Activation of Human T Cells Obtained Pre- and Post-Interleukin-2 (IL-2) Therapy by Anti-CD3 Monoclonal Antibody plus IL-2

Weil-Hillman, Gilda; Schell, Kathleen; Segal, David M.; Hank, Jacquelyn A.; Sosman, Jeffrey A.; Sondel, Paul M.

doi:10.1097/00002371-199108000-00005

Cited by 17 publications

(6 citation statements)

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“…Treatment of T cells with anti-CD3 antibody prior to IL2 exposure greatly increases T-cell cytolytic activity (6). Likewise, expansion of tumor-irfiltrating lymphocytes (TIL) by culture in the presence of high concentrations of IL2 with periodic target-cell stimulation leads to substantial increases in cytolytic activity (7).…”

mentioning

confidence: 99%

Antibody-targeted interleukin 2 stimulates T-cell killing of autologous tumor cells.

Gillies

Reilly

et al. 1992

Proc. Natl. Acad. Sci. U.S.A.

156

130

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A genetically engineered fusion protein consisting of a chimeric anti-ganglioside GD2 antibody (chl4.18) and interleukin 2 (IL2) was tested for its ability to enhance the killing of autologous GD2-expressing melanoma target cells by a tumor-infiltrating lymphocyte line (660 TIL). The fusion of IL2 to the carboxyl terminus of the immunoglobulin heavy chain did not reduce IL2 activity as measured in a standard proliferation assay using either mouse or human T-cell lines. Antigen-binding activity was greater than that of the native chimeric antibody. While IL2 treatment in vivo leads to increases in both natural killer and ADCC activities, the cytolytic activity of antigen-specific, major histocompatibility complex (MHC)-restricted T cells may actually be reduced (5). Treatment of T cells with anti-CD3 antibody prior to IL2 exposure greatly increases T-cell cytolytic activity (6). Likewise, expansion of tumor-irfiltrating lymphocytes (TIL) by culture in the presence of high concentrations of IL2 with periodic target-cell stimulation leads to substantial increases in cytolytic activity (7). Both approaches involve costimulation of IL2 and T-cell antigen receptors for expansion and maintenance of T-cell cytolytic activity. Thus, an optimal therapy might combine IL2 activation and tumor antigen presentation together with a tumor-specific antibody that mediates both complementdependent cytotoxicity (CDC) and ADCC activities. By combining a chimeric anti-ganglioside GD2 antibody (chl4.18) which has potent CDC and ADCC activities (8), with IL2, we hope to target this cytokine to tumors such as neuroblastoma (9, 10) and melanoma (11) expressing GD2. In this way, relatively large amounts of tumor antigens should be present during IL2 activation for expansion of cytotoxic T cells, since melanoma cell lines have been reported to express an average of 1.5 x 1 sites per cell for ch14.18 (8). Furthermore, the antibody would also be available to target Fc receptor-bearing cells that have been activated by the targeted IL2.The chl4.18 antibody used in this report has already been shown to mediate potent ADCC activity by IL2-activated peripheral blood mononuclear cells from cancer patients (12). We have focused on the ability of a chl4.18-IL2 fusion protein to stimulate the proliferation and cytolytic activity of a human T-cell line against autologous melanoma targets.This cell line, 660 TIL, is CD3', CD8', antigen-specific, and MHC class I-restricted and was originally obtained by outgrowth from a human metastatic melanoma (13). A melanoma line, 660 mel, was derived from the same tumor and serves as a source for antigen stimulation and as an autologous target for 660 TIL (14). Results of this study show that tumor cells coated with a fusion protein in which IL2 is at the carboxyl terminus of the heavy-chain constant region 3 (CH3) exon of ch14.18 (CH3-1L2) efficiently stimulate resting 660 TIL cells to kill autologous targets. These coated cells serve as a model for tumors that have been targeted in vivo. MATERIALS AND M...

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mentioning

confidence: 99%

Antibody-targeted interleukin 2 stimulates T-cell killing of autologous tumor cells.

Gillies

Reilly

et al. 1992

Proc. Natl. Acad. Sci. U.S.A.

156

130

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“…At the end of the assay, the supernatant, containing 51Cr released following lysis of 51Cr-labeled target cells by cytotoxic cells, is transferred to tubes or can be harvested with Skatron filters [12][13][14]. The Skatron system, based on harvesting 48 wells at once, is fast and easy to handle but is costly and requires each filter to be transferred individually into a tube for counting in a gamma counter.…”

Section: Discussionmentioning

confidence: 99%

“…To expand TIL, the lymphocytes were cultured at (2.5-5) x !05 cells/ml in 6-well culture plates in HS-RPMI, in the presence of 100 unitshnl IL-2 combined with 1000 units/ml TNF (TIL-TNF) [5]. In parallel, samples of TIL were stimulated with anti-CD3 mAb in plates coated with immobilized anti-CD3 mAb in the presence of 100 units/ml IL-2 (TIL-CD3) [5,14]. Cells were removed from anti-CD3-coated plates 3 days following initiation of culture, washed and cultured in 100 units/ml IL-2.…”

