Pre-clinical toxicity & immunobiological evaluation of DNA rabies vaccine & combination rabies vaccine in rhesus monkeys (Macaca mulatta).

BACKGROUND & OBJECTIVES: Pre-clinical toxicology evaluation of biotechnology products is a challenge to the toxicologist. The present investigation is an attempt to evaluate the safety profile of the first indigenously developed recombinant DNA anti-rabies vaccine [DRV (100 µg)] and combination rabies vaccine [CRV (100 µg DRV and 1.25 IU of cell culture-derived inactivated rabies virus vaccine)], which are intended for clinical use by intramuscular route in Rhesus monkeys. METHODS: As per the regulatory requirements, the study was designed for acute (single dose - 14 days), sub-chronic (repeat dose - 28 days) and chronic (intended clinical dose - 120 days) toxicity tests using three dose levels, viz. therapeutic, average (2x therapeutic dose) and highest dose (10 x therapeutic dose) exposure in monkeys. The selection of the model i.e. monkey was based on affinity and rapid higher antibody response during the efficacy studies. An attempt was made to evaluate all parameters which included physical, physiological, clinical, haematological and histopathological profiles of all target organs, as well as Tiers I, II, III immunotoxicity parameters. RESULTS: In acute toxicity there was no mortality in spite of exposing the monkeys to 10XDRV. In sub chronic and chronic toxicity studies there were no abnormalities in physical, physiological, neurological, clinical parameters, after administration of test compound in intended and 10 times of clinical dosage schedule of DRV and CRV under the experimental conditions. Clinical chemistry, haematology, organ weights and histopathology studies were essentially unremarkable except the presence of residual DNA in femtogram level at site of injection in animal which received 10X DRV in chronic toxicity study. No Observational Adverse Effects Level (NOAEL) of DRV is 1000 ug/dose (10 times of therapeutic dose) if administered on 0, 4, 7, 14, 28 th day. INTERPRETATION & CONCLUSIONS: The information generated by this study not only draws attention to the need for national and international regulatory agencies in formulating guidelines for pre-clinical safety evaluation of biotech products but also facilitates the development of biopharmaceuticals as safe potential therapeutic agents.

Nonclinical toxicology study of recombinant-plasmid DNA anti-rabies vaccines.

The absence of standard guidelines from National and International regulatory agencies for the safety evaluation of biotechnology products challenges the ingenuity of toxicologists. At present, the development of standard pre-clinical toxicology protocols for such products is on an individual case basis. The present investigation is an attempt to evaluate the safety profile of the first indigenously developed DNA based anti-rabies vaccine in India. The test compounds were DNA rabies vaccine [DRV (100 microg)] and combination rabies vaccine (CRV (100 microg DRV and 1/50 dose of cell culture vaccine)), intended for clinical use by intramuscular route on 1, 7, 14 and 28 day. As per the regular mandatory requirements, the study has been designed to undertake acute (single dose--10 days), sub-chronic (repeat dose--28 days) and chronic (intended clinical dose--120 days) toxicity tests using three dose levels viz. therapeutic, average (2 x therapeutic dose) and highest dose (10 x therapeutic dose) exposure in Swiss Albino mice. The selection of the rodent model viz. Swiss Albino mice is based on affinity and rapid higher antibody response during the efficacy studies. Apart from physical, physiological, clinical, hematological and histopathology profiles of all target organs, the tier-I immunotoxicity parameters have also been monitored. There were no observational adverse effects even at levels of 10x therapeutic dose administration of DRV and CRV. The procedure also emphasizes on the designing of protocols for the products developed by recombinant technique.

Local immunity in Indian women with bacterial vaginosis.

OBJECTIVES: To determine the immunoglobulin (Ig) and cytokine levels and degradation of Igs in the cervico-vaginal secretions (CVS) of non-pregnant Indian women of low socio economic status (LSES), with/without bacterial vaginosis (BV) and to assess the interactions among nutritional status, BV and local immunity. METHODS: A descriptive study in non-pregnant women of LSES attending the gynecology out patient clinic at a local government hospital, Hyderabad, India. Two hundred non-pregnant women underwent clinical, anthropometrical and gynecological examination and were screened for BV. In a sub-sample of 80 with/without BV, levels of IL-10 and IL-12, IgA, IgM and IgG were determined in the CVS by ELISA and degradation of IgA and IgM by Western blotting. Statistical significance among the groups was tested using the non-parametric Mann-Whitney U-test. RESULTS: Fifty seven percent of the women tested positive for BV. Women with BMI<16.0 had the highest BV positivity and the lowest IgA levels in CVS. Higher levels of IgA were observed in women with BMI>18.5. There was significant degradation of IgA and IgM in women with BV. IL-12 was undetectable while IL-10 was detected with higher means in CVS of women with BV. CONCLUSIONS: Severe under-nutrition appears to be relevant to BV positivity and local immunity in these women. Greater degradation of IgA and IgM in BV suggests impaired local immunity.

