Integrated Pharmacokinetic-Driven Approach to Screen Candidate Anticancer Drugs for Brain Tumor Chemotherapy

The goal of the study was to develop an effective screening strategy to select new agents for brain tumor chemotherapy from a series of low molecular weight anticancer agents [ON123x] by the combined use of in silico, in vitro cytotoxicity, and in vitro ADME profiling studies. The results of these studies were cast into a pipeline of tier 1 and tier 2 procedures that resulted in the identification of ON123300 as the lead compound. Of the 154 ON123xx compounds, 13 met tier 1 screening criteria based on physicochemical properties [i.e., MW < 450 Da, predicted log P between 2 and 3.5] and in vitro glioma cell cytotoxicity [i.e., IC50 < 10 μM] and were further tested in tier 2 assays. The tier 2 profiling studies consisted of metabolic stability, MDCK-MDR1 cell permeability and plasma and brain protein binding that were combined to globally assess whether favorable pharmacokinetic properties and brain penetration could be achieved in vivo. In vivo cassette dosing studies were conducted in mice for 12 compounds that permitted examination of in vitro/in vivo relationships that confirmed the suitability of the in vitro assays. A parameter derived from the in vitro assays accurately predicted the extent of drug accumulation in the brain based on the area under the drug concentration–time curve in brain measured in the cassette dosing study (r² = 0.920). Overall, the current studies demonstrated the value of an integrated pharmacokinetic-driven approach to identify potentially efficacious agents for brain tumor chemotherapy.

Electronic supplementary material

The online version of this article (doi:10.1208/s12248-012-9428-4) contains supplementary material, which is available to authorized users.KEY WORDS: brain tumor, CNS, drug development, pharmacokinetics, preclinical     

Discovery of novel DPP IV inhibitors: application of pharmacophore-based virtual screening

Dipeptidyl peptidase IV (DPP-IV) is a potential drug target for type-2 diabetes and DPP-IV inhibitors are known to efficiently improve glucose tolerance. In the present study, pharmacophore model for a set of 29 DPP-IV inhibitors was generated by ligand-based pharmacophore generation process. The best hypothesis, hypo 1, consisting of four chemical features, namely, one hydrogen bond donor, one hydrogen bond acceptor, one hydrophobe and one ring aromatic was validated by cost function analysis, test set prediction and Fischer test. The validated pharmacophore model was then used for searching new lead compounds from Maybridge and NCI database. Four compounds (CD01797, CD06202, CD02493, and AW01077) from Maybridge database and three compounds (NSC997, NSC2450, and NSC5815) from NCI database were identified as structurally diverse druggable novel leads with nM activity against DPP-IV.     

Topical delivery of Idebenone using nanostructured lipid carriers: evaluations of sun-protection and anti-oxidant effects

The objective of present study was to develop nanostructured lipid carriers (NLC) for topical delivery of antioxidant drug and evaluation of its sun protection efficacy. In the present study attempts have been made to formulate Idebenone loaded nanostructured lipid carriers (INLC) by using solvent precipitation method. Preformulation study evidenced for selection of Captex 500 P as an oil phase in which Idebenone has saturation solubility of 0.266 ± 0.032 g/ml. Compritol 888 ATO and ethanol were selected as solid lipid and solvent respectively. Surfactant and co-surfactant as Labrasol and Transcutol P have given stable formulations on the basis of HLB required for stabilization, respect to oil phase. INLC has particle size of 605 ± 4.01 nm and %EE of 82.58 ± 2.20 %. Optimized batches were subjected for crystallographic investigation, in vitro skin permeation study, drug deposition study, SPF determination and antioxidant activity. XRD, DSC studies illustrated that partial amorphization of Idebenone by molecularly dispersion within lipid blend leads for entrapment of drug. Permeation data showed that optimized INLC has flux value (Jss) of 7.87 μg cm^?2 h^?1. High significance (P < 0.001) of drug deposition in skin was observed between INLC and plain Idebenone gel. SPF value for INLC has 23 which represents that lipid nanocarriers have standards of blocking of 94–96 % of UVB rays. Such high skin deposition and SPF leads to more antioxidant effect of formulations. Hence lipid nanocarriers such as NLC have potential as an antioxidant and sun protection for topical drug delivery.     

Role of Ocimum sanctum leaf extract on dietary supplementation in the transgenic Drosophila model of Parkinson＇s disease

AIM： To evaluate the effect of Ocimum sanctum leaf extract on the dietary supplementation in the transgenic Drosophila model of Parkinson＇s disease. METHOD： The effect of Ocimum sanctum leaf extract was studied on the transgenic Drosophila model of flies expressing normal human alpha synuclein（h-αs） in the neurons. O. sanctum extract at final concentrations of 0.042 8 × 10-4, 0.87 × 10-4, and 1.85 × 10-4 g·mL-1 of diet were established and the flies were allowed to feed for 21 days. The climbing assay and lipid peroxidation were taken as parameters for the study. RESULTS： The supplementation of O. sanctum extract showed a dose-dependent significant delay in the loss of climbing ability and reduction in oxidative stress in the brain of PD model flies. CONCLUSION： The results of the present study showed that the O. sanctum extract is potent in reducing the PD symptoms in transgenic Drosophila model.     

