The major objective of this application has been to explore hormonal and nutritional regulation in experimental cancer, and to apply the knowledge obtained in animal studies towards the understanding and control of cancer in man. More specifically, these studies have examined the effects of thyroid hormones and of riboflavin deficiency upon synthesis and tissue concentrations of the flavin coenzymes, FMN and FAD, and their relation to survival of animals bearing transplants of Novikoff hepatoma. Riboflavin deficiency significantly prolongs the 50% survival time of tumor-bearing animals from 5.7 to 10.8 days. Recently completed studies indicate that in tumor from riboflavin deficient rats, the concentration of FAD is nearly identical to that in tumor from animals on a normal diet. By contrast, FMN concentrations are significantly lower in tumor from deficient than from control animals. Free riboflavin concentrations in tumors from deficient animals are decreased even further, and are undetectable in 1/3 of tumor samples assayed. FAD pyrophosphorylase activity is increased in liver but not in tumor from riboflavin deficient animals. Measurements of C14-Fad synthesis from C14-riboflavin have been performed employing newly devised methods of isotope dilution and ion exchange column chromatography with DEAE-Sephadex A-25. These data provide evidence that the rate of synthesis and/or turnover of FAD is much greater in tumor than in liver. To prepare for clinical studies in patients with cancer, a new assay for riboflavin in urine has been devised based upon the principle of competitive protein binding, and employing the protein from chicken egg white which binds riboflavin with high affinity and high specificity.