Search results for: M. tuberculosis inhibitors

Authors: Adebisi Adunola Demehin, Wanlaya Thamnarak, Jaruwan Chatwichien, Chatchakorn Eurtivong, Kiattawee Choowongkomon, Somsak Ruchirawat, Nopporn Thasana

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The epidermal growth factor receptor (EGFR) is a transmembrane glycoprotein involved in cellular signalling processes and, its aberrant activity is crucial in the development of many cancers such as lung cancer. Selaginellaceae are fern allies that have long been used in Chinese traditional medicine to treat various cancer types, especially lung cancer. Biflavonoids, the major secondary metabolites in Selaginellaceae, have numerous pharmacological activities, including anti-cancer and anti-inflammatory. For instance, amentoflavone induces a cytotoxic effect in the human NSCLC cell line via the inhibition of PARP-1. However, to the best of our knowledge, there are no studies on biflavonoids as EGFR inhibitors. Thus, this study aims to investigate the EGFR inhibitory activities of biflavonoids isolated from Selaginella siamensis and Selaginella bryopteris. Amentoflavone, tetrahydroamentoflavone, sciadopitysin, robustaflavone, robustaflavone-4-methylether, delicaflavone, and chrysocauloflavone were isolated from the ethyl-acetate extract of the whole plants. The structures were determined using NMR spectroscopy and mass spectrometry. In vitro study was conducted to evaluate their cytotoxicity against A549, HEPG2, and T47D human cancer cell lines using the MTT assay. In addition, a target-based assay was performed to investigate their EGFR inhibitory activity using the kinase inhibition assay. Finally, a molecular docking study was conducted to predict the binding modes of the compounds. Robustaflavone-4-methylether and delicaflavone showed the best cytotoxic activity on all the cell lines with IC50 (µM) values of 18.9 ± 2.1 and 22.7 ± 3.3 on A549, respectively. Of these biflavonoids, delicaflavone showed the most potent EGFR inhibitory activity with an 84% relative inhibition at 0.02 nM using erlotinib as a positive control. Robustaflavone-4-methylether showed a 78% inhibition at 0.15 nM. The docking scores obtained from the molecular docking study correlated with the kinase inhibition assay. Robustaflavone-4-methylether and delicaflavone had a docking score of 72.0 and 86.5, respectively. The inhibitory activity of delicaflavone seemed to be linked with the C2”=C3” and 3-O-4”’ linkage pattern. Thus, this study suggests that the structural features of these compounds could serve as a basis for developing new EGFR-TK inhibitors.

Keywords: anticancer, biflavonoids, EGFR, molecular docking, Selaginellaceae

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Authors: Harish Rajak, Preeti Patel

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The application of histone deacetylase inhibitors is a well-known strategy in prevention of cancer which shows acceptable preclinical antitumor activity due to its ability of growth inhibition and apoptosis induction of cancer cell. Molecular docking were performed using Histone Deacetylase protein (PDB ID:1t69) and prepared series of hydroxamic acid based HDACIs. On the basis of docking study, it was predicted that compound 1 has significant binding interaction with HDAC protein and three hydrogen bond interactions takes place, which are essential for antitumor activity. On docking, most of the compounds exhibited better glide score values between -8 to -10 which is close to the glide score value of suberoylanilide hydroxamic acid. The pharmacophore hypotheses were developed using e-pharmacophore script and phase module. The 3D-QSAR models provided a good correlation between predicted and actual anticancer activity. Best QSAR model showed Q2 (0.7974), R2 (0.9200) and standard deviation (0.2308). QSAR visualization maps suggest that hydrogen bond acceptor groups at carbonyl group of cap region and hydrophobic groups at ortho, meta, para position of R9 were favorable for HDAC inhibitory activity. We established structure activity correlation using docking, pharmacophore modeling and atom based 3D QSAR model for hydroxamic acid based HDACIs.

Keywords: HDACIs, QSAR, e-pharmacophore, docking, suberoylanilide hydroxamic acid

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Authors: Reena K., Mamatha H. G., Somshekarayya, P. Kumar

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Objectives: The annual proficiency testing of intermediate reference laboratories is conducted by the National Reference Laboratory (NRL) to assess the efficiency of the laboratories to correctly identify Mycobacterium tuberculosis and to determine its drug susceptibility pattern. The proficiency testing results from 2013 to 2017 were analyzed to determine laboratories that were consistent in reporting quality results and those that had difficulty in doing so. Methods: A panel of twenty cultures were sent out to each of these laboratories. The laboratories were expected to grow the cultures in their own laboratories, set up drug susceptibly testing by all the methods they were certified for and report the results within the stipulated time period. The turnaround time for reporting results, specificity, sensitivity positive and negative predictive values and efficiency of the laboratory in identifying the cultures were analyzed. Results: Most of the laboratories had reported their results within the stipulated time period. However, there was enormous delay in reporting results from few of the laboratories. This was mainly due to improper functioning of the biosafety level III laboratory. Only 40% of the laboratories had 100% efficiency in solid culture using Lowenstein Jensen medium. This was expected as a solid culture, and drug susceptibility testing is not used for diagnosing drug resistance. Rapid molecular methods such as Line probe assay and Genexpert are used to determine drug resistance. Automated liquid culture system such as the Mycobacterial growth indicator tube is used to determine prognosis of the patient while on treatment. It was observed that 90% of the laboratories had achieved 100% in the liquid culture method. Almost all laboratories had achieved 100% efficiency in the line probe assay method which is the method of choice for determining drug-resistant tuberculosis. Conclusion: Since the liquid culture and line probe assay technologies are routinely used for the detection of drug-resistant tuberculosis the laboratories exhibited higher level of efficiency as compared to solid culture and drug susceptibility testing which are rarely used. The infrastructure of the laboratory should be maintained properly so that samples can be processed safely and results could be declared on time.

Keywords: annual proficiency testing, drug susceptibility testing, intermediate reference laboratory, national reference laboratory

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Authors: Ncoza Dlova, Tivani Mashamba-Thompson

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Garcinia livingstonei (Clusiaceae) extracts, morelloflavone- 7″- sulphate and sargaol were shown to be effective against hyper-pigmentation through inhibition of tyrosinase enzyme, in vitro . The aim of this study is to elucidate the structural mechanism through which morelloflavone- 7″- sulphate and sargaol binds human tyrosinase. Implementing a homology model to construct a tyrosinase model using the crystal structure of a functional unit from Octopus hemocyanin (PDB: 1JS8) as a reference template enabled us to create a human tyrosinase model. Molecular dynamics and binding free energy calculations were optimized to enable molecular dynamics simulation of the copper dependent inhibitors. Results show the importance of the hydrogen bond formation morelloflavone- 7″- sulphate and sargaol between compound and active site residues. Both complexes demonstrated the metallic coordination between compound and arginine residue as well as copper ions within the active site. The comprehensive molecular insight gained from this study should be vital in understanding the binding mechanism morelloflavone- 7″- sulphate and sargaol. Moreover, these results will assist in the design of novel of metal ion dependent enzyme inhibitors as potential anti-hyper-pigmentation disorder therapies.