Curcumin and turmeric delay streptozotocin-induced diabetic cataract in rats.

PURPOSE: The purpose of this study was to investigate the effect of curcumin and its source, turmeric, on streptozotocin-induced diabetic cataract in rats. METHODS: Wistar-NIN rats were selected and diabetes was induced by streptozotocin (35 mg/kg body weight, intraperitoneally) and divided into four groups (group II-V). The control (group I) rats received only vehicle. Group I and II animals received an unsupplemented AIN-93 diet, and those in groups III, IV, and V received 0.002% and 0.01% curcumin and 0.5% turmeric, respectively, in an AIN-93 diet for a period of 8 weeks. Cataract progression due to hyperglycemia was monitored by slit lamp biomicroscope and classified into four stages. At the end of 8 weeks, the animals were killed and the biochemical pathways involved in the pathogenesis of cataract such as oxidative stress, polyol pathway, alterations in protein content and crystallin profile in the lens were investigated, to understand the possible mechanism of action of curcumin and turmeric. Blood glucose and insulin levels were also determined. RESULTS: Although, both curcumin and turmeric did not prevent streptozotocin-induced hyperglycemia, as assessed by blood glucose and insulin levels, slit lamp microscope observations indicated that these supplements delayed the progression and maturation of cataract. The present studies suggest that curcumin and turmeric treatment appear to have countered the hyperglycemia-induced oxidative stress, because there was a reversal of changes with respect to lipid peroxidation, reduced glutathione, protein carbonyl content and activities of antioxidant enzymes in a significant manner. Also, treatment with turmeric or curcumin appears to have minimized osmotic stress, as assessed by polyol pathway enzymes. Most important, aggregation and insolubilization of lens proteins due to hyperglycemia was prevented by turmeric and curcumin. Turmeric was more effective than its corresponding levels of curcumin. CONCLUSIONS: The results indicate that turmeric and curcumin are effective against the development of diabetic cataract in rats. Further, these results imply that ingredients in the study's dietary sources, such as turmeric, may be explored for anticataractogenic agents that prevent or delay the development of cataract.

Use of fermented foods to combat stunting and failure to thrive.

OBJECTIVES: With the adoption of vigorous child survival strategies, infant and child mortalities in India have declined significantly, even among the poorest, most undernourished segments of the population. Of the surviving infants and children, however, many remain stunted and undernourished. The present study was based on the hypothesis that failure to thrive is the result of damage to the gut epithelium incurred during repeated bouts of gastrointestinal infections. Promoting the regeneration of the damaged gut epithelium through the use of lactobacillus-rich fermented foods may yield beneficial results. This low-cost procedure can be widely used, even in poor communities. The objectives of this study were to investigate the effect in poor Indian communities of supplementation with a probiotic on the growth of children (aged 2 to 5 years) with growth retardation and assess the difference in morbidity between those receiving the supplement (n = 50) and the control group (n = 50), mainly with respect to the frequency, severity, and duration of diarrheal episodes. METHODS: One hundred children aged 2 to 5 y from an urban slum of New Delhi were matched for their age (+/-36 d), sex and weight (+/-1 kg) and assigned to one of two groups (experimental n= 50 and control n= 50). The experimental group received a probiotic supplement (50 ml curd containing Lactobacillus acidophilus) and the control group received an isocaloric supplement daily for 6 mo. Weight, height, and morbidity profile with respect to diarrhea, fever, cough, and cold was recorded. RESULTS: Increases in weight (P < 0.002) and height (P < 0.001) were significantly greater in the experimental group than in the control group. In addition, after 6 mo, of supplementation, there were fewer cases of diarrhea (P < 0.005) and fever (P < 0.001) in the intervention group then in the control group. CONCLUSIONS: From this study, it can be concluded that 6 mo of probiotic supplementation may be beneficial with respect to decrease in diarrheal morbidity and accelerated growth in the experimental group.

Field Level Evaluation of Aflatoxin Detection Kit.

An aflatoxin detection kit was evaluated at the field level for groundnut and groundnut meal. A total of 88 groundnut samples and 20 groundnut meal were analyzed with the help of the kit in the field. Aliquots of same samples were analyzed in the laboratory by the conventional BF-thin layer chromatographic method. In 89% of the groundnut samples and 100% groundnut meal, results of the both methods were comparable. Aflatoxins were overestimated in 5% of the groundnut samples, and in 6% of the samples, aflatoxins were underestimated.