Utilizing T-cell Activation Signals 1, 2, and 3 for Tumor-infiltrating Lymphocytes (TIL) Expansion: The Advantage Over the Sole Use of Interleukin-2 in Cutaneous and Uveal Melanoma (original) (raw)
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Expansion and characterization of human melanoma tumor-infiltrating lymphocytes (TILs)
PloS one, 2010
Background: Various immunotherapeutic strategies for cancer are aimed at augmenting the T cell response against tumor cells. Adoptive cell therapy (ACT), where T cells are manipulated ex vivo and subsequently re-infused in an autologous manner, has been performed using T cells from various sources. Some of the highest clinical response rates for metastatic melanoma have been reported in trials using tumor-infiltrating lymphocytes (TILs). These protocols still have room for improvement and furthermore are currently only performed at a limited number of institutions. The goal of this work was to develop TILs as a therapeutic product at our institution.
2014
Characteristics of the different stages of melanoma…………………………… 1.2. The long and winding road of Immunotherapy………………………………… 1.3. Different forms of adoptive T-cell therapy…………………………………….. 1.4. Schematic representation of the process of TIL expansion and TIL therapy for metastatic melanoma starting from tumor fragments……………………………….. 1.5. Stages of CD8+ T-cell differentiation found in melanoma CD8+ T-cell in TIL and phenotypic markers assessed by flow cytometry used to delineate these stages………………………………………………………………………………... 1.6. 4-1BB signaling pathways……………………………………………………… CHAPTER 2. 2.1. Detection of 4-1BB and OX40 on freshly-isolated CD8+ TIL………………… 2.2. Optimal concentration for anti-4-1BB antibody for pre-REP TIL expansion…. 2.3. Addition of anti-CTLA-4 antibody to the fragments increases the percentage and expansion of CD4+ TIL………………………………………………………… 2.4. Anti-4-1BB agonistic antibody increases TIL expansion in vitro……………… 2.5. CD8+ TIL percentage is increased with the addition of anti-4-1BB antibody to TIL cultures…………………………………………………………………………. 2.6. Activation of the 4-1BB pathway augments CD27 and CD70 expression…….. 2.7. TIL initially expanded with anti-4-1BB antibody continue to expand during secondary expansion in the REP……………………………………………………. xii 2.8. Activation of the 4-1BB pathway augments effector cell phenotype ………….. 2.9. Increased tumor reactivity in TIL expanded with anti-4-1BB antibody………..
BioDrugs : clinical immunotherapeutics, biopharmaceuticals and gene therapy, 2014
Cancer immunotherapy has become an important area for the future development of cancer therapy; this includes T-cell-based therapies that involve adoptive transfer of autologous T cells derived from the tumors or peripheral blood of cancer patients, vaccines, oncolytic virus therapy, and immunomodulatory antibodies and ligands. Here, we summarize the current approaches and clinical data in the field of adoptive T-cell transfer therapy using tumor-infiltrating lymphocytes (TILs) for metastatic melanoma. We also discuss current knowledge on the mechanism of transferred TILs in mediating tumor regression and the growing need for and recent advances in the identification of predictive biomarkers to better select patients for TIL therapy. The current technical limitations of current TIL expansion methods for out-scaling are discussed as well as how these are being addressed in order to further "industrialize" this form of cell therapy. Lastly, how TIL adoptive transfer can be i...
Cancer Immunology, Immunotherapy, 2011
Tumor-inWltrating lymphocytes (TILs) have been successfully used for adoptive cell transfer (ACT) immunotherapy; however, due to their scarce availability, this therapy is possible for a limited fraction of cutaneous melanoma patients. We assessed whether an eVective protocol for ex vivo T-cell expansion from peripheral blood mononuclear cells (PBMCs), suitable for ACT of both cutaneous and ocular melanoma patients, could be identi-Wed. PBMCs from both cutaneous and ocular melanoma patients were stimulated in vitro with autologous, irradiated melanoma cells (mixed lymphocyte tumor cell culture; MLTCs) in the presence of IL-2 and IL-15 followed by the rapid expansion protocol (REP). The functional activity of these T lymphocytes was characterized and compared with that of TILs. In addition, the immune inWltration in vivo of ocular melanoma lesions was analyzed. An eYcient in vitro MLTC expansion of melanoma reactive T cells was achieved from all PBMC's samples obtained in 7 cutaneous and ocular metastatic melanoma patients. Large numbers of melanoma-speciWc T cells could be obtained when the REP protocol was applied to these MLTCs. Most MLTCs were enriched in non-terminally diVerentiated T EM cells homogeneously expressing co-stimulatory molecules (e.g., NKG2D, CD28, CD134, CD137). A similar pattern of anti-tumor activity, in association with a more variable expression of co-stimulatory molecules, was detected on short-term in vitro cultured TILs isolated from the same patients. In these ocular melanoma patients, we observed an immune inWltrate with suppressive characteristics and a low rate of ex vivo growing TILs (28.5% of our cases). Our MLTC protocol overcomes this limitation, allowing the isolation of T lymphocytes with eVector functions even in these patients. Thus, anti-tumor circulating PBMC-derived T cells could be eYciently isolated from melanoma patients by our novel ex vivo enrichment protocol. This protocol appears suitable for ACT studies of cutaneous and ocular melanoma patients.