Keywords: hyper-pigmentation disorders, dyschromia African skin, morelloflavone- 7″- sulphate, sagoal

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Authors: Richa Sharma, Uma Nahar, Sadhna Sharma, Indu Verma

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Counteracting the deadly pathogen Mycobacterium tuberculosis (M. tb) effectively is still a global challenge. Scrutinizing alternative weapons like antimicrobial peptides to strengthen existing tuberculosis artillery is urgently required. Considering the antimycobacterial potential of Human Beta Defensin 1 (HBD-1) along with isoniazid, the present study was designed to explore the ability of HBD-1 to act against active and dormant M. tb. HBD-1 was screened in silico using antimicrobial peptide prediction servers to identify its short antimicrobial motif. The activity of both HBD-1 and its selected motif (Pep B) was determined at different concentrations against actively growing M. tb in vitro and ex vivo in monocyte derived macrophages (MDMs). Log phase M. tb was grown along with HBD-1 and Pep B for 7 days. M. tb infected MDMs were treated with HBD-1 and Pep B for 72 hours. Thereafter, colony forming unit (CFU) enumeration was performed to determine activity of both peptides against actively growing in vitro and intracellular M. tb. The dormant M. tb models were prepared by following two approaches and treated with different concentrations of HBD-1 and Pep B. Firstly, 20-22 days old M. tbH37Rv was grown in potassium deficient Sauton media for 35 days. The presence of dormant bacilli was confirmed by Nile red staining. Dormant bacilli were further treated with rifampicin, isoniazid, HBD-1 and its motif for 7 days. The effect of both peptides on latent bacilli was assessed by colony forming units (CFU) and most probable number (MPN) enumeration. Secondly, human PBMC granuloma model was prepared by infecting PBMCs seeded on collagen matrix with M. tb(MOI 0.1) for 10 days. Histopathology was done to confirm granuloma formation. The granuloma thus formed was incubated for 72 hours with rifampicin, HBD-1 and Pep B individually. Difference in bacillary load was determined by CFU enumeration. The minimum inhibitory concentrations of HBD-1 and Pep B restricting growth of mycobacteria in vitro were 2μg/ml and 20μg/ml respectively. The intracellular mycobacterial load was reduced significantly by HBD-1 and Pep B at 1μg/ml and 5μg/ml respectively. Nile red positive bacterial population, high MPN/ low CFU count and tolerance to isoniazid, confirmed the formation of potassium deficienybaseddormancy model. HBD-1 (8μg/ml) showed 96% and 99% killing and Pep B (40μg/ml) lowered dormant bacillary load by 68.89% and 92.49% based on CFU and MPN enumeration respectively. Further, H&E stained aggregates of macrophages and lymphocytes, acid fast bacilli surrounded by cellular aggregates and rifampicin resistance, indicated the formation of human granuloma dormancy model. HBD-1 (8μg/ml) led to 81.3% reduction in CFU whereas its motif Pep B (40μg/ml) showed only 54.66% decrease in bacterial load inside granuloma. Thus, the present study indicated that HBD-1 and its motif are effective antimicrobial players against both actively growing and dormant M. tb. They should be further explored to tap their potential to design a powerful weapon for combating tuberculosis.

Keywords: antimicrobial peptides, dormant, human beta defensin 1, tuberculosis

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Authors: Ananya Gupta, Sangeeta Bhaskar

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Mycobacterium indicus pranii (MIP) is a saprophytic mycobacterium. It is a predecessor of M. avium complex (MAC). Whole genome analysis and growth kinetics studies have placed MIP in between pathogenic and non-pathogenic species. It shares significant antigenic repertoire with M. tuberculosis and have unique immunomodulatory properties. MIP provides better protection than BCG against pulmonary tuberculosis in animal models. Immunization with MIP by aerosol route provides significantly higher protection as compared to immunization by subcutaneous (s.c.) route. However, mechanism behind differential protection has not been studied. In this study, using mice model we have evaluated and compared the M.tb specific immune response in lung compartments (airway lumen / lung interstitium) as well as spleen following MIP immunization via nasal (i.n.) and s.c. route. MIP i.n. vaccination resulted in increased seeding of memory T cells (CD4+ and CD8+ T-cells) in the airway lumen. Frequency of CD4+ T cells expressing Th1 migratory marker (CXCR3) and activation marker (CD69) were also high in airway lumen of MIP i.n. group. Significantly high ex vivo secretion of cytokines- IFN-, IL-12, IL-17 and TNF- from cells of airway luminal spaces provides evidence of antigen-specific lung immune response, besides generating systemic immunity comparable to MIP s.c. group. Analysis of T cell response on per cell basis revealed that antigen specific T-cells of MIP i.n. group were functionally superior as higher percentage of these cells simultaneously secreted IFN-gamma, IL-2 and TNF-alpha cytokines as compared to MIP s.c. group. T-cells secreting more than one of the cytokines simultaneously are believed to have robust effector response and crucial for protection, compared with single cytokine secreting T-cells. Adoptive transfer of airway luminal T-cells from MIP i.n. group into trachea of naive B6 mice revealed that MIP induced CD8 T-cells play crucial role in providing long term protection. Thus the study demonstrates that MIP intranasal vaccination induces M.tb specific memory T-cells in the airway lumen that results in an early and robust recall response against M.tb infection.

Keywords: airway lumen, Mycobacterium indicus pranii, Th1 migratory markers, vaccination

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Authors: Jakub Młoźniak, Adam Szymański, Gabriela Stondzik, Dagny Krankowska, Tomasz Mikuła

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Nontuberculous mycobacteria (NTM) are bacterial species that cause diversely manifesting diseases mainly in immunocompromised patients. In people with HIV, NTM infection is an AIDS-defining disease and usually appears when the lymphocyte T CD4 count is below 50 cells/μl. The usage of antiretroviral therapy has decreased the prevalence of NTM among people with HIV, but the disease can still be observed especially among patients with late HIV diagnosis. Common presence in environment, human colonization, clinical similarity with tuberculosis and slow growth on culture makes NTM especially hard to diagnose. The study aimed to analyze the epidemiology and clinical course of NTM among patients with HIV. This study included patients with NTM and HIV admitted to our department between 2017 and 2023. Medical records of patients were analyzed and data on age, sex, median time from HIV diagnosis to identification of NTM infection, median CD4 count at NTM diagnosis, methods of determining NTM infection, type of species of mycobacteria identified, clinical symptoms and treatment course were gathered. Twenty-four patients (20 men, 4 women) with identified NTM were included in this study. Among them, 20 were HIV late presenters. The patients' median age was 40. The main symptoms which patients presented were fever, weight loss and cough. Pulmonary disease confirmed with positive cultures from sputum/bronchoalveolar lavage was present in 18 patients. M. avium was the most common species identified. M. marinum caused disseminated skin lesions in 1 patient. Out of all, 5 people were not treated for NTM caused by lack of symptoms and suspicion of colonization with mycobacterium. Concomitant tuberculosis was present in 6 patients. The median diagnostic time from HIV to NTM infections was 3.5 months. The median CD4 count at NTM identification was 69.5 cells/μl. Median NTM treatment time was 16 months but 7 patients haven’t finished their treatment yet. The most commonly used medications were ethambutol and clarithromycin. Among analyzed patients, 4 of them have died. NTM infections are still an important disease among patients who are HIV late presenters. This disease should be taken into consideration during the differential diagnosis of fever, weight loss and cough in people with HIV with lymphocyte T CD4 count <100 cells/μl. Presence of tuberculosis does not exclude nontuberculous mycobacterium coinfection.

Keywords: mycobacteriosis, HIV, late presenter, epidemiology

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Authors: Katharigatta N. Venugopala, Melendhran Pillay, Bander E. Al-Dhubiab

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Tuberculosis (TB) infection is caused mainly by Mycobacterium tuberculosis (MTB) and it is one of the most threatening and wide spread infectious diseases in the world. Benzothiazole derivatives are found to have diverse chemical reactivity and broad spectrum of pharmacological activity. Some of the important pharmacological activities shown by the benzothiazole analogues are antitumor, anti-inflammatory, antimicrobial, anti-tubercular, anti-leishmanial, anticonvulsant and anti-HIV properties. Keeping all these facts in mind in the present investigation it was envisaged to synthesize a series of novel {2-(benzo[d]-thiazol-2-yl-methoxy)-substitutedaryl}-(substitutedaryl)-methanones (4a-f) and characterize by IR, NMR (1H and 13C), HRMS and single crystal x-ray studies. The title compounds are investigated for in vitro anti-tubercular activity against two TB strains such as H37Rv (ATCC 25177) and MDR-MTB (multi drug resistant MTB resistant to Isoniazid, Rifampicin and Ethambutol) by agar diffusion method. Among the synthesized compounds in the series, test compound {2-(benzo[d]thiazol-2-yl-methoxy)-5-fluorophenyl}-(4-chlorophenyl)-methanone (2c) was found to exhibit significant activity with MICs of 1 µg/mL and 2 µg/mL against H37Rv and MDR-MTB, respectively when compared to standard drugs. Single crystal x-ray studies was used to study intra and intermolecular interactions, including polymorphism behavior of the test compounds, but none of the compounds exhibited polymorphism behavior.

