Abstract
This phase I clinical trial evaluated safety, feasibility, and efficiency of nonviral intratumoral jet-injection gene transfer in patients with skin metastases from melanoma and breast cancer. Seventeen patients were enrolled. The patients received five jet injections with a total dose of 0.05 mg beta-galactosidase (LacZ)-expressing plasmid DNA (pCMVbeta) into a single cutaneous lesion. Clinical and laboratory safety monitoring were done. Systemic plasmid clearance was monitored by quantitative real-time PCR of blood samples throughout the study. All lesions were resected after 2 to 6 days. Intratumoral plasmid DNA load, DNA distribution, and LacZ expression was analyzed by quantitative real-time PCR, quantitative reverse transcription-PCR, Western blot, immunohistochemistry, and 5-bromo-4-chloro-3-indolyl-beta-D-galactoside staining. Jet injection of plasmid DNA was safely done in all patients. No serious side effects were observed. Thirty minutes after jet injection, peak plasmid DNA levels were detected in the blood followed by rapid decline and clearance. Plasmid DNA and LacZ mRNA and protein expression were detected in all treated lesions. Quantitative analysis revealed a correlation of plasmid DNA load and LacZ-mRNA expression confirmed by Western blot. Immunohistochemistry and 5-bromo-4-chloro-3-indolyl-beta-D-galactoside staining showed LacZ-protein throughout the tumor. Transfected tumor areas were found close and distant to the jet-injection site with varying levels of DNA load and transgene expression. Intratumoral jet injection of plasmid DNA led to efficient LacZ reporter gene expression in all patients. No side effects were experienced, supporting safety and applicability of this novel nonviral approach. A next step with a therapeutic gene product should determine antitumor efficacy of jet-injection gene transfer.
Highlights
This phase I clinical trial evaluated safety, feasibility, and efficiency of nonviral intratumoral jet-injection gene transfer in patients with skin metastases from melanoma and breast cancer
Quantitative analysis revealed a correlation of plasmid DNA load and LacZ-mRNA expression confirmed by Western blot
The major challenge in clinical gene therapy is the assurance of safety and transfer efficiency. In this phase I trial, we proved for the first time the safety and efficacy of nonviral intratumoral jet injection of naked LacZ-expressing plasmid DNA
Summary
The phase I trial evaluated safety and feasibility of the nonviral jet-injection transfer of the LacZ-expressing naked reporter plasmid DNA in skin metastases of breast cancer and melanoma. Patients with skin metastases from breast cancer or melanoma received five injections of naked plasmid DNA into the metastatic lesion Blood samples were collected before and 0.5, 3, 6, 24, 48, and 72 h and 4 weeks after jet injection for analysis of clinical laboratory variables and for quantitative analysis of plasmid DNA dissemination and clearance. Sections (7 Am) from shock-frozen specimens were used for plasmid DNA, mRNA, and protein analyses, for detection of LacZ expression by immunohistochemistry, and for detection of functional LacZ by 5-bromo-4-chloro-3-indolyl-h-D-galactoside (X-Gal) staining (Fig. 1B). Capillary gel electrophoresis of pCMVh plasmid was done to analyze plasmid DNA topology shortly before and after jet injection. Capillary gel electrophoresis analyses were done using a Beckman P/ACE MDQ instrument equipped with a LIF detector (488/520 nm) as described (24 – 26)
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
