Challenges in Cervical Cancer Prevention: Real-World Scenario in India.

Satinder KaurIn spite of global falling trends, cervical cancer remains a major healthcare challenge for India, South Asia Association for Regional Cooperation region, and other low- and middle-income countries. Our survey was to document the real-world challenges that still exist in India. A total of 316 eligible and complete responses to the 21 questions were analyzed. Screening of mothers and vaccinating their daughters was considered as the most important strategy to prevent cervical cancer by 65.8% (208/316). Screening was offered to all asymptomatic eligible females by 79% (250/316). Improvement in screening rates requires promoting the national program (67.7%; 214/316), strengthening existing infrastructure (62%; 196/316), regular training of primary healthcare workers (57.6%; 182/316), and increasing awareness among schools and colleges (57.9%; 183/316). Almost all responders (93%; 294/316) wanted to have human papillomavirus (HPV) vaccination included in the national immunization schedule. Cost of vaccine was considered a major roadblock. If it became available at INR 250 per dose, 96.8% (306/316) respondents would recommend it for all eligible patients. With the impending availability of this indigenous tetravalent HPV vaccine jointly produced by Department of Biotechnology, Govt of India and Serum Institute of India, the war against cervical cancer just got easier.

Molecular and cellular paradigms of multidrug resistance in cancer.

BACKGROUND: The acquisition of resistance to chemotherapy is a major hurdle in the successful application of cancer therapy. Several anticancer approaches, including chemotherapies, radiotherapy, surgery and targeted therapies are being employed for the treatment of cancer. However, cancer cells reprogram themselves in multiple ways to evade the effect of these therapies, and over a period of time, the drug becomes inactive due to the development of multi-drug resistance (MDR). MDR is a complex phenomenon where malignant cells become insensitive to anticancer drugs and attain the ability to survive even after several exposures of anticancer drugs. In this review, we have discussed the molecular and cellular paradigms of multidrug resistance in cancer. RECENT FINDINGS: An Extensive research in cancer biology revealed that drug resistance in cancer is the result of perpetuated intracellular and extracellular mechanisms such as drug efflux, drug inactivation, drug target alteration, oncogenic mutations, altered DNA damage repair mechanism, inhibition of programmed cell death signaling, metabolic reprogramming, epithelial mesenchymal transition (EMT), inherent cell heterogeneity, epigenetic changes, redox imbalance, or any combination of these mechanisms. An inevitable cross-link between inflammation and drug resistance has been discussed. This review provided insight molecular mechanism to understand the vulnerabilities of cancer cells to develop drug resistance. CONCLUSION: MDR is an outcome of interplays between multiple intricate pathways responsible for the inactivation of drug and development of resistance. MDR is a major obstacle in regimens of successful application of anti-cancer therapy. An improved understanding of the molecular mechanism of multi drug resistance and cellular reprogramming can provide a promising opportunity to combat drug resistance in cancer and intensify anti-cancer therapy for the upcoming future.

Molecular insights of NADPH oxidases and its pathological consequences.

Reactive oxygen species (ROS), formed by the partial reduction of oxygen, were for a long time considered to be a byproduct of cellular metabolism. Since, increase in cellular levels of ROS results in oxidative stress leading to damage of nucleic acids, proteins, and lipids resulting in numerous pathological conditions; ROS was considered a bane for aerobic species. Hence, the discovery of NADPH oxidases (NOX), an enzyme family that specifically generates ROS as its prime product came as a surprise to redox biologists. NOX family proteins participate in various cellular functions including cell proliferation and differentiation, regulation of genes and protein expression, apoptosis, and host defence immunological response. Balanced expression and activation of NOX with subsequent production of ROS are critically important to regulate various genes and proteins to maintain homeostasis of the cell. However, dysregulation of NOX activation leading to enhanced ROS levels is associated with various pathophysiologies including diabetes, cardiovascular diseases, neurodegenerative diseases, ageing, atherosclerosis, and cancer. Although our current knowledge on NOX signifies its importance in the normal functioning of various cellular pathways; yet the choice of ROS producing enzymes which can tip the scale from homeostasis toward damage, as mediators of biological functions remain an oddity. Though the role of NOX in maintaining normal cellular functions is now deemed essential, yet its dysregulation leading to catastrophic events cannot be denied. Hence, this review focuses on the involvement of NOX enzymes in various pathological conditions imploring them as possible targets for therapies. SIGNIFICANCE OF THE STUDY: The NOXs are multi-subunit enzymes that generate ROS as a prime product. NOX generated ROS are usually regulated by various molecular factors and play a vital role in different physiological processes. The dysregulation of NOX activity is associated with pathological consequences. Recently, the dynamic proximity of NOX enzymes with different molecular signatures of pathologies has been studied extensively. It is essential to identify the precise role of NOX machinery in its niche during the progression of pathology. Although inhibition of NOX could be a promising approach for therapeutic interventions, it is critical to expand the current understanding of NOX's dynamicity and shed light on their molecular partners and regulators.