Keywords: benzothiazole analogues, characterization, crystallography, anti-TB activity

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Authors: Mike Wei, Nebu Koshy, Koen van Besien, Giorgio Inghirami, Steven M. Horwitz

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T-cell prolymphocytic leukemia (T-PLL) is a rare hematologic disease characterized by a T-cell phenotype, rapid progression, and poor prognosis with median survival of less than a year. Alemtuzumab-based chemotherapy has increased the rate of complete remissions but these are often short-lived, and allogeneic transplant is considered the only curative therapy. In recent studies, JAK3 activating mutations have been identified in T-cell cancers, with T-PLL having the highest rate of JAK3 mutations (30 – 42%). As such, T-PLL is a model disease for evaluating the utility of JAK3 inhibitors. We present a case of a 64-year-old man with relapsed-refractory T-PLL. He was initially treated with alemtuzumab and obtained complete response and was consolidated with matched unrelated donor stem cell transplant. His disease stayed in remission for approximately 1.5 years before relapse, which was then treated with a clinical trial of romidepsin-lenalidomide (partial responses then progression at 6 months) and later alemtuzumab. Due to complications of myelosuppression and CMV reactivation, his treatment was interrupted leading to disease progression. The doubling time of lymphocyte count was approximately 20 days and over a span of 60 days the lymphocyte count rose from 8 x 109/L to 68 x 109/L. Exon sequencing showed a JAK3 mutation. The patient consented to and was treated with FDA-approved tofacitinib (initially 5 mg BID, increased to 10 mg BID after 15 days of treatment). An initial decrease in lymphocyte count was followed by progression. In vitro treatment of the patient’s cells showed modest effects of tofacitinib and ruxolitinib as single agents, in the range of doxorubicin, but synergy between the agents. After 40 days of treatment with tofacitinib and with a lymphocyte count of 150 x 109/L, ruxolitinib (5mg BID) was added. Over the 60 days since dual inhibition was started, the lymphocyte count has stabilized. The patient has remained completely asymptomatic during treatment with tofacitinib and ruxolitinib. Neutrophil count has remained normal. Platelet count and hemoglobin have however declined from ~50 x109/L to ~30 x109/L and from 11 g/dL to 8.1 g/dL respectively, since the introduction of ruxolitinib. The stabilization in lymphocyte count confirms the clinical activity of JAK inhibitors in T-PLL as suggested by the presence of JAK3 mutations and by in-vitro assays. It also suggests clinical synergy between ruxolitinib and tofacitinib in this setting. Prospective studies of JAK inhibitors in PLL patients with formal dose-finding studies are needed.

Keywords: tofacitinib, ruxolitinib, T-cell prolymphocytic leukemia, JAK3

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Authors: John Hang Leung, Shyh-Yau Wang, Hei-Tung Yip, Fion, Ho Tsung-chin, Agnes LF Chan

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Objectives: Platinum-resistant ovarian cancer has a short overall survival of 9–12 months and limited treatment options. The combination of immunotherapy and targeted therapy appears to be a promising treatment option for patients with ovarian cancer, particularly to patients with platinum-resistant recurrent ovarian cancer (PRrOC). However, there are no direct head-to-head clinical trials comparing their efficacy and toxicity. We, therefore, used a network to directly and indirectly compare seven newer immunotherapies or targeted therapies combined with chemotherapy in platinum-resistant relapsed ovarian cancer, including antibody-drug conjugates, PD-1 (Programmed death-1) and PD-L1 (Programmed death-ligand 1), PARP (Poly ADP-ribose polymerase) inhibitors, TKIs (Tyrosine kinase inhibitors), and antiangiogenic agents. Methods: We searched PubMed (Public/Publisher MEDLINE), EMBASE (Excerpta Medica Database), and the Cochrane Library electronic databases for phase II and III trials involving PRrOC patients treated with immunotherapy or targeted therapy plus chemotherapy. The quality of included trials was assessed using the GRADE method. The primary outcomes compared were progression-free survival, the secondary outcomes were overall survival and safety. Results: Seven randomized controlled trials involving a total of 2058 PRrOC patients were included in this analysis. Bevacizumab plus chemotherapy showed statistically significant differences in PFS (Progression-free survival) but not OS (Overall survival) for all interested targets and immunotherapy regimens; however, according to the heatmap analysis, bevacizumab plus chemotherapy had a statistically significant risk of ≥grade 3 SAEs (Severe adverse effects), particularly hematological severe adverse events (neutropenia, anemia, leukopenia, and thrombocytopenia). Conclusions: Bevacizumab plus chemotherapy resulted in better PFS as compared with all interested regimens for the treatment of PRrOC. However, statistical differences in SAEs as bevacizumab plus chemotherapy is associated with a greater risk for hematological SAE.

Keywords: platinum-resistant recurrent ovarian cancer, network meta-analysis, immune checkpoint inhibitors, target therapy, antiangiogenic agents

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Authors: Recep Keşli, Hayriye Tokay, Cengiz Demir, İsmail Ceyhan

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Objective: In many countries, tuberculosis (TB) is still one of the most important diseases. Tuberculosis is among top 10 causes of death worldwide. The early diagnosis of active tuberculosis still depends on the presence of acid resistant bacilli (ARB) in stained smears. In this study, we aimed to investigate the diagnostic performances of Erlich Ziehl Neelsen (EZN), Kinyoun and two different fluorescent stains. Methods: The specimens were obtained from the patients who applied to Chest Diseases Departments of Ankara Atatürk Chest Diseases and Thoracic Surgery Training and Research Hospital, and Afyon Kocatepe University, ANS Research and Practice Hospital. The study was carried out in the Medical Microbiology Laboratory, School of Medicine, Afyon Kocatepe University. All the non-sterile specimens were homogenized and decontaminated according to the EUCAST instructions. Samples were inoculated onto the Löwenstein-Jensen agars (bio-Merieux Marcy l'Etoile, France) and then incubated at 37˚C, for 40 days. Four smears were prepared from each specimen. Slides were stained with commercial EZN (BD, Sparks, USA), Kinyoun (SALUBRIS Istanbul, Turkey), Auramine (SALUBRIS Istanbul, Turkey) and Rhodamine (SALUBRIS Istanbul, Turkey) kit. While EZN and Kinyoun stainings were examined by light microscope, Auramine and Rhodamine slides were examined by fluorescence microscopy. Results: A total of 158 respiratory system samples (sputum, broncho alveolar lavage fluid…etc) were enrolled into the study. A hundred and two of the samples that processed were found as culture positive. The sensitivity, specificity, positive predictive, and negative predictive values were detected as 100%, 67.5%, 73.5%, and 100% for EZN, 100%, 70.9%, 77.4%, and 100% for Kinyoun, 100%,77.8%, 84.3%, 100% for Auramine, and 100%, 80% , 86.3%, and 100% for Rhodamine respectively. Conclusions: According to our study auramine and rhodamine staining methods showed the best diagnostic performance among the four investigated staining methods. In conclusion, the fluorochrome staining method may be accepted as the most reliable, rapid and useful method for diagnosis of the mycobacterial infections truly.

