Process validation and clinical evaluation of a protocol to generate gene-modified T lymphocytes for imunogene therapy for metastatic renal cell carcinoma: GMP-controlled transduction and expansion of patient's T lymphocytes using a carboxy anhydrase IX-specific scFv transgene

Background Adoptive transfer of autologous T cells that are gene-transduced to express Ag-specific receptors represents an experimental strategy to provide tumor-specific immunity to cancer patients. We studied this concept in patients with metastatic renal cell cancer (RCC) using retroviral transduction of T cells with a single-chain Ab-G250 chimeric receptor [scFv(G250)]. We describe the validation of our clinical protocol for gene transduction and expansion of human T lymphocytes. Methods A batch of scFv(G250) transgene-containing retrovirus was produced under conditions of good manufacturing practice ( GMP). In addition to quality control and safety testing of the virus batch, extensive potency testing was performed, i.e. assessment of its functional transduction efficiency in primary human T cells. Subsequently, the clinical gene transduction and cell-expansion protocol was subjected to a series of process validations and a clinical evaluation using T cells obtained from healthy donors and three RCC patients. Results The clinical batch of scFv(G250) transgene-containing retrovirus met the quality and safety control criteria. Small-scale transductions yielded 62-92% scFv(G250)(+) T cells and, at a clinical scale, 50- 84% transduction efficiencies were obtained. Patient and healthy donor T cells showed similar expansion potencies, and also yielded similar levels of scFv(G250)-mediated immune functions, i.e. specific cytolysis of G250-ligand expressing RCC cells and production of IFN-gamma upon stimulation with such cells. All T cell cultures were free of replication competent retroviruses. Discussion We have shown that the validated batch of scFv(G250) transgene-containing retrovirus in combination with our GMPT-cell transduction and expansion protocol successfully generates clinically relevant numbers of functional scFv(G250) gene-modified T cells for patient treatment.