This revised new NIH R01 application evaluates cyclin D1 as a molecular target for germ cell tumor (GCT) growth, maturation, and chemotherapy response. All-trans-retinoic acid (RA) induced a neuronal and non-tumorigenic state in the human GCT cell line, NT2/D1. We uncovered retinoic acid receptor (RARgamma) as regulating RA growth and differentiation in NT2/D1 cells and conferring RA resistance to NT2/D1-R1 cells. GCTs are useful to study tumor differentiation mechanisms. A unique clinical feature of GCTs is teratoma formation, reflecting their capacity to mature. An inverse relationship exists in GCTs between differentiation and tumorigenicity. To explore anti-neoplastic mechanisms in GCTs, NT2/D1 and NT2/D1-R1 cells were studied. NT2/D1 differentiation reproduced key features of teratomas in that RA-treatment caused these cells to mature and reduce tumorigenicity. A hallmark of this response is G1 arrest, at least partly through a previously unrecognized pathway, where retinoids trigger cyclin D1 proteolysis. This occurs partly through proteasomal degradation. RA effects on cyclin D1 are specific since cyclins D2 and D3 were not repressed. NT2/D1-R1 cells deregulated cyclin D1 expression. RARgamma transfection restored retinoid response and cyclin D1 repression to NT2/D1-R1 cells. Threonine 286 regulated cyclin D1 ubiquitination and proteasomal degradation by RA. Another relationship was that NT2/D1-R1 cells deregulated cyclin D1 and acquired cisplatin resistance, indicating a link between maturation and chemotherapy responses. Post-transcriptional and transcriptional retinoid mechanisms repressed cyclin D1. These will be studied comprehensively and compared to those of other cyclin D1 targeting agents. This is relevant for combination therapy. There is a need to uncover the cyclin D1 role in GCT biology these specific aims that: (1) elucidate proteolytic and transcriptional mechanisms that repress cyclin D1 during retinoid treatment of GCTs and learn whether other cyclin D1 targeting agents engage distinct or overlapping mechanisms; (2) validate whether cyclin D1 repression is required for GCT growth, maturation, or cisplatin response; and (3) extend findings to the clinical context by evaluating cyclin D1 as a target in a unique tissue bank with mature, immature, and chemotherapy resistant GCTs. These resources with pharmacologic, cell and molecular biological approaches would provide insights into cyclin D1 as a novel target for poor risk GCTs.
Showing the most recent 10 out of 38 publications