Assay in serum of exchangeable copper and total copper using inductively coupled plasma mass spectrometry (ICP-MS): development, optimisation and evaluation of a routine procedure.

The assay in serum of non-caeruloplasmin copper, as exchangeable copper after complexation with EDTA (ExCu) and total copper has been evaluated and compared in patients with varying c-reactive protein(CRP). Measurement of ExCu and total copper, range 0.2-47.2 µmol/L, was developed using ICP-MS. The chelating agents EDTA and TEPA were compared over 0.0-10 g/L after incubation with serum for 60 mins followed by ultrafiltration with Amicon 10 kDa filter. The assay for ExCu was optimised with EDTA 3 g/L (8.1 mmol/L) maintained at pH 7.0-8.0 before ultrafiltration. TEPA was not as selective in chelation of copper. Patients n = 82 were studied in relation to changes in inflammatory marker CRP and a group of patients n = 37 with normal CRP. The ExCu assay gave excellent recoveries (94-102 % but poor recovery for free uncomplexed copper), good repeatability, limit of quantitation 0.19 µmol/l with a provisional reference range 0.48 to 1.63 µmol/L (n = 37 patients). The range for relative exchangeable copper (exchangeable copper divided by total serum copper) was 2.49 to 9.96 %. ExCu was elevated in conditions with increased CRP greater than 100 mg/L suggesting an effect of inflammation on the free copper fraction. A reliable and reproducible assay for ExCu and total copper has been developed. The upregulated inflammatory state increases the ExCu suggesting excess free copper.

Structural insights into pharmacophore-assisted in silico identification of protein-protein interaction inhibitors for inhibition of human toll-like receptor 4 - myeloid differentiation factor-2 (hTLR4-MD-2) complex.

Toll-like receptor 4 (TLR4) is a member of Toll-Like Receptors (TLRs) family that serves as a receptor for bacterial lipopolysaccharide (LPS). TLR4 alone cannot recognize LPS without aid of co-receptor myeloid differentiation factor-2 (MD-2). Binding of LPS with TLR4 forms a LPS-TLR4-MD-2 complex and directs downstream signaling for activation of immune response, inflammation and NF-κB activation. Activation of TLR4 signaling is associated with various pathophysiological consequences. Therefore, targeting protein-protein interaction (PPI) in TLR4-MD-2 complex formation could be an attractive therapeutic approach for targeting inflammatory disorders. The aim of present study was directed to identify small molecule PPI inhibitors (SMPPIIs) using pharmacophore mapping-based approach of computational drug discovery. Here, we had retrieved the information about the hot spot residues and their pharmacophoric features at both primary (TLR4-MD-2) and dimerization (MD-2-TLR4*) protein-protein interaction interfaces in TLR4-MD-2 homo-dimer complex using in silico methods. Promising candidates were identified after virtual screening, which may restrict TLR4-MD-2 protein-protein interaction. In silico off-target profiling over the virtually screened compounds revealed other possible molecular targets. Two of the virtually screened compounds (C11 and C15) were predicted to have an inhibitory concentration in µM range after HYDE assessment. Molecular dynamics simulation study performed for these two compounds in complex with target protein confirms the stability of the complex. After virtual high throughput screening we found selective hTLR4-MD-2 inhibitors, which may have therapeutic potential to target chronic inflammatory diseases.