Keywords: acid resistant bacilli (ARB), auramine, Ehrlich-Ziehl-Neelsen (EZN), Kinyoun, Rhodamine

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Authors: Sharat Chandra, Zilong Wang, Ru-Rong Ji, Andrey Bortsov

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Chronic pain (CP) therapeutic approaches have limited efficacy. As a result, doctors are prescribing opioids for chronic pain, leading to opioid overuse, abuse, and addiction epidemic. Therefore, the development of effective and safe CP drugs remains an unmet medical need. Voltage-gated sodium (Nav) channels act as cardiovascular and neurological disorder’s molecular targets. Nav channels selective inhibitors are hard to design because there are nine closely-related isoforms (Nav1.1-1.9) that share the protein sequence segments. We are targeting the Nav1.7 found in the peripheral nervous system and engaged in the perception of pain. The objective of this project was to screen a 1.5 million compound library for identification of inhibitors for Nav1.7 with analgesic effect. In this study, we designed a protocol for identification of isoform-selective inhibitors of Nav1.7, by utilizing the prior information on isoform-selective antagonists. First, a similarity search was performed; then the identified hits were docked into a binding site on the fourth voltage-sensor domain (VSD4) of Nav1.7. We used the FTrees tool for similarity searching and library generation; the generated library was docked in the VSD4 domain binding site using FlexX and compounds were shortlisted using a FlexX score and SeeSAR hyde scoring. Finally, the top 25 compounds were tested with molecular dynamics simulation (MDS). We reduced our list to 9 compounds based on the MDS root mean square deviation plot and obtained them from a vendor for in vitro and in vivo validation. Whole-cell patch-clamp recordings in HEK-293 cells and dorsal root ganglion neurons were conducted. We used patch pipettes to record transient Na⁺ currents. One of the compounds reduced the peak sodium currents in Nav1.7-HEK-293 stable cell line in a dose-dependent manner, with IC50 values at 0.74 µM. In summary, our computer-aided analgesic discovery approach allowed us to develop pre-clinical analgesic candidate with significant reduction of time and cost.

Keywords: chronic pain, voltage-gated sodium channel, isoform-selective antagonist, similarity search, virtual screening, analgesics development

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Authors: Felicia Segeth, Florian G. Klein, Lea Berger, Andreas Kolk, Per S. Holm

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The entry of oncolytic viruses (OVs) into clinical application opens groundbreaking changes in current and future treatment regimens. However, despite their potent anti-cancer activity in vitro, clinical studies revealed limitations of OVs as monotherapy. The same applies to CDK 4/6 inhibitors (CDK4/6i) targeting cell cycle as well as bromodomain and extra-terminal domain inhibitors (BETi) targeting gene expression. In this study, the anti-tumoral effect of XVir-N-31, an YB-1 dependent oncolytic adenovirus, was evaluated in combination with Ribociclib, a CDK4/6i, and JQ1, a BETi. The head and neck squamous cell carcinoma (HNSCC) cell lines Fadu, SAS, and Cal-33 were used. DNA replication and gene expression of XVir-N-31 was measured by RT-qPCR, protein expression by western blotting, and cell lysis by SRB assays. Treatment with CDK4/6i and BETi increased viral gene expression, viral DNA replication, and viral particle formation. The data show that the combination of oncolytic adenovirus XVir-N-31 with CDK4/6i & BETi acts highly synergistic in cancer cell lysis. Furthermore, additional molecular analyses on this subject demonstrate that the positive transcription elongation factor P-TEFb plays a decisive role in this regard, indicating an influence of the combinational therapy on gene transcription control. The combination of CDK4/6i & BETi and XVir-N-31 is an attractive strategy to achieve substantial cancer cell killing and is highly suitable for clinical testing.

Keywords: adenovirus, BET, CDK4/6, HNSCC, P-TEFb, YB-1

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Authors: Neha Parekh, Divya Ladha, Poonam Wadhwani, Nisha Shah

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Nano scaled materials have attracted tremendous interest as corrosion inhibitor due to their high surface area on the metal surfaces. It is well known that the zinc oxide nanoparticles have higher reactivity towards aqueous acidic solution. This work presents a new method to incorporate zinc oxide nanoparticles with white sesame seeds extract (nano-green inhibitor) for corrosion protection of zinc in acidic medium. The morphology of the zinc oxide nanoparticles was investigated by TEM and DLS. The corrosion inhibition efficiency of the green inhibitor and nano-green inhibitor was determined by Gravimetric and electrochemical impedance spectroscopy (EIS) methods. Gravimetric measurements suggested that nano-green inhibitor is more effective than green inhibitor. Furthermore, with the increasing temperature, inhibition efficiency increases for both the inhibitors. In addition, it was established the Temkin adsorption isotherm fits well with the experimental data for both the inhibitors. The effect of temperature and Temkin adsorption isotherm revealed Chemisorption mechanism occurring in the system. The activation energy (Ea) and other thermodynamic parameters for inhibition process were calculated. The data of EIS showed that the charge transfer controls the corrosion process. The surface morphology of zinc metal (specimen) in absence and presence of green inhibitor and nano-green inhibitor were performed using Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM) techniques. The outcomes indicated a formation of a protective layer over zinc metal (specimen).

Keywords: corrosion, green inhibitor, nanoparticles, zinc

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Authors: A. J. S. P. Nileema, S. Kulatunga , S. H. Palihawadana

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Computer aided detection (CAD) system was developed for the diagnosis of pulmonary tuberculosis using digital chest X-rays with MATLAB image processing techniques using a statistical approach. The study comprised of 200 digital chest radiographs collected from the National Hospital for Respiratory Diseases - Welisara, Sri Lanka. Pre-processing was done to remove identification details. Lung fields were segmented and then divided into four quadrants; right upper quadrant, left upper quadrant, right lower quadrant, and left lower quadrant using the image processing techniques in MATLAB. Contrast, correlation, homogeneity, energy, entropy, and maximum probability texture features were extracted using the gray level co-occurrence matrix method. Descriptive statistics and normal distribution analysis were performed using SPSS. Depending on the radiologists’ interpretation, chest radiographs were classified manually into PTB - positive (PTBP) and PTB - negative (PTBN) classes. Features with standard normal distribution were analyzed using an independent sample T-test for PTBP and PTBN chest radiographs. Among the six features tested, contrast, correlation, energy, entropy, and maximum probability features showed a statistically significant difference between the two classes at 95% confidence interval; therefore, could be used in the classification of chest radiograph for PTB diagnosis. With the resulting value ranges of the five texture features with normal distribution, a classification algorithm was then defined to recognize and classify the quadrant images; if the texture feature values of the quadrant image being tested falls within the defined region, it will be identified as a PTBP – abnormal quadrant and will be labeled as ‘Abnormal’ in red color with its border being highlighted in red color whereas if the texture feature values of the quadrant image being tested falls outside of the defined value range, it will be identified as PTBN–normal and labeled as ‘Normal’ in blue color but there will be no changes to the image outline. The developed classification algorithm has shown a high sensitivity of 92% which makes it an efficient CAD system and with a modest specificity of 70%.

Keywords: chest radiographs, computer aided detection, image processing, pulmonary tuberculosis

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Authors: Fabiola Trejo Perez, Rolando Diaz Loving

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Sexual pleasure consists of the positively valued feelings induced by sexual stimuli, but it is also weighed down by pop-psychological baggage, and subjected to cross-cultural and cross-historical variation. Social and individual interpretations of what can or can’t be considered as pleasurable are intertwined with culture’s predominant values, norms and beliefs. For each culture, sexual norms work as a guide to be followed in order to model socially accepted behaviors. Therefore, cultural messages regarding sexuality are usually directed to restrict men and women from enjoyment, sexual satisfaction and specifically orgasm. Given that sexual pleasure hasn’t been recognized as an accepted topic of open discussion, particularly for women, people have to eventually complement their knowledge using their own experience filling in the blanks from what little has been said. Thus, this research aims to identify which are the particular social messages associated with the easing or inhibition of sexual pleasure. Three hundred Mexican men and women ages 25 to 35 years old answered a self-report survey composed by the Inventory of facilitators and inhibitors of sexual pleasure and the Sexual premises questionnaire via pencil-paper and online. Results show a high endorsement to double standard messages associated with higher levels of sexual pleasure inhibitors like feeling pressured to have sexual activity, guilt and inability to reach orgasm, in contrast with people who endorse more permissive norms and beliefs, feeling connected to their sexual partners and confident with themselves. These results illustrate that the shaping of sexuality, from experience to society, is comprised of an important relationship between culture and sexual pleasure.