Cloning and expression of an envelope gene of West Nile virus and evaluation of the protein for use in an IgM ELISA

West Nile virus (WNV) is a neurotropic flavivirus that has emerged globally as a significant cause of viral encephalitis. The early confirmatory diagnosis of WNV infections is important for timely clinical management and epidemiologic control in areas where multiple flaviviruses are endemic. The coexistence of WNV along with other members of flaviviruses like dengue and Japanese encephalitis in India has complicated the serodiagnosis due to cross-reactive antigens. In the present study, the development and evaluation of a highly sensitive and specific IgM enzyme-linked immunosorbent assay (ELISA) using the recombinant envelope protein (rWNV-Env) for rapid, early, and accurate diagnosis of WNV are reported. The gene coding for the envelope protein of WNV was cloned and expressed in pET 28a vector followed by purification of recombinant protein by affinity chromatography. An indirect IgM microplate ELISA using purified rWNV-Env protein was optimized having no cross reactivity with healthy human serum. Furthermore, the specificity of this assay was confirmed by cross checking with serum samples obtained from patients with dengue and Japanese encephalitis viruses. The comparative evaluation of this rWNV-Env protein–specific IgM ELISA with plaque reduction neutralization test assay using 105 acute phase of clinical samples revealed 95% concordance with sensitivity and specificity of 92% and 97%, respectively. The positive and negative predictive values of recombinant-based Env ELISA were 94% and 96%, respectively. The recombinant envelope protein–based WNV-specific ELISA reported in this study will be useful for rapid screening of large numbers of clinical samples in endemic areas during outbreaks.     

Apocrine (cutaneous) sweat gland carcinoma of axilla with signet ring cells: a diagnostic dilemma on fine-needle aspiration cytology

Carcinoma arising in the apocrine sweat glands is rare and there are few reports describing the cytological features of this neoplasm. We describe the cytological features of a histologically confirmed apocrine carcinoma occurring in a 55-year-old man who presented with an ulcerated mass in the right axilla. Fine-needle aspiration cytology revealed features of a signet ring adenocarcinoma. The significance of this infrequently encountered neoplasm lies in its potential for diagnostic confusion with more common lesions containing signet ring cells. In an axillary mass lesion, cytological features along with clinical correlation are essential to distinguish primary apocrine carcinoma from mammary neoplasms with signet ring cells and other metastatic adenocarcinomas.     

AMPA receptor-dependent H2O2 generation in striatal medium spiny neurons but not dopamine axons: one source of a retrograde signal that can inhibit dopamine release

Dopamine-glutamate interactions in the striatum are critical for normal basal ganglia-mediated control of movement. Although regulation of glutamatergic transmission by dopamine is increasingly well understood, regulation of dopaminergic transmission by glutamate remains uncertain given the apparent absence of ionotropic glutamate receptors on dopaminergic axons in dorsal striatum. Indirect evidence suggests glutamatergic regulation of striatal dopamine release is mediated by a diffusible messenger, hydrogen peroxide (H(2)O(2)), generated downstream from glutamatergic AMPA receptors (AMPARs). The mechanism of H(2)O(2)-dependent inhibition of dopamine release involves activation of ATP-sensitive K(+) (K(ATP)) channels. However, the source of modulatory H(2)O(2) is unknown. Here, we used whole cell recording, fluorescence imaging of H(2)O(2), and voltammetric detection of evoked dopamine release in guinea pig striatal slices to examine contributions from medium spiny neurons (MSNs), the principal neurons of striatum, and dopamine axons to AMPAR-dependent H(2)O(2) generation. Imaging studies of H(2)O(2) generation in MSNs provide the first demonstration of AMPAR-dependent H(2)O(2) generation in neurons in the complex brain-cell microenvironment of brain slices. Stimulation-induced increases in H(2)O(2) in MSNs were prevented by GYKI-52466, an AMPAR antagonist, or catalase, an H(2)O(2) metabolizing enzyme, but amplified by mercaptosuccinate (MCS), a glutathione peroxidase inhibitor. By contrast, dopamine release evoked by selective stimulation of dopamine axons was unaffected by GYKI-52466 or MCS, arguing against dopamine axons as a significant source of modulatory H(2)O(2). Together, these findings suggest that glutamatergic regulation of dopamine release via AMPARs is mediated through retrograde signaling by diffusible H(2)O(2) generated in striatal cells, including medium spiny neurons, rather than in dopamine axons.