Retinoblastoma (RB) gene is the prototype cancer suppressor gene allowing tumor induction to occur in the absence of the normal RB gene product (RB protein). Recent studies with DNA and antibody probes have revealed that in addition to the prototype retinoblastoma, many other tumor cells show RB gene/protein defects. These include osteosarcoma, breast carcinoma, small cell lung carcinoma, bladder carcinoma and a variety of soft tissue sarcomas. To understand the extent of RB gene/protein defects in various tumors, it is important to survey the expression of RB protein in established tumor cell lines as well as primary tumor tissues. It is also important to develop sensitive, specific, and reliable immunodetection methods for RB protein. We have successfully developed a panel of monoclonal antibodies specific for human RB protein. These antibodies can immunoprecipitate the authentic RB protein, have been used effectively in Western blotting, and immunostain RB protein in several cell lines. In our collaborative studies using these monoclonals as probes, RB protein has been demonstrated to complex with SV-40 large T transforming protein. It has also come to appear that RB protein functions in regulating cell cycle by differential phosphorylation. Thus, the monoclonals are very useful for assessing the function of RB protein in human neoplasia, and for diagnosing RB-related tumors.
The specific aims of this proposal are: 1. To generate a complete panel of monoclonal antibodies recognizing distinct epitopes on RB protein. 2. To develop sensitive, specific and reliable monoclonal antibody-based immunodetection systems for RB protein. 3. To study the extent of RB defects in established tumor cell lines. 4. To survey the extent of RB defects in various primary tumors.
