual/intended self-harm and/or attempted/completed suicide, and (ii) increased service utilisation associated with suicidal ideation/behaviour. All patients referred to Psychiatry for assessment prior to commencement of isotretinoin between 2014 and 2019 were examined. Inclusion criteria: >16 years of age, assessed for commencement of isotretinoin, complete clinical records. Data were collected by reviewing the Electronic Patient Records. Fifty-seven patients were eligible. We employed descriptive statistics, parametric/non-parametric/normality tests and logistic regression analysis, using socio-demographic and clinical characteristics as independent parameters, and suicidality risk as the dependent parameter. Results: Actual/intended self-harm/attempted suicide decreased significantly following assessment without significant change in service utilisation. Female gender, absence of protective factors and assessment by Consultation-Liaison Psychiatry were linked to increased suicidality risk, after controlling for age, ethnicity, recommendation for isotretinoin, and substance misuse. Conclusions: Psychiatric assessment is helpful before commencing isotretinoin. Female gender, and absence of ongoing psychopharmacological and/or psychological intervention and/or regular psychiatric follow-up predict increased suicidality risk among patients assessed for prescription of isotretinoin.

OBJECTIVE The aim of this study was to compare the University of Texas (UT) and Site, Ischemia, Neuropathy, Bacterial Infection, and Depth (SINBAD) foot ulcer scores in predicting ulcer outcome within a routine diabetes foot clinic. RESEARCH DESIGN AND METHODS From 2006 to 2018, data were collected from all patients attending an outpatient diabetes foot clinic with an active ulcer not healed within 4 weeks. UT and SINBAD were compared in predicting ulcer outcome. A unified numerical score for UT was constructed and compared with UT grade (depth) and stage scores. Outcomes included death, a healed ulcer, or a nonhealed ulcer, which included major or minor amputation and nonhealing chronic ulcers. RESULTS Outcomes were available from 1,645 ulcer outcomes in 1,068 patients (mean [SD] age 65.4 [4] years, 72% male), of which 1,108 (67%) healed. With exclusion of death as an adverse outcome, the c-statistic (area under operator curve) was 0.67 (95% CI 0.65–0.71) for UT grade/depth and 0.64 (0.61–0.67) for UT stage. The new unified UT score had an improved c-statistic of 0.71 (0.68–0.74). The c-statistic was 0.72 (0.69–0.75) for SINBAD. There was a stepwise decrease in the proportion of ulcers healed for each increased score on ulcer grading for both grading schemes. CONCLUSIONS This large and independent observational comparison, in a real-world clinical setting, demonstrated that the UT and SINBAD diabetes foot ulcer grading schemes had similar prognostic ability for predicting foot ulcer outcomes. We have devised and validated a unified numerical scoring system for UT.

Small-ring cage hydrocarbons are common bioisosteres for para-substituted benzene rings in drug design, exhibiting superior pharmacokinetic properties compared to the parent aromatics. In contrast, scaffolds mimicking meta-substituted arenes are lacking due to the challenge of synthesising saturated isosteres that accurately reproduce the substituent vectors of the corresponding arene. Here we report the use of [3.1.1]propellane as a convenient and scalable precursor to bicyclo[3.1.1]heptanes (BCHeps), hydrocarbons whose bridgehead substituents map precisely onto the geometry of meta-substituted benzenes. This precursor undergoes a range of radical-based transformations to generate medicinally-relevant carbon- and heteroatom-substituted BCHeps, including BCHep pharmaceutical analogues. Comparison of ADME properties of the latter reveals improved metabolic stability relative to their parent drugs, validating the potential of this meta-arene analogue as an sp3-rich motif in drug design.

The COVID-19 pandemic has highlighted the urgency for developing more efficient molecular discovery pathways. As exhaustive exploration of the vast chemical space is infeasible, discovering novel inhibitor molecules for emerging drug-target proteins is challenging, particularly for targets with unknown structure or ligands. We demonstrate the broad utility of a single deep generative framework toward discovering novel drug-like inhibitor molecules against two distinct SARS-CoV-2 targets — the main protease (Mpro) and the receptor binding domain (RBD) of the spike protein. To perform target-aware design, the framework employs a target sequence-conditioned sampling of novel molecules from a generative model. Micromolar-level in vitro inhibition was observed for two candidates (out of four synthesized) for each target. The most potent spike RBD inhibitor also emerged as a rare non-covalent antiviral with broad-spectrum activity against several SARS-CoV-2 variants in live virus neutralization assays. These results show that a broadly deployable machine intelligence framework can accelerate hit discovery across different emerging drug-targets.

