NHA1 is a cation/proton antiporter essential for the water-conserving functions of the rectal complex in Tribolium castaneum.

More than half of all extant metazoan species on earth are insects. The evolutionary success of insects is linked with their ability to osmoregulate, suggesting that they have evolved unique physiological mechanisms to maintain water balance. In beetles (Coleoptera)-the largest group of insects-a specialized rectal ("cryptonephridial") complex has evolved that recovers water from the rectum destined for excretion and recycles it back to the body. However, the molecular mechanisms underpinning the remarkable water-conserving functions of this system are unknown. Here, we introduce a transcriptomic resource, BeetleAtlas.org, for the exceptionally desiccation-tolerant red flour beetle Tribolium castaneum, and demonstrate its utility by identifying a cation/H+ antiporter (NHA1) that is enriched and functionally significant in the Tribolium rectal complex. NHA1 localizes exclusively to a specialized cell type, the leptophragmata, in the distal region of the Malpighian tubules associated with the rectal complex. Computational modeling and electrophysiological characterization in Xenopus oocytes show that NHA1 acts as an electroneutral K+/H+ antiporter. Furthermore, genetic silencing of Nha1 dramatically increases excretory water loss and reduces organismal survival during desiccation stress, implying that NHA1 activity is essential for maintaining systemic water balance. Finally, we show that Tiptop, a conserved transcription factor, regulates NHA1 expression in leptophragmata and controls leptophragmata maturation, illuminating the developmental mechanism that establishes the functions of this cell. Together, our work provides insights into the molecular architecture underpinning the function of one of the most powerful water-conserving mechanisms in nature, the beetle rectal complex.

Epigenetic modulation by targeting bromodomain containing protein 9 (BRD9): Its therapeutic potential and selective inhibition.

The bromodomain-containing protein 9, a component of the SWI/SNF chromatin remodeling complex, functions as an 'epigenetic reader' selectively recognizing acetyl-lysine marks. It regulates chromatin structure and gene expression by recruitment of acetylated transcriptional regulators and by modulating the function of remodeling complexes. Recent data suggests that BRD9 plays an important role in regulating cellular growth and it has been suggested to drive progression of several malignant diseases such as cervical cancer, and acute myeloid leukemia. Its role in tumorigenesis suggests that selective BRD9 inhibitors may have therapeutic value in cancer therapy. Currently, there has been increasing interest in developing small molecules that can specifically target BRD9 or the closely related bromodomain protein BRD7. Available chemical probes will help to clarify biological functions of BRD9 and its potential for cancer therapy. Since the report of the first BRD9 inhibitor LP99 in 2015, numerous inhibitors have been developed. In this review, we summarized the biological roles of BRD9, structural details and the progress made in the development of BRD9 inhibitors.

Synthesis of 2-Aminopyrimidine Derivatives and Their Evaluation as ß-Glucuronidase Inhibitors: In Vitro and In Silico Studies.

Currently the discovery and development of potent ß-glucuronidase inhibitors is an active area of research due to the observation that increased activity of this enzyme is associated with many pathological conditions, such as colon cancer, renal diseases, and infections of the urinary tract. In this study, twenty-seven 2-aminopyrimidine derivatives 1-27 were synthesized by fusion of 2-amino-4,6-dichloropyrimidine with a variety of amines in the presence of triethylamine without using any solvent and catalyst, in good to excellent yields. All synthesized compounds were characterized by EI-MS, HREI-MS and NMR spectroscopy. Compounds 1-27 were then evaluated for their ß-glucuronidase inhibitory activity, and among them, compound 24 (IC50 = 2.8 ± 0.10 µM) showed an activity much superior to standard D-saccharic acid 1,4-lactone (IC50 = 45.75 ± 2.16 µM). To predict the binding mode of the substrate and ß-glucuronidase, in silico study was performed. Conclusively, this study has identified a potent ß-glucuronidase inhibitor that deserves to be further studied for the development of pharmaceutical products.

