Activated ras oncogenes have been identified in a significant percentage of cancers including leukemias and carcinomas of the colon, pancreas, and lung. High affinity antibodies reactive with both normal and activated human p21ras are available. However, the use of ras antibodies to develop clinical diagnostic assay which specifically detect mutant ras have not been developed. A prototype immunometric assay developed under Phase I study for K-ras Asp-12 will be optimized for sensitivity and performance. Using an affinity chromatography procedure to immunoconcentrate p21ras from samples, the assay will be used to quantitate mutant p21ras from human tumors and sera. This will determine the feasibility of using a mutant-specific reporter antibodies reactive to p21ras containing Arg, Ser, Val, and Cys substitutions at position 12 will be generated. The corresponding recombinant proteins will be expressed and purified for antibody characterization and immunoassay development. An assay which detects several p21ras mutants will be developed, optimized, and evaluated using tumor extracts and serum. The ability to identify and quantitate activated p21ras proteins in tumors and serum may greatly facilitate the specific diagnosis and prognostic evaluation of human cancer.
