Genome and secretome insights: unravelling the lignocellulolytic potential of Myceliophthora verrucosa for enhanced hydrolysis of lignocellulosic biomass.

Lignocellulolytic enzymes from a novel Myceliophthora verrucosa (5DR) strain was found to potentiate the efficacy of benchmark cellulase during saccharification of acid/alkali treated bagasse by ~ 2.24 fold, indicating it to be an important source of auxiliary enzymes. The De-novo sequencing and analysis of M. verrucosa genome (31.7 Mb) revealed to encode for 7989 putative genes, representing a wide array of CAZymes (366) with a high proportions of auxiliary activity (AA) genes (76). The LC/MS QTOF based secretome analysis of M. verrucosa showed high abundance of glycosyl hydrolases and AA proteins with cellobiose dehydrogenase (CDH) (AA8), being the most prominent auxiliary protein. A gene coding for lytic polysaccharide monooxygenase (LPMO) was expressed in Pichia pastoris and CDH produced by M. verrucosa culture on rice straw based solidified medium were purified and characterized. The mass spectrometry of LPMO catalyzed hydrolytic products of avicel showed the release of both C1/C4 oxidized products, indicating it to be type-3. The lignocellulolytic cocktail comprising of in-house cellulase produced by Aspergillus allahabadii strain spiked with LPMO & CDH exhibited enhanced and better hydrolysis of mild alkali deacetylated (MAD) and unwashed acid pretreated rice straw slurry (UWAP), when compared to Cellic CTec3 at high substrate loading rate.

Effectiveness of WhatsApp based debunking reminders on follow-up visit attendance for individuals with hypertension: a randomized controlled trial in India.

BACKGROUND: Individuals with high blood pressure in India often miss essential follow-up visits. Missed visits contribute to gaps across the hypertension care continuum and preventable cardiovascular disease. Widespread misconceptions around hypertension care and treatment may contribute to low follow-up attendance rates, but to date, there is limited evidence of the effect of interventions to debunk such misconceptions on health-seeking behavior. We conducted a randomized controlled trial to measure whether combining information debunking commonly-held misconceptions with a standard reminder reduces missed follow-up visits among individuals with high blood pressure and investigated whether any observed effect was moderated through belief change. METHODS: We recruited 388 patients with uncontrolled blood pressure from the outpatient wards of two public sub-district hospitals in Punjab, India. Participants randomly assigned to the intervention arm received two WhatsApp messages, sent 3 and 1 days before their physician-requested follow-up visit. The WhatsApp message began with a standard reminder, reminding participants of their upcoming follow-up visit and its purpose. Following the standard reminder, we included brief debunking statements aimed at acknowledging and correcting common misconceptions and misbeliefs about hypertension care seeking and treatment. Participants in the control group received usual care and did not receive any messages. RESULTS: We did not find evidence that the enhanced WhatsApp reminders improved follow-up visit attendance (Main effect: 2.2 percentage points, p-value = 0.603), which remained low across both treatment (21.8%, 95% CI: 15.7%, 27.9%) and control groups (19.6%, 95% CI: 14.2%, 25.0%). Participants had widespread misconceptions about hypertension care but our debunking messages did not successfully correct these beliefs (p-value = 0.187). CONCLUSIONS: This study re-affirms the challenge of continuity of care for chronic diseases in India and suggests that simple phone-based health communication methods may not suffice for changing prevalent misconceptions and improving health-seeking behavior. TRIAL REGISTRATION: The trial began on July 18th. We registered the trial on July 18th (before recruitment began), including the main outcomes, on the German Clinical Trial Register [Identifier: DRKS00029712] and published a pre-analysis plan in the Open Science Framework [osf.io/67g35].

Developing endophytic Penicillium oxalicum as a source of lignocellulolytic enzymes for enhanced hydrolysis of biorefinery relevant pretreated rice straw.

