Abhisheak Sharma

Discovery of a New Class of Natural Product-Inspired Quinazolinone Hybrid as Potent Antileishmanial agents

The high potential of quinazolinone containing natural products and their derivatives in medicinal chemistry led us to discover four novel series of 53 compounds of quinazolinone based on the concept of molecular hybridization. Most of the synthesized analogues exhibited potent leishmanicidal activity against intracellular amastigotes (IC50 from 0.65 ± 0.2 to 7.76 ± 2.1 μM) as compared to miltefosine (IC50 = 8.4 ± 2.1 μM) and nontoxic toward the J-774A.1 cell line and Vero cells. Moreover, activation of Th1 type and suppression of Th2 type immune responses and induction in nitric oxide generation proved that 8a and 8g induce murine macrophages to prevent survival of parasites. Compounds 8a and 8g exhibited significant in vivo inhibition of parasite 73.15 ± 12.69% and 80.93 ± 10.50% against Leishmania donovani /hamster model. Our results indicate that compounds 8a, 8g, and 9f represent a new structural lead for this serious and neglected disease.

Dried blood spots: concepts, present status, and future perspectives in bioanalysis.

Over the past several years, dried blood spot (DBS) sampling technique has emerged as a pertinent method in both qualitative and quantitative bioanalysis context. In the DBS method, the blood sample is directly soaked on to a paper (with or without treatment). After drying it can be analyzed by modern analytical, immunological, or genomic detection systems. Several advantages of the DBS technique such as low blood volume requirement, transportation and storage without special treatment, better analytes stability, enhanced clinical cooperation in clinical trials, and reduced unforeseeable exposure of analysts to biohazards, make it the most appropriate blood sampling technique. This review illustrates the information available on the DBS method which may serve as a single window for investigators in the field of bioanalysis. Also, it explores the proficiency and appliance of the DBS method in pharmacokinetic (PK), therapeutic drug monitoring (TDM), toxicokinetic (TK), metabolomic, and disease diagnosis.

Synthesis and bioevaluation of novel 4- aminoquinoline-tetrazole derivatives as potent antimalarial agents

A series of novel tetrazole derivatives of 4-aminoquinoline were synthesized and screened for their antimalarial activities against both chloroquine-senstive (3D7) and chloroquine-resistant (K1) strains of Plasmodium falciparum as well as for cytotoxicity against VERO cell lines. Most of the synthesized compounds exhibited potent antimalarial activity as compared to chloroquine against K1-strain. Compounds with significant in vitro antimalarial activity were then evaluated for their in vivo efficacy in Swiss mice against Plasmodium yoelii following both intraperitoneal (ip) and oral administration, wherein compounds 20 and 23 each showed in vivo suppression of 99.99% parasitaemia on day 4.

Novel pre-clinical methodologies for pharmacokinetic drug-drug interaction studies: spotlight on "humanized" animal models.

Abstract Poly-therapy is common due to co-occurrence of several ailments in patients, leading to the elevated possibility of drug–drug interactions (DDI). Pharmacokinetic DDI often accounts for severe adverse drug reactions in patients resulting in withdrawal of drug from the market. Hence, the prediction of DDI is necessary at pre-clinical stage of drug development. Several human tissue and cell line-based in vitro systems are routinely used for screening metabolic and transporter pathways of investigational drugs and for predicting their clinical DDI potentials. However, ample constraints are associated with the in vitro systems and sometimes in vitro–in vivo extrapolation (IVIVE) fail to assess the risk of DDI in clinic. In vitro–in vivo correlation model in animals combined with human in vitro studies may be helpful in better prediction of clinical outcome. Native animal models vary remarkably from humans in drug metabolizing enzymes and transporters, hence, the interpretation of results from animal DDI studies is difficult. With the advent of modern molecular biology and engineering tools, novel pre-clinical animal models, namely, knockout rat/mouse, transgenic rat/mouse with humanized drug metabolizing enzymes and/or transporters and chimeric rat/mouse with humanized liver are developed. These models nearly simulate human-like drug metabolism and help to validate the in vivo relevance of the in vitro human DDI data. This review briefly discusses the application of such novel pre-clinical models for screening various type of DDI along with their advantages and limitations.

Novel, potent, orally bioavailable and selective mycobacterial ATP synthase inhibitors that demonstrated activity against both replicating and non-replicating M. tuberculosis.