Structure and ligand-based drug discovery of IL-4 inhibitors via interaction-energy-based learning approaches.

Interleukin-4 (IL-4), an anti-inflammatory cytokine plays significant in the development of various diseases especially asthmatic allergies. Previous structural and functional studies of IL-4 with its receptor bring forth different types of inhibitors to block their interaction but each of them failed in clinical trials. Since, no synthetic molecules have been identified against IL-4, so far. Therefore, 21 in-house tested IL-4 inhibitors were blindly docked over the entire surface of IL-4 to predict a suitable and druggable binding site as the crystal structure of IL-4 protein in complex with ligand has not been reported yet. After binding site prediction, both ligand-based and structure-based pharmacophore were generated to screen three ZINC libraries (24.5 M) i.e. purchasable, natural product and natural derivative. A total 5,800 top-scored compounds were further subjected towards score-based screening to find the potential leads. Following protein-ligand interaction fingerprints (PLIF) and molecular visualization of selected hits, six top-scored compounds (five from purchasable and one from natural product library) were further moved towards their stability dynamics, followed by their absolute binding free energy and residue-based energy decomposition calculation by MM-GBSA method. These efforts help us to reveal the key factors responsible for ligand binding that might help to improve the binding and stability of these newly discovered hits by structural modifications.Communicated by Freddie R. Salsbury.

Mechanistic insights into the inhibitory activity of FDA approved ivermectin against SARS-CoV-2: old drug with new implications.

The novel corona virus (Covid-19) has become a great challenge worldwide since 2019, as no drug has been reported yet. Different clinical trials are still under way. Among them is Ivermectin (IVM), an FDA approved drug which was recently reported as a successful candidate to reduce SARS-CoV-2 viral load by inhibiting Importin-α1 (IMP-α1) protein which subsequently affects nuclear transport of viral proteins but its basic binding mode and inhibitory mechanism is unknown. Therefore, we aimed to explore the inhibitory mechanism and binding mode of IVM with IMP-α1 via different computational methods. Initially, comparative docking of IVM was performed against two different binding sites (Nuclear Localization Signal (NLS) major and minor sites) of IMP-α1 to predict the probable binding mode of IVM. Then, classical MD simulation was performed (IVM/NLS-Major site and IVM/NLS-Minor site), to predict its comparative stability dynamics and probable inhibitory mechanism. The stability dynamics and biophysical analysis of both sites highlighted the stable binding of IVM within NLS-Minor site by establishing and maintaining more hydrophobic contacts with crucial residues, required for IMP-α1 inhibition which were not observed in NLS-major site. Altogether, these results recommended the worth of IVM as a possible drug to limit the SARS-CoV-2 viral load and consequently reduces its progression.Communicated by Ramaswamy H. Sarma.

Pregnant women perceptions regarding their husbands and in-laws' support during pregnancy: a qualitative study.

INTRODUCTION: pregnancy is a stressful condition during which women require family and in-laws´ support. This study was aimed to explore the women´s perceptions regarding their husband and in-law´s support during pregnancy. METHODS: by using qualitative exploratory design ten pregnant women in third trimester of pregnancy and living in joint family system were recruited through purposive sampling technique from a village of district Nowshehra, Khyber Pakhtunkhwa, Pakistan. Approval for conducting this study was obtained from Ethics Review Committee of Khyber Medical University. Data were collected from the recruited participants through face to face in-depth interviews. Data were analyzed through thematic analysis. One hundred open codes were generated from the data. Through axial coding, extra and unnecessary codes were omitted and then eleven categories were identified from open codes. RESULTS: the identified categories were kept under three salient themes of lack of comprehensive support mechanism, physical and mental strain, and barriers to antenatal services. Perceived support of husbands and in-laws, needs and barriers to maternal and child health were discussed by the participants. CONCLUSION: the study findings suggest that family relationship quality might not be improved by taking interventions i.e. making policies only but the incorporation of health professionals´ support with family member´s behavior can improve maternal health.