Endophytic fungi, as plant symbionts, produce an elaborate array of enzymes for efficient disintegration of lignocellulosic biomass into constituent monomeric sugars, making them novel source of lignocellulolytic CAZymes with immense potential in future biorefineries. The present study reports lignocellulolytic enzymes production potential of an endophytic halotolerant Penicillium oxalicum strain isolated from Citrus limon, under submerged and solid-state fermentation (SmF & SSF, respectively), in the presence and absence of salt (1 M NaCl). The comparative QTOF-LC/MS-based exoproteome analysis of the culture extracts unveiled differential expression of CAZymes, with the higher abundance of GH6 and GH7 family cellobiohydrolase in the presence of 1 M salt. The strain improvement program, employing cyclic mutagenesis and diploidization, was utilized to develop hyper-cellulase producing mutant strains of P. oxalicum. The enzyme production of the developed strain (POx-M35) was further enhanced through statistical optimization of the culture conditions utilizing glucose mix disaccharides (GMDs) as an inducer. This optimization process resulted in the lignocellulolytic cocktail that contained high titers (U/mL) of endoglucanase (EG) (146.16), cellobiohydrolase (CBHI) (6.99), ß-glucosidase (ß-G) (26.21), xylanase (336.05) and FPase (2.02 U/mL), which were 5.47-, 5.54-, 8.55-, 4.96-, and 4.39-fold higher when compared to the enzyme titers obtained in wild HP1, respectively. Furthermore, the lignocellulolytic cocktails designed by blending secretome produced by mutant POx-M35 with xylanases (GH10 and GH11) derived from Malbranchea cinnamomea resulted in efficient hydrolysis of unwashed acid pretreated (UWAP) rice straw slurry and mild alkali deacetylated (MAD) rice straw. This study underscores the potential of bioprospecting novel fungus and developing an improved strain for optimized production and constitution of lignocellulolytic cocktails that can be an important determinant in advancing biomass conversion technologies.

Axitinib in patients with advanced/metastatic soft tissue sarcoma (Axi-STS): an open-label, multicentre, phase II trial in four histological strata.

BACKGROUND: Axitinib is an oral vascular endothelial growth factor receptor inhibitor with anti-tumour activity in renal, thyroid, and pancreatic cancer. METHODS: Axi-STS was a pathologically-stratified, non-randomised, open-label, multi-centre, phase II trial of continuous axitinib treatment in patients ≥16 years, performance status ≤2, with pathologically-confirmed advanced/metastatic soft tissue sarcoma (STS). Patients were recruited within four tumour strata, each analysed separately: angiosarcoma, leiomyosarcoma, synovial sarcoma, or other eligible STSs. The primary outcome was progression-free survival at 12 weeks (PFS12). A Simon's two-stage design with activity defined as PFS12 rate of 40% determined a sample size of 33 patients per strata. RESULTS: Between 31-August-2010 and 29-January-2016, 145 patients were recruited: 38 angiosarcoma, 37 leiomyosarcoma, 36 synovial sarcoma, and 34 other subtypes. PFS12 rate for each stratum analysed was 42% (95% lower confidence interval (LCI); 29), 45% (95% LCI; 32), 57% (95% LCI; 42), and 33% (95% LCI; 21), respectively. There were 74 serious adverse events including two treatment-related deaths of pulmonary haemorrhage and gastrointestinal bleeding. Fatigue and hypertension were the most common grade 3 adverse events. CONCLUSIONS: Axitinib showed clinical activity in all STS strata investigated. The adverse event profile was acceptable, supporting further investigation in phase III trials. CLINICAL TRIAL REGISTRATION: ISRCTN 60791336.

TUXEDO: A phase I/II trial of cetuximab with chemoradiotherapy in muscle-invasive bladder cancer.

OBJECTIVE: To assess feasibility and preliminary efficacy of adding cetuximab to standard chemoradiotherapy for muscle-invasive bladder cancer. PATIENTS AND METHODS: TUXEDO was a prospective, single-arm, open-label, phase I/II trial conducted in six UK hospitals. Cetuximab was administered with an initial loading dose of 400mg/m2 on day 1 of week -1, and then 7-weekly doses of 250mg/m2 . Radiotherapy schedule was 64Gy/32F with day 1 mitomycin C (12g/m2 ) and 5-fluorouracil (500mg/m2 /day) over days 1-5 and 22-26. Patients with T2-4aN0M0 urothelial cancer and a performance status (PS) of 0-1 were eligible. Prior neoadjuvant therapy was permitted. The phase I primary outcome was impact on radiotherapy treatment completion and toxicity experienced during treatment. The phase II primary outcome was local control at three-months post-treatment. ISRCTN identifier: 80733590. RESULTS: Between Sept-2012 and Oct-2016, 33 patients were recruited; 7 in phase I, 26 in phase II. Three patients in phase II were subsequently deemed ineligible and received no trial therapy. Eight patients discontinued cetuximab due to adverse effects. Median age of patients was 70.1 years (range 60.6-75.1), 20 were PS 0, 27 male and 26 had already received neoadjuvant chemotherapy. In phase I, all patients completed planned radiotherapy, with no delays or dose reductions. Of the 30 evaluable patients in phase II, 25 had confirmed local control 3-months post treatment (77%, 95% CI: 58-90). During the trial there were 18 serious adverse events. The study was halted due to slow accrual. CONCLUSION: Phase I data demonstrate it is feasible and safe to add cetuximab to chemoradiotherapy. Exploratory analysis of phase II data provides evidence to consider further clinical evaluation of cetuximab in this setting.

