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© 2003 American Society for Clinical Oncology
Gene Therapy: The Challenges of Translating Laboratory Research Into Clinical Practice
University of Chicago, Chicago, IL AS OF early 2003, The Journal of Gene Medicine has tracked more than 400 completed, ongoing, or pending clinical trials that involve gene therapy for the treatment of cancer (http://www.wiley.co.uk/genetherapy/clinical/). Despite exciting preclinical studies and a few encouraging clinical reports, the majority of these trials have yet to produce clinically useful tumor responses.1 This apparent lack of efficacy is likely the result of inefficient gene transfer and inadequate therapeutic effects from genes that have been used. In this issue of Journal of Clinical Oncology, Pagliaro et al2 present a well-designed study aimed at testing the feasibility, safety, and activity of gene therapy for bladder cancer. Specifically, the investigators attempted adenoviral-mediated gene transfer of wild-type p53 in patients with locally advanced or extensive superficial transitional cell cancer. The replication-defective vector Ad5CMV-p53 was administered via repeated daily intravesical instillations. The study design included escalating doses of Ad5CMV-p53, reaching a maximum of 1012 viral particles/instillation for a total of 8 instillations. The treatment appears to have been well tolerated in all 13 patients. Most of the adverse events were limited to acute bladder inflammation and mild bladder spasms, which were observed predominantly in the higher titer dose groups. No dose-limiting toxicity was encountered, although only one patient received treatment in the highest dose group. The study was terminated earlier than planned because the investigators "did not expect to reach the maximum-tolerated dose." The results of this phase I trial are informative in terms of toxicity. The authors have also presented data regarding viral gene expression, apoptosis, tumor responses, and clinical outcomes. Reverse transcriptase polymerase chain reaction analysis of the RNA from biopsy samples demonstrated detectable p53 transgene expression in only two of 13 patients. This result is disappointing, but many of the biopsies could not be evaluated because insufficient quantities of RNA were extracted from the specimens. If only the tumor samples that did yield adequate amounts of RNA are considered, viral gene expression was detected in two of eight (25%) of all dose groups and in two of four (50%) of those in the 1012 particle/instillation treatment groups, indicating that the transfection of bladder tumor cells might be more efficient at the highest vector concentrations. Immunohistochemical staining did not detect any increase in p53 protein levels compared with pretreatment biopsies. This might be explained by the high background staining known to occur in cells that harbor p53 mutations. Similarly, the negative terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate-biotin nick end-labeling (TUNEL) data presented may have been due to high background levels of apoptosis. Although these factors may have limited the investigators’ ability to detect the action of a p53 transgene, poor viral transfection or poor expression of p53 are more probable explanations for the data. The primary goal of a phase I trial is to assess toxicity, and in this trial the investigators successfully demonstrated the safety and feasibility of intravesical Ad5CMV-p53, at least at the doses tested. However, the tumor response data that were presented are disappointing even at the highest vector concentrations. All patients with positive pretreatment urine cytologies remained positive after treatment. One patient was noted to have mucosal ulceration and tumor shrinkage after treatment, but the authors postulate that this effect was related to a nonspecific inflammatory (or bacille Calmette-Guérin–like) effect. No other clinical responses were observed, raising the question of whether the study of intravesical Ad5CMV-p53 should proceed to a phase II trial. It is possible that higher doses or more installations might lead to more efficient gene transfer and greater clinical responses, but testing this hypothesis would require additional phase I data to ensure the safety of higher vector concentrations. As the authors discuss, methods for enhancing viral transfer across the protective urothelial barrier of glycosaminoglycans may also improve the efficacy of Ad5CMV-p53. If adequate p53 expression could be achieved with Ad5CMV-p53, then one might consider combining this with proven cytotoxic therapies, such as cisplatin or ionizing radiation, which have produced encouraging results in tumor xenograft experiments.3–5 However, at present it seems unlikely that the regimen described by Pagliaro et al2 will achieve the necessary p53 expression in enough tumor cells to translate to a clinically relevant response. If inefficient gene transfer continues to be a persistent problem for gene therapy in bladder cancer, then other potential approaches could be considered. Viral transgenes that encode prodrug-converting enzymes may be attractive because these strategies induce bystander effects on regional tissue. For example, virus-directed expression of herpes simplex virus–thymidine kinase can be combined with systemic administration of the prodrug ganciclovir. The resulting metabolite of ganciclovir interferes with DNA replication, leading to chain terminations and single-strand DNA breaks. By this technique, infection of only 10% of the cells within a tumor can lead to significant tumor regression.6 Other