By scanning electron microscopy, sensilla type and distribution were studied on the antennae of ten species of Rhodnius. No sexual dimorphism was apparent. Intraspecific differences in sensillar pattern were shown to be potentially useful in species determination and to agree with results from biochemical studies and previous morphometric studies in terms of species affinities. © 1994 Wiley-Liss, Inc.

For many years, the development of insulin resistance has been seen as the core defect responsible for the development of Type 2 diabetes. However, despite extensive research, the initial factors responsible for insulin resistance development have not been elucidated. If insulin resistance can be overcome by enhanced insulin secretion, then hyperglycaemia will never develop. Therefore, a β-cell defect is clearly required for the development of diabetes. There is a wealth of evidence to suggest that disorders in insulin secretion can lead to the development of decreased insulin sensitivity. In this review, we describe the potential initiating defects in Type 2 diabetes, normal pulsatile insulin secretion and the effects that disordered secretion may have on both β-cell function and hepatic insulin sensitivity. We go on to examine evidence from physiological and epidemiological studies describing β-cell dysfunction in the development of insulin resistance. Finally, we describe how disordered insulin secretion may cause intracellular insulin resistance and the implications this concept has for diabetes therapy. In summary, disordered insulin secretion may contribute to development of insulin resistance and hence represent an initiating factor in the progression to Type 2 diabetes.

Insulin resistance is a characteristic of type 2 diabetes and is a major independent risk factor for progression to the disease. In particular, insulin resistance associates with increased body fat and almost certainly contributes to the dramatic increase in risk of type 2 diabetes associated with obesity. Therefore, in order to design truly effective insulin sensitising agents, targeted at the mechanism of disease development, we aimed to generate an obesity-related insulin resistant cell model. Rat hepatoma cells were grown in the presence of serum isolated from obese rodents or obese human volunteers, and the insulin sensitivity of the cells monitored over time by measuring a well-characterised insulin regulated gene promoter. Higher insulin concentrations were required to fully repress the gene in the cells grown in obese rodent serum compared with those grown in serum from lean rodents (almost a 10-fold shift in insulin sensitivity). This was reversed by restoration of normal growth medium, while the insulin resistance was prevented by pioglitazone or metformin. Meanwhile, growth of cells in serum collected from obese human volunteers with diabetes also reduced the insulin sensitivity of the rat cells. No clinical marker predicted the degree of insulin resistance that was generated by the human serum. We have developed a novel insulin resistant cell model for the study of the molecular development of obesity-linked insulin resistance, screen for compounds to overcome obesity-related insulin resistance and potentially search for novel serum biomarkers of insulin resistance.

Aims To ascertain which perifoveal changes on digital retinal screening in diabetes predict the need for subsequent macular grid or focal laser therapy. Methods Between 1 January 2004 and 31 December 2005, all consecutive retinal images where any lesion was within one disc diameter of the fovea were reviewed. Patients were categorized by lesion at screening as having microaneurysm, single blot haemorrhage, multiple blot haemorrhages and exudates or circinate exudates within one disc diameter of the fovea. We compared these retinal images with the findings on slit lamp examination and the related decision for laser photocoagulation. Results Four hundred and twenty-four retinal images were identified. Of these, 52 were excluded, principally because of an interval between photography and clinic attendance of greater than 120 days, leaving 372 retinal images in the study group (313 patients). No patients with a single blot haemorrhage required immediate laser therapy at ophthalmology review compared with 13 (23%) of those with multiple blot haemorrhages and 36 (16%) of those with exudates or circinate lesions (P

OBJECTIVE—To determine whether patients developing ulcers had previously been identified as being at high risk of ulceration using the Scottish Care Information–Diabetes Collaboration (SCI-DC) ulcer risk score and whether the risk score predicts ulcer healing. RESEARCH DESIGN AND METHODS—All patients attending the diabetes foot clinic with an ulcer had been assessed for foot ulcer risk using the SCI-DC risk calculator, which categorizes patients into low, moderate, or high risk of ulceration. Information on foot pulses, neuropathy, foot deformity, previous ulcer, ulcer site, depth, and presence of sepsis was recorded and related to ulcer outcome. Patients were followed up until outcome was achieved (median 3 months [range 1–33]). RESULTS—Of patients attending the clinic with a foot ulcer (mean [±SD] age 67.3 ± 12.7 years, 68% male), 68% were previously recognized to be at high risk of foot ulceration, and 98% were high or moderate risk. Of 221 ulcers, the healing rate was 75% overall but was lower for high-risk patients compared with other patients (68 vs. 93%; P < 0.0001). Of the remainder, 3% became chronic ulcers, 12% required minor or major amputation, and 10% died with their ulcer. In multivariate analysis, absent pulses, neuropathy, increased age, and deep ulcers were associated with poor healing. The combination of neuropathy and ischemia was particularly associated with poor outcome of an ulcer (61% healing). CONCLUSIONS—The Scottish foot ulcer risk score predicts both ulcer development and ulcer healing. The risk score can be a useful initial guide to determine the likelihood of poor healing. The individual criteria contributing to this overall risk are similar to other studies.