Exploiting Dengue Virus Protease as a Therapeutic Target: Current Status, Challenges and Future Avenues.

Dengue, the oldest and the most prevalent mosquito-borne illness, is caused by the dengue virus (DENV), from the family of Flaviviridae. It infects approximately 400 million individuals per annum, with approximately half of the global population residing in high-risk areas. The factors attributed to the geographic expansion of dengue, include urbanization, population density, modern means of transportation, international travels, habit modification, climate change, virus genetics, vector capacity, and poor vector control. Despite the significant progress made in the past against dengue, no effective antiviral therapy is currently available. Among the structural and non-structural proteins encoded by DENV genome, the NS2B-NS3 protease complex is amongst the extensively studied targets for the development of antiviral therapeutics owing to its multiple roles in virus life cycle. Furthermore, protease inhibitors were found to be successful in Hepatitis C Virus (HCV) and Human Immunodeficiency Virus (HIV). Likewise, several peptidic, peptide derived/peptidomimetic, and small molecules inhibitors have been identified as DENV protease inhibitors. Unfortunately, none of them have resulted in a clinically approved drug. Considering all the abovementioned facts, this review descriptively explains the molecular mechanism and therapeutic potential of DENV protease along with an up to date information on various competitive inhibitors reported against DENV protease. This review might be helpful for the researchers working in this area to understand the critical aspects of DENV protease that will help them develop effective and novel inhibitors against DENV to protect lives of millions of people worldwide.

Assessment of hospitalized patients awareness of their rights: a cross-sectional descriptive study.

INTRODUCTION: in this modern era and in the speedily changing atmosphere of health care, health care practice and patients has affected by many factors. These days, in many states the patient rights has become the pivot of the national attention in medical culture. Awareness of health rights is important to achieve the best level of health care. The study was aimed to assess the awareness of hospitalized patients about the rights. METHODS: a cross-sectional descriptive study was conducted in different wards of public sector tertiary care hospitals of Peshawar, over four months duration from February to May 2018. Hospitalized patients who were being admitted for at least two days from 17 to 70 years of age were included. Before asking the patients to answer the questionnaire, consent from the patients was acquired. A self-administered semi-structured questionnaire was adopted. Data analysis was done through SPSS version 22. RESULTS: among 200 participants 46% were male and 54% were female, 35.5% were from urban and 64.5% were from rural areas. Patients were less aware of their individual rights, like 65.5% of clients were not cognizant of the patients special bill of rights while 59% were aware to receive non-discriminatory and timely health services. CONCLUSION: most of the clients were not conscious of their individual health rights. About half of the sample knew that the patients will receive respectful care and the patients will receive care in clean and medically safe environment.

Late Diagnosis of Paracetamol Poisoning is Always Lethal in Young Adult.

Acetaminophen has a remarkable safety profile when prescribed in proper therapeutic doses, but hepatotoxicity can occur when misused or after an overdose. The principal toxic metabolite of acetaminophen is N-acetyl-p-benzoquinone imine (NAPQI). Toxicity should be considered in all suspicious cases because of the ubiquitous and initially asymptomatic nature of acetaminophen intoxication. A case of 29-year male soldier is discussed, who presented with pain in abdomen, vomiting and jaundice of sudden onset. The diagnosis of ischemic liver damage was made at initial presentation. Raised liver function tests and elevated prothrombin time were the first indication to this condition, which were proven by detection of acetaminophen in blood and urine by liquid chromatography-mass spectrometry. Further supportive evidence of hepatic necrosis was provided by an ultrasound abdomen, giving the final diagnosis of acetaminophen poisoning causing drug-induced liver injury. Key Words: Acetaminophen poisoning, Drug-induced liver injury, Fulminant hepatic failure, N-acetylcysteine, N-acetyl-p-benzoquinone imine (NAPQI).

