טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentBarnea Eilon
SubjectIdentification and Characterization of Cancer Cell MHC
Class-I Restricted Peptides
DepartmentDepartment of Biology
Supervisor Professor Arie Admon


Abstract

Thousands of different peptides are presented within the context of the MHC class-I on the surface of cancer cells. Many of these peptides originate from tumor associated antigens and therefore may serve as candidates for the development of cancer vaccines. Immuno-affinity purification of MHC molecules and peptides elution from them results in very complex mixtures, making the identification of individual peptides a demanding task. We reduced the complexity and increased the purified amounts of the individual peptides by transfecting human culture cancer cells with expression vectors to produce the MHCs as a soluble, secreted and native form. This was accomplished by truncated genes of HLA-A2 and HLA-B7. Soluble MHC molecules with the bound peptides were purified from two breast cancer (MDA-231 and MCF-7),  two Ovarian cancer (UCI-101 and UCI-107)  and prostate cancer (PC-3) cell lines. The peptides were eluted, resolved by capillary reverse-phase HPLC and analyzed by an electrospray ion-trap mass spectrometer. All of the identified peptides fit the known consensus and some were identical to known MHC peptides. We have identified peptides derived from the cancer antigen MAGE-B2 and MUC1  and new peptides originating from proteins associated with cancer, including DNA methyl transferase, fatty acid synthase and beta-cathenin. With this method, any MHC haplotype can be introduced into different cell lines, peptides can be identified and their patterns compared between cell lines or culture conditions. This analytical method may serve as an improved staging point for the human MHC peptides project.