Isolated Limb Perfusion for Local Gene Delivery (original) (raw)
Cancer Gene Therapy, 2003
The immune response is modulated by genetic adjuvants using plasmid vectors expressing cytokines. Skeletal muscle can express a foreign gene intramuscularly administered via a needle injection, and the potential of muscle as a target tissue for somatic gene therapy in treating cancer has been explored. In the present study, we investigated the efficacy of particle-mediated intramuscular transfection modified with a local anesthetic agent, bupivacaine, on luciferase and green fluorescent protein. The results indicate that these proteins are more efficiently expressed and persist longer in muscle modified in this way compared with the needleinjection method. Using an established rat sarcoma model, particle-mediated intramuscular gene-gun therapy with a combination of IL-12 and IL-18 cDNA was conducted. Growth of the distant sarcoma was significantly inhibited by particle-mediated intramuscular combination gene therapy, and the survival rate was also improved. Furthermore, the combination gene-gun therapy maintained significant levels of interferon-g and induced a high activity of tumor-specific cytotoxic T lymphocytes. These results suggest that the sustained local delivery of IL-12 and IL-18 cDNA using intramuscular gene-gun therapy modified with bupivacaine can induce long-term antitumor immunity, and can provide the great advantage of inhibiting the disseminated tumor.
Locoregional delivery of adenoviral vectors
Journal of Nuclear …, 2006
The overall median survival of patients with a malignant glioma is ,1 y. Because malignant gliomas rarely metastasize outside the skull, locoregional treatment strategies, such as gene therapy, are under investigation. Recently, convection-enhanced delivery (CED) has been presented as a method to improve delivery of large molecules. The goal of this study was to evaluate whether CED improves intratumoral delivery of adenoviral vectors and compare it with single injection (SI) and multiple injection (4•, MI). Methods: A replication-deficient adenoviral vector encoding the herpes simplex virus thymidine kinase (HSV-tk) and the human somatostatin receptor subtype 2 (sst 2) was administered into nude mice bearing subcutaneous U87 xenografts. Tumors were injected with 1.5 • 10 9 plaque-forming units of Ad5.tk.sstr by CED, SI, or MI. Three days later, [ 99m Tc-N 4 0-1 ,Asp 0 ,Tyr 3 ] octreotate (99m Tc-Demotate 2) was injected intravenously to monitor the virus-induced sst 2 expression. g-Camera imaging was performed for in vivo imaging, and the tumor uptake of 99m Tc-Demotate 2 was determined by g-counter. Furthermore, the tumor was sectioned and ex vivo autoradiography was performed. After decay of radioactivity, adjacent sections were submitted to in vitro autoradiography with 125 I-DOTA-Tyr 3-octreotate, which was used to calculate the transduced areas. Results: Transfected xenograft tissues showed high sst 2 expression and were clearly visualized with a g-camera. Accumulation of radioactivity was 2-fold higher in the tumors that were injected with MI compared with CED and SI (P 5 0.01). CED and SI resulted in equal uptake of radioactivity in the tumors. The measured areas of transduction in ex vivo and in vitro autoradiographs showed a high concordance (r 2 5 0.89, P , 0.0001). The maximum area of transfection was significantly larger after MI than after CED (P , 0.05) or SI (P 5 0.05). Also, the measured volume of distribution was twice as high after administration of Ad5.tk.sstr by MI (56.6 mm 3) compared with SI (25.3 mm 3) or CED (26.4 mm 3). Conclusion: CED does not increase adenoviral vector distribution in a glioma xenograft model compared with SI. Therefore, in the clinic MI is probably the most effective delivery method for the large adenoviral particle (70 nm) in malignant gliomas.