Human Toll-Like Receptor 4 (hTLR4): Structural and functional dynamics in cancer.

Toll-Like Receptors (TLRs) recognize a wide variety of pathogen-associated molecular patterns and are centrally involved in the initiation of the innate and adaptive immune responses. Extensive analysis of TLRs has shown specificity in terms of ligand recognition, expression and cellular localization in different cell types and tissues, and most importantly, its role in the pathogenesis of multiple chronic inflammatory diseases. In recent past extensive investigations showed that many TLRs are profoundly expressed in various types of cancers. This review is emphasized on human TLR4 structural and functional dynamics in cancer. The review was intended to explore the present understanding of the involvement of hTLR4 in different types of cancer and different danger signals that can affect the expression and function of TLR4 in both normal and cancer cells. Dual role of TLR4 in cancer has also been discussed along with therapeutic targeting, cellular response via signaling and the possible conformational changes that occur in response to agonist and antagonist. This review provides a comprehensive resource for designing and discovery of novel TLR4 ligands for therapeutic intervention.

Endocrine abnormalities in lithium toxicity.

Lithium toxicity can manifest as a variety of biochemical -abnormalities. This case report describes a patient -presenting to the emergency department with neuropsychiatric -symptoms on a background of bipolar disorder, for which she was prescribed lithium for 26 years previously. Cases of lithium toxicity are rare but can be severe and this case report -demonstrates to clinicians that they must be thorough in investigating patients with lithium toxicity, as there are many potential abnormalities that can manifest concurrently.

Optimisation of intravenous fluid prescribing: framework for changing practice through education and audits.

INTRODUCTION: Intravenous fluids are one of the most commonly prescribed drugs in the hospital setting and yet the practice continues to fall short of National Institute for Health and Care Excellence (NICE) guidelines, with significant gaps in staff knowledge exposing patients to heightened morbidity and mortality. AIM: Following the 2013 publishing of updated NICE guidelines on intravenous fluid prescribing, an intravenous fluid team was formed within the Royal Liverpool University Hospital (RLUH). Their role has been (and continues to be) to overhaul the culture of suboptimal intravenous fluid prescribing within the hospital and, ultimately, to improve patient outcomes. A framework to engender this change has been developed and is offered as an example to other Trusts within which improvement of guideline-compliant intravenous fluid prescribing remains stagnant. METHOD: There have been three principal stages of the project to this point which are best demonstrated in a chronological manner. The period of 2010-2014 allowed for assessment of the issue of intravenous fluid prescribing and analysis of its causes through serial audits and a staff-wide survey. From 2015, there has been implementation of several measures (educative, managerial, administrative and technological) within the hospital to foster reproducible and positive change with regards to intravenous fluid prescribing. Finally, between 2016 and 2017, three cycles of a rolling audit based on NICE guidelines have been completed to allow measurement of improvement in intravenous fluid prescribing practice. CONCLUSION: Results have demonstrated a significant improvement in the appropriateness of the intravenous maintenance and replacement fluids prescribed in the hospital since the March 2016 audit. Moreover, a 29-fold increase has been observed in the use of 4% dextrose/0.18% sodium chloride as maintenance fluid (gold standard as per NICE guidelines) since the staff-wide survey of 2015. Despite progress however, adherence to NICE guidelines remains below the recommended 100% and therefore further work remains to be done.

Management of a severe case of Gitelman syndrome with poor response to standard treatment.

Gitelman syndrome is an autosomal recessive distal renal tubular disorder caused by defective sodium chloride transporters. Biochemically, it presents with hypokalaemic metabolic alkalosis, hypomagnesaemia and hypocalciuria. It is usually managed with oral potassium supplements and potassium-sparing diuretics. We report a case of a 28-year-old woman whose condition worsened during pregnancy; she became resistant to standard management after delivery of her second child. She was managed in a specialist metabolic clinic through a comprehensive approach including perseverance with oral potassium supplement, weekly intravenous potassium and magnesium infusion, correction of vitamin D level and the offering of appropriate dietary advice; this controlled the patient's symptoms and prevented repeated hospital admissions. In this case report, we illustrate a patient's presentation and diagnosis with Gitelman syndrome, discuss triggers of exacerbation, review the relevant literature in terms of differential diagnoses and provide practical advice on the management of difficult cases in a specialist clinic.

