Rational design, synthesis, and structure-activity relationships of 5-amino-1H-pyrazole-4-carboxylic acid derivatives as protein tyrosine phosphatase 1B inhibitors.

A series of novel amino-carboxylic based pyrazole as protein tyrosine phosphatase 1B (PTP1B) inhibitors were designed on the basis of structure-based pharmacophore model and molecular docking. Compounds containing different hydrophobic tail (1,2-diphenyl ethanone, oxdiadizole and dibenzyl amines) were synthesized and evaluated in PTP1B enzymatic assay. Structure-activity relationship based optimization resulted in identification of several potent, metabolically stable and cell permeable PTP1B inhibitors.

Physiological and biochemical traits positively modulate tissue-specific withanolides and untargeted metabolites in Withania somnifera (L.) dunal under salinity stress.

Withania somnifera (L.) Dunal (Ashwagandha) has been used in herbal medicines worldwide and in the Indian traditional medicinal system for 3000 years. It is a member of the Solanaceae family distributed across Asia, Africa, Australia, and Europe. Its bioactive secondary metabolite (withanolide) biosynthesis is sensitive to salinity stress, though the mechanism remains unexplored. Therefore, we investigated the effect of Sodium chloride (NaCl) on growth, photosynthesis, biochemical traits, tissue-specific withanolide, and untargeted metabolites in W. somnifera. Ashwagandha plants were raised in pots containing soil mixture and treated with different NaCl concentrations (0 as control, 10, 30, and 50 mM) for one month inside the greenhouse. NaCl stress significantly enhanced withaferin A (WFA) (3.79 mg/g), withanolide A (WA) (0.51 mg/g), and withanone (WN) (0.022 mg/g) at 50 mM NaCl groups in the shoot. Similarly, in the root, a significant increase in WFA (0.19 mg/g) and WN (0.0016 mg/g) were observed at 10 mM, WA (0.059 mg/g) at 30 mM, and withanolide B (WB) (0.013 mg/g) at 50 mM NaCl groups compared to control. LC-MS-based untargeted metabolite profiling revealed 37 differentially accumulated metabolites in all groups. Maximum abundance of glycyl-hydroxyproline (8X) followed by tyrosyl-valine (2X) and 3-hydroxy-beta-ionone (2X) were recorded at 50 mM NaCl groups compared to the control. This study showed for the first time that low NaCl stress enhances the biosynthesis of tissue-specific withanolides through physio-biochemical and metabolites adjustment. Overall, we demonstrated a multifaceted approach for cultivating medicinal crops in salt-affected areas with enhanced bioactive metabolites for healthcare and pharmaceutical industries.

Nanoparticle-mediated amelioration of drought stress in plants: a systematic review.

Drought stress remains one of the most detrimental environmental constraints that hampers plant growth and development resulting in reduced yield and leading to economic losses. Studies have highlighted the beneficial role of carbon-based nanomaterials (NMs) such as multiwalled carbon nanotubes (MWNTs), single-walled carbon nanotubes (SWNTs), graphene, fullerene, and metal-based nanoparticles (NPs) (Ag, Au, Cu, Fe2O3, TiO2, and ZnO) in plants under unfavorable conditions such as drought. NPs help plants cope with drought by improving plant growth indices and enhancing biomass. It improves water and nutrient uptake and utilization. It helps retain water by altering the cell walls and regulating stomatal closure. The photosynthetic parameters in NP-treated plants reportedly improved with the increase in pigment content and rate of photosynthesis. Due to NP exposure, the activation of enzymatic and nonenzymatic antioxidants has reportedly improved. These antioxidants play a significant role in the defense system against stress. Studies have reported the accumulation of osmolytes and secondary metabolites. Osmolytes scavenge reactive oxygen species, which can cause oxidative stress in plants. Secondary metabolites are involved in the water retention process, thus improving plant coping strategies with stress. The deleterious effects of drought stress are alleviated by reducing malondialdehyde resulting from lipid peroxidation. Reactive oxygen species accumulation is also controlled with NP treatment. Furthermore, NPs have been reported to regulate the expression of drought-responsive genes and the biosynthesis of phytohormones such as abscisic acid, auxin, gibberellin, and cytokinin, which help plants defend against drought stress. This study reviewed 72 journal articles from 192 Google Scholar, ScienceDirect, and PubMed papers. In this review, we have discussed the impact of NP treatment on morphological, physio-biochemical, and molecular responses in monocot and dicot plants under drought conditions with an emphasis on NP uptake, transportation, and localization.