Proline based rationally designed peptide esters against dipeptidyl peptidase-4: Highly potent anti-diabetic agents.

With the target to develop small molecules based anti-diabetic agents, we, herein, report the design, synthesis and biological studies on Lys-Pro and Gly-Pro esters, and a Phe-Pro-Phe tripeptide inhibiting the activity of glycoprotein dipeptidyl peptidase-4 (DPP-4). Since DPP-4 cleaves the glucagon like peptide (GLP-1) and glucose dependent insulinotropic polypeptide (GIP) hormones which are responsible for inducing insulin secretion, the results of present studies could be significant in making control over glycemia. The structural analysis of DPP-4 and its binding mode with the substrate as well as the reported inhibitors provided the background for the design of new molecules. Among the 17 compounds screened against DPP-4, 14 compounds displayed IC50 better than the known drug Sitagliptin. Collectively, a highly encouraging set of molecules was identified that may prove as the clinical candidates for the treatment of diabetes.

Epoxides: Developability as active pharmaceutical ingredients and biochemical probes.

Epoxide functional group is a part of several natural as well as synthetic compounds. Irrespective of the role of epoxide group in drug efficacy and the use of epoxide compounds as tool molecules, uncertainty prevails over concerns associated with the reactivity of this functional group. Herein, we compile information about epoxide-based medicinal compounds and biochemical probes and look into the related advantages and challenges of the epoxide functional group. As a whole, this study is focussed on analyzing the strategies which have been adopted for the successful development of epoxide-based compounds within drug discovery programs.

Inflammation: Biochemistry, cellular targets, anti-inflammatory agents and challenges with special emphasis on cyclooxygenase-2.

Keeping in view the involvement of inflammation in the pathogenesis of several diseases including cancer, diabetes, neurodegenerative disorders and rheumatoid arthritis, herein, we review the processes for the initiation of inflammation and the treatment measures. While focusing on the cyclooxygenase mediated arachidonic acid metabolic pathways, biochemistry of inflammatory prostaglandins is discussed. The data corresponding to efficacy, pharmacokinetic profile and the side effects of the available natural and synthetic anti-inflammatory drugs is reviewed. Moreover, the given information for the drug-based design of new anti-inflammatory agents may help in the development of more potent and safe molecules.

Rationally designed TNF-α inhibitors: Identification of promising cytotoxic agents.

Design and synthesis of new indole derivatives as tumor growth inhibiting agents via inhibiting the TNF-α is described. The preliminary results showed the inhibition of LPS induced production of NO, TNF-α and IL-6 by these compounds out of which compounds 2d and 2g exhibited appreciable cytotoxicity against the 60 cell lines panel of human cancer. The rationale behind the design of the molecules and the results of their biological studies are presented. 2009 Elsevier Ltd. All rights reserved.

Transanal opening of the intersphincteric space: a novel sphincter-sparing procedure to treat 325 high complex anal fistulas with long-term follow-up.

AIM: Sepsis in the intersphincteric plane behaves like an abscess in a closed space and is present in most complex fistulas. Ignoring this sepsis is a cause of recurrence. Complex fistulas can be successfully managed by transanal opening of the intersphincteric space (TROPIS) into the anal canal. The long-term efficacy of the TROPIS procedure was analysed in an exclusive cohort of high complex anal fistulas. METHODS: All consecutive patients operated for a high complex fistula-in-ano were included prospectively. Preoperative MRI scans were obtained for all the patients. The intersphincteric space and internal opening was laid open into the anal canal while preserving the external sphincter. The external tracts were thoroughly curetted and cleaned. Fistula healing rate and objective incontinence scores (preoperatively and during long-term follow-up) were analysed. RESULTS: A total of 325 patients (age 39.9 ± 10.9 years, 292 men) were operated with TROPIS and had a follow-up of 7-67 months (median 36 months). In the cohort, 67.4% (219) had recurrent fistulas, 82.8% (269) had multiple tracts, 36.3% (118) had horseshoe tracts, 37.5% (122) had associated abscesses and 24% (78) were supralevator fistulas. Nineteen patients were excluded. Fistulas healed completely in 78.4% (240/306) of patients and did not heal in 21.6% (66/306) of patients. 36/66 of these patients were operated again and the fistulas healed in 28 patients. Thus, the overall healing rate was 87.6% (268/306). The mean preoperative and postoperative incontinence scores were 0.085 ± 0.35 and 0.119 ± 0.48 respectively (P = 0.38). The healing rate of fistulas with associated acute abscesses was similar to the fistulas without abscesses (87% vs. 88%, P = 0.85). CONCLUSION: Transanal opening of the intersphincteric space is a safe and highly effective sphincter-sparing procedure to manage high complex fistulas-in-ano.

