Value of large scale expansion of tumor infiltrating lymphocytes in a compartmentalised gas-permeable bag: interests for adoptive immunotherapy

<p>Abstract</p> <p>Background</p> <p>Adoptive cell therapy (ACT) has emerged as an effective treatment for patients with metastatic melanoma. However, there are several logistical and safety concerns associated with large-scale <it>ex vivo </it>expansion of tumour-specific T lymphocytes for widespread availability of ACT for cancer patients. To address these problems we developed a specific compartmentalised bag allowing efficient expansion of tumour-specific T lymphocytes in an easy handling, closed system.</p> <p>Methods</p> <p>Starting from lymph nodes from eight melanoma patients, we performed a side-by-side comparison of Tumour-Infiltrating Lymphocytes (TIL) produced after expansion in the compartmentalised bag versus TIL produced using the standard process in plates. Proliferation yield, viability, phenotype and IFNγ secretion were comparatively studied.</p> <p>Results</p> <p>We found no differences in proliferation yield and cell viability between both TIL production systems. Moreover, each of the cell products complied with our defined release criteria before being administered to the patient. The phenotype analysis indicated that the compartmentalised bag favours the expansion of CD8+ cells. Finally, we found that TIL stimulated in bags were enriched in reactive CD8+ T cells when co-cultured with the autologous melanoma cell line.</p> <p>Conclusions</p> <p>The stimulation of TIL with feeder cells in the specifically designed compartmentalised bag can advantageously replace the conventional protocol using plates. In particular, the higher expansion rate of reactive CD8+ T cells could have a significant impact for ACT.</p

Clinical Study Adoptive TIL Transfer in the Adjuvant Setting for Melanoma: Long-Term Patient Survival

Two first analyses of our clinical trial on TIL as adjuvant therapy for melanoma were published in 2002 and 2007. We present here an update of the clinical results after a 17-year median followup. In this trial, disease-free patients were randomly assigned to receive either TIL/IL-2 or IL-2. The relapse-free survival (RFS) was the primary objective. Eighty-eight patients were enrolled. A new analysis performed in May 2013 did not show significant changes in RFS or OS duration. However, our first finding on the association between the number of invaded lymph nodes and TIL effectiveness was strengthened. The Cox model adjusted on this interaction showed for the first time a significant treatment effect when considering the overall population, both on the RFS and OS. Patients treated with TIL had a longer RFS ( = 0.023) or OS ( = 0.020). This study being with a very long followup (17 years), confirmed the association between TIL effectiveness and the number of invaded lymph nodes, indicating that a low tumor burden could be a crucial factor enhancing the curative effect of TIL in possible microscopic residual disease. Moreover, we confirmed that a prolonged survival was associated with the presence of specific TIL and a decrease in Foxp3 expression

High-scale expansion of melanoma-reactive TIL by a polyclonal stimulus: predictability and relation with disease advancement

International audienceThe rationale of treating melanoma patients by infusion with tumor-infiltrating leukocytes (TIL) is to perform an adoptive therapy through injection of tumor-specific T cells. Nonetheless, methods currently used for ex vivo TIL expansion have not been evaluated for their efficacy to expand TAA-specific T cells. We have addressed this question here, using a culture method in which high TIL growth was induced by a polyclonal T cell stimulus. Intracellular cytokine assays were performed to measure the proportion of T cells responding to autologous tumor cells among the lymphocytes from lymph node biopsies (TIL) of 26 patients with stage III melanoma. The data show that TIL from 18 of these patients contained detectable amounts of tumor-specific T cells before expansion. Although they decreased somewhat in percent abundance during expansion, they were still present afterwards, ranging from 0.3 to 13.8%. Since a median number of 1.7 x 10(10) TIL was obtained from these patients (starting from 3.6 x 10(6) TIL), a total amount of tumor-reactive cytokine-secreting TIL of between 2.8 x 10(6) and 1.12 x 10(9) was obtained in each case from 18 patients. The TIL populations from 8 patients did not contain tumor-reactive T cells: neither before expansion, nor after expansion. Lack of tumor-reactive TIL only occurs for patients bearing several tumor-invaded lymph nodes (40%), but not for those having a single invaded lymph node. Therefore, high numbers of tumor-reactive T cells can be produced, through a polyclonal TIL stimulation, from most early stage III melanoma patients but from only about half of the patients with a more disseminated disease. For this last group, the possibility of getting tumor-reactive TIL can be predicted by checking the presence of these cells before expansion

Adoptive TIL Transfer in the Adjuvant Setting for Melanoma: Long-Term Patient Survival

Two first analyses of our clinical trial on TIL as adjuvant therapy for melanoma were published in 2002 and 2007. We present here an update of the clinical results after a 17-year median followup. In this trial, disease-free patients were randomly assigned to receive either TIL/IL-2 or IL-2. The relapse-free survival (RFS) was the primary objective. Eighty-eight patients were enrolled. A new analysis performed in May 2013 did not show significant changes in RFS or OS duration. However, our first finding on the association between the number of invaded lymph nodes and TIL effectiveness was strengthened. The Cox model adjusted on this interaction showed for the first time a significant treatment effect when considering the overall population, both on the RFS and OS. Patients treated with TIL had a longer RFS (P=0.023) or OS (P=0.020). This study being with a very long followup (17 years), confirmed the association between TIL effectiveness and the number of invaded lymph nodes, indicating that a low tumor burden could be a crucial factor enhancing the curative effect of TIL in possible microscopic residual disease. Moreover, we confirmed that a prolonged survival was associated with the presence of specific TIL and a decrease in Foxp3 expression