Keywords: culture, sexual double standard, sexual norms and beliefs, sexual pleasure

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Authors: A. A. Agbor, Jean Joel R. Bigna, Serges Clotaire Billong, Mathurin Cyrille Tejiokem, Gabriel L. Ekali, Claudia S. Plottel, Jean Jacques N. Noubiap, Hortence Abessolo, Roselyne Toby, Sinata Koulla-Shiro

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Background: Contributors to fatal outcomes in patients undergoing tuberculosis (TB) treatment in the setting of HIV co-infection are poorly characterized, especially in sub-Saharan Africa. Our study’s aim was to assess factors associated with death in TB/HIV co-infected patients during the first 6 months their TB treatment. Methods: We conducted a tertiary-care hospital-based retrospective cohort study from January 2006 to December 2013 at the Yaoundé Central Hospital, Cameroon. We reviewed medical records to identify hospitalized co-infected TB/HIV patients aged 15 years and older. Death was defined as any death occurring during TB treatment, as per the World Health Organization’s recommendations. Logistic regression analysis identified factors associated with death. Magnitudes of associations were expressed by adjusted odds ratio (aOR) with 95% confidence interval. A p value < 0.05 was considered statistically significant. Results: The 337 patients enrolled had a mean age of 39.3 (+/- 10.3) years and more (54.3%) were women. TB treatment outcomes included: treatment success in 60.8% (n=205), death in 29.4% (n=99), not evaluated in 5.3% (n=18), loss to follow-up in 5.3% (n=14), and failure in 0.3% (n=1) . After exclusion of patients lost to follow-up and not evaluated, death in TB/HIV co-infected patients during TB treatment was associated with: a TB diagnosis made before national implementation of guidelines regarding initiation of antiretroviral therapy (aOR = 2.50 [1.31-4.78]; p = 0.006), the presence of other AIDS-defining infections (aOR = 2.73 [1.27-5.86]; p = 0.010), non-AIDS comorbidities (aOR = 3.35 [1.37-8.21]; p = 0.008), not receiving co-trimoxazole prophylaxis (aOR = 3.61 [1.71-7.63]; p = 0.001), not receiving antiretroviral therapy (aOR = 2.45 [1.18-5.08]; p = 0.016), and CD4 cell counts < 50 cells/mm3 (aOR = 16.43 [1.05-258.04]; p = 0.047). Conclusions: The success rate of anti-tuberculosis treatment among hospitalized TB/HIV co-infected patients in our setting is low. Mortality in the first 6 months of treatment was high and strongly associated with specific clinical factors including states of greater immunosuppression, highlighting the urgent need for targeted interventions, including provision of anti-retroviral therapy and co-trimoxazole prophylaxis in order to enhance patient outcomes.

Keywords: TB/HIV co-infection, death, treatment outcomes, factors

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Authors: Francesco Pantano, Marta Fogolari, Michele Iuliani, Sonia Simonetti, Silvia Cavaliere, Marco Russano, Fabrizio Citarella, Bruno Vincenzi, Silvia Angeletti, Giuseppe Tonini

Abstract:

Background: Although immune checkpoint inhibitors (ICIs) have changed the treatment paradigm of non–small cell lung cancer (NSCLC), these drugs fail to elicit durable responses in the majority of NSCLC patients. The gut microbiota, able to regulate immune responsiveness, is emerging as a promising, modifiable target to improve ICIs response rates. Since the oral microbiome has been demonstrated to be the primary source of bacterial microbiota in the lungs, we investigated its composition as a potential predictive biomarker to identify and select patients who could benefit from immunotherapy. Methods: Thirty-five patients with stage IV squamous and non-squamous cell NSCLC eligible for an anti-PD-1/PD-L1 as monotherapy were enrolled. Saliva samples were collected from patients prior to the start of treatment, bacterial DNA was extracted using the QIAamp® DNA Microbiome Kit (QIAGEN) and the 16S rRNA gene was sequenced on a MiSeq sequencing instrument (Illumina). Results: NSCLC patients were dichotomized as “Responders” (partial or complete response) and “Non-Responders” (progressive disease), after 12 weeks of treatment, based on RECIST criteria. A prevalence of the phylum Candidatus Saccharibacteria was found in the 10 responders compared to non-responders (abundance 5% vs 1% respectively; p-value = 1.46 x 10-7; False Discovery Rate (FDR) = 1.02 x 10-6). Moreover, a higher prevalence of Saccharibacteria Genera Incertae Sedis genus (belonging to the Candidatus Saccharibacteria phylum) was observed in "responders" (p-value = 6.01 x 10-7 and FDR = 2.46 x 10-5). Finally, the patients who benefit from immunotherapy showed a significant abundance of TM7 Phylum Sp Oral Clone FR058 strain, member of Saccharibacteria Genera Incertae Sedis genus (p-value = 6.13 x 10-7 and FDR=7.66 x 10-5). Conclusions: These preliminary results showed a significant association between oral microbiota and ICIs response in NSCLC patients. In particular, the higher prevalence of Candidatus Saccharibacteria phylum and TM7 Phylum Sp Oral Clone FR058 strain in responders suggests their potential immunomodulatory role. The study is still ongoing and updated data will be presented at the congress.

Keywords: oral microbiota, immune checkpoint inhibitors, non-small cell lung cancer, predictive biomarker

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Authors: Asma Lamin, Anna H. Kaksonen, Ivan Cole, Paul White, Xiao-Bo Chen

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Microbiologically influenced corrosion (MIC) is a process in which microorganisms initiate, facilitate, or accelerate the electrochemical corrosion reactions of metallic components. Several reports documented that MIC accounts for about 20 to 40 % of the total cost of corrosion. Biofilm formation due to the presence of microorganisms on the surface of metal components is known to play a vital role in MIC, which can lead to severe consequences in various environmental and industrial settings. Quorum sensing (QS) system plays a major role in regulating biofilm formation and control the expression of some microbial enzymes. QS is a communication mechanism between microorganisms that involves the regulation of gene expression as a response to the microbial cell density within an environment. This process is employed by both Gram-positive and Gram-negative bacteria to regulate different physiological functions. QS involves production, detection, and responses to signalling chemicals, known as auto-inducers. QS controls specific processes important for the microbial community, such as biofilm formation, virulence factor expression, production of secondary metabolites and stress adaptation mechanisms. The use of QS inhibitors (QSIs) has been proposed as a possible solution to biofilm related challenges in many different applications. Although QSIs have demonstrated some strength in tackling biofouling, QSI-based strategies to control microbially influenced corrosion have not been thoroughly investigated. As such, our research aims to target the QS mechanisms as a strategy for mitigating MIC on metal surfaces in engineered systems.

Keywords: quorum sensing, quorum quenching, biofilm, biocorrosion

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Authors: Ankur Chaudhuri, Sibani Sen Chakraborty

