Gibbon ape leukemia virus poorly replicates in primary human T lymphocytes: implications for safety testing of primary human T lymphocytes transduced with GALV-pseudotyped vectors

• Published in: Journal of immunotherapy (1997) Vol. 32; no. 3; p. 272
• Main Authors: Lamers, Cor H J, Willemsen, Ralph A, van Elzakker, Pascal M M L, Gratama, Jan Willem, Debets, Reno
• Format: Journal Article
• Language: English
• Published: United States 01.04.2009
• Subjects: Animals; Biological Assay; Genetic Vectors - genetics; Genetic Vectors - physiology; Humans; Leukemia Virus, Gibbon Ape - genetics; Leukemia Virus, Gibbon Ape - physiology; Sensitivity and Specificity; T-Lymphocytes - virology; Transduction, Genetic; Virus Replication
• ISSN: 1537-4513
• DOI: 10.1097/CJI.0b013e318199840a

The Food and Drug Administration/Center for Biologics Evaluation and Research has defined that for retroviral gene therapy, the vector-producing cell, the vector preparation, and the ex vivo gene-transduced cells have to be tested for absence of replication-competent retrovirus (RCR) if the transduced cells are cultured for >4 days. We assessed the sensitivity of the "extended PG4(S+L-) assay" to detect gibbon ape leukemia virus (GALV) RCR, and applied this assay to measure GALV RCR spread in retrovirally transduced T cells. To this end, T cells were expanded for 12 days after transduction with a GALV-envelope pseudotyped retroviral vector expressing single chain variable fragment (anticarbonic anhydrase IX) in presence or absence of GALV RCR. Results showed that: (1) the "extended PG4(S+L-) assay" detects 1 focus-forming unit (ffu) GALV RCR and thus is applicable and sufficiently sensitive to screen human T-cell cultures for absence of infectious GALV RCR; (2) although GALV RCR infect human T cells, it very poorly replicate in T cells; (3) GALV RCR, when present at low levels immediately upon gene transduction (ie, 100 ffu/20x10 T cells in 100 mL), did not spread during a 12-day T-cell culture at clinical scale. Our observation that GALV RCR poorly spreads in primary human T-cell cultures questions the relevance of testing T-cell transductants for RCR on top of testing the vector-producing cells and the clinical vector batch for RCR and warrants evaluation of the current policy for safety testing of ex vivo retrovirally transduced T lymphocytes for GALV RCR.