Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase.

Malaria poses an enormous threat to human health. With ever increasing resistance to currently deployed drugs, breakthrough compounds with novel mechanisms of action are urgently needed. Here, we explore pyrimidine-based sulfonamides as a new low molecular weight inhibitor class with drug-like physical parameters and a synthetically accessible scaffold. We show that the exemplar, OSM-S-106, has potent activity against parasite cultures, low mammalian cell toxicity and low propensity for resistance development. In vitro evolution of resistance using a slow ramp-up approach pointed to the Plasmodium falciparum cytoplasmic asparaginyl-tRNA synthetase (PfAsnRS) as the target, consistent with our finding that OSM-S-106 inhibits protein translation and activates the amino acid starvation response. Targeted mass spectrometry confirms that OSM-S-106 is a pro-inhibitor and that inhibition of PfAsnRS occurs via enzyme-mediated production of an Asn-OSM-S-106 adduct. Human AsnRS is much less susceptible to this reaction hijacking mechanism. X-ray crystallographic studies of human AsnRS in complex with inhibitor adducts and docking of pro-inhibitors into a model of Asn-tRNA-bound PfAsnRS provide insights into the structure-activity relationship and the selectivity mechanism.

Laparoscopic deroofing of a recurrent giant simple hepatic cyst in a geriatric female: Case report.

ABSTRACT: Liver cysts often have no symptoms, are benign and are usually encountered during routine abdominal imaging for other conditions. Giant cysts have a higher risk of complications. Recurrence is common, and most research papers support laparoscopic deroofing as the initial management modality even in the recurrent cases as it avoids the morbidity of laparotomy and gives better short- and long-term post-operative outcomes. Here, we present a case of a 68-year-old female who presented to the outpatient department with complaints of right-sided dull-aching abdominal pain, heaviness, early satiety and significant weight loss for the past 2 months. A laparoscopic drainage and deroofing were done, and histopathological examination of the cystic wall was consistent with benign pathology. The patient has been doing well for the past year.

Case report on laparoscopic management of congenital peritoneal encapsulation: A rare cause of small bowel obstruction.

Peritoneal encapsulation (PE) is a rare congenital disorder described as an accessory peritoneal lining covering a part or whole of the small bowel. Some theorise the encapsulation is due to the formation of adhesion between the physiological hernia and the caudal duodenum. While others have stated it is a defect in the reduction of the physiological hernia. Patients usually present at different stages of intestinal obstruction at any point of time during life. There are also reports on post-humous diagnosis on autopsy. PE is a rare surgical entity, hence not much evidences are available on how to tackle this condition by minimally invasive approach. Here, we report a case of PE in a 43-year-old male who presented with features of intermittent sub-acute intestinal obstruction and was managed by laparoscopic surgery at our institute.

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase.

Malaria poses an enormous threat to human health. With ever increasing resistance to currently deployed drugs, breakthrough compounds with novel mechanisms of action are urgently needed. Here, we explore pyrimidine-based sulfonamides as a new low molecular weight inhibitor class with drug-like physical parameters and a synthetically accessible scaffold. We show that the exemplar, OSM-S-106, has potent activity against parasite cultures, low mammalian cell toxicity and low propensity for resistance development. In vitro evolution of resistance using a slow ramp-up approach pointed to the Plasmodium falciparum cytoplasmic asparaginyl tRNA synthetase (PfAsnRS) as the target, consistent with our finding that OSM-S-106 inhibits protein translation and activates the amino acid starvation response. Targeted mass spectrometry confirms that OSM-S-106 is a pro-inhibitor and that inhibition of PfAsnRS occurs via enzyme-mediated production of an Asn-OSM-S-106 adduct. Human AsnRS is much less susceptible to this reaction hijacking mechanism. X-ray crystallographic studies of human AsnRS in complex with inhibitor adducts and docking of pro-inhibitors into a model of Asn-tRNA-bound PfAsnRS provide insights into the structure activity relationship and the selectivity mechanism.

Potent acyl-CoA synthetase 10 inhibitors kill Plasmodium falciparum by disrupting triglyceride formation.

