Search

Catalysis with engineered enzymes has provided more efficient routes for the production of active pharmaceutical agents. However, the potential of biocatalysis to assist in early-stage drug discovery campaigns remains largely untapped. In this study, we have developed a biocatalytic strategy for the construction of sp3-rich polycyclic compounds via the intramolecular cyclopropanation of benzothiophenes and related heterocycles. Two carbene transferases with complementary regioisomer selectivity were evolved to catalyse the stereoselective cyclization of benzothiophene substrates bearing diazo ester groups at the C2 or C3 position of the heterocycle. The detailed mechanisms of these reactions were elucidated by a combination of crystallographic and computational analyses. Leveraging these insights, the substrate scope of one of the biocatalysts could be expanded to include previously unreactive substrates, highlighting the value of integrating evolutionary and rational strategies to develop enzymes for new-to-nature transformations. The molecular scaffolds accessed here feature a combination of three-dimensional and stereochemical complexity with rule-of-three properties, which should make them highly valuable for fragment-based drug discovery campaigns.

Lactam rings are found in many biologically active natural products and pharmaceuticals, including important classes of antibiotics. Methods for the asymmetric synthesis of these molecules are therefore highly desirable, particularly through the selective functionalization of unreactive aliphatic C-H bonds. Here we show the development of a strategy for the asymmetric synthesis of β-, γ-, and δ-lactams via hemoprotein-catalysed intramolecular C-H amidation reaction with readily available dioxazolone reagents. Engineered myoglobin variants serve as excellent biocatalysts for this transformation yielding the desired lactam products in high yields, high enantioselectivity, and on preparative scale. Mechanistic and computational studies elucidate the nature of the C-H amination and enantiodetermining steps and provide insights into protein-mediated control of regioselectivity and stereoselectivity. Additionally, an alkaloid natural product and a drug molecule were synthesized chemoenzymatically in much fewer steps (7-8 vs. 11-12) than previously reported, further demonstrating the power of biosynthetic strategy for the preparation of complex bioactive molecules.

Abstract Hemoproteins have recently emerged as promising biocatalysts for new-to-nature carbene transfer reactions. However, mechanistic understanding of the interplay between productive and unproductive pathways in these processes is limited. Using spectroscopic, structural, and computational methods, we investigate the mechanism of a myoglobin-catalyzed cyclopropanation reaction with diazoketones. These studies shed light on the nature and kinetics of key catalytic steps in this reaction, including the formation of an early heme-bound diazo complex intermediate, the rate-determining nature of carbene formation, and the cyclopropanation mechanism. Our analyses further reveal the existence of a complex mechanistic manifold for this reaction that includes a competing pathway resulting in the formation of an N-bound carbene adduct of the heme cofactor, which was isolated and characterized by X-ray crystallography, UV-Vis, and Mössbauer spectroscopy. This species can regenerate the active biocatalyst, constituting a non-productive, yet non-destructive detour from the main catalytic cycle. These findings offer a valuable framework for both mechanistic analysis and design of hemoprotein-catalyzed carbene transfer reactions.

Background: Methylmalonic Aciduria (MA) without homocystinuria (or isolated MA) is a group of rare inherited metabolic disorders which leads to the accumulation of methylmalonic acid (MMA), a toxic molecule that accumulates in blood, urine, and cerebrospinal fluid, causing acute and chronic complications including metabolic crises, acute kidney injury (AKI), and chronic kidney disease (CKD). Detailed Case Description: Herein, we report a case of a 39-year-old male with MA and stage IV CKD who experienced acute metabolic decompensation secondary to gastrointestinal infection. The patient underwent a single hemodialysis (HD) session to correct severe metabolic acidosis unresponsive to medical therapy and to rapidly remove MMA. The HD session resulted in prompt clinical improvement and shortening of hospitalization. Discussion: MMA accumulation in MA patients causes acute and life-threatening complications, such as metabolic decompensations, and long-term complications such as CKD, eventually leading to renal replacement therapy (RRT). Data reported in the literature show that, overall, all dialytic treatments (intermittent HD, continuous HD, peritoneal dialysis) are effective in MMA removal. HD, in particular, can be useful in the emergency setting to control metabolic crises, even with GFR > 15 mL/min. Kidney and/or liver transplantations are often needed in MA patients. While a solitary transplanted kidney can be rapidly affected by MMA exposure, with a decline in renal function even in the first year of follow-up, the combined liver–kidney transplantation showed better long-term results due to a combination of reduced MMA production along with increased urinary excretion. Conclusions: Early diagnosis, multidisciplinary management and preventive measures are pivotal in MA patients to avoid recurrent AKI episodes and, consequently, to slow down CKD progression.

