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Kumar A, Das SK, Emdad L, Fisher PB. Applications of tissue-specific and cancer-selective gene promoters for cancer diagnosis and therapy. Adv Cancer Res 2023; 160:253-315. [PMID: 37704290 DOI: 10.1016/bs.acr.2023.03.005] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 09/15/2023]
Abstract
Current treatment of solid tumors with standard of care chemotherapies, radiation therapy and/or immunotherapies are often limited by severe adverse toxic effects, resulting in a narrow therapeutic index. Cancer gene therapy represents a targeted approach that in principle could significantly reduce undesirable side effects in normal tissues while significantly inhibiting tumor growth and progression. To be effective, this strategy requires a clear understanding of the molecular biology of cancer development and evolution and developing biological vectors that can serve as vehicles to target cancer cells. The advent and fine tuning of omics technologies that permit the collective and spatial recognition of genes (genomics), mRNAs (transcriptomics), proteins (proteomics), metabolites (metabolomics), epiomics (epigenomics, epitranscriptomics, and epiproteomics), and their interactomics in defined complex biological samples provide a roadmap for identifying crucial targets of relevance to the cancer paradigm. Combining these strategies with identified genetic elements that control target gene expression uncovers significant opportunities for developing guided gene-based therapeutics for cancer. The purpose of this review is to overview the current state and potential limitations in developing gene promoter-directed targeted expression of key genes and highlights their potential applications in cancer gene therapy.
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Affiliation(s)
- Amit Kumar
- Department of Human and Molecular Genetics, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Institute of Molecular Medicine, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States
| | - Swadesh K Das
- Department of Human and Molecular Genetics, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Institute of Molecular Medicine, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Massey Comprehensive Cancer Center, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States
| | - Luni Emdad
- Department of Human and Molecular Genetics, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Institute of Molecular Medicine, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Massey Comprehensive Cancer Center, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States
| | - Paul B Fisher
- Department of Human and Molecular Genetics, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Institute of Molecular Medicine, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States; VCU Massey Comprehensive Cancer Center, Virginia Commonwealth University, School of Medicine, Richmond, VA, United States.
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Wang L, Dong J, Wei M, Wen W, Gao J, Zhang Z, Qin W. Selective and augmented β-glucuronidase expression combined with DOX-GA3 application elicits the potent suppression of prostate cancer. Oncol Rep 2015; 35:1417-24. [PMID: 26648021 DOI: 10.3892/or.2015.4454] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2015] [Accepted: 10/19/2015] [Indexed: 11/05/2022] Open
Abstract
The present study was carried out to evaluate the specific and amplified β-glucuronidase (βG) expression in prostate cancer cells by using a prostate‑specific antigen (PSA) promoter-controlled bicistronic adenovirus and to evaluate the specific killing of prostate cancer cells after the application of the prodrug DOX‑GA3. Bicistronic adenoviral expression vectors were constructed, and the effectiveness of specific and amplified expression was evaluated using luciferase and EGFP as reporter genes. βG expression was detected in LNCaP cells after they were infected with the βG‑expressing PSA promoter-controlled bicistronic adenovirus. MTT assays were conducted to evaluate the cytoxicity on the infected cells after the application of the prodrug DOX‑GA3. Tumor growth inhibition was also evaluated in nude mice after treatment with the βG‑expressing adenovirus and DOX‑GA3. Selective and amplified expression was observed in the PSA-producing LNCaP cells, but not in the PSA‑non‑producing DU145 cells. Potent cytotoxity and a strong bystander effect were observed in the LNCaP cells after infection with the βG‑expressing adenovirus and the application of DOX‑GA3. Intravenous injection of a GAL4 regulated bicistronic adenovirus vector constructed to express βG under the control of the PSA promoter (Ad/PSAP‑GV16‑βG) and the application of DOX‑GA3 strongly inhibited tumor growth and prolonged the survival time of tumor‑bearing nude mice. Selective and amplified βG expression together with the prodrug DOX‑GA3 had an increased antitumor effect, showing great potential for prostate cancer therapy.
