MICRONUCLEI AND CARCINOGEN-DNA ADDUCTS AS INTERMEDIATE END-POINTS IN NUTRIENT INTERVENTION TRIAL OF PRECANCEROUS LESIONS IN THE ORAL CAVITY

In cancer chemoprevention trials, biomarkers as intermediate end point s have gained importance. A variety of biomarkers have been proposed a s intermediate end points for upper aerodigestive tract cancers. This study was aimed at studying the frequency of micronucleated cells and carcinogen DNA adducts as indicators of DNA damage and intervention en d points in chemoprevention trials. Reverse smokers of chutta (rolled tobacco) from four villages numbering 298 in total were selected. Out of these, 150 were supplemented with four nutrients (vitamin A, ribofl avin, zinc and selenium) and 148 controls received placebo, one capsul e twice a week for 1 year. Slides of buccal smears were prepared and s tained with Fuelgen reaction and counterstained with Fast Green and ex amined microscopically for the presence of micronucleated cells. Oral cell washings were collected and centrifuged. The DNA adducts were eva luated by the P-32 post-labelling assay method. Protein and RNA free D NA (adducted) isolated from the cells was digested with MN/SPD and the DNA adducts isolated by the butanol enrichment procedure. The DNA add ucts were identified and quantitated by multidimensional chromatograph y on PEI-TLC sheets by screen enhanced autoradiography and presented a s RAL (relative adduct labelling) values. Both the micronuclei and DNA adducts were significantly elevated in subjects with lesions. At the end of 1 year the frequency of micronuclei decreased significantly (P < 0.001) in the supplemented subjects with or without lesions. The DNA adducts in the supplement group at the end of 1 year also reduced sig nificantly. The adducts decreased by 95% in subjects with all categori es of lesions and by 72% in subjects without lesions. No such effects were noted in the placebo group. The two biomarkers investigated in th e case study appear to be modifiable by the administration of micronut rient supplements. The results are in agreement with the clinical resp onse and suggest that suppression of genetic damage was consistent wit h clinical remission. In the study, a cocktail of micronutrients was a dministered and as such no comments can be made as to the relative ben efit of each of the nutrients. However, these biomarkers used in addit ion to the clinical response of the precancerous lesions can be valuab le measures to arrive at beneficial impacts.