Role of MRI to confirm healing in complex high cryptoglandular anal fistulas: long-term follow-up of 151 cases.

AIM: Complex fistula-in-ano can recur even after complete clinical healing has occurred. 'Radiological healing' of fistula on MRI correlates well with long-term healing rates but no study has yet objectively quantified this. The aim of this study was to assess the accuracy of anal fistula healing as documented on MRI and to correlate it with long-term healing as evidenced on long-term follow-up. METHODS: Patients with clinically healed anal fistulas who also had radiological healing checked by postoperative MRI were included in the study. RESULTS: Three hundred and twenty-five patients operated for high complex fistula-in-ano were followed up for 14-68 months (median 38 months). Postoperative MRI was done to assess radiological healing of the fistula in 151 patients, and they were included in the study. The mean age was 39.4 ± 10.5 years (116 men). Five patients were lost to follow-up. The fistulas did not heal radiologically (on MRI) in 20 patients and recurred in all these patients. The fistulas healed radiologically (on MRI) in 126 patients. On long-term follow-up, 124/126 patients remained healed while 2/126 had a recurrence. In the first patient, the fistula recurred 40 months after complete radiological healing. In the second patient, the fistula recurred 10 months after complete radiological healing but pus from the fistula tested positive for tuberculosis (by real-time polymerase chain reaction) and he was excluded from the analysis. Thus, there was only one (1/125) recurrence on long-term follow-up. CONCLUSIONS: Radiological healing on MRI correlates well with long-term healing in complex fistula-in-ano.

Analysing the expenditure on childbearing: a community-based cross-sectional study in rural areas of Punjab (India).

BACKGROUND: A vast array of literature has established that high maternity expenditure precludes women from accessing health services. Further, this maternity expenditure takes catastrophic form, forcing individuals or households to significantly lower their standard of living now or at some time in future. The present study analyses expenditure on childbearing in rural areas of one of the richest and top performer states on health parameters in India, namely Punjab along with examining the determinants of catastrophic expenditure. It also attempts to examine the implementation of Janani Shishu Suraksha Karyakaram (JSSK) which entitles pregnant women to free maternity services in public health facilities. METHODS: A cross-sectional study was conducted in rural areas of Punjab involving 420 recently delivered women, who were questioned about their socio-economic attributes and expenditure incurred in the process of childbearing using face to face, semi-structured interviews. Employing logistic regression, an attempt has been made to understand the determinants of catastrophic maternity expenditure, i.e., expenditure exceeding 10% of annual household income. RESULTS: Of the 420 respondents surveyed, 96.7% reported bearing expenditure on childbearing, irrespective of the type of health facility used and 25% respondents spent catastrophically. On an average, respondents have spent US$62.87 on antenatal care, US$112.86 on delivery and US$6.55 on postnatal care. The results of multivariable analysis reveal that respondents belonging to general category (non reserve category), lower wealth quintiles and using private health facilities have higher odds of incurring catastrophic expenditure. At the same time, poor quality of care at government hospitals and inability of public health staff to provide timely treatment are the driving forces for utilizing private health facilities. Even in the presence of free maternity scheme at government hospitals, respondents on an average spent US$55.22 on availing maternity services. CONCLUSION: The study shows that risk of bearing catastrophic expenditure and being pushed down to abject poverty is higher for respondents who are already at the bottom of wealth quintiles. The policy imperative has to swing towards upgrading the creaky health infrastructure and addressing the issues of poor accountability and corruption at government hospitals, along with thwarting unregulated expansion of private health sector.

Fear vs. hope in India: Finding the silver lining amid the dark clouds of COVID-19.

COVID-19 has been declared a global pandemic, and the associated high rates of morbidity and mortality have made individuals susceptible to mental health problems that affect their psychological well-being. Although individual strengths can shield the negative impact of adverse conditions, their protective role in the context of COVID-19 has not received much attention. This study examines the relationship between fear of COVID-19 and mental health via rumination through the lens of hope as a personal psychological strength. This study employed a two-wave longitudinal design. Data was collected from 412 Indian participants with a time interval of three months and analyzed using a two-step approach to structural equation modelling. Fear of COVID-19 was found to a have negative effect on mental health through rumination. However, results from moderation analysis support the role of hope as a buffer against the indirect negative impact of fear of COVID-19 on mental health outcomes. As one of the first studies to demonstrate the role of psychological strengths of individuals in coping with the direct and indirect psychological ramifications of COVID-19 over a period of time, it contains important implications for the development of mental health interventions in the face of this global crisis.