The mechanical analysis of photovoltaics and building integrated photovoltaics is a key step for their optimal design and certification, and requires careful consideration, alongside solar power, durability and functionality issues. The solar cells are encapsulated in thin interlayers that are usually composed of a viscoelastic Ethylene–Vinyl Acetate compound, and protected by thin glass and/or plastic layers. This paper investigates the out-of-plane bending response of a full-scale commercial PV module and focuses attention on the shear bonding efficiency of the thin encapsulant for quasi-static and dynamic mechanical considerations. The parametric analytical analysis, carried out in this study for a laminated glass plate, highlights the possible consequences of the viscoelastic shear coupling on the cross-section load-bearing demand in the covers. As a direct effect of severe operational conditions (i.e., ageing, non-uniform/cyclic thermal gradients, humidity, extreme mechanical/thermal loads, etc.) the shear rigidity and adhesion of these films can suffer from repeated/progressive modification and even degradation, and thus induce major stress and deflection effects in the out-of-plane mechanical response of the PV module components. The minimum shear bond efficiency required to prevent mechanical issues is calculated for various configurations of technical interest. Accordingly, it is shown how the quasi-static and dynamic mechanical performance of the system modifies as a function of a more rigid or weak shear coupling.

Background: Wilms tumor (nephroblastoma) is a malignant pediatric tumor that arises from mesodermal embryological precursors of renal parenchyma. Lung metastasis as the first presentation of nephroblastoma occurs in 20% of cases. However, haematogenous spread to the lungs or to regional lymphnodes from a previously diagnosed nephroblastoma occurs in of 85% of cases. Case report: A 3-year old girl presented at children emergency unit with cough and weight loss. Examination revealed an emaciated child, with a huge, right sided abdominal mass. Abdominal ultrasound scan showed a heterogenous, spherical mass with irregular anechoic areas and echogenic solid areas in kidney. Chest radiograph revealed bilateral cannon ball opacities. A diagnosis of nephroblastoma with lung metastasis was made. She underwent combined therapy but relapsed one year later.

Summary: Background: Lack of specific definitions of clinical characteristics, disease severity, and risk and preventive factors of post-COVID-19 syndrome (PCS) severely impacts research and discovery of new preventive and therapeutics drugs. Methods: This prospective multicenter cohort study was conducted from February 2020 to June 2022 in 5 countries, enrolling SARS-CoV-2 out- and in-patients followed at 3-, 6-, and 12-month from diagnosis, with assessment of clinical and biochemical features, antibody (Ab) response, Variant of Concern (VoC), and physical and mental quality of life (QoL). Outcome of interest was identification of risk and protective factors of PCS by clinical phenotype, setting, severity of disease, treatment, and vaccination status. We used SF-36 questionnaire to assess evolution in QoL index during follow-up and unsupervised machine learning algorithms (principal component analysis, PCA) to explore symptom clusters. Severity of PCS was defined by clinical phenotype and QoL. We also used generalized linear models to analyse the impact of PCS on QoL and associated risk and preventive factors. CT registration number: NCT05097677. Findings: Among 1796 patients enrolled, 1030 (57%) suffered from at least one symptom at 12-month. PCA identified 4 clinical phenotypes: chronic fatigue-like syndrome (CFs: fatigue, headache and memory loss, 757 patients, 42%), respiratory syndrome (REs: cough and dyspnoea, 502, 23%); chronic pain syndrome (CPs: arthralgia and myalgia, 399, 22%); and neurosensorial syndrome (NSs: alteration in taste and smell, 197, 11%). Determinants of clinical phenotypes were different (all comparisons p

