Abstract B05: Differential autoimmune response to glycolysis and plasminogen antigens from aggressive prostate cancer cells in African American and European American men with prostate cancer

Prostate cancer (PCa) is the most commonly diagnosed cancer and the second leading cause of cancer-related male deaths in the U.S. This cancer is more aggressive in African American (AA) men, who are twice as likely to die from the disease compared to other racial groups. Despite racial disparities in PCa mortality, immunoproteomic studies profiling sera from thousands of men for the presence of autoantibodies to tumor-associated antigens (TAA) have mostly used sera from European or Asian ancestry. These autoantibodies can serve as both cancer biomarkers and potential tools in anti-tumor immunotherapy. In this study we profiled sera of AA and European American (EA) men with (N=157) and without (N=183) PCa against protein lysates from aggressive PCa cell lines using one and two-dimensional Western blotting. Sera from AA-PCa patients showed stronger immunoreactivity to proteins in PC3 cell lysates compared to EA-PCa sera. Common bands of immunoreactivity were observed around 37, 40, 50, 55, 60, and 70 kD from the sera of multiple patients. Several AA-PCa sera showed common immunoreactivity against a 50 kD protein band (16 out of 24, 67%) which was identified by serological proteomic analysis (SERPA) as alpha-enolase (ENO1), an enzyme that participates in both the glycolysis and plasminogen pathways. ELISA using purified ENO1 was conducted on 360 sera and showed significantly elevated frequency in men with PCa compared to non-PCa controls. Interestingly, SERPA analysis of other strongly immunoreactive spots outside the 50 kD region recognized of the other highly reactive PCa sera revealed additional candidate TAAs involved in the glycolysis and plasminogen pathways. These included key glycolytic enzymes GAPDH, fructose bisphosphate aldolase, phosphoglycerate kinase and lactate dehydrogenase, and other members of the plasminogen system such as GRP78 and annexin A2. Autoantibodies to stress survival proteins HSP60 and STIP1 were also identified in multiple PCa sera. We then probed by immunoblotting representative sera recognizing some of these candidate TAAs against a panel of 12 PCa and non-PCa cell lines. Anti-STIP1 sera showed elevated immunoreactivity predominantly in docetaxel-resistant cell lines. In the anti-ENO1 positive sera, we observed a race-related differential immune response, with anti-ENO1 sera from AA-PCa men showing elevated immunoreactivity mostly in metastatic cell lines, and anti-ENO1 sera from EA-PCa men showing uniform immunoreactivity across the entire panel of prostate cell lines. Interestingly, while several anti-ENO1 positive sera from AA-PCa men had increased immunoreactivity to this protein in the metastatic PCa cell lines LNCaP, MDA-PC2b, PC3 and DU145, they lacked reactivity against ENO1 in the docetaxel-resistant PC3-DR and DU145-DR cells, or purified ENO1 used in the ELISA. Anti-ENO1 EA-PCa sera did not display this differential pattern of immunoreactivity. To better understand these differences in anti-ENO1 reactivity, we interrogated the pattern of ENO1 post-translational modifications (PTMs) from PC3, PC3-DR and purified ENO1. Several different acetylated, methylated, phosphorylated, citrullinated, and glycosylated ENO1 PTMs were identified in the PC3 cells that were not present in the PC3-DR cells nor purified ENO1 which might explain the differential reactivity of AA-PCa sera against these three sources of ENO1. Taken together, these data suggest the anti-ENO1 autoantibodies from AA-PCa men may target a modified form of ENO1 that is different from that recognized by EA-PCa men. These discrepancies point to racial differences in immune response to TAAs in prostate tumors, and may have implications for identifying immunobiological factors contributing to PCa health disparities. Citation Format: Tino W. Sanchez, Guangyu Zhang, Jitian Li, Liping Dai, Rowaid Kellow, Saied Mirshahidi, Nathan R. Wall, Clayton Yates, Colwick Wilson, Susanne Montgomery, Jian-Ying Zhang, Carlos Casiano. Differential autoimmune response to glycolysis and plasminogen antigens from aggressive prostate cancer cells in African American and European American men with prostate cancer. [abstract]. In: Proceedings of the Ninth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2016 Sep 25-28; Fort Lauderdale, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2017;26(2 Suppl):Abstract nr B05.