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T-Cell Culture in G-Rex

Accelerated Production of Antigen-specific T Cells for Preclinical and Clinical Applications Using Gas-permeable Rapid Expansion Cultureware (Vera et al., J. Immunother, 2009)

By careful review of this excellent publication, you will gain knowledge of the underlying principles that make G-Rex the best culture technology for production of antigen-specific T cells. It provides extensive scientific analysis of how to optimize culture conditions in G-Rex to provide many benefits.

Link www.ncbi.nlm.nih.gov/pubmed/20445351

Click on the video link below to watch a specific G-Rex protocol currently being used to produce clinical grade cytotoxic T cells.

Center for Cell and Gene Therapy, Baylor College of Medicine: Ulrike Gerdemann, Juan F. Vera, Cliona M. Rooney, Ann M. Leen

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

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

Journal of Immunotherapy: April 2012 - Volume 35 - Issue 3 - p 283–292
doi: 10.1097/CJI.0b013e31824e801f

Abstract
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.

Review the PDF below for a discussion of why proper selection of the culture device at the research stage of a project is critical for later stage efficiency.

Good manufacturing practice-grade cytotoxic T lymphocytes specific for latent membrane proteins (LMP)-1 and LMP2 for patients with Epstein-Barr virus-associated lymphoma.

Bollard CM, Gottschalk, Huls HM, Leen AM, Gee AP, Rooney CM. Cytotherapy, 2011; 13:518-533.

Challenges of T Cell Therapies for Virus-associated Diseases after Hematopoietic Stem Cell Transplantation (Leen et al. Expert Opin Biol Ther 2010)

Link http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2818535/

Immunotherapy of Human Cancers Using Gene Modified T Lymphocytes (Vera et al., Curr Gene Ther, 2009)

Link www.ncbi.nlm.nih.gov/pubmed/19860654

Nucleofection of DCs to Generate Multivirus-specific T cells for Prevention or Treatment of Viral Infections in the Immunocompromised Host (Vera et al., Curr Gene Ther, 2009)

Link www.ncbi.nlm.nih.gov/pubmed/19584818

Small Business Innovative Research Grant for TIL Therapy

We have received a grant, under which we will expend $2MM towards more efficient production of Tumor Infiltrating Lymphocytes for TIL Therapy. We will be working with the Surgery Branch of the NCI. The grant will optimize our GRex platform cell culture technology for the most practical and efficient configuration and method of TIL production. The PDF below shows that intended research plan, which was submitted in the grant application. Feel free to provide feedback and ideas to improve the plan. It is provided as a MS Word document to make it easier to mark up and comment on. Email comments to john.wilson@wilsonwolf.com. Thanks.

Optimization of Manufacturing of Virus and Tumor Specific T Cells Using G-Rex

Although ex vivo expanded T cells are currently widely used in pre-clinical and clinical trials, the complexity of manufacture is a major impediment for broader application. In this review, Baylor College of Mediine discusses traditional protocols for the ex vivo expansion of virus and tumor specific T cells and describes how G-Rex devices greatly improved and optimized their manufacturing process.

Link http://www.hindawi.com/journals/sci/2011/434392/