Rationale: A U-shaped relationship should exist between lung volume and pulmonary vascular resistance (PVR), with minimal PVR at FRC. Thus, positive end-expiratory pressure (PEEP) in patients with acute respiratory distress syndrome (ARDS) should increase PVR if it induces significant lung distension compared with recruitment. However, this has never been proved in patients. Objectives: To study the effects of PEEP on PVR according to lung recruitability, evaluated by the recruitment-to-inflation (R/I) ratio. Methods: In patients with ARDS, we measured hemodynamic (pulmonary artery catheter), echocardiographic, and ventilatory variables (including esophageal pressure) at both low PEEP and higher PEEP by 10 cm H2O. Preload responsiveness was assessed by the passive leg-raising test at high PEEP. Measurements and Main Results: We enrolled 23 patients, including 10 low recruiters (R/I <0.5) and 13 high recruiters (R/I ⩾0.5). Raising PEEP from 4 (2–5) to 14 (12–15) cm H2O increased PVR in low recruiters (from 160 [120–297] to 243 [166–380] dyn·s/cm5; P < 0.01), whereas PVR was unchanged in high recruiters (from 224 [185–289] to 235 [168–300] dyn·s/cm5; P = 0.55). Right-to-left ventricular end-diastolic area ratio simultaneously increased in low recruiters (from 0.54 [0.50–0.59] to 0.64 [0.56–0.70]; P < 0.01) while remaining stable in high recruiters (from 0.70 [0.65–0.79] to 0.68 [0.58–0.80]; P = 0.48). Raising PEEP decreased cardiac index only in preload responsive patients. Conclusions: PEEP increases PVR only when it induces significant lung distension compared with recruitment according to the R/I ratio. Tailoring PEEP on this recruitability index should mitigate its hemodynamic effects. [ABSTRACT FROM AUTHOR]

Fluorinated cyclopropanes are highly desired pharmacophores in drug discovery owing to the rigid nature of the cyclopropane ring and the beneficial effects of C−F bonds on the pharmacokinetic properties, cell permeability, and metabolic stability of drug molecules. Herein a biocatalytic strategy for the stereoselective synthesis of mono‐fluorinated and gem‐difluoro cyclopropanes is reported though the use of engineered myoglobin‐based catalysts. In particular, this system allows for a broad range of gem‐difluoro alkenes to be cyclopropanated in the presence of diazoacetonitrile with excellent diastereo and enantiocontrol (up to 99 : 1 d.r. and 99 % e.e.), thereby enabling a transformation not currently accessible with chemocatalytic methods. The synthetic utility of the present approach is further exemplified through the gram‐scale synthesis of a key gem‐difluorinated cyclopropane intermediate useful for the preparation of fluorinated bioactive molecules. [ABSTRACT FROM AUTHOR]

Background: Methylmalonic Aciduria (MA) without homocystinuria (or isolated MA) is a group of rare inherited metabolic disorders which leads to the accumulation of methylmalonic acid (MMA), a toxic molecule that accumulates in blood, urine, and cerebrospinal fluid, causing acute and chronic complications including metabolic crises, acute kidney injury (AKI), and chronic kidney disease (CKD). Detailed Case Description: Herein, we report a case of a 39-year-old male with MA and stage IV CKD who experienced acute metabolic decompensation secondary to gastrointestinal infection. The patient underwent a single hemodialysis (HD) session to correct severe metabolic acidosis unresponsive to medical therapy and to rapidly remove MMA. The HD session resulted in prompt clinical improvement and shortening of hospitalization. Discussion: MMA accumulation in MA patients causes acute and life-threatening complications, such as metabolic decompensations, and long-term complications such as CKD, eventually leading to renal replacement therapy (RRT). Data reported in the literature show that, overall, all dialytic treatments (intermittent HD, continuous HD, peritoneal dialysis) are effective in MMA removal. HD, in particular, can be useful in the emergency setting to control metabolic crises, even with GFR > 15 mL/min. Kidney and/or liver transplantations are often needed in MA patients. While a solitary transplanted kidney can be rapidly affected by MMA exposure, with a decline in renal function even in the first year of follow-up, the combined liver–kidney transplantation showed better long-term results due to a combination of reduced MMA production along with increased urinary excretion. Conclusions: Early diagnosis, multidisciplinary management and preventive measures are pivotal in MA patients to avoid recurrent AKI episodes and, consequently, to slow down CKD progression. [ABSTRACT FROM AUTHOR]