Identifying how small molecules act to kill malaria parasites can lead to new "chemically validated" targets. By pressuring Plasmodium falciparum asexual blood stage parasites with three novel structurally-unrelated antimalarial compounds (MMV665924, MMV019719 and MMV897615), and performing whole-genome sequence analysis on resistant parasite lines, we identify multiple mutations in the P. falciparum acyl-CoA synthetase (ACS) genes PfACS10 (PF3D7_0525100, M300I, A268D/V, F427L) and PfACS11 (PF3D7_1238800, F387V, D648Y, and E668K). Allelic replacement and thermal proteome profiling validates PfACS10 as a target of these compounds. We demonstrate that this protein is essential for parasite growth by conditional knockdown and observe increased compound susceptibility upon reduced expression. Inhibition of PfACS10 leads to a reduction in triacylglycerols and a buildup of its lipid precursors, providing key insights into its function. Analysis of the PfACS11 gene and its mutations point to a role in mediating resistance via decreased protein stability.

The anticancer human mTOR inhibitor sapanisertib potently inhibits multiple Plasmodium kinases and life cycle stages.

Compounds acting on multiple targets are critical to combating antimalarial drug resistance. Here, we report that the human "mammalian target of rapamycin" (mTOR) inhibitor sapanisertib has potent prophylactic liver stage activity, in vitro and in vivo asexual blood stage (ABS) activity, and transmission-blocking activity against the protozoan parasite Plasmodium spp. Chemoproteomics studies revealed multiple potential Plasmodium kinase targets, and potent inhibition of Plasmodium phosphatidylinositol 4-kinase type III beta (PI4Kß) and cyclic guanosine monophosphate-dependent protein kinase (PKG) was confirmed in vitro. Conditional knockdown of PI4Kß in ABS cultures modulated parasite sensitivity to sapanisertib, and laboratory-generated P. falciparum sapanisertib resistance was mediated by mutations in PI4Kß. Parasite metabolomic perturbation profiles associated with sapanisertib and other known PI4Kß and/or PKG inhibitors revealed similarities and differences between chemotypes, potentially caused by sapanisertib targeting multiple parasite kinases. The multistage activity of sapanisertib and its in vivo antimalarial efficacy, coupled with potent inhibition of at least two promising drug targets, provides an opportunity to reposition this pyrazolopyrimidine for malaria.

Reaction hijacking of tyrosine tRNA synthetase as a new whole-of-life-cycle antimalarial strategy.

Aminoacyl transfer RNA (tRNA) synthetases (aaRSs) are attractive drug targets, and we present class I and II aaRSs as previously unrecognized targets for adenosine 5'-monophosphate-mimicking nucleoside sulfamates. The target enzyme catalyzes the formation of an inhibitory amino acid-sulfamate conjugate through a reaction-hijacking mechanism. We identified adenosine 5'-sulfamate as a broad-specificity compound that hijacks a range of aaRSs and ML901 as a specific reagent a specific reagent that hijacks a single aaRS in the malaria parasite Plasmodium falciparum, namely tyrosine RS (PfYRS). ML901 exerts whole-life-cycle-killing activity with low nanomolar potency and single-dose efficacy in a mouse model of malaria. X-ray crystallographic studies of plasmodium and human YRSs reveal differential flexibility of a loop over the catalytic site that underpins differential susceptibility to reaction hijacking by ML901.

Combatting future variants of SARS-CoV-2 using an in-silico peptide vaccine approach by targeting the spike protein.

The far-reaching effects of the SARS-CoV-2 pandemic have crippled the progress of the world today. With the introduction of newer and newer mutated variants of the virus, it has become necessary to have a vaccine that remains useful against all the mutated strains of SARS-CoV-2. In this regard, peptide vaccines turn out to be a cheap alternative to the traditionally designed vaccines owing to their much quicker and computationally easier, and more robust design procedures. Here, in this article, we hypothesize that there are three possible peptide vaccine regions that can be targeted to prevent the surge of SARS-CoV-2. The candidates that were selected, were surface-exposed and were not sequestered by any neighbouring amino acids. They were also found to be capable of generating both B-cell and T-cell immune responses. Most importantly, none of them contains any spike protein mutation of the currently prevailing variants of SARS-CoV-2. From these findings, we have therefore concluded that these three regions can be used in wet labs for peptide vaccine design against the upcoming strains of SARS-CoV-2.