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In human, glutaminyl cyclase activity is highly abundant in neuronal and secretory tissues and is preferentially restricted to hypothalamus and pituitary. The N-terminal modification of β-amyloids (Aβs) peptides by the generation of a pyro-glutamyl (pGlu) modified Aβs (pE-Aβs) is an important process in the initiation of the formation of neurotoxic plaques in Alzheimer’s disease (AD). This process is catalyzed by glutaminyl cyclase (QC). The expression of QC is characteristically up-regulated in the early stage of AD, and the hallmark of the inhibition of QC is the prevention of the formation of pE-Aβs and plaques. A computer-aided drug design (CADD) process was employed to give an idea for the designing of potentially active compounds to understand the inhibitory potency against human glutaminyl cyclase (QC). This work elaborates the ligand-based and structure-based pharmacophore exploration of glutaminyl cyclase (QC) by using the known inhibitors. Three dimensional (3D) quantitative structure-activity relationship (QSAR) methods were applied to 154 compounds with known IC50 values. All the inhibitors were divided into two sets, training-set, and test-sets. Generally, training-set was used to build the quantitative pharmacophore model based on the principle of structural diversity, whereas the test-set was employed to evaluate the predictive ability of the pharmacophore hypotheses. A chemical feature-based pharmacophore model was generated from the known 92 training-set compounds by HypoGen module implemented in Discovery Studio 2017 R2 software package. The best hypothesis was selected (Hypo1) based upon the highest correlation coefficient (0.8906), lowest total cost (463.72), and the lowest root mean square deviation (2.24Å) values. The highest correlation coefficient value indicates greater predictive activity of the hypothesis, whereas the lower root mean square deviation signifies a small deviation of experimental activity from the predicted one. The best pharmacophore model (Hypo1) of the candidate inhibitors predicted comprised four features: two hydrogen bond acceptor, one hydrogen bond donor, and one hydrophobic feature. The Hypo1 was validated by several parameters such as test set activity prediction, cost analysis, Fischer's randomization test, leave-one-out method, and heat map of ligand profiler. The predicted features were then used for virtual screening of potential compounds from NCI, ASINEX, Maybridge and Chembridge databases. More than seven million compounds were used for this purpose. The hit compounds were filtered by drug-likeness and pharmacokinetics properties. The selective hits were docked to the high-resolution three-dimensional structure of the target protein glutaminyl cyclase (PDB ID: 2AFU/2AFW) to filter these hits further. To validate the molecular docking results, the most active compound from the dataset was selected as a reference molecule. From the density functional theory (DFT) study, ten molecules were selected based on their highest HOMO (highest occupied molecular orbitals) energy and the lowest bandgap values. Molecular dynamics simulations with explicit solvation systems of the final ten hit compounds revealed that a large number of non-covalent interactions were formed with the binding site of the human glutaminyl cyclase. It was suggested that the hit compounds reported in this study could help in future designing of potent inhibitors as leads against human glutaminyl cyclase.

Keywords: glutaminyl cyclase, hit lead, pharmacophore model, simulation

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Authors: Susan Scott, Dina Hanna, Bassel Zebian, Gary Ruiz, Sreena Das

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Although the number of cases of TB in England has fallen over the last 4 years, it remains an important public health burden with 1 in 20 cases dying annually. The vast majority of cases present in non-UK born individuals with social risk factors. We present a case of non-pulmonary TB presenting in a healthy child born in the UK to professional parents. We present a case of a healthy 10 year old boy who developed acute back pain during school PE. Over the next 5 months, he was seen by various health and allied professionals with worsening back pain and kyphosis. He became increasing unsteady and for the 10 days prior to admission to our hospital, he developed fevers. He was admitted to his local hospital for tonsillitis where he suffered two falls on account of his leg weakness. A spinal X-ray revealed a pathological fracture and gibbus formation. He was transferred to our unit for further management. On arrival, the patient had lower motor neurone signs of his left leg. He underwent spinal fixture, laminectomy and decompression. Microbiology samples taken intra-operatively confirmed Mycobacterium Tuberculosis. He had a positive Mantoux and T-spot and treatment were commenced. There was no evidence of immune compromise. The patient was born in the UK, had a BCG scar and his only travel history had been two years prior to presentation when he travelled to the Phillipines for a short holiday. The patient continues to have issues around neuropathic pain, mobility, pill burden and mild liver side effects from treatment. Discussion: There is a paucity of case reports on spinal TB in paediatrics and diagnosis is often difficult due to the non-specific symptomatology. Although prognosis on treatment is good, a delayed diagnosis can have devastating consequences. This case highlights the continued need for higher index of suspicion and diagnosis in a world with changing patterns of migration and increase global travel. Surgical intervention is limited to the most serious cases to minimise further neurological damage and improve prognosis. There remains the need for a multi-disciplinary approach to deal with challenges of treatment and rehabilitation.

Keywords: tuberculosis, non-pulmonary TB, public health burden, diagnostic challenge

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Authors: Swati Gautam, Amita Jain, Shyampyari Jaiswar

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Objective: To elucidate the role of (ApaI&TaqI) VDR gene polymorphism in the pathogenesis of female genital tuberculosis (FGTB) cases. Background: Female genital TB represents about 15-20% of total extra-pulmonary TB (EPTB). Female subjects with vitamin D deficiency have been shown to be at higher risk of pulmonary TB as well as FGTB. In same context few functional polymorphism in vitamin D receptor (VDR) gene has been considered as an important genetic risk factor that modulate the development of FGTB. Therefore we aimed, to elucidate the role of (ApaI&TaqI) VDR gene polymorphism in the pathogenesis of FGTB. Study design: Case-Control study. Sample size: Cases (60) and Controls (60). Study site: Department of Obstetrics & Gynecology & Department of Microbiology, K.G.M.U. Lucknow, (UP). Inclusion criteria: Cases: Women with age group 20-35 years, premenstrual endometrial aspiration collected and included in the study, those were positive with acid-fast bacilli (AFB)/ TB-PCR/ LJ culture/ liquid culture. Controls: Women with age group 20-35 years having no history of ATT and all test negative for TB recruited as control. Exclusion criteria: -Women with endometriosis, polycystic ovaries (PCOD), positive on Chlamydia & gonorrhea, already on anti-tubercular therapy (ATT) excluded. Materials and Methods: Blood samples were collected in EDTA tubes from cases and controls stored at -20ºC. Genomic DNA extraction was carried out by salting-out method. Genotyping of VDR gene (ApaI&TaqI) polymorphism was performed by using single amplification refractory mutation system (ARMS) PCR technique. PCR products were analyzed by electrophoresis on 2% agarose gel. Statistical analysis was done by SPSS16.3 software & computing odds ratio (OR) with 95% CI. Results: Increased risk of female genital tuberculosis was observed in AA genotype (OR =1.1419-6.212 95% CI, P*<0.036) and A allele (OR =1.255-3.518, 95% CI, P* < 0.006) in FGTB as compared to controls. Moreover A allele was found more frequent in FGTB patients. No significant difference was observed in TaqI gene polymorphism of VDR gene. Conclusion: The ApaI polymorphism is significantly associated with etiology of FGTB and plays an important role as a genetic risk factor in FGTB women.

Keywords: ARMS, ATT, EPTB, FGTB, VDR

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Authors: Tivani Phosa Mashamba-Thompson, Mahmoud E. S. Soliman

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To date, the cause of neurodegeneration is not well understood and diseases that stem from neurodegeneration currently have no known cures. Cathepsin B (CB) enzyme is known to be involved in the production of peptide neurotransmitters and toxic peptides in neurodegenerative diseases (NDs). CA-074Me is a membrane-permeable irreversible selective cathepsin B (CB) inhibitor as confirmed by in vivo studies. Due to the lack of the crystal structure, the binding mode of CA-074Me with the human CB at molecular level has not been previously reported. The main aim of this study is to gain an insight into the binding mode of CB CA-074Me to human CB using various computational tools. Herein, molecular dynamics simulations, binding free energy calculations and per-residue energy decomposition analysis were employed to accomplish the aim of the study. Another objective was to identify novel CB inhibitors based on the structure of CA-074Me using fragment based drug design using scaffold hoping drug design approach. Results showed that two of the designed ligands (hit 1 and hit 2) were found to have better binding affinities than the prototype inhibitor, CA-074Me, by ~2-3 kcal/mol. Per-residue energy decomposition showed that amino acid residues Cys29, Gly196, His197 and Val174 contributed the most towards the binding. The Van der Waals binding forces were found to be the major component of the binding interactions. The findings of this study should assist medicinal chemist towards the design of potential irreversible CB inhibitors.