Discovery of novel and potent heterocyclic carboxylic acid derivatives as protein tyrosine phosphatase 1B inhibitors.

A series of novel heterocyclic carboxylic acid based protein tyrosine phosphatase 1B (PTP1B) inhibitors with hydrophobic tail have been synthesized and characterized. Structure-activity relationship (SAR) optimization resulted in identification of several potent, selective (over the highly homologous T-cell protein tyrosine phosphatase, TCPTP) and metabolically stable PTP1B inhibitors. Compounds 7a, 19a and 19c showed favorable cell permeability and pharmacokinetic properties in mouse with moderate to very good oral (% F=13-70) bio-availability.

Differential Gene Expression and Withanolides Biosynthesis During in vitro and ex vitro Growth of Withania somnifera (L.) Dunal.

Ashwagandha (Withania somnifera L. Dunal) is a medicinally important plant with withanolides as its major bioactive compounds, abundant in the roots and leaves. We examined the influence of plant growth regulators (PGRs) on direct organogenesis, adventitious root development, withanolide biosynthetic pathway gene expression, withanolide contents, and metabolites during vegetative and reproductive growth phases under in vitro and ex vitro conditions. The highest shooting responses were observed with 6-benzylaminopurine (BAP) (2.0 mg L-1) + Kinetin (KIN) (1.5 mg L-1) supplementation. Furthermore, BAP (2.0 mg L-1) + KIN (1.5 mg L-1) + gibberellic acid (GA3) (0.5 mg L-1) exhibited better elongation responses with in vitro flowering. Half-strength MS medium with indole-3-butyric acid (IBA) (1.5 mg L-1) exhibited the highest rooting responses and IBA (1.0 mg L-1) with highest fruits, and overall biomass. Higher contents of withaferin A (WFA) [∼8.2 mg g-1 dry weight (DW)] were detected in the reproductive phase, whereas substantially lower WFA contents (∼1.10 mg g-1 DW) were detected in the vegetative phase. Cycloartenol synthase (CAS) (P = 0.0025), sterol methyltransferase (SMT) (P = 0.0059), and 1-deoxy-D-xylulose-5-phosphate reductase (DXR) (P = 0.0375) genes resulted in a significant fold change in expression during the reproductive phase. The liquid chromatography-mass spectrometry (LC-MS) analysis revealed metabolites that were common (177) and distinct in reproductive (218) and vegetative (167) phases. Adventitious roots cultured using varying concentrations of indole-3-acetic acid (IAA) (0.5 mg L-1) + IBA (1.0 mg L-1) + GA3 (0.2 mg L-1) exhibited the highest biomass, and IAA (0.5 mg L-1) + IBA (1.0 mg L-1) exhibited the highest withanolides content. Overall, our findings demonstrate the peculiarity of withanolide biosynthesis during distinct growth phases, which is relevant for the large-scale production of withanolides.

Red laser-mediated alterations in seed germination, growth, pigments and withanolide content of Ashwagandha [Withania somnifera (L.) Dunal].

