Erste Ergebnisse einer Phase-I-Studie zur nicht-viralen Gentherapie mittels Jet-Injektion in Hautmetastasen des Mammakarzinoms sowie Intransit-Metastasen des Malignen Melanoms

Abstract

Background: Nonviral gene delivery technologies are of increasing importance because of their easy application and the comparable low risk profile. The jet-injection technology allows efficient gene transfer of naked DNA and is based on generation of jets possessing the force to pass the skin and penetrate deeper tissues leading to transfection of the affected area. We established a phase-I-clinicaltrial to apply a LacZ-expressing reporter-plasmid by intratumoral jet-injection (DeReGe 62) to evaluate tolerance and feasibility as well as efficacy of the gene transfer. Here we present the results from the first patients. Methods: We used a jet-injector prototype (EMS Medical Systems, SA Nyon, Switzerland) and applied a pCMVβ-plasmid produced under GMP conditions. Between September 2005 and September 2006 14 patients with metastatic malignant melanoma or metastatic breast cancer were treated. All patients received 5 injections a 10μL of plasmid-DNA (1 μg/μL) into a single metastatic lesion. 72 to 96 hours after application the tumour lesion was surgically removed. In addition to the clinical and laboratory monitoring, the following molecular parameters were assessed: Detection of LacZ-expression at RNA-level by quantitative real-time PCR (qRT-PCR), and demonstration of LacZ-protein expression within the tissue (by X-Gal-staining and western blot), plasmidload within the tumour tissue by quantitative PCR (qPCR) and plasmid-clearance within the patients’ blood (30 min, 3, 6, 24, 48 and 72 hours after jet-injection). Results: All 14 patients were treated according to the protocol. No problems related to the technology and handling procedures were reported. Besides from very small bleeding signs at the injection sites, no adverse events or complications were observed during the whole treatment. The laboratory and clinical monitoring revealed no sign of systemic inflammation or any other significant findings. The treated lesions were removed and the pathologist confirmed the previous diagnoses (metastatic melanoma or breast cancer). Within all tumours plasmid-DNA (by qPCR) and LacZ-RNA (by qRT-PCR) was detected. We could prove protein expression in all tissues. The qPCR-analysis of the patients’ blood revealed a quick increase of plasmid-DNA 30 min. after jet-injection in all patients. Already 3 hours after injection, the clearance was finished and no plasmid-DNA was detected. Conclusion: The first results from this phase-I-clinical- trial clearly demonstrate the safety and feasibility of nonviral gene transfer. Within all treated tumours plasmid-DNA could be detected. This correlated well with the expression at RNA- and protein-level. The preliminary data from this trial is in accordance with previous animal experiments. For further therapeutic application increase of DNA-amount would be necessary to achieve better distribution and higher level of transgene expression. A new phase-I/II-trial for the therapeutic application of TNF-expressing-DNA has already been initiated.