Keywords: cathepsin B, scaffold hopping, docking, molecular dynamics, binding-free energy, neurodegerative diseases

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Authors: Ismail Celik, Gulgun Ayhan Kilcigil, Berna Guven, Zumra Kara, Arzu Onay-Besikci

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Epidermal Growth Factor Receptor is the cell-surface receptor of the ErbB (erythroblastic leukemia viral oncogene homologue receptors) family of tyrosine kinases. It plays a vital role in regulating the proliferation and differentiation of cells. However, a variety of mechanisms, such as EGFR expression, mutation, and ligand-dependent receptor dimerization, are associated with the development of various activated EGFR tumors. EGFR is highly expressed in most solid tumors, including breast, head and neck cancer, non-small cell lung cancer (NSCLC), renal, ovarian, and colon cancers. Thus, specific EGFR inhibition plays one of the key roles in cancer treatment. The compounds used in the treatment as tyrosine kinase inhibitors are known to contain the benzimidazole isosterium indole, pazopanib, and axitinibin indazole rings. In addition, benzimidazoles have been shown to exhibit protein kinase inhibitory activity in addition to their different biological activities.Based on these data, it was planned and synthesized of some oxadiazole bearing benzimidazole derivatives [N-cyclohexyl-5-((2-phenyl/substitutedphenyl-1H-benzo[d]imidazole-1-yl) methyl)-1,3,4-oxadiazole-2-amine]. EGFR kinase inhibitory efficiency of the synthesized compounds was determined by comparing them with a known kinase inhibitor erlotinib in vitro, and two of the compounds bearing phenyl (19a) and 3,4-dibenzyloxyphenyl (21a) ring exhibited significant activities.

Keywords: benzimidazole, EGFR kinase inhibitory, oxadiazole, synthesis

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Authors: Sushil Kumar Singh, Ashok Kumar, Ankit Ganeshpurkar, Ravi Singh, Devendra Kumar

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In spectrum of neurodegenerative diseases, Alzheimer’s disease (AD) is characterized by the presence of amyloid β plaques and neurofibrillary tangles in the brain. It results in cognitive and memory impairment due to loss of cholinergic neurons, which is considered to be one of the contributing factors. Donepezil, an acetylcholinesterase (AChE) inhibitor which also inhibits butyrylcholinesterase (BuChE) and improves the memory and brain’s cognitive functions, is the most successful and prescribed drug to treat the symptoms of AD. The present work is based on designing of the selective BuChE inhibitors using computational techniques. In this work, machine learning models were trained using classification algorithms followed by screening of diverse chemical library of compounds. The various molecular modelling and simulation techniques were used to obtain the virtual hits. The amide derivatives of 4-(phenylsulfonamido) benzoic acid were synthesized and characterized using 1H & 13C NMR, FTIR and mass spectrometry. The enzyme inhibition assays were performed on equine plasma BuChE and electric eel’s AChE by method developed by Ellman et al. Compounds 31, 34, 37, 42, 49, 52 and 54 were found to be active against equine BuChE. N-(2-chlorophenyl)-4-(phenylsulfonamido)benzamide and N-(2-bromophenyl)-4-(phenylsulfonamido)benzamide (compounds 34 and 37) displayed IC50 of 61.32 ± 7.21 and 42.64 ± 2.17 nM against equine plasma BuChE. Ortho-substituted derivatives were more active against BuChE. Further, the ortho-halogen and ortho-alkyl substituted derivatives were found to be most active among all with minimal AChE inhibition. The compounds were selective toward BuChE.

Keywords: Alzheimer disease, butyrylcholinesterase, machine learning, sulfonamides

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Authors: Anushka Naidoo, Veron Ramsuran, Maxwell Chirehwa, Paolo Denti, Kogieleum Naidoo, Helen McIlleron, Nonhlanhla Yende-Zuma, Ravesh Singh, Sinaye Ngcapu, Nesri Padayatachi

Abstract:

Background: Despite efforts to introduce new drugs and shorter drug regimens for drug-susceptible tuberculosis (TB), the standard first-line treatment has not changed in over 50 years. Rifampicin, isoniazid, and pyrazinamide are critical components of the current standard treatment regimens. Some studies suggest that microbiologic failure and acquired drug resistance are primarily driven by low drug concentrations that result from pharmacokinetic (PK) variability independent of adherence to treatment. Wide between-patient pharmacokinetic variability for rifampin, isoniazid, and pyrazinamide has been reported in prior studies. There may be several reasons for this variability. However, genetic variability in genes coding for drug metabolizing and transporter enzymes have been shown to be a contributing factor for variable tuberculosis drug exposures. Objective: We describe the pharmacokinetics of first-line TB drugs rifampicin, isoniazid, and pyrazinamide and assess the effect of genetic variability in relevant selected drug metabolizing and transporter enzymes on pharmacokinetic parameters of isoniazid and rifampicin. Methods: We conducted the randomized-controlled Improving retreatment success TB trial in Durban, South Africa. The drug regimen included rifampicin, isoniazid, and pyrazinamide. Drug concentrations were measured in plasma, and concentration-time data were analysed using nonlinear-mixed-effects models to quantify the effects of relevant covariates and single nucleotide polymorphisms (SNP’s) of drug metabolizing and transporter genes on rifampicin, isoniazid and pyrazinamide exposure. A total of 25 SNP’s: four NAT2 (used to determine acetylator status), four SLCO1B1, three Pregnane X receptor (NR1), six ABCB1 and eight UGT1A, were selected for analysis in this study. Genotypes were determined for each of the SNP’s using a TaqMan® Genotyping OpenArray™. Results: Among fifty-eight patients studied; 41 (70.7%) were male, 97% black African, 42 (72.4%) HIV co-infected and 40 (95%) on efavirenz-based ART. Median weight, fat-free mass (FFM), and age at baseline were 56.9 kg (interquartile range, IQR: 51.1-65.2), 46.8 kg (IQR: 42.5-50.3) and 37 years (IQR: 31-42), respectively. The pharmacokinetics of rifampicin and pyrazinamide was best described using one-compartment models with first-order absorption and elimination, while for isoniazid two-compartment disposition was used. The median (interquartile range: IQR) AUC (h·mg/L) and Cmax (mg/L) for rifampicin, isoniazid, and pyrazinamide were; 25.62 (23.01-28.53) and 4.85 (4.36-5.40), 10.62 (9.20-12.25) and 2.79 (2.61-2.97), 345.74 (312.03-383.10) and 28.06 (25.01-31.52), respectively. Eighteen percent of patients were classified as rapid acetylators, and 34% and 43% as slow and intermediate acetylators, respectively. Rapid and intermediate acetylator status based on NAT 2 genotype resulted in 2.3 and 1.6 times higher isoniazid clearance than slow acetylators. We found no effects of the SLCO1B1 genotypes on rifampicin pharmacokinetics. Conclusion: Plasma concentrations of rifampicin, isoniazid, and pyrazinamide were low overall in our patients. Isoniazid clearance was high overall and as expected higher in rapid and intermediate acetylators resulting in lower drug exposures. In contrast to reports from previous South African or Ugandan studies, we did not find any effects of the SLCO1B1 or other genotypes tested on rifampicin PK. However, our findings are in keeping with more recent studies from Malawi and India emphasizing the need for geographically diverse and adequately powered studies. The clinical relevance of the low tuberculosis drug concentrations warrants further investigation.

Keywords: rifampicin, isoniazid pharmacokinetics, genetics, NAT2, SLCO1B1, tuberculosis

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Authors: Milad Gholizadeh

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Asthma and chronic obstructive pulmonary disease (COPD) are very shared inflammatory diseases of the airlines. They togethercause airway tapering and are cumulative in occurrence throughout the world, imposing huge burdens on health care. It is currently recognized that some asthmatic inflammation is neutrophilic, controlled by the TH17 subset of helper T cells, and that some eosinophilic inflammation is controlled by type 2 innate lymphoid cells (ILC2 cells) temporary together with basophils. Patients who have plain asthma or are asthmatic patients who smoke with topographies of COPD-induced inflammation and might advantage from treatments targeting neutrophils, countingmacrolides, CXCR2 antagonists, phosphodiesterase 4 inhibitors, p38 mitogen-activating protein kinase inhibitors, and antibodies in contradiction of IL-1 and IL-17.Viral and bacterial infections, not only reason acute exacerbations of COPD, but also intensify and continue chronic inflammation in steady COPD through pathogen-associated molecular patterns. Present treatment plans are absorbed on titration of inhaled therapies such as long-acting bronchodilators, with cumulative interest in the usage of beleaguered biologic therapies meant at the underlying inflammatory devices. Educationssuggest that the mucosal IgA reply is abridged in COPD, and a lacking conveyance of IgA across the bronchial epithelium in COPD has been recognized, perhaps involving neutrophil proteinases, which may damage the Ig receptor mediating this transepithelialdirection-finding. Future instructions for investigation will emphasis elucidating the diverse inflammatory signatures foremost to asthma and chronic obstrucive, the development of reliable analytic standards and biomarkers of illness, and refining the clinical organization with an eye toward targeted therapies.