Characterization of cryptic allosteric site at IL-4Rα: New paradigm towards IL-4/IL-4R inhibition.

Interleukin-4(IL-4), an anti-inflammatory cytokine, plays significant role in pathogenesis of various diseases such as asthma, tumors, and HIV infections. These responses are mediated by expression of IL-4R (receptor) on various hematopoietic and non-hematopoietic cells surfaces. To date, the X-ray crystal structure of unbound (i.e. free) IL-4R is not reported which hampers active research on the molecular interaction mechanism between IL-4 and IL-4R. To investigate the missing gaps about stable binding mode of IL-4 and drug-ability of IL-4R active site, modelling and molecular dynamics (MD) simulation of IL-4/IL-4R complex was performed. Drug-ability of the target protein changed after modelling the loop region near C-terminal of IL-4R protein. This led to the identification of a novel druggable site other than the reported interfacial site. Our analysis showed that the modelled residues Ser111 and Ser164-Lys167 are part of newly discovered allosteric site, which underwent major fluctuation after association with its ligand protein (IL-4). The results indicated possible role of this cryptic allosteric site in IL-4/IL-4R signaling pathway that might help us to block IL-4/IL-4R association to prevent various allergic and malignant diseases.

2-Oxo-1,2,3,4-tetrahydropyrimidines Ethyl Esters as Potent ß- Glucuronidase Inhibitors: One-pot Synthesis, In vitro and In silico Studies.

BACKGROUND: Glucuronidation is essential for the metabolism and excretion of toxic substances. ß-Glucuronidase enzyme slows down the process of glucuronidation, and thus plays an important role in the on-set of colorectal carcinoma, and many other diseases. Inhibition of ß- glucuronidase activity is thus identified as an important approach for the treatment of several diseases. OBJECTIVE: Current study was aimed to synthesize a library of 2-oxo-1,2,3,4-tetrahydropyrimidine and to evaluate their ß-glucuronidase inhibitory activity, and their mode of enzyme inhibition. METHOD: We synthesized a series of 2-oxo-1,2,3,4-tetrahydropyrimidines 1-25 by fusing urea, ethyl acetoacetate, and a variety of aldehydes using copper nitrate trihydrate as catalyst. All synthesized compounds were evaluated for their in vitro ß-glucuronidase inhibitory activity. In addition, molecular docking studies were also performed by using MOE docking tools. RESULTS: Eighteen compounds showed inhibitory activity better than the standard D-saccharic acid 1,4-lactone, a well known ß-glucuronidase inhibitor (IC50 = 45.75 ± 2.16 µM). Compound 20 (IC50 = 1.36 ± 0.03 µM) showed an excellent inhibitory activity, thirty-five folds superior to the standard. Docking results highlighted the role of various chemical moieties at different positions on 2- oxo-1,2,3,4-tetrahydropyrimidine skeleton in enzyme inhibitory activity. CONCLUSION: This study has identified a class of potent ß-glucuronidase inhibitors with the potential to be investigated further.

Computational and biological characterization of fusion proteins of two insecticidal proteins for control of insect pests.

Sucking pests pose a serious agricultural challenge, as available transgenic technologies such as Bacillus thuringiensis crystal toxins (Bt) are not effective against them. One approach is to produce fusion protein toxins for the control of these pests. Two protein toxins, Hvt (ω-atracotoxin from Hadronyche versuta) and onion leaf lectin, were translationally fused to evaluate the negative effects of fusion proteins on Phenacoccus solenopsis (mealybug), a phloem-feeding insect pest. Hvt was cloned both N-terminally (HL) and then C-terminally (LH) in the fusion protein constructs, which were expressed transiently in Nicotiana tabacum using a Potato Virus X (PVX) vector. The HL fusion protein was found to be more effective against P. solenopsis, with an 83% mortality rate, as compared to the LH protein, which caused 65% mortality. Hvt and lectin alone caused 42% and 45%, respectively, under the same conditions. Computational studies of both fusion proteins showed that the HL protein is more stable than the LH protein. Together, these results demonstrate that translational fusion of two insecticidal proteins improved the insecticidal activity relative to each protein individually and could be expressed in transgenic plants for effective control of sucking pests.

