Gwyther, Jacqueline Mary; (2000) Molecular analysis and gene therapy of X-linked severe combined immunodeficiency. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

X-linked severe combined immunodeficiency disease (XSCID), characterised by the failure of cellular and humoral immunity, is caused by differential and survival defects within the T and natural killer cell lineages of the haematopoietic system. The γc chain protein encoded by IL2RG, the gene mutated in this monogenic disorder, is an integral component of a number of cytokine receptors including interleukin-2 (IL-2), IL-4, IL-7. IL-9 and IL-15. This study documents a diverse range of interesting disease-causing IL2RG mutations and their effects on protein expression and stability. The importance of γc chain functionality in various haematopoietic lineages was addressed. On analysis of XSCID patients treated by bone marrow transplantation (BMT), significantly higher levels of γc chain expression and greater kinetics of engraftment were observed in the T cell lineage compared to B-lymphocytes. This reflects the strong selective growth advantage of γc chain expressing over non-expressing T cells whereas B cell precursors experience no such competitive effect. Normal levels of γc chain expression were detected on the monocytes of XSCID carrier females. This suggests that non-γc chain expressing monocytes were subjected to extinction so that only those cells expressing normal γc chain persisted. As an alternative to BMT and as a potentially superior treatment of XSCID, the feasibility of moloney murine leukaemia retroviral (MoMLV) vector mediated gene transfer of the IL2RG gene to patients' cells was explored. Higher efficiencies of lacZ reporter gene transfer were achieved with virus pseudotyped in the novel feline endogenous viral (RD114) envelope, compared to the commonly used MoMLV- amphotropic envelope, in primary cells and EBV transformed B cell lines. Therefore RD114 pseudotyped viruses bearing the IL2RG gene were used to restore γc chain expression in EBV transformed B-lymphoblastoid cells derived from an XSCID individual and to integrate the IL2RG gene into γc chain-deficient primary cells.

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