GeneTargeter: Automated In Silico Design for Genome Editing in the Malaria Parasite, Plasmodium falciparum.

Functional characterization of the multitude of poorly described proteins in the human malarial pathogen, Plasmodium falciparum, requires tools to enable genome-scale perturbation studies. Here, we present GeneTargeter (genetargeter.mit.edu), a software tool for automating the design of homology-directed repair donor vectors to achieve gene knockouts, conditional knockdowns, and epitope tagging of P. falciparum genes. We demonstrate GeneTargeter-facilitated genome-scale design of six different types of knockout and conditional knockdown constructs for the P. falciparum genome and validate the computational design process experimentally with successful donor vector assembly and transfection. The software's modular nature accommodates arbitrary destination vectors and allows customizable designs that extend the genome manipulation outcomes attainable in Plasmodium and other organisms.

Identification of Generalized Peptide Regions for Designing Vaccine Effective for All Significant Mutated Strains of SARS-CoV-2.

BACKGROUND: Coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2 infection has become a worldwide pandemic and created an utmost crisis across the globe. To mitigate the crisis, the design of vaccine is the crucial solution. The frequent mutation of the virus demands generalized vaccine candidates, which would be effective for all mutated strains at present and for the strains that would evolve due to further new mutations in the virus. OBJECTIVE: The objective of this study is to identify more frequently occurring mutated variants of SARS-CoV-2 and to suggest peptide vaccine candidates effective against the viral strains considered. METHODS: In this study, we have identified all currently prevailing mutated strains of SARS-CoV-2 through 2D Polar plot and Quotient Radius characterization descriptor. Then, by considering the top eight mutation strains, which are significant due to their frequency of occurrence, peptide regions suitable for vaccine design have been identified with the help of a mathematical model, 2D Polygon Representation, followed by the evaluation of epitope potential, ensuring that there is no case of any autoimmune threat. Lastly, in order to verify whether this entire approach is applicable for vaccine design against any other virus in general, we have made a comparative study between the peptide vaccine candidates prescribed for the Zika virus using the current approach and a list of potential vaccine candidates for the same already established in the past. RESULTS: We have finally suggested three generalized peptide regions which would be suitable as sustainable peptide vaccine candidates against SARS-CoV-2 irrespective of its currently prevailing strains as well any other variant of the same that may appear in the future. We also observed that during the comparative study using the case of E protein of Zika virus, the peptide regions suggested using the new approach that matches with the already established results. CONCLUSION: The study, therefore, illustrates an approach that would help in developing peptide vaccine against SARS-CoV-2 by suggesting those peptide regions which can be targeted irrespective of any mutated form of this virus. The consistency with which this entire approach was also able to figure out similar vaccine candidates for Zika virus with utmost accuracy proves that this protocol can be extended for peptide vaccine design against any other viruses in the future.

The Plasmodium falciparum ABC transporter ABCI3 confers parasite strain-dependent pleiotropic antimalarial drug resistance.

Widespread Plasmodium falciparum resistance to first-line antimalarials underscores the vital need to develop compounds with novel modes of action and identify new druggable targets. Here, we profile five compounds that potently inhibit P. falciparum asexual blood stages. Resistance selection studies with three carboxamide-containing compounds, confirmed by gene editing and conditional knockdowns, identify point mutations in the parasite transporter ABCI3 as the primary mediator of resistance. Selection studies with imidazopyridine or quinoline-carboxamide compounds also yield changes in ABCI3, this time through gene amplification. Imidazopyridine mode of action is attributed to inhibition of heme detoxification, as evidenced by cellular accumulation and heme fractionation assays. For the copy-number variation-selecting imidazopyridine and quinoline-carboxamide compounds, we find that resistance, manifesting as a biphasic concentration-response curve, can independently be mediated by mutations in the chloroquine resistance transporter PfCRT. These studies reveal the interconnectedness of P. falciparum transporters in overcoming drug pressure in different parasite strains.