Section: Lymphokine-activated Killer (Lak) Cells To Generate Lak Celmentioning

confidence: 99%

Application of the direct beta counter Matrix 96 for cytotoxic assays: Simultaneous processing and reading of 96 wells using a51Cr-retention assay

Roessler

et al. 1993

Cancer Immunol Immunother

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To assess the cytotoxic activity of immune cells, we have developed a 51Cr-retention assay in which the radioactivity retained by 51Cr-labeled target cells, following coincubation with cytotoxic cells, is monitored using the automated Matrix 96 beta counter. The Matrix 96 is designed for simultaneously counting 96 samples isolated from a 96-well microplate. It uses 96 uniform and independent detectors operating on the principle of avalanche gas ionization in the Geiger-Muller mode. Samples must be dry because the detectors are of the open-window type. Therefore, samples from the 96 wells of the microplate are simultaneously harvested onto a filter using the MicroMate 196, a 96-well cell harvester, dried and quantified in the Matrix 96. Usually the 51Cr isotope is measured by the detection of gamma radiation in gamma counters. The Matrix 96, however, monitors Auger electrons, which are also emitted by 51Cr. We have shown that the retention assay can be used to monitor the cytotoxic activity of activated lymphocytes including lymphokine-activated killer cells and tumor-infiltrating lymphocytes against various tumor cell lines. This assay is most suitable for experiments in which low E/T ratios are sufficient to detect highly cytotoxic cells, such as clone screening in cloning assays or in limiting-dilution analysis assays. These assays involve processing and reading large numbers of microplates. In this case, the retention assay monitored in the Matrix 96 will improve the work flow and decrease the amount of radioactive waste.

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“…Only one study so far [5] has reported immunological changes during s.c. rIL-2 therapy. After comparing data before and after therapy (mostly after 3-7 weeks), it was concluded that the natural killer (NK) cells form the predominantly stimulated subpopulation [2,16] and that T cell function becomes impaired after rIL-2 therapy [4,15,17].…”

Section: Introductionmentioning

confidence: 99%

HLA-Dr-expressing CD8bright cells are only temporarily present in the circulation during subcutaneous recombinant interleukin-2 therapy in renal cell carcinoma patients

Janssen¹,

Straatsma²,

Heijn³

et al. 1993

Cancer Immunol Immunother

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The effect of subcutaneous recombinant interleukin-2 (rIL-2) therapy on the "activation status" of peripheral blood lymphocytes (PBL) of 17 renal cell carcinoma patients was investigated in a longitudinal study. The expression of the activation markers HLA-Dr and CD25 on cytotoxic T cells, helper T cells, and natural killer (NK) cells, was analysed using two-colour flow cytometry of whole-blood samples. In addition, the ability of isolated PBL to proliferate in vitro in response to various stimuli was investigated. The absolute amounts of NK cells and HLA-DR-expressing NK cells increased continuously during the whole course of therapy. The absolute amounts of T cells and HLA-Dr-expressing T cells, however, showed an early increase only during the first 1 or 2 weeks of therapy, after which the absolute amounts of HLA-Dr-expressing T cells decreased. In particular, the absolute amount of HLA-Dr-expressing CD8bright+ T cells was significantly lowered in the second half of therapy. PBL collected on day 7 of therapy (post-cycle-1 PBL) showed, as compared to those collected prior to therapy (pretherapy PBL), a decreased proliferative response in vitro after stimulation with phytohaemagglutinin, concanavalin A, soluble CD3 mAb (WT32) or rIL-2. This decreased in vitro response of post-cycle-1 PBL was also reflected in a decrease in the percentage of CD8bright+ T cells expressing HLA-Dr in cultures with rIL-2 or CD3 mAb, in contrast to cultures of pretherapy PBL, which showed an increase of this percentage. We conclude that T cells are the predominantly stimulated subpopulation during the first 2 weeks of subcutaneous rIL-2 therapy. The significant decrease in the absolute amounts of HLA-Dr-expressing T cells in the peripheral blood during the second half of therapy may partly be explained by a decreased responsiveness to rIL-2, but a selective redistribution of HLA-Dr-expressing cells may also be involved.

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Activation of Human T Cells Obtained Pre- and Post-Interleukin-2 (IL-2) Therapy by Anti-CD3 Monoclonal Antibody plus IL-2

Cited by 17 publications

References 0 publications

Antibody-targeted interleukin 2 stimulates T-cell killing of autologous tumor cells.

Antibody-targeted interleukin 2 stimulates T-cell killing of autologous tumor cells.

Application of the direct beta counter Matrix 96 for cytotoxic assays: Simultaneous processing and reading of 96 wells using a51Cr-retention assay

HLA-Dr-expressing CD8bright cells are only temporarily present in the circulation during subcutaneous recombinant interleukin-2 therapy in renal cell carcinoma patients

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