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Research ProfileS
Immunoproteomics for early cancer diagnoses and cancer therapies
from 20 ovarian cancer patients and 20 control subjects. Samples from four subjects were pooled to create five composites for each group. To harvest TAAs, tumor cell lysates were prepared from two ovarian cancer cell lines. Autoantibodies that recognized tumor proteins were immunoprecipitated, and these antibody–antigen complexes were run on a size-exclusion column. Next, the
composites. Of these cancer-specific TAA humoral targets, eight appeared on the list of MHC-associated TAAs. Early in tumor development, the imDeveloping a diagnostic assay is a key mune system sometimes does strange goal. All of the proteins that are shared things. In many cases, B cells produce by the two immune pathways are good antibodies against tumor-associated candidates for early-detection biomarkantigens (TAAs). What makes this proers. Even those that were only identified cess so odd is that TAAs are produced as cancer-specific autoantibody targets by cancer patients themselves, and the could serve as diagnostics. “We have to antibodies (autoantibodweed out the ones that are ies) are directed against shared between cancers,” Humoral immunity Cellular immunity these self-antigens. Besays Philip. “That list, Serum sample Tumor lysate cause these autoantibodeven if it is not shared by Ag2 ies are detectable in the MHC–peptide data sets, Ag4 sera of patients with some is still useful because it is Ag3 Ag6 Tumor cell types of cancer well before specific for that diagnosis.” Ag1 Ab symptoms arise, they She and her team plan to Ag5 Immunoprecipitation Ag3 MHC–peptide could be used as earlypursue protein-array and Ag2 complexes detection biomarkers. nanotechnology-based Ag6 Ag5 Ag1 Ag4 If autoantibodies are platforms. MHC-associated produced, then why can’t Specificity for a particuAg2 Ag3 Ag5 peptides the human body simply lar cancer is less important LC/MS/MS LC/MS/MS eliminate tumors on its when therapeutics are AutoantibodyTumor-associated own? “That’s the $64,000 developed. “Overlapping reactive antigens Ag2 Ag5 MHC-associated peptides question!” exclaims RamiMHC-associated peptide Common autoantigens la Philip of Immunotope, presentation is very good Inc. Researchers don’t because you can use a Let’s share. Immunoprecipitation methods were used to identify antigens know which antibodies cocktail of antigens for (Ag) involved in cancer immune pathways (Ab = antibody). indicate a bad or a good various cancers. You can prognosis, she explains. In generate a cancer vaccine addition, the B-cell antibody response antigens were separated by HPLC, dithat is broad-based,” explains Philip. may not be enough to kill a tumor; gested with trypsin, and analyzed with “In fact, the cancer vaccine trials we are enlisting the help of T cells for a cella second LC step and MS/MS. conducting now include the same set mediated immune response may be reIn parallel with these experiments, of antigens for both ovarian and breast the researchers also searched for anti quired. A cancer vaccine that includes cancer.” To develop a potential vaccine, TAAs that are processed into peptides gens presented on the surfaces of tumor the researchers will choose proteins and presented on the surfaces of tumor cells by MHC proteins. Lysates from that are autoantibody targets and are cells by the major histocompatibility the same cell lines were immunopreinvolved in processes that are imporcomplex (MHC) could activate the T cipitated with an antibody to MHCs tant for cancer metastasis and cell surcells to recognize and destroy a tumor. that was coupled to Protein A/G beads. vival. That way, if a tumor tries to evade TAAs shared by the humoral and MHC–peptide complexes were eluted a T-cell response by eliminating that cell-mediated pathways would be ideal from the antibody and run on a size-exantigen, it will die or stop metastasizing targets because patients who produce clusion column. Isolated peptides were anyway, she says. autoantibodies against these antigens separated by HPLC, then analyzed with The researchers are very pleased have immune systems that already cona second LC step and MS/MS. with the results they obtained. “When sider the antigens to be foreign. As deThe researchers identified parent we thought about this 2 years ago and scribed in this issue of JPR (2509–2517), proteins of the MHC-associated pepwhen we were doing it, we didn’t really Philip and co-workers at Immunotope tides as well as proteins that were spebelieve that we would end up with this took a two-pronged approach to discifically recognized by autoantibodies nice set of antigens shared between cover possible TAA targets in ovarian from the patients. They discovered that diagnostics and therapeutics,” says cancer. ~100 TAAs were targeted by autoanPhilip. “That was a pleasant surprise To study the humoral response, the tibodies in all five patient-composite for us.” researchers obtained serum samples samples but not by those in control —Katie Cottingham
2408 Journal of Proteome Research • Vol. 6, No. 7, 2007
© 2007 American Chemical Society
