Immunology, National Institute on Aging, National Instituteof Health, Baltimore, MD, Michael Wade, Student,Laboratory of Immunology, National Institute on Aging,National Institute of Health, Baltimore, MD, Sherry Yang,Student, Laboratory of Immunology, National Institute onAging, National Institute of Health, Baltimore, MD, PolokoLeotlela, Post doctoral Fellow, Laboratory of Immunology,National Institute on Aging National Institute of Health,Baltimore, MD, Dennis Taub, Chief, Laboratory ofImmunology, Laboratory of Immunology, National Instituteon Aging, National Institute of Health, Baltimore, MD, AdamRiker, Chief of Surgical Oncology, Mitchell cancer Institute,University of South Alabama, North Mobile, AL, BrianNickoloff, Professor pathology, Loyola University MedicalCenter, Maywood, IL, Ashani Weeraratna, Staff Scientist,Laboratory of Immunology, National Institute on Aging,National Institute of Health, Baltimore, MD

Wnt5a increases melanoma cell motility via activation ofProtein Kinase C (PKC). Microarray analysis showed aninverse relationship between Wnt5a and MART-1. Using PKCactivation and inhibition studies, as well as recombinantWnt5a and Wnt5a RNAi, we demonstrate here that Wnt5a-activated PKC affects the expression of MART-1 via thephosphorylation of STAT3. STAT3 is a transcriptional factorknown to inhibit the binding of MART-1 transcriptionalactivators MITF, SOX10 and PAX3. These effects are mediatedby Wnt5a upstream of PKC, as overexpression of Wnt5aincreases pSTAT3 and Wnt5a RNAi treatment decreasespSTAT3 levels. The decrease in pSTAT3 corresponds to anincrease in MART1 and vice versa. Translocation of pSTAT3from the cytosol to the nucleus upon transfection of Wnt5a,indicated activation of STAT3 by Wnt5a, while the reversewas observed for cells knocked down for Wnt5a. Further,steady increases in PAX3 and MART-1 protein levels wereobserved in cells treated with Wnt5a RNAi, possibly due tosuppressing the effects of STAT3. MART1 positive melanomacells are able to activate CTL, while addition of recombinantWnt5a to these cells could not, presumably due to the down-regulation of MART-1. RNA from human melanoma biopsiesanalyzed for Wnt5a by real-time PCR shows an inverserelationship between Wnt5a levels and patient survival time,suggesting that cells with higher MART-1 have a betteroutcome. Collectively, our data suggest that Wnt5a, via PKC,results in the phosphorylation of STAT-3 and a subsequentdown-regulation MART1, thereby rendering melanoma cellsable to escape immune detection.

doi:10.1016/j.clim.2007.03.502

Sa.116 Discovery of Cancer and AutoimmuneBiomarkers Utilizing Serum Antibody Profiling onProtein MicroarraysJennifer Cannon, Scientist, Invitrogen Corporation,Carlsbad, CA, Michael Snyder, Professor, Yale University,New Haven, CT, Michael Hudson, Scientist, Yale University,New Haven, CT, Gregory Michaud, Scientist, InvitrogenCorporation, Carlsbad, CA, Michael Smith, Scientist,Invitrogen Corporation, Carlsbad, CA, Barry Schweitzer,Research and Development Director, Invitrogen

Corporation, Carlsbad, CA, Guillermo Mor, AssociateProfesor, OB/GYN, New Haven, CT

It has been well established that serum autoantibodieshave important diagnostic value for many diseases, includingcancer and autoimmune diseases. Identifying the antigenswhich elicit an autoimmune response can yield sets ofbiomarkers that provide classifiers for particular diseasesand disease stages as well as predictors of patient outcomesand patient responses to therapeutics. Protein microarraysare valuable tools that have been successfully applied toinvestigate the circulating antibody profile in several diseasestates. To discover potential autoantibody biomarkersspecific to ovarian cancer, sera from 30 ovarian cancerpatients and 30 healthy patients were profiled on proteinmicroarrays. Patient and control sera were diluted andapplied to protein microarrays containing over 5000 purifiedproteins expressed in a baculovirus expression system. Over95 candidate biomarkers were identified that exhibitedenhanced reactivity in sera from cancer patients relative tothat of the control individuals. Antibodies to four antigenswere tested for differential reactivity in tissue samples ofcancer patients relative to healthy patients using immuno-blot analysis and tissue microarrays. Three biomarkersexhibited elevated expression in the cancer tissue relativeto controls. Signal observed for the biomarker antigensappeared significantly stronger than signal corresponding tothe existing ovarian cancer antigen, CA-125. Additionalstudies focused on identifying biomarkers associated withautoimmune diseases, such as Rheumatoid Arthritis andSystemic Lupus Erythematosus, have recently utilized ahigher content protein microarray including over 8000unique human proteins. The application of protein micro-array technology for serum antibody profiling will bediscussed along with data from on-going validation studies.

doi:10.1016/j.clim.2007.03.503

Sa.117 Phenotype and Function Analysis ofDendritic Cells From Patients with ProstateCarcinoma After Radical ProstatectomyIvo Minarik, PhD Student, Department of Immunology, 2ndMedical School, Charles University, Prague 5, CzechRepublic, Zuzana Tobiasova, PhD Student, Department ofImmunology, 2nd Medical School, Charles University, Prague5, Czech Republic, Jirina Bartunkova, Head of Department,Department of Immunology, 2nd Medical School, CharlesUniversity, Prague 5, Czech Republic, Ivan Kawaciuk, Headof Department, Department of Urology, 2nd Medical School,Charles University, Prague 5, Czech Republic

Prostate carcinoma represents the 3rd most frequentmalignancy in the Czech male population with theincidence of 68.3/100000 and mortality 28.2/100000.About 35% of patients experience biochemical relapse(PSA increase) any time after radical prostatectomy.Hence, alternative treatment is highly required. Dendriticcell (DC)-based immunotherapy seems to be a promisingapproach for the elimination of relapse. In our study we aimto compare function and phenotype of dendritic cells from

S114 Abstracts