Transformation of human T-cell clones by Herpesvirus saimiri: intact antigen recognition by autonomously growing myelin basic protein-specific T cells.

Abstract

Herpesvirus saimiri has recently been shown to immortalize human T cels. It was unknown, however, whether Herpesvirus saimiri transformation affects T-cell receptor (TCR) expression and signal transduction. In the present study, we have transformed CD4+ human T-cell clones speciffc for human myelin basic protein. The transformed T cells were grown in interleukin 2 and divided in the absence of antigen and antigen-presenting ceUls. They retained the membrane phenotype of activated T ceUs and secreted the cytokines interferon y and lymphotoxin, but interleukin 4 was not detected. Further, the transformed T cells continued to express the original TCR as demonstrated by TCR variable-region-V(&specific monoclonal antibodies and TCR sequencing. Antigen-specific recognition and signal transduction by the TCR were demonstrated by myelin-basic-protein-induced HLA-DRrestricted secretion of interferon y and lymphotoxin and by myelin-basic-protein-speciflc proliferation. Antigen specificity and reactivity have been maintained for >1 year after transformation. Transformation with Herpesvirus saimiri now allows the production of virtually unlimited numbers of (auto)antigen-specific T cells expressing functional TCR and a stable membrane phenotype. This technology will facilitate studies of the pathogenesis of putative autoimmune diseases, such as multiple sclerosis, and may be of help in TCR-targeted immunotherapy.Autoantigen-specific autoaggressive T-cell clones have become indispensable tools to study the cellular and molecular mechanisms of autoimmunity. For example, studies using myelin-specific T-cell lines have provided insights into the details of demyelinating autoimmune responses and into the general organization of intracerebral immune reactivity and seem to provide a basis for additional therapeutic strategies (1). Once isolated from the donor organism, autoimmune T cells can be cultured for a few weeks in interleukin (IL) 2-containing medium in the absence of antigen. For prolonged culture, the cells must be restimulated at regular intervals with antigen and antigen-presenting cells (APCs) (2, 3). Even under optimal culture conditions, most human and rodent T-cell lines have finite life spans. This and the requirement of fresh human APCs form major obstacles limiting the duration and dimension of human T-cell studies.These problems could be overcome by virus transformation of antigen-specific T cells, in analogy to Epstein-Barr virus transformation of B cells. To be useful for broad application, the ideal T-cell-transforming agent should lead to permanent APC-and antigen-independent growth, whileThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.conserving the essential features of the nontransformed parental T cells. We show here that Herpesvirus saimiri (HVS), a lymphotropic -.2 herpesvirus (4), fulfills most, if not all, of these criteria. Using ce...

Functional long-term thymidine kinase suicide gene expression in human T cells using a herpesvirus saimiri vector

“…Among many preserved relevant functions of normal T cells, the MHC-restricted antigen specificity of parental T cell clones is the most important feature. [35][36][37][38][39] A broad variety of T cell subtypes can be transformed, either CD4 + or CD8 + cells, and either ␣␤-or ␥␦-T cells. 32,37,[44][45][46][47] When polyclonal bulk cultures had been infected with the virus, a broad spectrum of V␤ chains was represented in the resulting transformed cultures.…”

“…32 In contrast to leukemic cell lines or to HTLV-1 immortalized T cells, the herpesvirus-transformed human T lymphocytes retain a high degree of functionality including the MHC-restricted antigen-specific reactivity of parental non-infected T cell clones. [35][36][37][38][39] There is only little virus gene expression and no virion production in these T cells. 33,40 The C488-transformed human T cells appear to be non-tumorigenic.…”

Peptide Binding Inhibits Aggregation of Soluble MHC Class II in Solution

“…The Herpesvirus saimiri-transformed SS8T human T cell clone (22) restricted by DRB5*0101 and MBP(84-102) was cultured in RPMI 1640 medium supplemented with 2 mM L-glutamine, 100 units/ml penicillin, 100 l g/ml streptomycin, 10% fetal bovine serum (Hyclone), and 50 units/ml human recombinant interleukin-2 (from ABI) at 37 ± C. Every alternate day the cells were transferred to fresh medium. Various LMW and HMW complex preparations were coated onto a microtiter tissue culture plate, and the cells were inoculated at a density of 20 000/well in 200 l l of medium without interleukin-2.…”

Efficient lentiviral transduction of Herpesvirus saimiri immortalized T cells as a model for gene therapy in primary immunodeficiencies

“…3,4 More recently, it has been reported that certain strains of Herpesvirus saimiri (HVS) immortalize human T lymphocytes (HVS-T), thus yielding long-term cell lines with stable surface markers and functional phenotypes. 5,6 This strategy has provided a suitable, convenient experimental approach to examine T-cell function. HVS is non-pathogenic in its natural host, the squirrel monkey, but provokes extremely aggressive lymphomas and lymphocytic leukemias in other primates, 7 where it enters the cell and remains as nonintegrated viral episomes.…”

“…HVS is non-pathogenic in its natural host, the squirrel monkey, but provokes extremely aggressive lymphomas and lymphocytic leukemias in other primates, 7 where it enters the cell and remains as nonintegrated viral episomes. 5,6 Although HVS encodes two antiapoptotic proteins, 8,9 all evidence suggests that it is not interference with apoptotic programs that promotes HVS immortalization of T cells but rather interactions between viral proteins and normal T-cell signaling pathways. 10 HVS-T cells are potentially important in the study of both primary and acquired immunodeficiencies.…”