Repositioning and Characterization of 1-(Pyridin-4-yl)pyrrolidin-2-one Derivatives as Plasmodium Cytoplasmic Prolyl-tRNA Synthetase Inhibitors.

Prolyl-tRNA synthetase (PRS) is a clinically validated antimalarial target. Screening of a set of PRS ATP-site binders, initially designed for human indications, led to identification of 1-(pyridin-4-yl)pyrrolidin-2-one derivatives representing a novel antimalarial scaffold. Evidence designates cytoplasmic PRS as the drug target. The frontrunner 1 and its active enantiomer 1-S exhibited low-double-digit nanomolar activity against resistant Plasmodium falciparum (Pf) laboratory strains and development of liver schizonts. No cross-resistance with strains resistant to other known antimalarials was noted. In addition, a similar level of growth inhibition was observed against clinical field isolates of Pf and P. vivax. The slow killing profile and the relative high propensity to develop resistance in vitro (minimum inoculum resistance of 8 × 105 parasites at a selection pressure of 3 × IC50) constitute unfavorable features for treatment of malaria. However, potent blood stage and antischizontal activity are compelling for causal prophylaxis which does not require fast onset of action. Achieving sufficient on-target selectivity appears to be particularly challenging and should be the primary focus during the next steps of optimization of this chemical series. Encouraging preliminary off-target profile and oral efficacy in a humanized murine model of Pf malaria allowed us to conclude that 1-(pyridin-4-yl)pyrrolidin-2-one derivatives represent a promising starting point for the identification of novel antimalarial prophylactic agents that selectively target Plasmodium PRS.

Base Distribution in Dengue Nucleotide Sequences Differs Significantly from Other Mosquito-Borne Human-Infecting Flavivirus Members.

INTRODUCTION: Among the mosquito-borne human-infecting flavivirus species that include Zika, West Nile, yellow fever, Japanese encephalitis and Dengue viruses, the Zika virus is found to be closest to Dengue virus, sharing the same clade in the Flavivirus phylogenetic tree. We consider these five flaviviruses and on closer examination in our analyses, the nucleotide sequences of the Dengue viral genes (envelope and NS5) and genomes are seen to be quite widely different from the other four flaviviruses. We consider the extent of this distinction and determine the advantage and/or disadvantage such differences may confer upon the Dengue viral pathogenesis. METHODS: We have primarily used a 2D graphical representation technique to show the differences in base distributions in these five flaviviruses and subsequently, obtained quantitative estimates of the differences. Similarity/dissimilarity between the viruses based on the genes were also determined which showed that the differences with the Dengue genes are more pronounced. RESULTS: We found that the Dengue viruses compared to the other four flaviviruses spread rapidly worldwide and became endemic in various regions with small alterations in sequence composition relative to the host populations as revealed by codon usage biases and phylogenetic examination. CONCLUSION: We conclude that the Dengue genes are indeed more widely separated from the other aforementioned mosquito-borne human-infecting flaviviruses due to excess adenine component, a feature that is sparse in the literature. Such excesses have a bearing on drug and vaccine, especially peptide vaccine, development and should be considered appropriately.

Robotic-assisted minimally invasive oesophagectomy for cancer: An initial experience.

