Bio-data of

Design & development of bio-activity guided fractionated plant/microbial extracts as Radioprotectors and studies on their mode of action at the molecular level (Project CHARAK)

Radioprotective drugs belong to the category of prophylactic modalities which are administered to alleviate radiation-induced oxidative stress. The high-altitude region plants of the Himalayan region have proved to be a useful resource for the discovery of new and potent bioactive molecules with a diverse array of properties. Antioxidants available as secondary metabolites in the Himalayan herbs, i.e., Podophyllum hexandrum, Hippophae rhamnoides and Rhodiola imbricata, were screened for their radioprotective effects. Bio-activity guided fractionation has been performed to identify potent fractions.
The different extracts/fractions and synthetic antioxidants or isolated molecules were evaluated in vitro for their free radical scavenging ability, DNA protective ability, membrane protective ability and radioprotective efficacy using standardized protocols. Our endeavor to characterize the bioactive molecule(s) using modern analytical techniques like HPLC/LC-MS/NMR has yielded two new molecules. The protective effects of antioxidant(s) are also being evaluated at ex vivo and in vivo level in lower animal models. Using the antioxidants in appropriate therapeutically effective doses can repair the damage to the critical biomolecules like DNA, proteins and lipids. Effect of natural antioxidants on oxidative stress (induced both by chemical stressors or ionizing radiation) DNA damage and repair of simple as well as complex DNA damage alteration is being studied using damage specific enzymes. Electrophoretic as well as PCR based methodologies have been employed for studying random as well as gene specific damage and repair kinetics. Apart from that effect of oxidative stress on induction and repair of gene specific mutations is being evaluated both in nuclear and mitochondrial genomes.
Mechanistic studies are in progress to find out the exact mechanism of action at cellular / molecular level. Techniques like COMET assay, Western blotting, Flow cytometry, spectroflourimetry and spectrometry, Fluorescence microscopy etc. will be utilized. The transcription Factor [(Early growth responsive Factor-1 (EGR-1)] is expressed instantly following radiation-induced oxidative stress. Utilizing EGR-1 regulated the test drug will be evaluated for its antioxidative / radioprotective properties. Molecular biomarkers (gene/gene products) of dose-dependent oxidative stress are being studied.
An endophytic fungus name Trametes hirusta was isolated from the rhizomes of P. hexandrum having a mycoryzzal association with the host plant. The active radioprotective molecules including podophylotoxin and its derivatives like demethylpodophylotoxin and podophylotoxin D-glucoside was isolated from this endophytic fungus. It could be used as an alternative source of these lead molecules in higher quantity for further drug development utilizing synthentic chemical interventions.
Three research scholars have received doctorate degree in Biotechnology discipline (Titles of Ph.D. Thesis are i). Radiation-induced apoptosis and its modulation by some herbal preparations; ii). Modulatory effects of some herbal preparations on gamma radiation-induced DNA damage and repair, and iii). Influence of Podophyllum hexandrum on molecular regulation of genes responsible for radioprotection in vivo and in vitro) in May 2007, July 2007 & July 2008 respectively and one research fellow received Ph.D. in Toxicology discipline (Title of Thesis: Radiation response in mammalian model and its modulation by certain natural plant product(s)) in September 2007 while working in the above project.

3. Development of Behavioural radioprotectors

Ionising radiation is one of the causative factors in inducing nausea, vomiting and malaise. Performance decrement produced by radiation-induced emesis is an important operational factor in Defence Services. An ideal radioprotector should also possess the ability to reduce radiation-induced performance decrement. Extract of Zingiber officinale (Ginger) and Mentha piperata (Mint) appears promising in the preliminary studies based on radiation –induced Conditioned Taste Aversion studies in mice model system. Further studies are underway for fractionation and isolation of active metabolites responsible for such behavior.

 

4. Development of Radiation Biomakers for Radiation biodosimetry

Radiation induces biochemical, molecular, genetic changes and physiological alternations including various immunological responses. Risk and threat analysis revealed a requirement for novel approaches to reduce the time for analysis and improvement of sensitivity of biodosimetry techniques utilizing specific and moderately-specific biomarkers. Some of the lead areas have been identified in this direction are chromosomal related changes, specific enzyme levels, levels of Interleukin, IFN-, NF-Kappa and C-reactive compliment proteins etc. It is essential to develop integrated approach for identifiying different biomarkers followed by development of biodosimetic assessment techniques. EPR studies on hydroxyapatite, and HPLC studies on uric acid & allantoin, are in progress. A CSIR research scholar is presently working in this field under my supervision.

3. DESIGN & DEVELOPMENT OF RADIOPHARMACEUTICALS

I spearheaded Radiopharmaceutical research programme at INMAS from 1999 till Feb 2006. Besides working on development of SPECT radiopharmaceuticals for nuclear scintigraphy & therapy, I initiated work on development of magnetopharmaceuticals for contrast enhancement in Magnetic Resonance Imaging. Studies on Novel Drug Delivery Systems were commenced and pharmacoscintigraphic techniques & animal models of disease were developed for their preclinical evaluation in active collaboration with academic and industrial partners. Pathbreaking research in emerging technologies in some new R & D areas like inflammation / infection scintigraphy, PET for biopharmaceutics studies, nuclear decorporation and molecular nuclear medicine, Stem Cell Research etc., related to charter of duties of INMAS were nurtured. Details are as follows:

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