Minimally Cultured Tumor-infiltrating Lymphocytes Display Optimal Characteristics for Adoptive Cell Therapy

Abstract

Adoptive cell therapy (ACT) with tumor reactive lymphocytes in patients with refractory melanoma can result in tumor regression and prolonged survival. Generating tumor reactive lymphocyte cultures is technically difficult and resource intensive; these limitations have restricted the widespread application of ACT. Tumor infiltrating lymphocytes (TIL) from melanoma contain tumor antigen reactive cells. The “standard” method for producing TIL cultures for clinical administration requires extended in vitro expansion in interleukin-2, then identification of tumor reactive cells by immunological assays. We show here that limitations in reagents and methods during screening under-represent the actual reactivity of TIL cultures. Furthermore, the extended culture times necessitated by the screening assays resulted in telomere shortening and reduced expression of CD27 and CD28 in the TIL cultures, properties that our prior studies showed are correlated within vivo persistence and clinical response. We have thus developed an alternative “young” TIL method that demonstrated superior in vitro attributes compared with standard TIL. This approach utilizes the entire resected tumor to rapidly expand TIL for administration without in vitro testing for tumor recognition. Our observations suggest that younger TIL can have an undetermined but high level of antigen reactivity, and other advantageous attributes such as long telomeres and high levels of CD27 and CD28. We suggest that minimally cultured, unselected lymphocytes represent an alternative strategy for generating TIL cultures suitable for use in ACT therapy that, if effective in vivo, may facilitate the widespread application of this approach to a broader population of patients with melanoma.

Adoptive Transfer of Cytotoxic T Lymphocytes Targeting Two Different Antigens Limits Antigen Loss and Tumor Escape

“…There have been many attempts to identify the optimal phenotype of, and culture conditions for, adoptively transferred T cells (Zeh et al, 1999;Klebanoff et al, 2005;Janicki et al, 2008;Tran et al, 2008;Hinrichs et al, 2009;Caserta et al, 2010;Garcia-Hernandez et al, 2010;Pouw et al, 2010). Our results here suggest that therapy may also be significantly enhanced by combining multiple populations, with distinct characteristics, including the targeting of different antigens.…”

Chemokine receptor engineering of T cells with CXCR2 improves homing towards subcutaneous human melanomas in xenograft mouse model

“…When cell counts of >50 × 10 6 T cells were reached (10 cultures, expansion time-range; 22–65 days in culture), from here designated “Young” TILs. 21 Flow cytometry was performed on 2 × 10 5 cells from each Young TIL culture. The remaining Young TILs were cryopreserved according to protocol described above.…”

Tumor Regression in Patients With Metastatic Synovial Cell Sarcoma and Melanoma Using Genetically Engineered Lymphocytes Reactive With NY-ESO-1

“…1,2 The observation that melanoma-reactive TILs could be generated from only 50% of resected samples 3 and the difficulty in generating tumorreactive TILs from other cancer types have prompted cell transfer studies using autologous T cells that have been genetically engineered to express T-cell receptors (TCRs) directed against shared tumor antigens. In a recent trial targeting the MART-1 melanocyte differentiation antigen, an objective response rate of 30% was observed.…”