Keywords: imminology, asthma, COPD, CXCR2 antagonists

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Authors: Gabriela Wyszogrodzka, Przemyslaw Dorozynski, Barbara Gil, Maciej Strzempek, Bartosz Marszalek, Piotr Kulinowski, Wladyslaw Piotr Weglarz, Elzbieta Menaszek

Abstract:

MOFs (Metal-Organic Frameworks) belong to a new group of porous materials with a hybrid organic-inorganic construction. Their structure is a network consisting of metal cations or clusters (acting as metallic centers, nodes) and the organic linkers between nodes. The interest in MOFs is primarily associated with the use of their well-developed surface and large porous. Possibility to build MOFs of biocompatible components let to use them as potential drug carriers. Furthermore, forming MOFs structure from cations possessing paramagnetic properties (e.g. iron cations) allows to use them as MRI (Magnetic Resonance Imaging) contrast agents. The concept of formation of particles that combine the ability to transfer active substance with imaging properties has been called theranostic (from words combination therapy and diagnostics). By building MOF structure from iron cations it is possible to use them as theranostic agents and monitoring the distribution of the active substance after administration in real time. In the study iron-MOF: Fe-MIL-101-NH2 was chosen, consisting of iron cluster in nodes of the structure and amino-terephthalic acid as a linker. The aim of the study was to investigate the possibility of applying Fe-MIL-101-NH2 as inhalable theranostic particulate system for the first-line anti-tuberculosis antibiotic – isoniazid. The drug content incorporated into Fe-MIL-101-NH2 was evaluated by dissolution study using spectrophotometric method. Results showed isoniazid encapsulation efficiency – ca. 12.5% wt. Possibility of Fe-MIL-101-NH2 application as the MRI contrast agent was demonstrated by magnetic resonance tomography. FeMIL-101-NH2 effectively shortening T1 and T2 relaxation times (increasing R1 and R2 relaxation rates) linearly with the concentrations of suspended material. Images obtained using multi-echo magnetic resonance imaging sequence revealed possibility to use FeMIL-101-NH2 as positive and negative contrasts depending on applied repetition time. MOFs micronization via ultrasound was evaluated by XRD, nitrogen adsorption, FTIR, SEM imaging and did not influence their crystal shape and size. Ultrasonication let to break the aggregates and achieve very homogeneously looking SEM images. MOFs cytotoxicity was evaluated in in vitro test with a highly sensitive resazurin based reagent PrestoBlue™ on L929 fibroblast cell line. After 24h no inhibition of cell proliferation was observed. All results proved potential possibility of application of ironMOFs as an isoniazid carrier and as MRI contrast agent in inhalatory treatment of tuberculosis. Acknowledgments: Authors gratefully acknowledge the National Science Center Poland for providing financial support, grant no 2014/15/B/ST5/04498.

Keywords: imaging agents, metal-organic frameworks, theranostics, tuberculosis

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Authors: Ved Vrat Verma, Rani Gupta, Manisha Goel

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γ-glutamyltranspeptidase (GGT: EC 2.3.2.2) is an N-terminal nucleophile hydrolase conserved in all three domains of life. GGT plays a key role in glutathione metabolism where it catalyzes the breakage of the γ-glutamyl bonds and transfer of γ-glutamyl group to water (hydrolytic activity) or amino acids or short peptides (transpeptidase activity). GGTs from bacteria, archaea, and eukaryotes (human, rat and mouse) are homologous proteins sharing >50% sequence similarity and conserved four layered αββα sandwich like three dimensional structural fold. These proteins though similar in their structure to each other, are quite diverse in their enzyme activity: some GGTs are better at hydrolysis reactions but poor in transpeptidase activity, whereas many others may show opposite behaviour. GGT is known to be involved in various diseases like asthma, parkinson, arthritis, and gastric cancer. Its inhibition prior to chemotherapy treatments has been shown to sensitize tumours to the treatment. Microbial GGT is known to be a virulence factor too, important for the colonization of bacteria in host. However, all known inhibitors (mimics of its native substrate, glutamate) are highly toxic because they interfere with other enzyme pathways. However, a few successful efforts have been reported previously in designing species specific inhibitors. We aim to leverage the diversity seen in GGT family (pathogen vs. eukaryotes) for designing specific inhibitors. Thus, in the present study, we have used DIVERGE software to identify sites in GGT proteins, which are crucial for the functional and structural divergence of these proteins. Since, type II divergence sites vary in clade specific manner, so type II divergent sites were our focus of interest throughout the study. Type II divergent sites were identified for pathogen vs. eukaryotes clusters and sites were marked on clade specific representative structures HpGGT (2QM6) and HmGGT (4ZCG) of pathogen and eukaryotes clade respectively. The crucial divergent sites within 15 A radii of the binding cavity were highlighted, and in-silico mutations were performed on these sites to delineate the role of these sites on the mechanism of catalysis and protein folding. Further, the amino acid network (AAN) analysis was also performed by Cytoscape to delineate assortative mixing for cavity divergent sites which could strengthen our hypothesis. Additionally, molecular dynamics simulations were performed for wild complexes and mutant complexes close to physiological conditions (pH 7.0, 0.1 M ionic strength and 1 atm pressure) and the role of putative divergence sites and structural integrities of the homologous proteins have been analysed. The dynamics data were scrutinized in terms of RMSD, RMSF, non-native H-bonds and salt bridges. The RMSD, RMSF fluctuations of proteins complexes are compared, and the changes at protein ligand binding sites were highlighted. The outcomes of our study highlighted some crucial divergent sites which could be used for novel inhibitors designing in a species-specific manner. Since, for drug development, it is challenging to design novel drug by targeting similar protein which exists in eukaryotes, so this study could set up an initial platform to overcome this challenge and help to deduce the more effective targets for novel drug discovery.

Keywords: γ-glutamyltranspeptidase, divergence, species-specific, drug design

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Authors: Gajanan M. Sonwane

Abstract:

The computational studies on 2-phenazinamines with their protein targets have been carried out to design compounds with potential anticancer activity. This strategy of designing compounds possessing selectivity over specific tyrosine kinase has been achieved through G-QSAR and molecular docking studies. The objective of this research has been to design newer 2-phenazinamine derivatives as Bcr-Abl tyrosine kinase inhibitors by G-QSAR, molecular docking studies followed by wet-lab studies along with evaluation of their anticancer potential. Computational chemistry was done by using VLife MDS 4.3 and Autodock 4.2 followed by wet-lab experiments for synthesizing 2-phenazinamine derivatives. The chemical structures of ligands in 2D were drawn by employing Chemdraw 2D Ultra 8.0 and were converted into 3D. These were optimized by using a semi-empirical method called MOPAC. The protein structure was retrieved from RCSC protein data bank as a PDB file. The binding interactions of protein and ligands were done by using PYMOL. The molecular properties of the designed compounds were predicted in silico by using Osiris property explorer. The parent compound 2-phenazinamine was synthesized by reduction of 2, 4-dinitro-N-phenyl-benzenamine in the presence of tin chloride followed by cyclization in the presence of nitrobenzene and magnesium sulfate. The derivatization at the amino function of 2-phenazinamine was performed by treating parent compound with various aldehydes in the presence of dicyclohexylcarbodiimide (DCC) and urea to afford 2-(2-chlorophenyl)-3-(phenazine-2-yl) thiazolidine-4-one. Synthesized 39 novel derivatives of 2-phenazinamine and performed antioxidant activity, anti antiproliferative on the bulb of onion and anticancer activity on cell line showing significant competition with marked blockbuster drug imatinib.

Keywords: computer-aided drug design, tyrosin kinases, anticancer, docking

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