T-cell response against human malignant melanoma.

thor Straten, Per (1998). T-cell response against human malignant melanoma. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.00010204


The present Thesis is introduced with a discussion of reverse transcription polymerase chain reaction (RT-PCR) based analysis of TCRBV regions, including the description of a panel of primers. Using this primer panel, we analysed for TCR clonality in progressive versus regressive parts of partially regressive primary melanoma lesions (Paper 1). The main conclusion drawn from this study is that clonotypic T cells were present in both regressive and progressive parts of the same lesions. However, sequencing of “non clonal” BV-regions demonstrated that most of them were not polyclonal by nature but oligoclonal e.g. that in some cases all transcripts were of the same length or only a limited number of BJ regions were used. This indicated that some T-cell clones were left undetected by direct sequencing. To facilitate full and detailed analysis of T-cell clonotypes in the infiltrates we aimed at establishing a suitable method for the detection of clonally expanded T cells. Denaturing gradient gel electrophoresis (DGGE) was chosen because this method is highly sensitive and excludes laborious steps that are obligatory in almost all other methods dealing with the detection of TCR clonality (Paper 2). We applied the DGGE based method to analyse for in situ T-cell clonality in 6 subcutaneous melanoma lesions from two patients. The results demonstrated two important characteristics of in situ TIL in melanoma. First, the infiltrate constitutes an exceedingly high number of different T-cell clones, and second, the T-cell response appears to be executed by local T cells that do not enter the periphery (Paper 3). The final study deals with a comparative analysis of in situ T cells and T cells propagated in vitro, demonstrating that standard in vitro culture conditions do not support the growth of in vivo expanded T-cell clones (Paper 4).

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