Adenovirus-mediated delivery of antiangiogenic genes as an antitumor approach
Cancer Gene Therapy, 2000
Based on the observation that the growth of solid tumors is dependent on the formation of new blood vessels, therapeutic strategies aimed at inhibiting angiogenesis have been proposed. A number of proteins with angiostatic activity have been described, but their development as therapeutic agents has been hampered by difficulties in their production and their poor pharmacokinetics. These limitations may be resolved using a gene therapy approach whereby the genes are delivered and expressed in vivo. Here we compared adenoviral delivery of endostatin, proliferin -related protein ( PRP ) , and interferon -inducible protein 10 ( IP10 ) genes. Recombinant adenoviruses carrying the three angiostatic genes express biologically active gene products as determined in vitro in endothelial cell proliferation and migration assays, and in vivo by inhibition of neoangiogenesis in rat chambers. Eradication of established tumors in vivo, in the murine B16F10 melanoma model in immunocompetent mice, was not achieved by intratumoral injection of the different vectors. However, the combination of intravenous plus intratumoral injections allowed rejection of tumors. Ad -PRP or Ad -IP10 were significantly more efficient than Ad -endostatin, leading to complete tumor rejection and prolonged survival in a high proportion of treated animals. These data support the use of in vivo gene delivery approaches to produce highcirculating and local levels of antiangiogenic agents for the therapy of local and metastatic human tumors. Cancer Gene Therapy ( 2001 ) 8, 45 ± 54
Gene therapy prospects--intranasal delivery of therapeutic genes
Advances in clinical and experimental medicine : official organ Wroclaw Medical University
Gene therapy is recognized to be a novel method for the treatment of various disorders. Gene therapy strategies involve gene manipulation on broad biological processes responsible for the spreading of diseases. Cancer, monogenic diseases, vascular and infectious diseases are the main targets of gene therapy. In order to obtain valuable experimental and clinical results, sufficient gene transfer methods are required. Therapeutic genes can be administered into target tissues via gene carriers commonly defined as vectors. The retroviral, adenoviral and adeno-associated virus based vectors are most frequently used in the clinic. So far, gene preparations may be administered directly into target organs or by intravenous, intramuscular, intratumor or intranasal injections. It is common knowledge that the number of gene therapy clinical trials has rapidly increased. However, some limitations such as transfection efficiency and stable and long-term gene expression are still not resolved. Co...
Surgery, 2001
Background. Many gene therapy strategies would benefit from efficient, regional organ delivery of therapeutic genes. Methods. Regional perfusions of lung, liver, or bladder were performed to determine if rapid and efficient gene transfer can be accomplished in vivo, and to determine if in vivo gene transfer can be limited to the organ of interest. In addition, herpes simplex virus tumor necrosis factor (HSVtnf), carrying the human tumor necrosis factor α gene was used as a treatment for methylcholanthrene sarcoma in a syngeneic lung metastases model in Fisher rats.
Journal of General Virology, 2004
There is a need for improvement of the commonly used adenovirus vectors based on serotype 5. This study was performed on three adenovirus serotypes with a CAR-binding motif (Ad4p, Ad5p and Ad17p) and three non-CAR-binding serotypes (Ad11p, Ad16p and Ad21p). The capacity of these alternative adenovirus vector candidates to deliver DNA into low-passage glioma cell lines from seven different donors was evaluated. The non-CAR-binding serotype Ad16p was the most efficient serotype with regard to import of its DNA, as well as initiation of hexon protein expression. Ad16p established hexon expression in 60-80 % of the cell population in gliomas from all donors tested. The other non-CAR-binding serotypes, Ad11p and Ad21p, showed hexon expression in 25-60 and 40-80 % of cells, respectively. The corresponding figure for the best CAR-binding serotype, Ad5p, was only 25-65 %, indicating greater variability between cells from different donors than serotype Ad16p had. The other CAR-binding serotypes, Ad4p and Ad17p, were refractory to some of the gliomas, giving a maximum of only 45 and 40 % hexon expression, respectively, in the most permissive cells. Interestingly, the transduction capacity of the CAR-binding serotypes was not correlated to the level of CAR expression on the cells.
Nonreplicating Adenoviral Vectors: Improving Tropism and Delivery of Cancer Gene Therapy
Cancers, 2021
Recent preclinical and clinical studies have used viral vectors in gene therapy research, especially nonreplicating adenovirus encoding strategic therapeutic genes for cancer treatment. Adenoviruses were the first DNA viruses to go into therapeutic development, mainly due to well-known biological features: stability in vivo, ease of manufacture, and efficient gene delivery to dividing and nondividing cells. However, there are some limitations for gene therapy using adenoviral vectors, such as nonspecific transduction of normal cells and liver sequestration and neutralization by antibodies, especially when administered systemically. On the other hand, adenoviral vectors are amenable to strategies for the modification of their biological structures, including genetic manipulation of viral proteins, pseudotyping, and conjugation with polymers or biological membranes. Such modifications provide greater specificity to the target cell and better safety in systemic administration; thus, a ...
