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Phoenix-ampho outperforms PG13 as retroviral packaging cells to transduce human T cells with tumor-specific receptors: implications for clinical immunogene therapy of cancer.
- Source :
-
Cancer gene therapy [Cancer Gene Ther] 2006 May; Vol. 13 (5), pp. 503-9. - Publication Year :
- 2006
-
Abstract
- We have designed a transgene that encodes a scFv(G250) chimeric receptor, which is specific for carboxyanhydrase IX (G250-ligand, G250L), a molecule overexpressed by renal cell cancer (RCC). Retroviral transduction of this transgene into primary human T lymphocytes confers these cells with specific functional responses towards G250L-positive RCC cells. In preparation of a clinical phase (I/II) study in RCC patients, we set up a protocol for gene transduction and expansion of primary human T cells. For this purpose, we directly compared two packaging cell lines, that is, the GALV-pseudotyped MLV producing cell line PG13, and the MLV-A-producing cell line Phi-NX-Ampho (a.k.a. Phoenix-A). We generated and characterized stable scFv(G250)-positive clones of both PG13 and Phoenix cells and optimized the retrovirus production conditions. Transductions of primary human T cells yielded 30-60% scFv(G250)+ T cells using PG13-derived retrovirus versus up to 90% scFv(G250)+ T cells using Phoenix-derived retrovirus. The median number of transgene integrations per scFv(G250)+ T cell differed only 1.5-fold as determined by real-time PCR (mean number of integrations per T cell 2.6 and 3.7 for PG13 and Phoenix-based transductions, respectively). In addition, T cells transduced with Phoenix-derived retrovirus showed, on a per cell basis, 10-30% higher levels of scFv(G250)-mediated TNFalpha production and cytolysis of G250L+ RCC cells than T cells transduced with PG13-derived retrovirus. The improved functional transduction efficiency together with a limited increase in the number of integrations per recipient cell, made us select Phoenix clone 58 for our clinical immunogene therapy study.
- Subjects :
- Carbonic Anhydrase IX
Cell Line
Cells, Cultured
Combined Modality Therapy
Cytotoxicity, Immunologic
Genetic Therapy methods
Humans
Immunotherapy, Adoptive methods
Retroviridae genetics
Single-Chain Antibodies
Antigens, Neoplasm genetics
Carbonic Anhydrases genetics
Immunoglobulin Variable Region genetics
Kidney Neoplasms therapy
Receptors, Immunologic genetics
T-Lymphocytes immunology
T-Lymphocytes transplantation
Transduction, Genetic methods
Virus Assembly
Subjects
Details
- Language :
- English
- ISSN :
- 0929-1903
- Volume :
- 13
- Issue :
- 5
- Database :
- MEDLINE
- Journal :
- Cancer gene therapy
- Publication Type :
- Academic Journal
- Accession number :
- 16282986
- Full Text :
- https://doi.org/10.1038/sj.cgt.7700916