TO THE EDITOR: Adoptive transfer of autologous T-lymphocytes that are gene transduced to express antigen-specific receptors represents an experimental therapy to provide tumor-specific immunity to cancer patients. We studied safety and the proof of this concept in patients with metastatic renal cell carcinoma (RCC), and have encountered toxicity that is likely to be antigen specific. We have constructed a single-chain antibody-type (scFv)–receptor based on murine monoclonal antibody (mAb) G250. 1 This mAb recognizes an epitope on carboxy-anhydrase-IX (CAIX), which is frequently overexpressed on clear cell RCC. Following retroviral introduction of the scFv (G250) transgene into primary human T-cells, the scFv (G250) receptor is expressed on the surface of these cells, which enables them to recognize CAIX and to exert antigen-specific effector functions, such as cytokine production after exposure to CAIX and the killing of CAIX RCC cell lines. 2, 3 We treated patients with scFv (G250)-transduced T-cells in an inpatient dose-escalation scheme of intravenous (IV) doses of 2 107 cells at day 1; 2 108 cells at day 2; 2 109 cells at days 3 through 5 (treatment cycle 1); and 2 109 cells at days 17 to 19 (treatment cycle 2), in combination with human recombinant interleukin-2 (IL-2; Chiron Corporation, Amsterdam, the Netherlands), subcutaneously, 5 105 U/m2 twice daily administered at days 1 to 10 and days 17 to 26. This protocol was approved by the governmental regulatory authorities and the institutional medical ethical review board. Adaptations to this protocol were implemented only after approval by these boards. Written informed consent was obtained from all patients. In this letter, we report on the clinical experiences of the first three patients. The patients had CAIX metastatic clear cell RCC, had undergone tumor nephrectomy, and had progressive disease after 6 to 17 months of interferon alfa (IFN-) treatment. From all, we successfully generated functional scFv (G250) T-cells ex vivo (Table 1). Infusions of these gene-modified T-cells were initially well-tolerated. However, after four to five infusions, liver enzyme disturbances reaching National Cancer Institute Common Toxicity Criteria grades 2 to 4 developed. These toxicities necessitated the cessation of treatment in patient 1 and patient 3, corticosteroid treatment in patient 1, and reduction of the maximal T-cell dose to 2108 T-cells in patient 2 and patient 3. After treatment, patients showed progressive disease between 36 and 106 days. In order to elucidate the underlying mechanismsaccountingforthelivertoxicity, aliverbiopsywasperformedon patient 1, showing a discrete cholangitis with T-cell infiltration around the bile ducts, and CAIX expression on the bile duct epithelial cells. Although technical limitations prohibited direct identification of scFv (G250) T-cells in these sections, these findings strongly suggest that the liver toxicity is caused by a specific attack of the scFv (G250) T-cells against the CAIX bile duct epithelial cells. We transiently detected both scFv (G250) T-cells and scFv (G250) DNA copies in the circulation of all three patients from day 3 of treatment onward, using flow cytometry and quantitative real-time polymerase chain reaction (PCR). The time period during which the transduced cells were detected in the circulation depended on the method used, that is, up to 32 days by flow cytometry and up to 53 days by PCR4 (Table 1).

Before treatment, peripheral blood mononuclear cells did not show scFv (G250)-mediated functions, that is, specific cytolysis of CAIX target cells and production of IFN-on stimulation with such cells. After infusions of scFv (G250)-transduced T cells, these …