OBJECTIVE—We sought to compare the risk of mortality and hospitalization between patients with and without diabetes following incident lower-extremity amputation (LEA). RESEARCH DESIGN AND METHODS—We performed a retrospective data-linkage review of all incident amputations between 1 January 1992 and 31 December 1995. Patients were categorized according to their diabetes status. Follow-up for mortality was until 1 January 2005 and until 31 March 1996 for hospitalization. RESULTS—Of 390 major-incident LEAs performed during the study period, 119 (30.5%) were in patients with diabetes and 271 (69.5%) were in nondiabetic subjects. The median time to death was 27.2 months in patients with diabetes compared with 46.7 months for patients without (P = 0.01). Diabetic subjects had a 55% greater risk of death than those without diabetes. The risk of developing congestive cardiac failure with diabetes was 2.26 (95% CI 1.12–4.57) and of further amputation was 1.95 (1.14–3.33) times that of a patient without diabetes after incident LEA. CONCLUSIONS—After LEA, patients with diabetes have an increased risk of death compared with nondiabetic patients. Efforts should be made to minimize these risks with aggressive treatment of cardiovascular risk factors and management of cardiac failure.

Aims and methodThe assessment of capacity is integral to all areas of medical practice; however, research has shown that doctors in many specialties have inadequate knowledge about both the Mental Capacity Act and the Mental Health Act. In this study, 39 trainee doctors and 49 fourth-year medical students completed an e-learning package, which included a pre- and post-evaluation of knowledge of the Mental Capacity Act and confidence in the use of this act.ResultsIn comparison to pre-intervention scores, trainee doctors and students displayed a statistically significant improvement in knowledge and subjective confidence.Clinical implicationsThe e-learning package is an effective educational tool for improving knowledge about the Mental Capacity Act for trainee doctors and could be included as part of induction training for all foundation doctors as well as being considered for use in the induction programme of other trainee doctors. It is an effective tool for educating clinical medical students about the Mental Capacity Act.

SummaryOver many years and with various pieces of new legislation there are significant gaps in doctors' knowledge about mental health law. It is time to ensure that doctors know the law and can apply it to the patients they see. Practising legally and not detaining or allowing people to leave hospital inappropriately should be a mandatory part of training for every doctor no matter what the specialty. Medical schools, deaneries, training programme directors and the General Medical Council should take up the challenge and ensure good-quality training for all doctors to ensure good-quality care in this area is given to all patients.

Hypocalcaemia due to hypoparathyroidism following thyroidectomy is a relatively common occurrence. Standard treatment is with oral calcium and vitamin D replacement therapy; lack of response to oral therapy is rare. Herein we describe a case of hypoparathyroidism following thyroidectomy unresponsive to oral therapy in a patient with a complex medical history. We consider the potential causes in the context of calcium metabolism including: poor adherence, hungry bone syndrome, malabsorption, vitamin D resistance, bisphosphonate use and functional hypoparathyroidism secondary to magnesium deficiency. Malabsorption due to intestinal hurry was likely to be a contributory factor in this case and very large doses of oral therapy were required to avoid symptomatic hypocalcaemia.

OBJECTIVE Guidelines on the clinical use of growth hormone therapy in adults were issued by the UK National Institute for Clinical Excellence (NICE) in August 2003. We conducted a retrospective clinical audit on the use of growth hormone (GH) in Scotland to evaluate the use of these guidelines and their impact on clinical practice. The audit had two phases. In phase I, the impact of NICE criteria on specialist endocrine practice in starting and continuing GH replacement was assessed. In phase II, the reasons why some adults in Scotland with growth hormone deficiency were not on replacement therapy were evaluated. METHODS A retrospective cross-sectional case note review was carried out of all adult patients being followed up for growth hormone deficiency during the study period (1 March 2005 to 31 March 2008). Phase I of the audit included 208 patients and phase II 108 patients. RESULTS Sellar tumours were the main cause of GH deficiency in both phases of the audit. In phase I, 53 patients (77%) had an AGHDA-QoL score >11 documented before commencing GH post-NICE guidance, compared with 35 (25%) pre-NICE guidance. Overall, only 39 patients (18%) met the full NICE criteria for starting and continuing GH (pre-NICE, 11%; post-NICE, 35%). Phase II indicated that the main reasons for not starting GH included perceived satisfactory quality of life (n = 47, 43%), patient reluctance (16, 15%) or a medical contraindication (16, 15%). CONCLUSIONS Although the use of quality of life assessments has increased following publication of the NICE guidelines, most adults on GH in Scotland did not fulfil the complete set of NICE criteria. The main reason for not starting GH therapy in adult GH-deficient patients was perceived satisfactory quality of life.