Synthesis, antimicrobial activity, pharmacophore modeling and molecular docking studies of new pyrazole-dimedone hybrid architectures.

BACKGROUND: Design and synthesis of pyrazole-dimedone derivatives were described by one-pot multicomponent reaction as new antimicrobial agents. These new molecular framework were synthesized in high yields with a broad substrate scope under benign conditions mediated by diethylamine (NHEt2). The molecular structures of the synthesized compounds were assigned based on different spectroscopic techniques (1H-NMR, 13C-NMR, IR, MS, and CHN). RESULTS: The synthesized compounds were evaluated for their antibacterial and antifungal activities against S. aureus ATCC 29213, E. faecalis ATCC29212, B. subtilis ATCC 10400, and C. albicans ATCC 2091 using agar Cup plate method. Compound 4b exhibited the best activity against B. subtilis and E. faecalis with MIC = 16 µg/L. Compounds 4e and 4l exhibited the best activity against S. aureus with MIC = 16 µg/L. Compound 4k exhibited the best activity against B. subtilis with MIC = 8 µg/L. Compounds 4o was the most active compounds against C. albicans with MIC = 4 µg/L. CONCLUSION: In-silico predictions were utilized to investigate the structure activity relationship of all the newly synthesized antimicrobial compounds. In this regard, a ligand-based pharmacophore model was developed highlighting the key features required for general antimicrobial activity. While the molecular docking was carried out to predict the most probable inhibition and binding mechanisms of these antibacterial and antifungal agents using the MOE docking suite against few reported target proteins.

Interleukin-4 receptor signaling and its binding mechanism: A therapeutic insight from inhibitors tool box.

Studies on Interlukin-4 (IL-4) disclosed great deal of information about its various physiological and pathological roles. All these roles depend upon its interaction and signaling through either type-I (IL-4Rα/common γ-chain) or type-II (IL-4Rα/IL-13Rα) receptors. Another cytokine, IL-13, shares some of the functions of IL-4, because both cytokines use a common receptor subunit, IL-4Rα. Here in this review, we discuss the structural details of IL-4 and IL-4Rα subunit and the structural similarities between IL-4 and IL-13. We also describe detailed chemistry of type-I and type-II receptor complexes and their signaling pathways. Furthermore, we elaborate the strength of type-II hetero dimer signals in response to IL-4 and IL-13. These cytokines are prime players in pathogenesis of allergic asthma, allergic hypersensitivity, different cancers, and HIV infection. Recent advances in the structural and binding chemistry of these cytokines various types of inhibitors were designed to block the interaction of IL-4 and IL-13 with their receptor, including several IL-4 mutant analogs and IL-4 antagonistic antibodies. Moreover, different targeted immunotoxins, which is a fusion of cytokine protein with a toxin or suicidal gene, are the new class of inhibitors to prevent cancer progression. In addition few small molecular inhibitors such as flavonoids have also been developed which are capable of binding with high affinity to IL-4Rα and, therefore, can be very effective in blocking IL-4-mediated responses.

Synthesis, bioactivity, molecular docking and POM analyses of novel substituted thieno[2,3-b]thiophenes and related congeners.

Several series of novel substituted thienothiophene derivatives were synthesized by reacting the synthone 1 with different reagents. The newly synthesized compounds were characterized by means of different spectroscopic methods such as IR, NMR, mass spectrometry and by elemental analyses. The new compounds displayed significant activity against both Gram-positive and Gram negative bacteria, in addition to fungi. Molecular docking and POM analyses show the crucial role and impact of substituents on bioactivity and indicate the unfavorable structural parameters in actual drug design: more substitution doesn't guaranty more efficiency in bioactivity.