COLON-CANCER-SPECIFIC GENES AS PROBES

DUE TO AN INCOMPLETE UNDERSTANDING OF THE DISEASE, THE PROGNOSIS OF COLON CANCER, A HIGH-INCIDENCE CANCER IN WESTERN COUNTRIES, HAS NOT IMPROVED SIGNIFICANTLY. MOST OF THE STUDIES IN THE MOLECULAR BIOLOGY OF COLON CANCER HAVE FOCUSED ON THE STUDY OF A LIMITED SET OF APPROXIMATELY 25-40GENES, TERMED ONCOGENES, THAT CAN ONLY BE DETECTED IF THEY HAVE A COUNTERPART IN RETROVIRUSES, OR IF THEY CAN ONCOGENICALLY TRANSFORM SPECIALIZED CELL LINES IN TRANSFECTION EXPERIMENTS. OTHER STUDIES HAVE INDICATED THAT COLON CANCER MAY BE ASSOCIATED WITH THE LOSS OF A GENE,BUT A SINGLE GENE CHANGE DOES NOT EXPLAIN THE EXTENSIVE TISSUE CHANGES THAT OCCUR WITH COLON CANCER. GIVEN THE POTENTIALLY LARGE CHANGES IN GENE EXPRESSION THAT MAY OCCUR WHEN TISSUES BECOME MALIGNANT, IT BECOMES IMPERATIVE THAT WEARE ABLE TO IDENTIFY ALL GENES THAT ARE UNIQUELY EXPRESSED IN THE CANCEROUS STATE BY A MEANS THAT DOES NOT LIMIT WHICH GENES CAN BE DETECTED. TO ADDRESS THE PROBLEM OF IDENTIFYING AND ISOLATING THE COMPLETE SET OF GENES THAT AREEXPRESSED UNIQUELY IN THE MALIGNANT STATE, WE PROPOSE TO USEOUR NATIVE-STATE CULTURE TECHNOLOGY THAT ALLOWS CULTURED NORMAL AND CANCEROUS TISSUE TO GROW INTACT THREE-DIMENSIONALLY IN VITRO WHILE MAINTAINING IN VIVO-LIKE TISSUE ORGANIZATION AND FUNCTION IN ORDER TO ISOLATE INTACT RNA FROM CULTURED NORMAL AND CANCEROUS COLON TISSUES. THIS RNA WILL BE USED FOR SYNTHESIS OF CANCER-SPECIFIC CDNA LIBRARIES. THE LIBRARIES WILL BE SCREENED BY CANCER-SPECIFIC CDNA PROBES WHICH ARE SYNTHESIZED ON CANCEROUS MRNA, WITH THE SUBSEQUENT REMOVAL OF GENE PRODUCTSCOMMON TO TUMOR AND NORMAL TISSUE BY SELECTIVE HYBRIDIZATIONWITH NORMAL-TISSUE MRNA AND ABSORPTION BY HYDROXY APATITE CHROMATOGRAPHY. THE PROBES WHICH ARE HIGHLY ENRICHED FOR SEQUENCES EXPRESSED SPECIFICALLY IN COLON CANCER WILL HYBRIDIZE TO CANCER-SPECIFIC CDNA CLONES, THEREBY IDENTIFYING THEM AND ALLOWING THEIR ISOLATION. IF THE ISOLATED CLONES ARE FOUND TO BE COMMON AND SPECIFIC TO A LARGE PERCENTAGE OF COLON CANCERS, THEY COULD SUBSEQUENTLY BE USED AS CANCER-SPECIFIC HYBRIDIZATION PROBES IN THE DETERMINATION OF SUSCEPTIBILITY, DIAGNOSIS, STAGING, FOLLOWING THE COURSE OF AND PROGNOSIS OF COLON CANCER WHICH WILL BE CARRIED OUT IN PHASE II.