Cancer Biology & Therapy, 2005
Successful adenoviral (Ad) vector-mediated strategies for breast cancer gene therapy and virotherapy have heretofore been hindered by low transduction efficiency. This has recently been understood to result from a relative paucity of expression of the primary adenovirus receptor, coxsackie-adenovirus-receptor (CAR), on primary tumor cells. To further investigate this issue, we evaluated the expression of CAR on breast cancer cell lines as well as primary breast cancer cells. With the exception of one patient sample, CAR expression was notably higher in the tumor cells from patients compared to CAR expression in the tumor cell lines. Furthermore, we explored CAR-independent targeting strategies to breast cancer tissue by exploring a panel of infectivity-enhanced Ad vectors, which contain CAR-independent targeting motifs for their utility in breast cancer gene therapy and virotherapy. These targeting motifs included Ad 3 knob (Ad5/3), canine Ad serotype 2 knob (Ad5CAV-2), RGD (Ad5.RGD), polylysine (Ad5.pK7), or both RGD and polylysine (Ad5.RGD.pK7), and were tested using the breast cancer tissue slice model, which is the most stringent substrate system available. Of all the tested tropism modified Ad vectors, Ad5/3 exhibited the highest transductional efficiency in breast cancer. These preclinical results suggest that Ad5/3 is the most useful modification to achieve higher clinical efficacy of breast cancer gene therapy and virotherapy.
Molecular Therapy: Clinical Applications. Intra-Arterial Adenoviruses Administration
Journal of experimental & clinical cancer research: CR
patient with ornithine-cytosine transferase deficiency died as a direct consequence of gene therapy at the University of Pennsylvania. A first generation replication-defective adenovirus administered trough the hepatic artery was used to transduce the DNA sequence of the missing enzyme. The high viral load initiated an out-of-control immune response with a final releasing of cytokines: fever, followed by coagulopathy and signs of liver dysfunction: because the underlying metabolic deficiency, ammonia accumulatd in the blood followed by coma and ultimately multi-organ failure with adult respiratory distress syndrome. This incident raised serious concerns about the safety of arterial adenovirus administration (2). Data on the feasibility of intra-arterial adenovirus was needed. Adenoviruses are doublestranded DNA viruses.At least three regions of the viral genome can accept insertions or substitutions of DNA to generate an helper indipendent virus: a region I E1, a region in E3 and a short region between E4 and the end of the genome. In the first-generation vectors, the E1 region was removed to introduce the therapeutic transgene and to prohibit viral replication. However there was low-level transcription of the remaining 47 Gene therapy involves the introduction of foreign DNA into somatic cells to produce a therapeutic effect. The therapeutic gene is transferred into the tumor cells using a vector. Transfer may either be in vivo in which the DNA and vector are directly introduced into the body, or ex vivo, in which cells are removed from the body, transfected with DNA and then reintroduced into the patients. The mode of gene transfer can be classified into chemical, physical and viral (1). Viruses are the most popular vectors in clinical trials because they invade cells and manipulate the cell's machinery to make viral protein; but the immune response they provoke can rapidly destroy the viral vector or the infected cells, blocking production of the useful protein.Most nonviral vector fly under the radar of immune system, but most of them have not been as efficient as viruses in shuttling genes into cells and the genes that were delivered didn't remain active for long. Intra-arterial administration can have advantages over intravenous, and intralesion routes.
Transduction of murine and human tumors using recombinant adenovirus vectors
Annals of Surgical Oncology, 1997
Background: Most cytokine-based cancer gene therapy clinical trials have used laborintensive, retrovirus-mediated strategies resulting in unpredictable gene expression. Recombinant AdV vectors were evaluated for easier, more reproducible gene transfer into 12 human melanoma, 2 murine fibrosarcomas, and 8 other tumor cell lines. Methods: AdV vectors contained a reporter (Escherichia coli 13-galactosidase or firefly luciferase) or cytokine gene (human interleukin-2 [IL-2] or IL-7). Transduction efficiencies and expression levels were assessed by histochemical staining, flow cytometry, polymerase chain reaction, fluorometry, and enzyme-linked immunosorbent assay. Tumorigenicity was determined by subcutaneous injection of cells into syngeneic mice. Results: All cell lines studied were transduced with AdV. Most cell lines exhibited 100% transduction efficiencies (by flow cytometry) at multiplicities of infection (MOI) el0. Gene expression correlated linearly with MOI, but a cytopathic effect was observed at MOI > 100 with all vectors. Nanogram gene expression levels were routinely achieved. Irradiation (30 Gy) minimally affected expression levels. Tumorigenicity of AdV-IL-2-transduced fibrosarcoma cells in mice was inversely related to IL-2 production. A majority of mice that rejected their tumor challenge were immune to tumor rechallenge. Conclusions: El-deleted AdV vectors may prove useful in generating tumor vaccines ex vivo with high, transient cytokine expression levels.