Metformin is the most commonly used pharmacological therapy for type 2 diabetes. We report a genome-wide association study for glycemic response to metformin in 1,024 Scottish individuals with type 2 diabetes with replication in two cohorts including 1,783 Scottish individuals and 1,113 individuals from the UK Prospective Diabetes Study. In a combined meta-analysis, we identified a SNP, rs11212617, associated with treatment success (n = 3,920, P = 2.9 P×-9, odds ratio = 1.35, 95% CI 1.22-1.49) at a locus containing ATM, the ataxia telangiectasia mutated gene. In a rat hepatoma cell line, inhibition of ATM with KU-55933 attenuated the phosphorylation and activation of AMP-activated protein kinase in response to metformin. We conclude that ATM, a gene known to be involved in DNA repair and cell cycle control, plays a role in the effect of metformin upstream of AMP-activated protein kinase, and variation in this gene alters glycemic response to metformin.

Aims To assess the changing rate of amputation in patients with diabetes over a 7-year period. Methods All patients undergoing lower extremity amputation in Tayside, Scotland between 1 January 2000 and 31 December 2006 were identified. Temporal linkage of cases to the diabetes database was used to ascertain which amputations were in patients with diabetes. Results The incidence of major amputations fell from 5.1 [95% confidence interval (CI) 3.8-6.4] to 2.9 (95% CI 1.9-3.8) per 1000 patients with diabetes (P < 0.05). There is a clear linear trend in the adjusted incidence of major amputation (P = 0.023 and 0.027 for age- and sex-adjusted, and duration- and sex-adjusted incidences, respectively). The adjusted incidence of total amputations followed decreased linear regression trend over the whole study period when adjusted for age and sex or diabetes duration and sex (P = 0.002). Conclusions There has been a significant reduction in the incidence of major lower extremity amputation in patients with diabetes over the 7-year period. Diabet. Med. 26, 773-777 (2009).

This thesis reports the work done on two research projects which were carried out independently of each other but converge on the central theme of β-lactam biosynthesis. Chapter 1 provides an overview of biosynthesis in secondary metabolism, with special emphasis on current knowledge about the β-lactams. The first project, covered from Chapters 2 to 5, was part of our group's continuing effort to understand the structure and mechanism of Ring Expandase-Hydroxylase (REXH), an enzyme involved in the biosynthesis of cephalosporin C in Cephalosporium acremonium. REXH is a bifunctional enzyme, converting penicillin N to DAOC and thence to DAC. [diagram omitted from transcription] Chapter 3 discusses the investigation of purification protocols for native REXH and soluble recombinant REXH, as well as an improved refolding method for recombinant REXH expressed as inclusion bodies. Chapter 4 describes two new alternative substrates for REXH, viz. carba-DAOC and DAC, whilst the y-lactam analogue of penicillin N was not found to be a substrate for REXH. Chapter 5 summarises some structural investigations of REXH employing methods such as electrospray mass spectrometry, selective proteolysis and inhibition kinetics. [diagram omitted from transcription] The second project, covered from Chapters 6 to 9, represents the first biosynthetic studies on valclavam, an antifungal produced by Streptomyces antibioticus. Valclavam belongs to the family of clavams which includes clavulanic acid as its most well studied member. [diagram omitted from transcription] Chapter 7 details the development of methods for the bioassay, fermentation and isolation of valclavam. It also describes the isolation of a stable degradation fragment of valclavam which led to the revision of the structures of valclavam and Tü 1718B (another metabolite from the same organism). Chapter 8 gives an account of the whole-cell feeding experiments which strongly suggest that the primary metabolic precursors for valclavam are L-valine, L-arginine, L-methionine and glycerol. Chapter 9 reports the discovery of two enzymic activities, belonging to those of clavaminic acid synthase and proclavaminic acid amidino hydrolase, which are likely to be involved in the biosynthesis of valclavam. Together, the results of Chapters 8 and 9 point to an extensive overlap between the clavulanic acid pathway in Streptomyces clavuligerus and the valclavam pathway in Streptomyces antibioticus.