Journal of Biomedical Science, 2003
P14 TCR transgenic CD8+ T cells (LCMV gp33-specific) were activated by antigen in the presence of either IL-2 or IL-2+IL-4 to generate effector cytotoxic T lymphocytes (CTLs). The therapeutic effectiveness of such IL-2- or IL-2+IL-4-grown CTLs was tested in mice that had received intravenous inoculations of B16.gp33 melanoma cells 7 days previously. Administration of P14 CTLs activated by antigen +IL-2+IL-4 was significantly more effective at reducing melanoma colony formation in the lung than those grown in the presence of antigen +IL-2. Highly significant improvement in survival was observed with 80% of B16.gp33-inoculated mice showing long-term survival after therapy with 10×106 antigen +IL-2+IL-4-activated P14 CTLs. Similar therapeutic effectiveness of antigen +IL-2+IL-4-activated P14 CTLs against subcutaneously inoculated B16.gp33 melanoma cells was also found. There was significant reduction in P14 CD8+ T cells in the peripheral blood of B16.gp33-inoculated mice than in mice that did not receive B16.gp33 melanoma cells, indicating possible homing of P14 CD8+ T cells to the site of tumor growth or antigen-induced apoptotic cell death. These results may have implications in tumor therapy using CTLs grown ex vivo, especially during early stages of tumor formation. They also support the concept that the therapeutic effectiveness of CTLs can be governed by the cytokine context in which they are activated.
Cancer Immunology Immunotherapy, 1995
The therapeutic potential of adoptive therapy using tumour-infiltrating lymphocytes (TIL) has been demonstrated in a number of clinical trials. However, freshly isolated tumour-infiltrating lymphocytes (TIL) are often impaired in their proliferative and cytotoxic responses, which limits their use in immunotherapy. Several hypotheses with regard to the poor effector function of TIL have been postulated, including the production of immunosuppressive factors by tumour cells. In a previous paper we reported the efficient expansion of immunoreactive TIL from a variety of solid tumours by stimulation with a combination of monoclonal antibodies (mAbs) against CD3 and CD28. In the present study we analysed whether this protocol would be improved by the removal of tumour cells at the start of the culture. We tested a highly immunogenic tumour, melanoma, and a poorly immunogenic tumour, colon carcinoma. Removal of tumour cells highly improved anti-CD3/CD28 stimulated expansion of TIL from colon carcinoma, resulting in a significantly higher percentage of potentially tumour-specific CD8-positive Tcells and a reduced CD4/CD8 ratio compared to expansion in the presence of tumour cells. In contrast, expansion and CD4/CD8 ratio of melanoma-derived TIL was not significantly influenced by the removal of autologous tumour cells. CD3/CD28-stimulated melanoma TIL cultured in the absence of tumour cells showed specific lysis of autologous tumour cells comparable to melanoma TIL cultured in highdose IL2. However, no cytotoxicity could be detected in colon TIL irrespective of the culture conditions used. On the other hand, 3/8 colon carcinoma TIL cultures and 9/12 melanoma-derived TIL cultures showed IFN? secretion upon stimulation with autologous tumour cells. We conclude that stimulation of TIL with a combination of mAbs to CD3 and CD28 in the absence of tumour cells induces efficient expansion of potentially tumour-specific cells from a highly and a poorly immunogenic tumour. Removal of tumour cells does not have a negative influence on the generation of tumour-specific T cells, while cell yield improves. Therefore, for large-scale cultures this protocol can efficiently induce the outgrowth of tumour-specific TIL, at the same time providing a useful source of autologous tumour cells that can be stored and used to direct or test antitumour specificity.
Clinical Cancer Research, 2011
Purpose: Clinical trials on adoptive T-cell therapy (ACT) using expanded tumor-infiltrating lymphocytes (TIL) have shown response rates of over 50% in refractory melanoma. However, little is known how clinical and pathologic features impact TIL outgrowth isolated from metastatic melanoma tumors. Experimental Design: We analyzed the impact of clinical and pathologic features on initial TIL outgrowth in 226 consecutive patients undergoing tumor resection. Successful initial TIL outgrowth was defined as ≥40 million viable lymphocytes harvested from all tumor fragments in a 5-week culture. To normalize for the different size of resected tumors and thus available tumor fragments, we divided the number of expanded TIL by the starting number of tumor fragments (TIL/fragment). Results: Overall, initial TIL outgrowth was successful in 62% of patients, with patients ≤30 years of age (94%; P = 0.01) and female patients (71% vs. 57% for males; P = 0.04) having the highest rate of success. Syste...
Surgery, 2012
INTRODUCTION-Adoptive immunotherapy for patients with metastatic melanoma has yielded encouraging results. However, methods to expand melanoma-specific T-cells from Stage III are limited. The objective of this study is to determine whether melanoma-specific T-cells could be generated from the melanoma-draining lymph nodes (MDLN) of Stage III patients. METHODS-Stage III patients undergoing completion lymphadenectomy were enrolled onto an IRB-approved protocol. MDLN cells were tested for ability to undergo cryopreservation, expand ex vivo in IL-2 or IL-2 and IL-7 and mediate melanoma-specific antitumor responses in vitro. RESULTS-Cryopreservation produced no significant differences from fresh cultures in terms of cell growth and cellular phenotype. IL-2 and IL-2/IL-7 cultures resulted in similar growth rates, and functional studies revealed the presence of T cells which secreted interferon gamma in response to melanoma antigen peptides. Both IL-2 and IL-2/IL-7 cultured MDLN cells mediated significant apoptosis of human melanoma cell lines as compared to breast and brain tumor lines in vitro. Overall there did not seem to be a benefit of adding IL-7. Both CD4+ and CD8+ T-cells appear to mediate tumor cell apoptosis. CONCLUSION-This study demonstrates that melanoma antigen-specific T-cells can be generated from regional melanoma-draining lymph nodes and expanded ex vivo from patients with Stage III disease.