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Simplified Method of the Growth of Human Tumor Infiltrating Lymphocytes in Gas-permeable Flasks to Numbers Needed for Patient Treatment

2012; Lippincott Williams & Wilkins; Volume: 35; Issue: 3 Linguagem: Inglês

10.1097/cji.0b013e31824e801f

1537-4513

Jianjian Jin, Marianna Sabatino, Robert Somerville, John R. Wilson, Mark E. Dudley, David F. Stroncek, Steven A. Rosenberg,

Virus-based gene therapy research

Adoptive cell therapy of metastatic melanoma with autologous tumor infiltrating lymphocytes (TIL) is clinically effective, but TIL production can be challenging. Here we describe a simplified method for initial TIL culture and rapid expansion in gas-permeable flasks. TIL were initially cultured from tumor digests and fragments in 40 mL capacity flasks with a 10 cm² gas-permeable silicone bottom, G-Rex10. A TIL rapid expansion protocol (REP) was developed using 500 mL capacity flasks with a 100 cm² gas-permeable silicone bottom, G-Rex100. TIL growth was successfully initiated in G-Rex10 flasks from tumor digests from 13 of 14 patients and from tumor fragments in all 11 tumor samples tested. TIL could then be expanded to 8-10×10⁹ cells in a 2-step REP that began by seeding 5×10⁶ TIL into a G-Rex100 flask, followed by expansion at day 7 into 3 G-Rex100 flasks. To obtain the 30-60×10⁹ cells used for patient treatment, we seeded 6 G-Rex100 flasks with 5×10⁶ cells and expanded into 18 G-Rex100 flasks. Large-scale TIL REP in gas-permeable flasks requires approximately 9-10 L of media, about 3-4 times less than other methods. In conclusion, TIL initiation and REP in gas-permeable G-Rex flasks require fewer total vessels, less media, less incubator space, and less labor than initiation and REP in 24-well plates, tissue culture flasks, and bags. TIL culture in G-Rex flasks will facilitate the production of TIL at the numbers required for patient treatment at most cell processing laboratories.