Association of Beta 2 adrenergic receptor (Thr164Ile) polymorphism with Salbutamol refractoriness in severe asthmatics from Indian population.

BACKGROUND: Thr164Ile polymorphism in the ADRB2 gene encoding ß2 adrenergic receptor (ß2AR) has its functional consequence in declining ligand-receptor interactions and depressed coupling of ß2AR to adenylcyclase. In the present study, we sought to evaluate the possible association of Thr164Ile polymorphism with asthma susceptibility, pharmacogenetic response to Salbutamol and varying degrees of severity. METHODS: Three hundred and ninety eight clinically diagnosed patients and four hundred and fifty six healthy controls were enrolled in the study. Patients were classified into severity classes according to the Global Initiative for Asthma (GINA) guidelines. To assess bronchodilator response, spirometry was performed before and 15min after Salbutamol (200µg) delivery. Responders to Salbutamol were categorized if percentage reversibility was greater than or equal to 12% in them, while those showing reversibility <12% were classified as non-responders. Further, responding phenotypes were stratified into severity groups. Genotyping was carried out by ARMS-PCR technique. Statistical methods were applied to test the significance of the results. RESULTS: In the present study, polymorphism was not associated with disease susceptibility however; there was significant association with non-responding asthmatics. In case of severity subsets, the polymorphism was not associated with milder subtypes; although, notable association was observed with moderate and severe asthma subtypes. In addition, the polymorphism was significantly associated with non-responding patients with severe asthma. CONCLUSIONS: In south Indian population, the ADRB2 Thr164Ile polymorphism may not form susceptible variant to develop asthma, however, it can form a predictive maker for bronchodilator (Salbutamol) response in severe asthmatics.

Molecular Docking studies of FKBP12-mTOR inhibitors using binding predictions.

UNLABELLED: Mammalian target of rapamycin (mTOR) is a key regulator of cell growth, proliferation and angiogenesis. mTOR signaling is frequently hyper activated in a broad spectrum of human cancers thereby making it a potential drug target. The current drugs available have been successful in inhibiting the mTOR signaling, nevertheless, show low oral bioavailability and suboptimal solubility. Considering the narrow therapeutic window of the available inhibitors, through computational approaches, the present study pursues to identify a compound with optimal oral bioavailability and better solubility properties in addition ensuing high affinity between FKBP12 and FRB domain of mTOR. Current mTOR inhibitors; Everolimus, Temsirolimus Deforolimus and Echinomycin served as parent molecules for similarity search with a threshold of 95%. The query molecules and respective similar molecules were docked at the binding cleft of FKBP12 protein. Aided by MolDock algorithm, high affinity compounds against FKBP12 were retrieved. Patch Dock supervised protein-protein interactions were established between FRB domain of mTOR and ligand (query and similar) bound and free states of FKBP12. All the similar compounds thus retrieved showed better solubility properties and enabled better complex formation of mTOR and FKBP12. In particular Everolimus similar compound PubChem ID: 57284959 showed appreciable drugs like properties bestowed with better solubility higher oral bioavailability. In addition this compound brought about enhanced interaction between FKBP12 and FRB domain of mTOR. In the study, we report Everolimus similar compound PubChem ID: 57284959 to be potential inhibitor for mTOR pathway which can overcome the affinity and solubility concerns of current mTOR drugs. ABBREVIATIONS: mTOR - Mammalian Target of Rapamycin, FRB domain - FKBP12-rapamycin associated protein, FKBP12 - FK506-binding protein 12, OPLS - Optimized Potentials for Liquid Simulations, Akt - RAC-alpha serine/threonine-protein kinase, PI3K - phosphatidylinositide 3-kinases.

Analysis of ADRB2 (Arg16Gly) Gene Variant with Susceptibility, Pharmacogenetic Response and Disease Severity in South Indian Asthmatics.

ß2-Adrenergic receptor (ß2-AR) plays a crucial role in asthma pathophysiology by regulating, processes of the lung function, and clinical response to bronchodilators. The +46G>A- Gly16Arg polymorphism in the gene encoding ß2 adrenergic receptor (ADRB2) has been associated with receptor non-responsiveness after ß2-agonist exposure. In the present study, we sought to evaluate the possible association of Gly16Arg polymorphism with asthma susceptibility, pharmacogenetic response to Salbutamol, and varying degrees of disease severity. Three hundred ninety-eight clinically diagnosed patients and 456 healthy controls were enrolled for the study. Patients were classified into severity classes according to Global Initiative for Asthma guidelines. To assess bronchodilator response, spirometry was performed before and 15 min after Salbutamol (200 µg) delivery. Responders to Salbutamol were categorized if percentage reversibility was greater than or equal to 12% in them, while those showing reversibility less than 12% were classified as non-responders. Genotyping was carried out by ARMS-PCR technique. Statistical methods were applied to test for the significance of the results. In the present study, there was lack of significant association of polymorphism with disease susceptibility as well as with bronchodilator response. The polymorphism was not associated with mild and moderate asthma subtypes; however, there was a notable association with severe asthma subtype. In addition, the polymorphism was associated with severe asthma compared to subtypes of mild and moderate asthma combined. In a South Indian population, the ADRB2 Arg/Gly may not form a susceptible variant to develop asthma nor can be a standard predictive marker to bronchodilator response; nevertheless, the patterns in asthma severity can be predicted by analyzing this variant.