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Affiliation(s)
- Longxin Wang
- Department of Urology, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China
| | - Jie Dong
- Department of Urology, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China
| | - Ming Wei
- Department of Urology, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi 710032, P.R. China
| | - Weihong Wen
- State Key Laboratory of Cancer Biology, Department of Immunology, Fourth Military Medical University, Xi'an, Shaanxi 710032, P.R. China
| | - Jianping Gao
- Department of Urology, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China
| | - Zhengyu Zhang
- Department of Urology, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China
| | - Weijun Qin
- Department of Urology, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi 710032, P.R. China
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Zhou PH, Zheng JB, Wei GB, Wang XL, Wang W, Chen NZ, Yu JH, Yao JF, Wang H, Lu SY, Sun XJ. Lentivirus-mediated RASSF1A expression suppresses aggressive phenotypes of gastric cancer cells in vitro and in vivo. Gene Ther 2015; 22:793-801. [PMID: 26005859 PMCID: PMC4598615 DOI: 10.1038/gt.2015.49] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2014] [Revised: 03/18/2015] [Accepted: 05/12/2015] [Indexed: 12/15/2022]
Abstract
Loss of Ras association domain family protein 1 isoform A (RASSF1A) expression is associated with the development of a variety of human cancers and the expression of carcinoembryonic antigen (CEA) frequently occurs in gastric cancer. This study investigated the effects of RASSF1A expression restoration using a hypoxia-inducible CEA promoter-driven vector on xenograft tumor growth in nude mice and on the in-vitro regulation of gastric cancer cell viability, cell cycle distribution, apoptosis, colony formation and invasion capacity. The data showed that the level of CEA mRNA and protein was much higher in gastric cancer SGC7901 cells than in a second gastric cancer cell line, MKN28, or in the MCF-10A normal epithelial breast cell line. RASSF1A expression was restored in SGC7901 cells compared with the negative control virus-infected SGC7910 cells. RASSF1A expression restoration significantly inhibited gastric cancer cell viability, colony formation and invasion capacity, but induced cell cycle arrest and apoptosis in vitro, especially under hypoxic culture conditions. At the gene level, restoration of RASSF1A expression under hypoxic culture conditions significantly suppressed matrix metalloproteinase-2 expression and prevented cyclinD1 expression. A nude mouse xenograft assay showed that the restoration of RASSF1A expression reduced gastric cancer xenograft formation and growth. In conclusion, the restoration of RASSF1A expression using a hypoxia-inducible and CEA promoter-driven vector suppressed aggressive phenotypes of gastric cancer cells in vitro and in vivo. These results suggest that LV-5HRE-CEAp-RASSF1A gene therapy may be a promising novel approach to treat advanced gastric cancer.
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Affiliation(s)
- P-H Zhou
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - J-B Zheng
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - G-B Wei
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - X-L Wang
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - W Wang
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - N-Z Chen
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - J-H Yu
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - J-F Yao
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - H Wang
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - S-Y Lu
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
| | - X-J Sun
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, China
- Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, 277 West Yanta Road, Xi'an 710061, Shanxi, China.
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Pouliot F, Sato M, Jiang ZK, Huyn S, Karanikolas BD, Wu L. A molecular imaging system based on both transcriptional and genomic amplification to detect prostate cancer cells in vivo. Mol Ther 2012; 21:554-60. [PMID: 23247102 DOI: 10.1038/mt.2012.259] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022] Open
Abstract
An imaging modality that can accurately discern prostate cancer (PCa) foci would be useful to detect PCa early or guide treatment. We have engineered numerous adenoviral vectors (Ads) to carry out reporter gene-based imaging using specific promoters to express a potent transcriptional activator, which in turn activates the reporter gene in PCa. This two-step transcriptional amplification (TSTA) method can boost promoters' activity, while maintaining cell specificity. Here, we examined a dual TSTA (DTSTA) approach, which utilizes TSTA not only to express the imaging reporter, but also to direct viral genome replication of a conditionally replicating Ad (CRAd) to further augment the expression levels of the reporter gene by genomic amplification supported in trans by coadministered CRAd. In vitro studies showed up to 50-fold increase of the reporter genome by DTSTA. Compared with TSTA reporter alone, DTSTA application exhibited a 25-fold increase in imaging signal in PCa xenografts. DTSTA approach is also beneficial for a combination of two TSTA Ads with distinct promoters, although amplification is observed only when TSTA-CRAd can replicate. Consequently, the DTSTA approach is a hybrid method of transcriptional and genomic augmentation that can provide higher level reporter gene expression potentially with a lower dose of viral administration.