The mycobacterial F0F1-ATP synthase (ATPase) is a validated target for the development of tuberculosis (TB) therapeutics. Therefore, a series of eighteen novel compounds has been designed, synthesized and evaluated against Mycobacterium smegmatis ATPase. The observed ATPase inhibitory activities (IC50) of these compounds range between 0.36 and 5.45μM. The lead compound 9d [N-(7-chloro-2-methylquinolin-4-yl)-N-(3-((diethylamino)methyl)-4-hydroxyphenyl)-2,3-dichlorobenzenesulfonamide] with null cytotoxicity (CC50>300μg/mL) and excellent anti-mycobacterial activity and selectivity (mycobacterium ATPase IC50=0.51μM, mammalian ATPase IC50>100μM, and selectivity >200) exhibited a complete growth inhibition of replicating Mycobacterium tuberculosis H37Rv at 3.12μg/mL. In addition, it also exhibited bactericidal effect (approximately 2.4log10 reductions in CFU) in the hypoxic culture of non-replicating M. tuberculosis at 100μg/mL (32-fold of its MIC) as compared to positive control isoniazid [approximately 0.2log10 reduction in CFU at 5μg/mL (50-fold of its MIC)]. The pharmacokinetics of 9d after p.o. and IV administration in male Sprague-Dawley rats indicated its quick absorption, distribution and slow elimination. It exhibited a high volume of distribution (Vss, 0.41L/kg), moderate clearance (0.06L/h/kg), long half-life (4.2h) and low absolute bioavailability (1.72%). In the murine model system of chronic TB, 9d showed 2.12log10 reductions in CFU in both lung and spleen at 173μmol/kg dose as compared to the growth of untreated control group of Balb/C male mice infected with replicating M. tuberculosis H37Rv. The in vivo efficacy of 9d is at least double of the control drug ethambutol. These results suggest 9d as a promising candidate molecule for further preclinical evaluation against resistant TB strains.

Role of enterohepatic recirculation in drug disposition: cooperation and complications

Abstract Enterohepatic recirculation (EHC) concerns many physiological processes and notably affects pharmacokinetic parameters such as plasma half-life and AUC as well as estimates of bioavailability of drugs. Also, EHC plays a detrimental role as the compounds/drugs are allowed to recycle. An in-depth comprehension of this phenomenon and its consequences on the pharmacological effects of affected drugs is important and decisive in the design and development of new candidate drugs. EHC of a compound/drug occurs by biliary excretion and intestinal reabsorption, sometimes with hepatic conjugation and intestinal deconjugation. EHC leads to prolonged elimination half-life of the drugs, altered pharmacokinetics and pharmacodynamics. Study of the EHC of any drug is complicated due to unavailability of the apposite model, sophisticated procedures and ethical concerns. Different in vitro and in vivo methods for studies in experimental animals and humans have been devised, each having its own merits and demerits. Involvement of the different transporters in biliary excretion, intra- and inter-species, pathological and biochemical variabilities obscure the study of the phenomenon. Modeling of drugs undergoing EHC has always been intricate and exigent models have been exploited to interpret the pharmacokinetic profiles of drugs witnessing multiple peaks due to EHC. Here, we critically appraise the mechanisms of bile formation, factors affecting biliary drug elimination, methods to estimate biliary excretion of drugs, EHC, multiple peak phenomenon and its modeling.

A combination of complexation and self-nanoemulsifying drug delivery system for enhancing oral bioavailability and anticancer efficacy of curcumin.

Abstract Objective: Curcumin, the golden spice from Indian saffron, has shown chemoprotective action against many types of cancer including breast cancer. However, poor oral bioavailability is the major hurdle in its clinical application. In the recent years, self-nanoemulsifying drug delivery system (SNEDDS) has emerged as a promising tool to improve the oral absorption and enhancing the bioavailability of poorly water-soluble drugs. In this context, complexation with lipid carriers like phospholipid has also shown the tremendous potential to improve the solubility and therapeutic efficacy of certain drugs with poor oral bioavailability. Methods: In the present investigation, a systematic combination of both the approaches is utilized to prepare the phospholipid complex of curcumin and facilitate its incorporation into SNEDDS. The combined use of both the approaches has been explored for the first time to enhance the oral bioavailability and in turn increase the anticancer activity of curcumin. Results: As evident from the pharmacokinetic studies and in situ single pass intestinal perfusion studies in Sprague–Dawley rats, the optimized SNEDDS of curcumin–phospholipid complex has shown enhanced oral absorption and bioavailability of curcumin. The cytotoxicity study in metastatic breast carcinoma cell line has shown the enhancement of cytotoxic action by 38.7%. The primary tumor growth reduction by 58.9% as compared with the control group in 4T1 tumor-bearing BALB/c mice further supported the theory of enhancement of anticancer activity of curcumin in SNEDDS. Conclusion: The developed formulation can be a potential and safe carrier for the oral delivery of curcumin.