Rational modification of semaxanib and sunitinib for developing a tumor growth inhibitor targeting ATP binding site of tyrosine kinase.

Analysis of the crystal structure of tyrosine kinase in complexation with an ATP analogue, supplemented with the molecular docking studies of semaxanib and sunitinib in the ATP binding site of the enzyme enabled us to make design of a series of tyrosine kinase inhibitors. The combination of pyrrole and indolinone in one molecule and placement of appropriate substituent thereof made the molecule compatible for the hydrophobic sub-pocket of the enzyme. Screening of the compounds over 60 cell line panel of human tumor cell lines identified compound 3a that exhibited GI50 35 nM and 63 nM against MCF7 and MDA-MB-468 cell lines of breast cancer.

A bisubstrate reagent orchestrating adenosine triphosphate and l-tyrosine and making tyrosyl adenylate: partial mimicking of tyrosyl-tRNA synthetase.

We report the development of a bisubstrate reagent that, similar to tyrosyl t-RNA synthetase (TyrTS), provides a surface for ATP and l-Tyr to render a pseudo-intramolecular reaction forming 5'-tyrosyl adenylate (tyrAd). The presence of the reagent in solution with TyrTS marred the enzymatic reaction and, noticeably, tyrAd formed under the catalytic mode of the biomodel reagent was not picked up by TyrTS and hence was not transferred to tRNA. A potential application of this reagent, which doesn't allow the formation of tyrosyl tRNA, may lie in an emerging therapeutic targeting the translation machinery of cells without inhibiting the normal workings of enzymes.

Magnetic Resonance Imaging (MRI): Operative Findings Correlation in 229 Fistula-in-Ano Patients.

BACKGROUND: To correlate the operative findings of patients with fistula-in-ano with preoperative MRI and quantify the information added with MRI. METHODS: All consecutive fistula-in-ano patients operated between July 2013 and May 2015 were prospectively enrolled. Preoperative MRI was done in every patient. The details of tracts, internal opening and "complex parameters" (additional tract or additional internal opening, horseshoe tract, associated abscess and supralevator extension) found at surgery were compared to the findings determined by MRI. RESULTS: A total of 229 patients (424 tracts) with mean age-49.0 ± 11.3 years were included. M/F 198/31. James hospital classification: Type I 58, II 20, III 49, IV 86 and V 16. The sensitivity and specificity of MRI in diagnosing fistula tracts were 98.8 and 99.7%, respectively, and in identifying internal opening were 97.7 and 98.6%, respectively. MRI added significant information in 46.7% (107/229) patients which was presence of additional tracts in 71 (66.3%), horseshoe tract in 63 (58.8%), supralevator extension in 16 (14.9%), unsuspected abscess in 11 (10.3%) and multiple internal openings in one patient (1%). The proportion of simple/complex fistula (based on history and clinical examination alone) was 32.8/67.2% which changed to 21.4/78.6% after the MRI scan. MRI added significant information about unsuspecting complex parameters which were missed on history and clinical examination in more than one-third (26/75: 34.6%) of simple fistulae and more than half (81/154: 52.5%) of already known complex fistulae. CONCLUSIONS: MRI is highly accurate in diagnosing fistula-in-ano and added significant information about unsuspected complex parameters in over one-third (34.6%) of simple and in half (52.5%) of complex fistula-in-ano.

Small Peptides Targeting BACE-1, AChE, and A-ß Reversing Scopolamine-Induced Memory Impairment: A Multitarget Approach against Alzheimer's Disease.

Based on the biochemical understanding of Alzheimer's disease, here, we report the design, synthesis, and biological screening of a series of compounds against this neuro-disorder. Adopting the multitarget approach, the catalytic processes of BACE-1 and AChE were targeted, and thereby, compounds 15, 22, 25, 26, 27, and 30 were identified with IC50 in the submicromolar range against these two enzymes. Further, compounds 15 and 25 displayed more than 50% inhibition of ß-amyloid aggregation. Implying their physiological use, the compounds exhibited appreciable biological membrane permeability as observed through the parallel artificial membrane permeability experiment. Supporting these results, treatment of the mice with the test compounds reversed their scopolamine-affected memory impairment, where the highest healing effect was seen in the case of compound 25. Overall, the combination of molecular modeling and experimental studies provided highly effective molecules against Alzheimer's disease.