Ligand based virtual screening to find novel inhibitors against plant toxin Ricin by using the ZINC database.

Ricin is known as a potent toxin against animals. It consists of two chains, Ricin Toxin A (RTA) and Ricin Toxin B (RTB). The toxic effect is known to be caused by RTA. Inhibitors for RTA with less efficiency have been reported. Hence, it is of interest to identify new inhibitors. Virtual screening methods (computer aided drug designing) to find similar molecules in drug database were used for screening new inhibitors against RTA. We used the structure of RTA in complex with Pteroic acid (PDB code: 1BR6) as target molecule. Ligand based virtual screening approach was used in which the known inhibitory molecule Pteroic acid (PTA) served as a template to identify similar ligands from the ZINC database. These ligands were docked inside the binding pocket of RTA by using the MVD (Molegro Virtual Docker). This approach successfully identified six novel compounds. These docked ligands interacted with Asn78, Ala79, Val81, Gly121 and Ser176 amino acids, which are key residues of the RTA active site. Three compounds in particular, ZINC05156321 (6, 7 diphenylpteridin-4-ol), ZINC05156324 (6, 7-bis (3-fluorophenyl) pteridin-4-ol) and ZINC08555900 (6, 7-bis (4-fluorophenyl)-1H-pteridin-4-one), showed higher binding affinity in comparison to PTA, with high interaction energy, better space fitting and electrostatic interactions. These molecules should be tested for in vitro and in vivo activities in future for consideration as effective inhibitors.

Oxidative stress and role of antioxidant supplementation in critical illness.

Sepsis or systemic inflammatory response (SIRS) to infection or to non-infectious stimuli such as trauma, surgery, pancreatitis or ischemia, is an increasingly common cause of morbidity and mortality in patients on intensive therapy unit (ITU). In critically ill patients, this accounts for 10% to 50% of all deaths. Oxidative stress has an important role in the development and manifestations of SIRS. Oxidative stress is an imbalance between the free radical production and the antioxidant defense. In critical illness, overwhelming inflammatory mediator response to infective or non-infective stimuli results in excessive production of free radicals (FR). The action of FR is normally limited by the antioxidant defense system of the body, but in critically ill patients the antioxidant capacity is likely to be compromised. Hence, provision of antioxidants to critically ill patients may help in removing the FR and therefore improving the clinical outcome. However, no study has yet provided conclusive evidence of the beneficial effect of antioxidant supplementation in critically ill patients. The clinical evidence provided so far shows that there are several factors which might determine the efficacy of antioxidant supplementation in critically ill patients. There is a need for large multicentre prospective randomized control trials to assess the effects of different types and doses of antioxidant supplementation in selected groups of patients with different types of critical illness.

Effect of selenium supplementation on biochemical markers and outcome in critically ill patients.

BACKGROUND & AIMS: This study aimed to assess the effect of high dose selenium (Se) supplementation on Se status in blood, oxidative stress, thyroid function and possible effects on requirement for renal replacement therapy (RRT) in severely septic patients admitted to the intensive care unit (ICU). METHODS: This prospective single-centre study was carried out in 40 septic ICU patients who were randomized to high dose Se (Se+ group, N=18 (474, 316, 158 microg/day), each for 3 consecutive days followed by a standard dose of 31.6 microg/day of Se given as sodium selenite whereas the control group (Se-, N=22) received only the standard dose of Se. Plasma Se, glutathione peroxidase (GSH-Px), F2 isoprostanes, thyroid function tests (total T4 and total T3), C-reactive protein (CRP) and red blood cell (RBC) GSH-Px were estimated on day 0, 3, 7, 14. RESULTS: In the Se+ group, plasma Se increased by day 3 and 7 (P<0.0001) and day 14 (P=0.02), plasma GSH-Px increased by day 3 and 7 (P=0.01) as compared to Se- group. There was a significant negative correlation between plasma Se and SOFA (sepsis related organ failure assessment) (r=-0.36, P=0.03) along with low plasma Se and high CRP at the time of admission. Requirement for renal replacement therapy was not significantly different between the groups. CONCLUSION: Although high dose Se supplementation increased plasma Se and GSH-Px activity, it did not reduce oxidative damage or the requirement for RRT. Se levels in blood are influenced by redistribution and severity of illness and therefore should be interpreted with caution.

