Abnormalities of HLA and β₂microglobulin expression on tumour cells

Bicknell, David Charles (2003). Abnormalities of HLA and β₂microglobulin expression on tumour cells. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.0000f57c

Abstract

This thesis describes the genetic events that can lead to the loss of expression of HLA class 1 on tumour cells. Earlier studies, in colorectal cancer, have shown both complete loss of surface HLA A, B & C expression and loss of single allele products using antibodies recognizing either monomorphic or polymorphic HLA determinants. Data presented demonstrates that complete loss of HLA class 1 expression correlates with mutations in β2 microglobulin. In a study of 52 colorectal cell lines, 8 showed mutations in β2 microglobulin leading to loss or reduced HLA expression. In fresh colorectal tumours 9/147 had mutations in β2 microglobulin, occurring at single or di-nucleotide repeat sequences. From these tumours 71 were analyzed for microsatellite instability, associated with the loss of DNA mismatch repair, and 7 (10%) found to be unstable. β2 microglobulin mutations were identified in 5 of these tumors. Therefore, mutations in β2 microglobulin occur more frequently in mismatch repair defective tumours than in colorectal tumours in general (p<0.01).

The colorectal cell line HCA-7 lacks expression of HLA-A*0101 based on studies with the polymorphic antibody 142.2, but the normal B cell line (EVA-1224) from the same patient expresses A*0101. This thesis shows that the A*0101 gene, in HCA-7, contains an insertion of a cytidine in a cytidine repeat sequence in exon 4. This mutated A*0101 gene has the similar sequence to that of a rare A*0104'null' allele reported to lack A*0104 expression.

In a model system a β2 microglobulin-HLA-A*0201 construct was transfected, using an ecdysone inducible expression system, into CHO cells and expression shown to be under the control of the inducer, Ponasterone A. Functional studies demonstrated the construct to be capable of presenting the flu virus peptide GILGFVFTL to a clone of human A*0201 restricted cytotoxic T cells and for the T cells to efficiently lyse the CHO cells.