Preclinical Assessment of Autologous Tolerogenic Dendritic Cells From End-stage Renal Disease Patients

International audienceBackground: Kidney transplantation is the therapeutic of choice for patients with kidney failure. While immunosuppressive drugs can control graft rejection, their use is associated with increased infections and cancer, and they do not effectively control chronic graft rejection. Cell therapy is an attractive strategy to minimize the use of pharmacological drugs.Methods: We recently developed a protocol to generate human monocyte-derived autologous tolerogenic dendritic cells (ATDCs) from healthy volunteers. Herein, we transferred the ATDC manufacturing protocol to a Good Manufacturing Practice (GMP)-compliant facility. Furthermore, we compared the phenotype and in vitro functions of ATDCs generated from patients with end-stage renal disease to those generated from healthy volunteers.Results: We describe the critical steps for GMP-compliant production of ATDCs and define the quality criteria required to allow release of the cell products. Furthermore, we showed that ATDCs generated from healthy volunteers and patients with kidney failure display the same tolerogenic profile based on their phenotype, resistance to maturation, and ability to modulate T-cell responses.Conclusions: Together, these results allowed us to define the production process and the quality criteria for the release of ATDCs before their administration in patients receiving a kidney transplant

Randomized trial of adoptive transfer of melanoma tumor-infiltrating lymphocytes as adjuvant therapy for stage III melanoma

International audienceThe aim of this study was to demonstrate the interest of using tumor-infiltrating lymphocytes (TIL) as adjuvant therapy for stage III (regional lymph nodes) melanoma. After lymph node excision, patients without any detectable metastases were randomly assigned to receive either TIL plus interleukin-2 (IL-2) for 2 months, or IL-2 only. The primary endpoint was determination of the duration of the relapse-free interval. Eighty-eight patients determined as eligible for treatment were enrolled in the study. After a median follow-up of 46.9 months, for the study population the analysis did not show a significant extension of the relapse-free interval or overall survival. However, a significant interaction (P<0.001) was found between the treatment and the number of invaded lymph nodes. In the group with only one invaded lymph node, the estimated relapse rate was significantly lower (P adjusted =0.0285) and the overall survival was increased (P adjusted =0.039) in the TIL+IL-2 arm compared with the IL-2 only arm. No differences between the two arms, either as regards the duration of disease-free survival or overall survival, were noted in the group with more than one invaded lymph node whatever the number of invaded lymph nodes. Treatment was compatible with normal daily activity. This study demonstrates for the first time that the efficiency of TIL in stage III melanoma (AJCC) is directly related to the number of invaded lymph nodes, indicating that tumor burden might be a crucial factor in the efficacy and/or in vitro expansion of T cells specific for autolo-gous tumor antigen, a finding which could be of value in future vaccine development for the treatment of melanoma

Long-term follow-up of patients treated by adoptive transfer of melanoma tumor-inWltrating lymphocytes as adjuvant therapy for stage III melanoma

International audienceThe first analysis of our clinical trial on interest of using tumor-infiltrating lymphocytes (TIL) as adjuvant therapy for stage III (regional lymph nodes) melanoma was published in 2002 [5]. The aim of this paper is to update clinical results of 7 years of follow-up after the last treated patient. In the trial conducted between December 1993 and January 1999, patients without any detectable metastases after lymph node excision were randomly assigned to receive either TIL plus interleukin-2 (IL-2) for 2 months, or IL-2 only. The duration of the relapse-free interval was the primary objective. Eighty-eight patients were enrolled in the study. Currently, the last analysis performed in June 2006, after a median follow-up of 114.8 months, did not show change of non-signiWcant extension of the relapse-free interval or overall survival. However, this second analysis strengthens our first hypothesis about the relationship between number of invaded lymph nodes and TIL treatment eVectiveness. In the group with only one invaded lymph node, the estimated relapse rate was signiWcantly lower (P adjusted = 0.0219) and the overall survival was increased (P adjusted = 0.0125) in the TIL+IL-2 arm compared with the IL-2 only arm. No diVerences between the two arms, either with regard to the duration of disease-free survival (P adjusted = 0.38) or overall survival (P adjusted = 0.43), were noted in the group with more than one invaded lymph node, whatever the number of invaded lymph nodes. Treatment was compatible with normal daily activity. This study, with a very long follow up (median of almost 10 years), postulates for the Wrst time relationship between TIL eYciency in stage III melanoma (AJCC) and number of invaded lymph nodes, indicating that tumor burden might be a crucial factor in the production of an eVective in vitro expansion of T cells speciWc for autologous tumor antigen, a finding which could be of value in future vaccine development for the treatment of melanoma

A phase I/II feasibility vaccine study by autologous leukemic apoptotic corpse-pulsed dendritic cells for elderly AML patients

International audienceThis was a phase I/II study testing the feasibility of a vaccine by autologous leukemic apoptotic corpse-pulsed dendritic cells (DC) in elderly acute myeloid leukemia (AML) patients in first or second complete remission. Pulsed DC were administered at doses of 9 × 106 subcutaneously (1 mL) and 1 × 106 intra-dermally (0.1 mL). Five doses of vaccine were planned on days +1 + 7 + 14 + 21 and +35. Five DC-vaccines were produced and injected for all five patients included in the study. No severe adverse event was documented. Larger Phase 2 studies are now required to precise the role of DC-vaccines with leukemic apoptotic bodies in older as well as younger AML populations