The isolation and identification of MHC-bound CTL peptide epitopes expressed by tumour cells

Hill, S.C., 2000. The isolation and identification of MHC-bound CTL peptide epitopes expressed by tumour cells. PhD, Nottingham Trent University.

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Cytotoxic T lymphocytes are an essential component of the anti-tumour immune response and they are activated by binding of the T cell receptor to a peptide-MHC complex on the tumour cell surface. These surface-bound peptides originate from intracellular proteins that have been degraded into peptides by the proteasome and presented by MHC class I.

Many tumours inappropriately express mutated or overexpressed self-proteins (p53, ras), tissue specific antigens (MAGE, MelanA) or tumour-specific fusion proteins (bcr-abl) which, due to their tumour association can be used as targets for anti-tumour immunotherapy.

This study presents a technique to isolate and identify surface-MHC bound tumour antigen peptides from tumour cells. Peptide isolation was achieved using a mild acid buffer to destabilise surface MHC class I molecules; concentration and purification of the peptides was facilitated by ion-exchange-HPLC and subsequent peptide sequence information was generated by electrospray mass spectrometry. The sequence data was then entered into a database to determine the protein origin.

Peptides were isolated from an HLA-A3 transfected K562 cell line model, which naturally expresses bcr-abl but no other HLA class I molecules, to determine whether peptides from the fusion region were naturally presented by HLA-A3. PBMC from HLA- A3 positive CML patients was then analysed in the same way. The identification of bcr-abl fusion region peptides from the K562 transfectants and HLA-A3 positive CML patient material is, to the best of my knowledge, the first conclusive demonstration that the bcr-abl fusion protein is processed intracellularly resulting in the surface expression of peptides from the fusion region. These peptides represent an entirely tumour specific antigen and there is potential for their use as a target in anti-tumour immunotherapeutic strategies.

Item Type: Thesis
Creators: Hill, S.C.
Date: 2000
ISBN: 9781369316599
Divisions: Schools > School of Science and Technology
Record created by: Linda Sullivan
Date Added: 30 Sep 2020 10:23
Last Modified: 12 Sep 2023 14:47

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