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Affiliation(s)
- Frédéric Pouliot
- David Geffen School of Medicine, University of California, Los Angeles, CA, USA
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Targeting different types of human meningioma and glioma cells using a novel adenoviral vector expressing GFP-TRAIL fusion protein from hTERT promoter. Cancer Cell Int 2011; 11:35. [PMID: 22035360 PMCID: PMC3283457 DOI: 10.1186/1475-2867-11-35] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2011] [Accepted: 10/28/2011] [Indexed: 01/23/2023] Open
Abstract
Objective The objective of this study was to evaluate the anti-tumor effects of Ad/gTRAIL (an adenoviral vector in which expression of GFP and TRAIL is driven by a human telomerase reverse transcriptase promoter, hTERT) on malignant meningiomas and gliomas. Background Gliomas and meningiomas are the two most common types of human brain tumors. Currently there is no effective cure for recurrent malignant meningiomas or for gliomas. Ad/gTRAIL has been shown to be effective in killing selected lung, colon and breast cancer cells, but there have been no studies reporting its antitumor effects on malignant meningiomas. Therefore, we tested the antitumor effect of Ad/gTRAIL for the first time in human malignant meningioma and glioma cell lines, and in intracranial M6 and U87 xenografts. Methods Materials and Methods: Human malignant meningioma and glioma cells were infected with adenoviruses, Ad/gTRAIL and Ad/CMV-GFP. Cell viability was determined by proliferation assay. FACS analysis and quantification of TRAIL were used to measure apoptosis in these cells. We injected Ad/gTRAIL viruses in intracranial M6 and U87 xenografts, and measured the brain tumor volume, quantified apoptosis by TUNEL assay in the brain tumor tissue. Results Our studies demonstrate that in vitro/in vivo treatment with Ad/gTRAIL virus resulted in significant increase of TRAIL activity, and elicited a greater tumor cell apoptosis in malignant brain tumor cells as compared to treatment with the control, Ad/CMV-GFP virus without TRAIL activity. Conclusions We showed for the first time that adenovirus Ad/gTRAIL had significant antitumor effects against high grade malignant meningiomas as well as gliomas. Although more work needs to be done, our data suggests that Ad/gTRAIL has the potential to be useful as a tool against malignant brain tumors.
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Arendt ML, Nasir L, Morgan IM. A novel two-step transcriptional activation system for gene therapy directed toward epithelial cells. Mol Cancer Ther 2010; 8:3244-54. [PMID: 19952120 DOI: 10.1158/1535-7163.mct-09-0543] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
The two-step transcriptional activation (TSTA) mechanism in gene therapy amplifies cell type-specific promoter activity, allowing for increased levels of gene expression in target tissues. In this system, the specific promoter drives expression of a strong transcriptional activator that binds to DNA target sequences located upstream from a second promoter controlling the expression of the therapeutic gene. The majority of previous studies have exploited a fusion between the DNA binding domain of the yeast transcriptional activator Gal4 fused to the VP16 activation domain of herpes simplex virus 1 as the transcriptional activator. In this report, an alternative to this system is described based on a fusion protein containing the DNA binding domain of the bovine papillomavirus 1 transcriptional activator E2 fused to VP16 that induces target gene expression following binding to a minimal bovine papillomavirus 4 promoter containing upstream E2 binding sites and only 3 bp of promoter sequence upstream from the TATA box. VP16-E2 is superior to Gal4-VP16 as the transcriptional activator in a TSTA system driven by either of the two potentially cancer-specific promoters telomerase RNA and telomerase reverse transcriptase in several cell lines. Results also suggest that this new system has an advantage in epithelial cells and is therefore ideal for potential targeting of carcinomas. By incorporating the TRAIL gene as a transgene in the VP16-E2 TSTA system, selective killing of telomerase-positive cells occurs. We propose that our new system should be considered in future TSTA, particularly when targeting epithelial-derived cells.
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Affiliation(s)
- Maja L Arendt
- Institute of Comparative Medicine, Division of Pathological Sciences, University of Glasgow Faculty of Veterinary Medicine, Bearsden Road, Glasgow G61 1QH, United Kingdom
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Boulaire J, Balani P, Wang S. Transcriptional targeting to brain cells: Engineering cell type-specific promoter containing cassettes for enhanced transgene expression. Adv Drug Deliv Rev 2009; 61:589-602. [PMID: 19394380 DOI: 10.1016/j.addr.2009.02.007] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2008] [Accepted: 02/05/2009] [Indexed: 12/16/2022]
Abstract
Transcriptional targeting using a mammalian cellular promoter to restrict transgene expression to target cells is often desirable for gene therapy. This strategy is, however, hindered by relatively weak activity of some cellular promoters, which may lead to low levels of gene expression, thus declining therapeutic efficacy. Here we outline the advances accomplished in the area of transcriptional targeting to brain cells, with a particular focus on engineering gene cassettes to augment cell type-specific expression. Among the effective approaches that improve gene expression while retaining promoter specificity are promoter engineering to change authentic sequences of a cellular promoter and the combined use of a native cellular promoter and other cis-acting elements. Success in achieving high level and sustained transgene expression only in the cell types of interest would be of importance in allowing gene therapy to have its impact on patient treatment.