Markers of oxidative damage, antioxidant status and clinical outcome in critically ill patients.

BACKGROUND: Oxidative stress is a consequence of critical illness, and may have an impact on survival. We studied markers of oxidative damage and antioxidant (AO) protection and compared them with clinical scores and outcome. METHODS: Blood sampling and clinical scoring was carried out on 60 consecutively admitted intensive therapy unit (ITU) patients within 24 h of admission and then every three days of ITU stay. The patients included 30 surgical and 30 medical patients, of whom 46 survived their stay in ITU. Clinical scoring was by Acute Physiology and Chronic Health Evaluation (APACHE) II score, multiple organ dysfunction (MOD) score and sepsis rating. Oxidative damage was assessed by measurement of plasma malondialdehyde (MDA) and F2 isoprostanes (F2 IsoPs). AO protection was assessed by measurement of plasma total AO status, AO gap, ascorbic acid and the enzymes glutathione peroxidase and superoxide dismutase. RESULTS: Both clinical markers, APACHE II and MOD, and oxidative damage markers MDA and F2 IsoPs were significantly higher in non-survivors (NS) than in survivors (S) at the time of admission. Median (interquartile ranges) were (APACHE II), 14[12--17] (S), 20.5[16.7--22.2] (NS),P<0.0001; (MOD), 3.0[2.0--5.0] (S), 8.0[4.7--9.2] (NS), P<0.0005; (MDA, mumol/L), 0.22[0.19--0.27] (S), 0.25[0.20--0.34] (NS), P=0.04 and (F2 IsoPs, pg/mL), 9.7[6.0--9.9] (S), 11.0[9.0--12.0] (NS), P=0.01. Oxidative damage markers reduced (improved) in the survivors but increased in the non-survivors. There was little difference between the groups in AO protection markers. There was a significant positive correlation between MOD and markers of oxidative damage at the time of admission (r=0.40, P=0.003, F2 IsoPs; r=0.28, P=0.035, MDA) and between the oxidative damage markers themselves (r=0.32, P=0.017). CONCLUSION: Increased oxidative stress is associated with poor outcome in critically ill patients, and may be a prognostic indicator. Oxidative damage markers are more useful than AO protection markers in predicting outcome.

Comparison of spectrophotometric and enhanced chemiluminescent assays of serum antioxidant capacity.

BACKGROUND: Over recent years, interest in total antioxidant capacity measurement in biological fluids has increased. A number of assays are now available, and we wished to compare an enhanced chemiluminescence (ECL) method to a spectrophotometric method, the total antioxidant status (TAS) assay. METHODS: Serum urate concentration, ECL and TAS were measured in 34 healthy subjects. Additionally, 10 subjects participated in a two-way, randomised crossover study, and received urate 1000 mg or vitamin C 1000 mg intravenously over 1 h. Serum ECL and TAS were measured at 0, 15, 30, 45, 60, 90 and 120 min after commencing infusion. RESULTS: Baseline measurements were poorly correlated between ECL and TAS assays, and between serum urate concentration and each antioxidant assay. There was good correlation between the change in antioxidant capacity detected by both assays during urate infusion (R=0.79, p<0.001, n=60), but not vitamin C infusion. CONCLUSIONS: ECL and TAS measures of serum antioxidant capacity correlate poorly in a healthy population, although both are sensitive to increases in circulating urate concentrations. Therefore, ECL and TAS appear sensitive to different factors. The comparative strengths and weaknesses of various antioxidant assays should be reviewed.