Isolation and in vitro growth of glioma-infiltrating lymphocytes, and an analysis of their surface phenotypes
✓ The present investigation was conducted in order to examine the feasibility of isolating and growing gliomainfiltrating lymphocytes in vitro as possible effector cells for use in new adoptive immunotherapy. Eight surgical specimens obtained from patients with malignant astrocytomas were treated by enzyme dispersion; the cells were separated on a density gradient and grown in the presence of human recombinant interleukin-2. The cultured lymphocytes were tested for cell-surface markers by using monoclonal antibodies in a flow cytometric analysis. In all cases the glioma-derived lymphocytes were grown in culture for several weeks with substantial increases in cell numbers (at least 5 × 108 cells). The mature T cell population (CD3, 89%) was found to have an increased proportion of the cytotoxic/suppressor phenotype CD8 (55%) as compared to peripheral blood lymphocytes (PBL's). Eighty-six percent of the cultivated lymphocytes expressed HLA-DR. The IL-2 receptor was predominantly expressed on the helper subset (CD4-positive). Otherwise, anti-CD16, which specifically reacts with natural killer (NK) cells, did not stain significantly more of the cultured glioma-derived lymphocytes compared with lymphocyte-activated PBL's. These results corroborate the observations made with conventional immunohistochemical examination. It has been demonstrated that T lymphocytes isolated from human cancers are enriched for specific reactivity to their autochthonous tumor cells. These experiments support the possible use of glioma-infiltrating lymphocytes as a new treatment for patients with malignant glioma.
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