Withania somnifera (L.) Dunal, generally well-known as Ashwagandha, is part of Indian traditional medicinal systems like Ayurveda, Siddha, and Unani for over 3000 years for treating an array of disorders. The chief bioactive component of this plant is the withanolides, a group of C28-steroidal lactone triterpenoids. These compounds are present in very low concentrations and hence cell culture methods have been used to enhance their production. Low-level laser irradiation has been reported to have elicited the seed germination, agronomical characters, biosynthesis of bioactive compounds in some plants. Therefore, the objective of the study was to investigate the effect of red (He-Ne) laser irradiation on seed germination, growth characters, pigment contents and withanolide content in W. somnifera. The seeds were inoculated onto two different combinations of Murashige and Skoog (MS) media and incubated for germination. The highest germination percentage was observed in ½ MS with pH 6.5 and GA3 presoaking followed by ½ MS with different pH. Four different doses of Helium-Neon (He-Ne) laser (10, 15, 20 and 25 J/cm2) were used to irradiate the seeds at 632.8 nm and germinated in vitro on ½ MS with pH 6.5. The maximum germination percentage, 63.88% was noted from seeds irradiated with 25 J/cm2 (P = 0.04). The highest total length of 13.33 cm was observed in the seedlings irradiated with 25 J/cm2 groups (P = 0.008). The highest total chlorophyll content of 329.5 µg/g fresh weight (FW) was observed for seedlings irradiated with 15 J/cm2 (P = 0.02) and the highest carotenoid content of 49.6 µg/g FW was observed for 25 J/cm2 treated seedlings. Further, primary root length was measured and found to be highest (11.14 cm) in seedlings irradiated with 10 J/cm2 and the highest number of lateral roots were observed for 15 and 25 J/cm2 groups. The significant amount of Withanolide A (WA) 0.52 µg/g dry weight (DW) and 0.60 µg/g DW was noted in 15 (P = 0.01) and 20 J/cm2 (P = 0.002) groups, respectively than control. The present investigation thus reveals the positive impact of red laser on the germination of seeds, growth characters and withanolide contents under in vitro environment.

A2B adenosine receptor antagonists: Design, synthesis and biological evaluation of novel xanthine derivatives.

A2BAdoR is a low affinity adenosine receptor that functions by Gs mediated elevation of cAMP and subsequent downstream signaling. The receptor has been implicated in lung inflammatory disorders like COPD and asthma. Several potent and selective A2BAdoR antagonists have been reported in literature, however most of the compounds suffer from poor pharmacokinetic profile. Therefore, with the aim to identify novel, potent and selective A2BAdoR antagonists with improved pharmacokinetic properties, we first explored more constrained form of MRS-1754 (4). To improve the metabolic stability, several linker modifications were attempted as replacement of amide linker along with different phenyl or other heteroaryls between C8 position of xanthine head group and terminal phenyl ring. SAR optimization resulted in identification of two novel A2BAdoR antagonists, 8-{1-[5-Oxo-1-(4-trifluoromethyl-phenyl)-pyrrolidin-3-ylmethyl]-1H-pyrazol-4-yl}-1,3-dipropyl-xanthine (31) and 8-(1-{2-Oxo-2-[4-(3-trifluoromethyl-phenyl)-piperazin-1-yl]-ethyl}-1H-pyrazol-4-yl)-1,3-dipropyl-xanthine (65), with high binding affinity (Ki = 1 and 1.5 nM, respectively) and selectivity for A2BAdoR with very good functional potency of 0.9 nM and 4 nM, respectively. Compound 31 and 65 also displayed good pharmacokinetic properties in mice with 27% and 65% oral bioavailability respectively. When evaluated in in vivo mice model of asthma, compound 65 also inhibited airway inflammation and airway reactivity in ovalbumin induced allergic asthma at 3 mpk dose.

Discovery of Potent and Selective A2A Antagonists with Efficacy in Animal Models of Parkinson's Disease and Depression.

