Discovery and validation of small-molecule heat-shock protein 90 inhibitors through multimodality molecular imaging in living subjects

Up-regulation of the folding machinery of the heat-shock protein 90 (Hsp90) chaperone protein is crucial for cancer progression. The two Hsp90 isoforms (α and β) play different roles in response to chemotherapy. To identify isoform-selective inhibitors of Hsp90(α/β)/cochaperone p23 interactions, we developed a dual-luciferase (Renilla and Firefly) reporter system for high-throughput screening (HTS) and monitoring the efficacy of Hsp90 inhibitors in cell culture and live mice. HTS of a 30,176 small-molecule chemical library in cell culture identified a compound, N-(5-methylisoxazol-3-yl)-2-[4-(thiophen-2-yl)-6-(trifluoromethyl)pyrimidin-2-ylthio]acetamide (CP9), that binds to Hsp90(α/β) and displays characteristics of Hsp90 inhibitors, i.e., degradation of Hsp90 client proteins and inhibition of cell proliferation, glucose metabolism, and thymidine kinase activity, in multiple cancer cell lines. The efficacy of CP9 in disrupting Hsp90(α/β)/p23 interactions and cell proliferation in tumor xenografts was evaluated by non-invasive, repetitive Renilla luciferase and Firefly luciferase imaging, respectively. At 38 h posttreatment (80 mg/kg × 3, i.p.), CP9 led to selective disruption of Hsp90α/p23 as compared with Hsp90β/p23 interactions. Small-animal PET/CT in the same cohort of mice showed that CP9 treatment (43 h) led to a 40% decrease in 18F-fluorodeoxyglucose uptake in tumors relative to carrier control-treated mice. However, CP9 did not lead to significant degradation of Hsp90 client proteins in tumors. We performed a structural activity relationship study with 62 analogs of CP9 and identified A17 as the lead compound that outperformed CP9 in inhibiting Hsp90(α/β)/p23 interactions in cell culture. Our efforts demonstrated the power of coupling of HTS with multimodality molecular imaging and led to identification of Hsp90 inhibitors

Illuminating the life of GPCRs

The investigation of biological systems highly depends on the possibilities that allow scientists to visualize and quantify biomolecules and their related activities in real-time and non-invasively. G-protein coupled receptors represent a family of very dynamic and highly regulated transmembrane proteins that are involved in various important physiological processes. Since their localization is not confined to the cell surface they have been a very attractive "moving target" and the understanding of their intracellular pathways as well as the identified protein-protein-interactions has had implications for therapeutic interventions. Recent and ongoing advances in both the establishment of a variety of labeling methods and the improvement of measuring and analyzing instrumentation, have made fluorescence techniques to an indispensable tool for GPCR imaging. The illumination of their complex life cycle, which includes receptor biosynthesis, membrane targeting, ligand binding, signaling, internalization, recycling and degradation, will provide new insights into the relationship between spatial receptor distribution and function. This review covers the existing technologies to track GPCRs in living cells. Fluorescent ligands, antibodies, auto-fluorescent proteins as well as the evolving technologies for chemical labeling with peptide- and protein-tags are described and their major applications concerning the GPCR life cycle are presented

Effect of the subterranean termite used in the South Indian folk medicine

376-379The present investigation reports the effect of the termite, Odontotermes formosanus Shiraki, most commonly used by the South Indian tribes as food for enhancing lactation in women, on growth and reproduction in Swiss albino mice, Mus musculus Linn. Dietary supplementation with termite to Swiss albino mice significantly increased the percentage growth rate and number of litters delivered, when compared to the control groups fed with normal rodent pellets. The percentage growth rate of experimental male and female mice fed on Odontotermes formosanus Shiraki showed a significant increase (P<0.01), when compared to the respective control groups. Results on litter production revealed a 22% increase in the experimental groups supplemented with termite, when compared to the control groups. The present results suggest that termite can be used as a viable protein rich feed. Studies on termite culture on a commercial scale and its use, as an alternative protein rich feed for poultry would probably go a long way

Antibacterial activity of subterranean termites used in South Indian folk medicine

559-562The investigation reports antimicrobial activity of the termite species most commonly used by the South Indian tribes for treating diseases likely to be associated with microorganisms. The antibacterial activities of 90% alcohol extracts of three species of subterranean termites, viz: Microtermes obes Holmgren, Macrotermes estherae (Desneux), and Odontotermes formosanus Shiraki, their mounds and nearby soil extracts collected from three different sites of South India were assayed against various bacterial strains. The antibacterial activity was most apparent in Odontotermes formosanus Shiraki, and its mound extracts, which inhibited all the bacterial strains studied. The highest antibacterial activity was encountered in Odontotermes formosanus (12.6 ±0.5 mm) and its mound extracts (14.3 ± 1.1 mm) against E. coli BL21. The MIC values also varied with the extracts and test organisms used and ranged from 10 µg/disc to 50 µg/disc. No activity was observed in the negative control as well as soil extracts collected from near by the termite mounds. Since, the traditional knowledge of indigenous people throughout the world has played an important role in identifying natural resources worthy of commercial exploitation and the search for new pharmaceuticals from naturally occurring biological material has been guided by ethnobiological data, the results are of significance to modern medicine

Antigenotoxic activity of the subterranean termite on Swiss albino mice

406-411The paper reports the antigenotoxic potential of the termite (Odontotermes formosanus Shiraki) supplemented food on acephate and endosulfan induced toxicity in the Swiss albino mice (Mus musculus). Dietary supplementation with termite to male and female mice Mus musculus treated with acephate and endosulfan significantly decreased the percentage of chromosomal aberration and micronuclei in the bone marrow cells than in pesticide treated F0 and F1 generations (P < 0.01). The chromosomal aberrations induced by acephate and endosulfan included minute, gap, inversion, ring, pulverization and tetraploids. The experimental groups treated with acephate/endosulfan and supplemented with termite food did not show critical genotoxic aberration markers like pulverization and ploidy suggesting a strong antigenotoxic effect of the termite components