Identification of high affinity bioactive Salbutamol conformer directed against mutated (Thr164Ile) beta 2 adrenergic receptor.

Salbutamol forms an important and widely administered ß2 agonist prescribed in the symptomatic treatment of bronchial asthma. Unfortunately, a subset of patients show refractoriness to it owing to ADRB2 gene variant (rs 1800888). The variant substitutes Thr to Ile at the position 164 in the ß2 adrenergic receptor leading to sub-optimal binding of agonists. The present study aims to associate the Salbutamol response with the variant and select the bioactive conformer of Sabutamol with optimal binding affinity against mutated receptor by in silico approaches. To assess bronchodilator response spirometry was performed before and 15 min after Salbutamol (200 mcg) inhalation. Responders to Salbutamol were categorized if percentage reversibility was greater than or equal to 12%, while those showing FEV1 reversibility less than 12% were classified as non-responders. Among the 344 subjects screened, 238 were responders and 106 were non-responders. The frequency of mutant allele "T" was significantly higher in case of non-responders (p < 0.05). In silico process involved generation of Salbutamol conformer ensembles supported by systematic search algorithm. 4369 conformers were generated of which only 1882 were considered bioactive conformers (threshold RMSD≤1 in reference to normalized structure of salbutamol). All the bioactive conformers were evaluated for the binding affinity against (Thr164 Ile) receptor through MolDock aided docking algorithm. One of the bioactive conformer (P.E. = -57.0038, RMSD = 0.6) demonstrated 1.54 folds greater affinity than the normal Salbutamol in the mutated receptor. The conformer identified in the present study may be put to pharmacodynamic and pharmacokinetic studies in future ahead.

Association of transforming growth factor-Beta 1 promoter variant -509 c/t with bronchial asthma in South Indian population.

Transforming growth factor-beta 1 (TGF-ß1) is a multifunctional cytokine that plays a pivotal role in airway remodeling observed in the asthmatic airways. C to T base substitution at -509 promoter position in the TGF-ß1 gene leads to its increased expression which contributes to airway remodeling in bronchial asthma. We sought to evaluate the association of TGF-ß1 -509 C/T promoter variant with clinical asthma and varying degrees of disease severity. Three hundred and eighty-two clinically diagnosed asthma patients and 410 healthy controls were enrolled for the study. Patients were classified into severity classes according to the Global Initiative for Asthma (GINA) guidelines. TGF-ß1 -509 C/T genotyping was carried out by amplification refractory mutation system polymerase chain reaction (ARMS-PCR) technique. In the present study, we found significantly higher frequency of TT genotype in asthma patients compared to controls (for TT vs. CC, p = 0.020). In addition, a significant difference was observed in the frequency of C and T allele in patients and controls (for T vs. C, p = 0.029). The heterozygous "CT" genotype was higher in moderate and severe asthmatics compared to mild subset of patients (for mild vs. moderate, p = 0.037). However, there was no significant distribution and association of variant allele with the severity subsets.

Structure based virtual screening of ligands to identify cysteinyl leukotriene receptor 1 antagonist.

Montelukast and Zafirlukast are known leukotriene receptor antagonists prescribed in asthma treatment. However, these fall short as mono therapy and are frequently used in combination with inhaled glucocorticosteroids with or without long acting beta 2 agonists. Therefore, it is of interest to apply ligand and structure based virtual screening strategies to identify compounds akin to lead compounds Montelukast and Zafirlukast. Hence, compounds with structures having 95% similarity to these compounds were retrieved from NCBI׳s PubChem database. Compounds similar to lead were grouped and docked at the antagonist binding site of cysteinyl leukotriene receptor 1. This exercise identified compounds UNII 70RV86E50Q (Pub Cid 71587778) and Sure CN 9587085 (Pub Cid 19793614) with higher predicted binding compared to Montelukast and Zafirlukast. It is shown that the compound Sure CN 9587085 showed appreciable ligand receptor interaction compared to UNII 70RV86E50Q. Thus, the compound Sure CN 9587085 is selected as a potent antagonist to cysteinyl leukotriene receptor 1 for further consideration in vitro and in vivo validation.