TY - JOUR
T1 - Development and implementation of a direct detection, quantitation and validation system for class I MHC self-peptide epitopes
AU - Weidanz, Jon A.
AU - Piazza, Paolo
AU - Hickman-Miller, Heather
AU - Woodburn, David
AU - Nguyen, Tiffany
AU - Wahl, Angela
AU - Neethling, Francisca
AU - Chiriva-Internati, Maurizio
AU - Rinaldo, Charles R.
AU - Hildebrand, William H.
N1 - Funding Information:
This work was supported by the Advanced Technology Program, NIST grant # 70NANB4H3048 (JAW), NIH contracts HHSN266200400027C and NO1-AI-95360 (WHH), OCAST grant AR04-028 (WHH), NIH Immunology Training Grant #A1007633-006 (AW) and NIH grants U01-AI35041 and R37-AI41870 (CRR).
PY - 2007/1/10
Y1 - 2007/1/10
N2 - Gene and protein expression studies demonstrate that viral-infected and malignant cells undergo a complex series of transcriptional and translational changes. As class I MHC molecules reflect the proteome (and changes therein) by presenting intracellular peptide epitopes, the development of a direct discovery and validation technology for the identification of these epitopes is needed. We developed our technology using HIV-1-infected cells as a model. A combination of hollow fiber class I HLA protein production and mass spectrometric epitope analysis indicated a 3-fold increase in the host-peptide VLMTEDIKL720-728, [eIF4G(720)] presented by the HLA-A*0201 of HIV-1-infected cells. This peptide is derived from the host-protein translation of eukaryotic initiation factor 4-gamma (eIF4G) that plays a pivotal role in cellular protein synthesis. Direct confirmation of expression of this self-encoded antigen was performed through development of a T cell receptor mimic (TCRm) monoclonal antibody (mAb). The resulting 4F7 TCRm demonstrated specific recognition of the eIF4G(720)-A*0201 complex. Staining of normal PBMCs with 4F7 showed only low levels of endogenous eIF4G(720) presentation by HLA-A*0201, while 4F7 staining of HIV-1-infected PBMCs revealed a ∼ 3-fold increase in eIF4G(720)-A*0201. The MHC-peptide complex was initially detectable by 4F7 at 3 days post-infection, with a steady increase through day 8. We therefore demonstrate the successful development and implementation of an integrated discovery and validation technology system for direct identification and confirmation of class I MHC-peptide epitopes on cells.
AB - Gene and protein expression studies demonstrate that viral-infected and malignant cells undergo a complex series of transcriptional and translational changes. As class I MHC molecules reflect the proteome (and changes therein) by presenting intracellular peptide epitopes, the development of a direct discovery and validation technology for the identification of these epitopes is needed. We developed our technology using HIV-1-infected cells as a model. A combination of hollow fiber class I HLA protein production and mass spectrometric epitope analysis indicated a 3-fold increase in the host-peptide VLMTEDIKL720-728, [eIF4G(720)] presented by the HLA-A*0201 of HIV-1-infected cells. This peptide is derived from the host-protein translation of eukaryotic initiation factor 4-gamma (eIF4G) that plays a pivotal role in cellular protein synthesis. Direct confirmation of expression of this self-encoded antigen was performed through development of a T cell receptor mimic (TCRm) monoclonal antibody (mAb). The resulting 4F7 TCRm demonstrated specific recognition of the eIF4G(720)-A*0201 complex. Staining of normal PBMCs with 4F7 showed only low levels of endogenous eIF4G(720) presentation by HLA-A*0201, while 4F7 staining of HIV-1-infected PBMCs revealed a ∼ 3-fold increase in eIF4G(720)-A*0201. The MHC-peptide complex was initially detectable by 4F7 at 3 days post-infection, with a steady increase through day 8. We therefore demonstrate the successful development and implementation of an integrated discovery and validation technology system for direct identification and confirmation of class I MHC-peptide epitopes on cells.
KW - CTL
KW - Epitope discovery and validation
KW - Human leukocyte antigen
KW - Major hiscompatability complex
KW - Mass spectrometry
KW - Monoclonal antibody
KW - Peptide epitope
KW - TCR mimics
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U2 - 10.1016/j.jim.2006.09.019
DO - 10.1016/j.jim.2006.09.019
M3 - Article
C2 - 17134715
AN - SCOPUS:33845866856
SN - 0022-1759
VL - 318
SP - 47
EP - 58
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
IS - 1-2
ER -