therapeutic genes that exert a regional effect on tumor tissue could also be considered. For example, viral-directed expression of tumor necrosis factor alpha within tumors appears to disrupt tumor vasculature and target tumor cells.7 Targeted gene therapies using tumor-selective viruses may also improve the efficiency of gene transfer in urothelial cancers. For example, recent research has focused on the fibers and globular knobs extending from adenoviral capsids. These knobs promote cellular attachment and infection by interacting with coxsackie-adenovirus receptors (CAR) on cell surfaces. CAR expression is known to vary widely in human bladder cancer cell lines, and CAR expression correlates closely with cellular susceptibility to adenoviral infection.8 A recent pathologic analysis of bladder cancer specimens showed detectable CAR expression to be particularly low in invasive high-grade tumors.9 To avoid the problem of low CAR expression and to improve the selectivity of gene therapy vectors, several groups are working to replace the adenoviral knobs with novel ligands that are more likely to interact with tumor cells. For example, investigators at the University of Alabama (Birmingham, AL) genetically modified an adenovirus vector to express an arginine-glycine-aspartate–containing peptide within the fiber knob domain. This modification resulted in CAR-independent gene transfer, particularly to cells that highly express arginine-glycine-aspartate–binding integrins.10,11 Perhaps this type of strategy could be applied to gene therapy for bladder cancer to exploit a normal interaction between an abundantly expressed membrane protein and a known ligand. Finally, the patient inclusion criteria of this study are worthy of discussion. Eligibility for enrollment required that patients must have experienced primary therapy failure (bacille Calmette-Guérin–based or cisplatin-based treatment) and were not considered candidates for cystectomy. It was not specifically noted whether patients could have been treated by combined-modality therapy with concurrent radiation and chemotherapy. This is important because these regimens are associated with 10-year disease-specific survival rates of almost 60% in select patients with locally advanced bladder cancer.12 Despite the selection criteria used in these trials, combined radiation and chemotherapy can potentially cure patients who do not qualify for cystectomy. This treatment option should be considered before patients are offered treatment on phase I trials. The administration of an adenoviral vector by intravesical instillation is a novel application of gene therapy. It avoids many of the clinical hurdles associated with systemic viral delivery or direct injection of primary tumors located in inaccessible sites. However, additional research is needed to maximize this evolving technology and to optimize the chances of developing effective new therapies. NOTES
* R.R.W. is a consultant for and has an equity interest in GenVec, Inc. R.R.W. is a consultant for MediGene, Inc. REFERENCES 1. Kufe DW, Advanti S, Weichselbaum RR: Principles of gene therapy, in Holland JF, Frei E, Bast RC, et al (eds): Cancer Medicine (ed 5). London, BC Decker Inc, 2000, pp 876–889
2. Pagliaro LC, Keyhani A, Williams D, et al: Repeated intravesical instillations of an adenoviral vector in patients with locally advanced bladder cancer: A phase I study of p53 gene therapy. J Clin Oncol 21:2247–2253, 2003
3. Fujiwara T, Grimm EA, Mukhopadhyay T, et al: Induction of chemosensitivity in human lung cancer cells in vivo by adenovirus-mediated transfer of the wild-type p53 gene. Cancer Res 54:2287–2291, 1994
4. Gallardo D, Drazan KE, McBride WH: Adenovirus-based transfer of wild-type p53 gene increases ovarian tumor radiosensitivity. Cancer Res 56:4891–4893, 1996 5. Spitz FR, Nguyen D, Skibber JM, et al: Adenoviral-mediated wild-type p53 gene expression sensitizes colorectal cancer cells to ionizing radiation. Clin Cancer Res 2:1665–1671, 1996[Abstract]
6. Freeman SM, Abboud CN, Whartenby KA, et al: The "bystander effect": Tumor regression when a fraction of the tumor mass is genetically modified. Cancer Res 53:5274–5283, 1993 7. Weichselbaum RR, Kufe DW, Hellman S, et al: Radiation-induced tumour necrosis factor-alpha expression: Clinical application of transcriptional and physical targeting of gene therapy. Lancet Oncol 3:665–671, 2002[CrossRef][Medline]
8. Li Y, Pong RC, Bergelson JM, et al: Loss of adenoviral receptor expression in human bladder cancer cells: A potential impact on the efficacy of gene therapy. Cancer Res 59:325–330, 1999 9. Sachs MD, Rauen KA, Ramamurthy M, et al: Integrin alpha(v) and coxsackie adenovirus receptor expression in clinical bladder cancer. Urology 60:531–536, 2002[CrossRef][Medline]
10. Dmitriev I, Krasnykh V, Miller CR, et al: An adenovirus vector with genetically modified fibers demonstrates expanded tropism via utilization of a coxsackievirus and adenovirus receptor-independent cell entry mechanism. J Virol 72:9706–9713, 1998
11. Kasono K, Blackwell JL, Douglas JT, et al: Selective gene delivery to head and neck cancer cells via an integrin targeted adenoviral vector. Clin Cancer Res 5:2571–2579, 1999 12. Shipley WU, Kaufman DS, Zehr E, et al: Selective bladder preservation by combined modality protocol treatment: Long-term outcomes of 190 patients with invasive bladder cancer. Urology 60:62–68, 2002[Medline]
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Copyright © 2003 by the American Society of Clinical Oncology, Online ISSN: 1527-7755. Print ISSN: 0732-183X
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