Adenosine A2A receptor (A2AAdoR) antagonism is a nondopaminergic approach to Parkinson's disease treatment that is under development. Earlier we had reported the therapeutic potential of 7-methoxy-4-morpholino-benzothiazole derivatives as A2AAdoR antagonists. We herein described a novel series of [1,2,4]triazolo[5,1-f]purin-2-one derivatives that displays functional antagonism of the A2A receptor with a high degree of selectivity over A1, A2B, and A3 receptors. Compounds from this new scaffold resulted in the discovery of highly potent, selective, stable, and moderate brain penetrating compound 33. Compound 33 endowed with satisfactory in vitro and in vivo pharmacokinetics properties. Compound 33 demonstrated robust oral efficacies in two commonly used models of Parkinson's disease (haloperidol-induced catalepsy and 6-OHDA lesioned rat models) and depression (TST and FST mice models).

Design and synthesis of novel xanthine derivatives as potent and selective A2B adenosine receptor antagonists for the treatment of chronic inflammatory airway diseases.

Adenosine induces bronchial hyperresponsiveness and inflammation in asthmatics through activation of A2B adenosine receptor (A2BAdoR). Selective antagonists have been shown to attenuate airway reactivity and improve inflammatory conditions in pre-clinical studies. Hence, the identification of novel, potent and selective A2BAdoR antagonist may be beneficial for the potential treatment of asthma and Chronic Obstructive Pulmonary Disease (COPD). Towards this effort, we explored several prop-2-ynylated C8-aryl or heteroaryl substitutions on xanthine chemotype and found that 1-prop-2-ynyl-1H-pyrazol-4-yl moiety was better tolerated at the C8 position. Compound 59, exhibited binding affinity (Ki) of 62 nM but was non-selective for A2BAdoR over other AdoRs. Incorporation of substituted phenyl on the terminal acetylene increased the binding affinity (Ki) significantly to <10 nM. Various substitutions on terminal phenyl group and different alkyl substitutions on N-1 and N-3 were explored to improve the potency, selectivity for A2BAdoR and the solubility. In general, compounds with meta-substituted phenyl provided better selectivity for A2BAdoR compared to that of para-substituted analogs. Substitutions such as basic amines like pyrrolidine, piperidine, piperazine or cycloalkyls with polar group were tried on terminal acetylene, keeping in mind the poor solubility of xanthine analogs in general. However, these substitutions led to a decrease in affinity compared to compound 59. Subsequent SAR optimization resulted in identification of compound 46 with high human A2BAdoR affinity (Ki = 13 nM), selectivity against other AdoR subtypes and with good pharmacokinetic properties. It was found to be a potent functional A2BAdoR antagonist with a Ki of 8 nM in cAMP assay in hA2B-HEK293 cells and an IC50 of 107 nM in IL6 assay in NIH-3T3 cells. Docking study was performed to rationalize the observed affinity data. Structure-activity relationship (SAR) studies also led to identification of compound 36 as a potent A2BAdoR antagonist with Ki of 1.8 nM in cAMP assay and good aqueous solubility of 529 µM at neutral pH. Compound 46 was further tested for in vivo efficacy and found to be efficacious in ovalbumin-induced allergic asthma model in mice.

Design, Synthesis of Novel, Potent, Selective, Orally Bioavailable Adenosine A2A Receptor Antagonists and Their Biological Evaluation.

Our initial structure-activity relationship studies on 7-methoxy-4-morpholino-benzothiazole derivatives featured by aryloxy-2-methylpropanamide moieties at the 2-position led to identification of compound 25 as a potent and selective A2A adenosine receptor (A2AAdoR) antagonist with reasonable ADME and pharmacokinetic properties. However, poor intrinsic solubility and low to moderate oral bioavailability made this series unsuitable for further development. Further optimization using structure-based drug design approach resulted in discovery of potent and selective adenosine A2A receptor antagonists bearing substituted 1-methylcyclohexyl-carboxamide groups at position 2 of the benzothiazole scaffold and endowed with better solubility and oral bioavailability. Compounds 41 and 49 demonstrated a number of positive attributes with respect to in vitro ADME properties. Both compounds displayed good pharmacokinetic properties with 63% and 61% oral bioavailability, respectively, in rat. Further, compound 49 displayed oral efficacy in 6-OHDA lesioned rat model of Parkinson diseases.