Large scale expansion of gamma 9 delta 2 T lymphocytes: Innacell gamma delta cell therapy product.

gamma9delta2 T lymphocytes are non-conventional lymphocytes presenting a direct cytotoxic effect against a broad range of tumour targets. These cells also secrete inflammatory cytokines that can boost the other components of the immune system. In contrast to conventional CD8(+) T cells, the cytotoxic effect of gamma9delta2 T lymphocytes does not depend on the expression of major histocompatibility complex molecules by target tumour cells. INNACELL gammadeltatrade mark is a cell therapy product obtained by ex vivo amplification of mononuclear cells. The stimulation is achieved by a specific synthetic agonist of gamma9delta2 T lymphocytes, bromohydrin pyrophosphate (BrHPP). After a single stimulation with BrHPP, gamma9delta2 T lymphocytes are expanded for 2 weeks in a closed system in culture medium with interleukin-2 (IL-2). On day 15, cells are washed and harvested in 4% human serum albumin. In this manufacturing process, the total cell population is expanded by approximately 10-fold and gamma9delta2 T lymphocytes undergo a specific 1000-fold expansion, corresponding to a gamma9delta2 T lymphocyte enrichment of more than 70% at the end of the culture. This manufacturing process is much simpler than most current cellular therapy approaches using conventional CD8(+) T-cell lines or clones: there is no final or initial separation, no purification step and no use of feeder cells; the specific T-cell receptor-mediated signal provided by BrHPP is sufficient to trigger the IL-2-dependent expansion of the gamma9delta2 subset, which then becomes predominant in the cell culture in large amounts.

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

The 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-significant 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 effectiveness. In the group with only one invaded lymph node, the estimated relapse rate was significantly 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 differences 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 first time relationship between TIL efficiency 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 effective in vitro expansion of T cells specific for autologous tumor antigen, a finding which could be of value in future vaccine development for the treatment of melanoma.

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

The 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 autologous tumor antigen, a finding which could be of value in future vaccine development for the treatment of melanoma.