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A Closed Culture System For the Ex Vivo Transduction and Expansion of Human T Lymphocytes

  1. Author:
    Robinet, E.
    Certoux, J. M.
    Ferr, C.
    Maples, P.
    Hardwick, A.
    Cahn, J. Y.
    Reynolds, C. W.
    Jacob, W.
    Herve, P.
    Tiberghien, P.
    1. Year: 1998
  2. Journal: Journal of Hematotherapy
    1. 7
    2. 3
    3. Pages: 205-215
  4. Type of Article: Article
  1. Abstract:

    A phase I clinical trial is currently being performed at our institution, with the aim of evaluating the feasibility and toxicity related to the administration of herpes simplex thymidine kinase gene-expressing human primary T lymphocytes following allogeneic hematopoietic stem cell transplantation. The need for safe and standardized preparation conditions for gene-modified cells is crucial. We describe the closed culture system used in the current trial for ex vivo retroviral-mediated gene transfer and transduced cell selection. Cell handling is performed in closed systems using a sterile connection device that avoids opening the culture system. Cell numbers during the production process increased from 93 +/- 16 on day 0 to 440 +/- 92 x 10(6) on day 12 (7.2 +/- 1.4-fold increase) (n = 11). Transduction efficiency before and after G418 resistance-based selection was 13.5 +/- 3.8% and 90.0 +/- 1.4%, respectively. Safety and efficacy testing included a search for replication-competent retrovirus, endotoxins, Mycoplasm, and bacterial contamination (n = 0/9), PCR-DNA, % CD3 + cells (91 +/- 2%), and viability after thawing (82 +/- 3%). Effective working time from day 0 to day 12 is similar to 20 h. The closed system we developed allows for safe and reproducible ex vivo preparation of gene-modified primary T lymphocytes for clinical use. [References: 11]

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