Gender-related pharmacokinetics and bioavailability of a novel anticancer chalcone, cardamonin, in rats determined by liquid chromatography tandem mass spectrometry

A reversed phase liquid chromatography tandem mass spectrometry method was developed and validated for quantification of cardamonin, a potential anticancer chalcone, in rat serum. Curcumin was used as an internal standard. Following liquid-liquid extraction using n-hexane and ethyl acetate (60:40, v/v), the processed samples were chromatographed on a C18 column using acetonitrile and ammonium acetate buffer (0.01 M, pH 4.5) (85:15, v/v) as mobile phase at a flow rate of 0.6 mL min(-1). Mass spectrometric detection was performed in the negative electrospray ionization mode by multiple reaction monitoring (m/z 269→122 and 367→217 for cardamonin and curcumin, respectively). The method was validated in terms of selectivity, accuracy, precision, sensitivity, reproducibility, dilution integrity and stability. The linearity was established in the range of 1-200 ng mL(-1) (r≥0.999). The recovery of cardamonin from spiked serum was always >90%. The intra- and inter-day precision (%RSD) and accuracy (%bias) were well within the acceptable limits. The method was applied for single oral and intravenous dose pharmacokinetics in male and female Sprague Dawley rats. Following oral dose, cardamonin showed peak serum concentration that occurred at ∼2 h with very low bioavailability in both male (0.6%) and female (4.8%) rats. Cardamonin exhibited a significant gender influence on pharmacokinetics and bioavailability in rats.

Preclinical Evaluation of DMA, a Bisbenzimidazole as Radioprotector: Toxicity, Pharmacokinetics and Biodistribution Studies in Balb/c Mice

Radiotherapy, a therapeutic modality of cancer treatment, nonselectively damages normal tissues as well as tumor tissues. The search is ongoing for therapeutic agents that selectively reduce radiation-induced normal tissue injury without reducing tumoricidal effect, thereby increasing the therapeutic ratio of radiation therapy. Our laboratory established 5-(4-methylpiperazin-1-yl)-2-[2ʹ-(3,4-dimethoxyphenyl)-5′benzimidazolyl] benzimidazole (DMA) as noncytotoxic radioprotector in mammalian cells. DMA showed an excellent radioprotection in mice at single nontoxic oral dose by a dose-reduction factor of 1.28. An oxygen radical absorbing capacity assay confirmed its free-radical quenching ability. Single bolus dose and 28-days of repeated administration of DMA in mice for toxicity studies determined an LD50 of >2000 mg/kg body weight (bw) and 225 mg/kg bw, respectively, suggesting DMA is safe. Histopathology, biochemical parameters, and relative organ weight analysis revealed insignificant changes in the DMA-treated animals. The pharmacokinetic study of DMA at oral and intravenous doses showed its Cmax = 1 hour, bioavailability of 8.84%, elimination half-life of 4 hours, and an enterohepatic recirculation. Biodistribution study in mice with Ehrlich ascites tumors showed that 99mTc-DMA achieved its highest concentration in 1 hour and was retained up to 4 hours in the lungs, liver, kidneys, and spleen, and in a low concentration in the tumor, a solicited property of any radioprotector to protect normal cells over cancerous cells. We observed that the single-dose treatment of tumor-bearing mice with DMA 2 hours before 8 Gy total body irradiation showed an impressive rescue of radiation-induced morbidity in terms of weight loss and mortality without a change in tumor response.

Identification of novel phenyl butenonyl C-glycosides with ureidyl and sulfonamidyl moieties as antimalarial agents.

A new series of C-linked phenyl butenonyl glycosides bearing ureidyl(thioureidyl) and sulfonamidyl moieties in the phenyl rings were designed, synthesized, and evaluated for their in vitro antimalarial activities against Plasmodium falciparum 3D7 (CQ sensitive) and K1 (CQ resistant) strains. Among all the compounds screened the C-linked phenyl butenonyl glycosides bearing sulfonamidyl moiety (5a) and ureidyl moiety in the phenyl ring (7d and 8c) showed promising antimalarial activities against both 3D7 and K1 strains with IC50 values in micromolar range and low cytotoxicity offering new HITS for further exploration.