Discovery of biomarkers for early detection of prostate cancer: Identification of biomarkers that are capable of detecting prostate cancer early and more accurately than currently possible is necessary to prevent prostate cancer-related deaths. Ideally, early detection biomarkers should be present in easily obtainable body fluids, such as blood, which are available in a procedure acceptable to an asymptomatic population. With these goals in mind, we developed RT-PCR differential display (DD) technology-based methodology called averaged differential expression (ADE) capable of detecting protein-coding (mRNA) and non-coding RNA (ncRNA) transcripts whose expression is altered, i.e., overexpressed or downregulated, in a majority of patients, and established proof-of-principle that the level of these transcripts in blood can discriminate between prostate cancer patients and healthy controls. We are currently using this ADE methodology, which is capable of interrogating ~96% of the transcriptome, to identify differentially expressed RNA transcripts (DE-transcripts) in tumor vs. patient-matched nontumor tissues. The goal of our studies is to identify DE-transcripts that can detect prostate cancer in blood earlier and more accurately than currently possible. Our study will also determine whether markers of early disease differ between African American (AA) and European American (EA) men. Together, these studies are expected to discover a battery of RNA transcripts that are cancer-specific and establish a "signature" or "barcode" of DE-transcripts for early detection of prostate cancer in AA and EA men that is curable with currently available therapies.
Novel targets to incapacitate androgen receptor for the treatment of prostate cancer: Androgen, by activating androgen receptor (AR), plays a critical role in promoting proliferation and preventing death of prostate cancer (PCa) cells (Fig). Thus, androgen-ablation by pharmacological or surgical castration has been the frontline therapy for treatment of advanced PCa; however, it is not curative, and does not prevent the development of castration-resistant (CR) disease, which continues to depend on AR. Therefore, developing a curative strategy for the treatment of CR disease requires either identifying new targets that can disrupt AR-dependent proliferation of PCa cells or finding effective means to eliminate AR altogether from PCa cells. My laboratory is pursuing both these avenues to develop new therapeutic targets and treatment strategies that can render AR incapable of supporting proliferation and viability of PCa cells.
Role of AR in progression of PCa cells from G1 to S phase: Despite the use of androgen ablation to treat prostate cancer, little is known about the mechanism by which androgen regulates PCa cell proliferation. Elucidating the mechanism could provide new insights into treatment. We have established an experimental system in which PCa cells can be reversibly blocked in G0/G1 phase and then followed as they synchronously progress from G1 into S phase. We found that the AR level increased during mid-G1. Confocal microscopy revealed that AR colocalized with newly replicated DNA and DNA polymerase-alpha in the nuclei of S phase cells. Biochemical fractionation studies revealed that AR cosedimented with the DNA polymerase-alpha-containing multienzyme complex called replitase. Taken together, these observations led us to hypothesize that AR regulates PCa cell proliferation at least in part by participating in the assembly and/or function of the DNA replication machinery. We are investigating structural composition of AR-associated replication machinery to identify differences in protein composition of complexes isolated from androgen-sensitive vs. androgen-independent PCa cells that both express and require AR for proliferation. Such proteins may contribute to hormone-refractory proliferation and prove to be viable targets for the treatment of hormone-refractory PCa.
Targeting calmodulin to eliminate AR from PCa cells: We reported that calmodulin (CaM) binds directly to AR and regulates AR activity and stability in PCa cells. We also reported that, like most other CaM-binding proteins, AR undergoes Ca++-dependent, calpain-mediated proteolysis in PCa cells. Our most recent studies demonstrated that CaM plays a direct role in protecting AR from calpain-mediated proteolysis both in-vitro and in intact cells. These seminal observations formed the basis for our hypothesis that CaM binding to AR regulates AR transcriptional activity and AR protein stability. Con¬sequ¬ently, disruption of CaM-AR interaction inhibits AR activity and exposes AR to calpain-mediated breakdown in PCa cells. We are investigating this possibility by using molecular approaches to identify amino acid sequence in AR that binds to CaM. AR peptide containing CaM-binding sequence will then be used in crystallography and molecular modeling studies to design small molecules that can selectively disrupt CaMAR interaction and thereby render AR susceptible to proteolytic breakdown. Thus, these studies examine for the first time potential involvement a major Ca++-binding protein CaM in regulation of AR activity and AR protein stability, and lead to the development of a targeted approach for the treatment of disseminated PCa by disrupting CaMAR interaction.
1992, 1993, Site Visit Committe Member for NIH, NCI
1993, Special Review Committee Member for NIH, NCI (RFA CA-92-22)
1993, Special Review Committee Member for NIH, NIDDK (RFA DK-94-05)
1984 -present, Member of American Society of Biological Chemists 1
982 -present, Member of American Association for the Advancement of Science
2002 -present, Member of American Association for Cancer Research
2004-present, Member of Editorial Board of the Journal of Cellular Biochemistry