BACKGROUND: The morbidity related to radical oesophagectomy can be reduced by adopting minimally invasive techniques. Over 250 thoraco-laparoscopic oesophagectomy (TLE) was done in our centre over the last 15 years, before adopting robotic surgery as the latest innovation in the field of minimally invasive surgery. Here, we share our initial experience of robotic-assisted minimally invasive oesophagectomy (RAMIE) for carcinoma oesophagus. METHODS: A prospective observational study conducted from February to December 2017. A total of 15 patients underwent RAMIE in this period. Data regarding demography, clinical characteristics, investigations, operating techniques, and post-operative outcome were collected in detail. RESULTS: There were 10 (66.7%) male patients and the median age of all patients was 62.9 (range 36-78) years. The median body mass index was 24.4 (range 15-32.8) kg/m2. Twelve (80.0%) patients had squamous cell carcinoma (SCC) of the oesophagus and 3 (20%) patients had adenocarcinoma (AC). Five (33.3%) patients received neoadjuvant therapy. All 15 patients underwent RAMIE. Patients with SCC underwent McKeown's procedure, and those with AC underwent Ivor Lewis procedure. Extended two-field lymphadenectomy (including total mediastinal lymphadenectomy) was done for all the patients. The median operating time was 558 (range 390-690) min and median blood loss was 145 (range 90-230) ml. There were no intra-operative adverse events, and none of them required conversion to open or total thoracolaparoscopic procedure. The most common post-operative complications were recurrent laryngeal nerve paresis (3 patients, 20.0%) and pneumonia (2 patients, 13.3%). The median hospital stay was 9 (range 7-33) days. In total, 9 (60%) patients required adjuvant treatment. CONCLUSION: Adequate experience in TLE can help minimally invasive surgeons in easy adoption of RAMIE with satisfactory outcome.

Epidemics and Peptide Vaccine Response: A Brief Review.

We briefly review the situations arising out of epidemics that erupt rather suddenly, threatening life and livelihoods of humans. Ebola, Zika and the Nipah virus outbreaks are recent examples where the viral epidemics have led to considerably high degree of fatalities or debilitating consequences. The problems are accentuated by a lack of drugs or vaccines effective against the new and emergent viruses, and the inordinate amount of temporal and financial resources that are required to combat the novel pathogens. Progress in computational, biological and informational sciences have made it possible to consider design of synthetic vaccines that can be rapidly developed and deployed to help stem the damages. In this review, we consider the pros and cons of this new paradigm and suggest a new system where the manufacturing process can be decentralized to provide more targeted vaccines to meet the urgent needs of protection in case of a rampaging epidemic.

First robotic pylorus preserving pancreaticoduodenectomy for Frantz's tumour in an adolescent girl.

Solid pseudopapillary tumour (SPT) is one of the uncommon benign cystic neoplasms of pancreas occurring predominantly in young females. Being benign in nature, surgical resection is the treatment of choice with excellent 5-year survival. A 14-year-old girl presented with pain abdomen for 1 week. On evaluation, she was found to have a large SPT involving head and uncinate process of Pancreas She underwent robotic pylorus preserving pancreaticoduodenectomy (R-PPPD) with da Vinci® Si Robotic System (Intuitive Surgical, Sunnyvale, CA, USA). The total operating time was 480 min. Her postoperative recovery was uneventful and she was discharged on postoperative day 6. In the era of minimally invasive surgery, robotic pancreatic resection and reconstruction are becoming more acceptable. Although the operating domain is small in younger age group, the precise movement of robotic arm and high quality magnified three-dimensional view allows the surgeons to perform PPPD on younger patients also. Young female patients suffering from SPTs can electively undergo R-PPPD with minimal morbidity and mortality. R-PPPD can become the treatment of choice for SPTs involving pancreatic head region even in paediatric and adolescent age group.

A case report of modified laparoscopic keyhole plus repair for parastomal hernia following ileal conduit.

Parastomal hernia is one of the most common but challenging complication after stoma formation. Modified Sugarbaker technique is the recommended procedure for repair parastomal hernia, however, keyhole repair technique had also been used in certain instances. In cases of parastomal hernia following ileal conduit procedure, the Sugarbaker technique is been described, although with associated theoretical risk of conduit failure. We are reporting a case of post-radical cystectomy with ileal conduit presented with symptomatic large parastomal hernia. Laparoscopic modified keyhole plus repair has been done successfully in this patient with no recurrence in 2 years of follow-up. The purpose of our case report is to describe our novel modification of the laparoscopic keyhole technique which can be a feasible and acceptable alternative surgical method in these types of patients.