Incidence and mortality rates of prostate cancer vary substantially among different geographic areas and ethnic groups. Notably, African-American men have the highest risk of developing prostate cancer and, due to a more aggressive disease, their mortality rate is twice that of European-American men. While socioeconomic factors contribute to this health disparity, these factors do not fully explain the differences in prostate cancer incidence, aggressiveness, and mortality among different ethnic groups in the U.S. We hypothesized that gene expression profiles of prostate tumors from African-American and European-American men may reveal biological differences between the two groups that could explain the more aggressive cancer phenotype in African-American men. We obtained gene expression profiles of primary prostate tumors resected from 33 African-American and 36 European-American patients. These tumors were matched on clinical parameters. The resulting datasets were analyzed for expression differences on the gene and pathway level, comparing these patients. Our analysis revealed a significant number of genes to be differently expressed among the two patient groups. Using disease association analysis, we identified a significant relationship among these transcripts with autoimmunity and inflammation. These findings were corroborated on the pathway level, with numerous differently expressed genes clustering in the immune and stress response, cytokine signaling, and chemotaxis pathways. Specifically, an interferon gene signature was present in tumors of African-American patients, suggesting a possible involvement of viral infections in disease pathology. We hypothesize that the altered immune response in African-American men is a predisposing factor for tumor progression and metastasis. We do not yet know why prostate tumors from African-American patients have a different immunologic profile than tumors from European-American patients. A follow-up study of breast cancer in our laboratory showed that some of the same differences between African-American and European-American patients can be detected in breast tumors, e.g. an interferon signature in African-American tumors. Possible causes include environmental factors (e.g., infections), genetic factors (e.g., variations in immune regulatory genes), or the interactions of both. Our current and future studies are examining the involvement of these factors in the tumor biology of African-Americans. Based on these findings, we started two projects exploring the contribution of: 1) viral infections, and 2) intrinsic expression differences in immunoregulatory genes to the existing prostate cancer health disparity between African-American and European-American men. In FY09, we continued to pursue the hypothesis that viral infectious agents cause the interferon signature in tumors from African-American patients. We completed a study that used a proprietary microarray chip that can detect all known mammalian viruses (650) in total RNA preparations. We analyzed tumors from African-American (n = 26) and European-American patients (n = 37), and benign prostate hyperplasia tissue (n = 9). We detected a number of viral sequences that were enriched in prostate tumors but were not present in benign prostate hyperplasia resections. Some of these sequences refer to viruses that one would not expect in the human prostate (e.g., bovine popular stomatitis virus, equine rhinitis B virus, Ljungan virus), while others point to the presence of either known human viruses (e.g., herpesvirus 1, 2, 7) or human endogenous retroviral sequences (e.g., ERV9, HERV 4-1) in the tissues. We will validate the microarray results by qRT-PCR analysis, and if this analysis confirms the chip results and indicates the presence of viral sequences in prostate tumor extracts, we will proceed with in situ hybridization to identify the location of the sequence and determine whether it is present in prostate tumors with an epithelial and/or stromal distribution. Future experiments are planned and will include the screening of blood samples for viral seroprevalence in our case control study of prostate cancer in the context of race/ethnicity. Screening will be performed for those viruses which we have validated as being present in prostate tissue. To prepare for these experiments, we have begun isolating total RNA and DNA from white blood cells of case subjects and controls from the Baltimore study. This is necessary as immunoassays are not currently available for all known viruses, and therefore RNA and DNA techniques are used as an alternative of examining viral prevalence. The gene expression profiles of prostate tumors revealed prominent differences in tumor immunobiology between African-American and European-American men. These differences may occur because of differences in the tumor immune cell profile between the two patient groups. We are currently pursuing this hypothesis in collaboration with Dr. Arthur Hurwitz (Laboratory of Molecular Immunoregulation, CCR). We are collecting tumors from surgery at the NCI (collaboration with Drs. Peter Pinto and Marston Linehan, Urologic Oncology Branch) and at the University of Maryland (under the IRB-approved protocol, 05-C-N021). This study is evaluating immune cell profiles (T cells, dendritic cells, myeloid cells) of cancerous tissues and surrounding non-cancerous tissues from African-Americans and cancerous tissue and surrounding non-cancerous tissues from European-Americans. A pilot study has been completed showing that immune cells can be isolated, profiled, and cultured from surgically resected prostate tissue at Dr. Hurwitz's group after shipment of the tissues from the site of collection to his laboratory by courier. The microarray analysis of African-American and European-American prostate tumors identified a two-gene tumor signature that accurately differentiated between African-American and European-American cancer patients. PSPHL is included in this two-gene signature. PSPHL was the most highly up-regulated gene in prostate tumors from African-American patients. Little is known about the function of PSPHL. Interestingly, PSPHL is located on chromosome 7q11.2, a chromosomal region known to have gain of function related to advanced tumor stage in prostate cancer. Despite its chromosomal location, there are no studies linking PSPHL expression to cancer progression. In collaboration with Drs. Jun Luo and William Isaacs at Johns Hopkins University, we are examining the genomic region of PSPHL. In addition, we generated siRNA that targets PSPHL and constructed expression vectors for the gene. We have several expression vectors for tagged and untagged PSPHL. These tools have been validated and are currently used in phenotypic assays to elucidate the function of PSPHL. Preliminary data indicate that PSPHL may influence endoplasmic reticulum function, protein ubiquitination, O-linked N-acetylglucosamine metabolism and the TGF-beta signaling pathway. Attempts to develop an antibody against PSPHL have been unsuccessful. PSPHL encodes a cysteine-rich small protein of 72 amino acids (predicted size 7.6 kDa) that has been very difficult to purify. We are continuing our efforts to raise an anti-PSPHL antibody. Recently, a two hybrid screen with PSPHL as bait has been completed through an arrangement with Myriad Genomics. Only one binding partner was identified, COPS5. This protein, also known as CSN5 or JAB1, is a subunit of the COP9 signalosome, a highly conserv [summary truncated at 7800 characters]
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