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E1A, E1B double-restricted replicative adenovirus at low dose greatly augments tumor-specific suicide gene therapy for gallbladder cancer. Cancer Gene Ther 2008; 16:126-36. [PMID: 18818710 DOI: 10.1038/cgt.2008.67] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Abstract
Combination therapy with replicative oncolytic viruses is a recent topic in innovative cancer therapy, but few studies have examined the efficacy of oncolytic adenovirus plus replication-deficient adenovirus carrying a suicide gene. We aim to evaluate whether an E1A, E1B double-restricted oncolytic adenovirus, AxdAdB-3, can improve the efficacy for gallbladder cancers (GBCs) of the replication-deficient adenovirus-based herpes simplex virus thymidine kinase (HSVtk)/ganciclovir (GCV) therapy directed by the carcinoembryonic antigen (CEA) promoter. Cytopathic effects of AxdAdB-3 plus AxCEAprTK (an adenovirus expressing HSVtk directed by CEA promoter) or AxCAHSVtk (an adenovirus expressing HSVtk directed by a nonspecific CAG promoter) with GCV administration were examined in several GBC lines and normal cells. Efficacy in vivo was tested in severe combined immunodeficiency disease mice with GBC xenografts. Addition of AxdAdB-3 (1 multiplicity of infection, MOI) significantly enhanced the cytopathic effects of AxCEAprTK (10 MOI)/GCV on GBC cells. The augmented effect was attributable to the replication of the AxCEAprTK and also to the enhanced CEA promoter activity, which was presumably transactivated by E1A. In normal cells, AxdAdB-3 (20 MOI) plus AxCEAprTK (200 MOI)/GCV was not cytopathic, whereas AxdAdB-3 (1 MOI) plus AxCAHSVtk (10 MOI)/GCV was significantly toxic. Low-dose AxdAdB-3 (2 x 10(7) PFU, plaque-forming unit) plus AxCEAprTK (2 x 10(8) PFU)/GCV significantly suppressed the growth of GBC xenografts as compared with either AxdAdB-3 (2 x 10(7) PFU)/GCV or AxCEAprTK (2 x 10(9) PFU)/GCV alone. E1A, E1B double-restricted replicating adenovirus at low dose significantly augmented the efficacy of CEA promoter-directed HSVtk/GCV therapy without obvious toxicity to normal cells, suggesting a potential use of this combination for treating GBC and other CEA-producing malignancies.
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Guo ZS, Li Q, Bartlett DL, Yang JY, Fang B. Gene transfer: the challenge of regulated gene expression. Trends Mol Med 2008; 14:410-8. [PMID: 18692441 DOI: 10.1016/j.molmed.2008.07.003] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2008] [Revised: 07/04/2008] [Accepted: 07/04/2008] [Indexed: 01/04/2023]
Abstract
Gene therapy is expected to have a major impact on human healthcare in the future. However, precise regulation of therapeutic gene expression in vivo is still a challenge. Natural and synthetic enhancer-promoters (EPs) can be utilized to drive gene transcription in a temporal, spatial or environmental signal-inducible manner in response to heat shock, hypoxia, radiation, chemotherapy, epigenetic agents or viral infection. To allow tightly regulated expression, a regulatable gene-expression system can also be implemented. Most of these systems are based on small molecule (drug)-responsive artificial transactivators. In this review, we aim to provide a brief overview of the classes of EPs and regulatable systems, along with lessons learned from these studies. We highlight the potential applications in gene transfer, gene therapy for cancer and genetic disease and the future challenges for clinical applications.
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Affiliation(s)
- Z Sheng Guo
- Division of Surgical Oncology, University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
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Nakamura S, Watanabe S, Ohtsuka M, Maehara T, Ishihara M, Yokomine T, Sato M. Cre-loxP system as a versatile tool for conferring increased levels of tissue-specific gene expression from a weak promoter. Mol Reprod Dev 2008; 75:1085-93. [PMID: 18163444 DOI: 10.1002/mrd.20847] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
Abstract
Attempts to image reporter gene expression driven by weak promoters are often hampered by the poor transcriptional activity of such promoters. Most tissue-specific promoters are weak compared with stronger but constitutively expressing viral promoters. In this study, we validated methods of enhancing the transcriptional activity of weak promoters using a Cre-loxP system in vitro and in vivo. We constructed a tester vector, pCTL, which carries a strong systemic cytomegalovirus enhancer/chicken beta-actin promoter (CAG), loxP-flanked CAT, and firefly luciferase (luc) cDNAs. Herpes simplex virus-thymidine kinase (HSV-tk) promoter was used as a weak and systemic promoter and ligated to Cre for construction of pTC. Luc activity was higher (about 10-fold enhancement) in co-transfected (with pCTL and pTC) than in singly (with HSV-tk promoter-driven luc expression vector pTL) transfected NIH3T3 cells. In vivo electroporation-mediated gene delivery of both pCTL and pTC into murine oviductal epithelium yielded results (about 16-fold enhancement) similar to those obtained with in vitro-transfected NIH3T3 cells. To evaluate tissue-specific enhancement of gene expression, podocyte (glomerular visceral epithelial cell)-specific nephrin promoter was ligated to the Cre gene or luc cDNA to create pNC and pNL, respectively. We achieved 2.4-fold improvement of luc gene expression in the mouse kidney in vivo when pCTL and pNC were co-transfected via the tail vein via the lipoplex method. The combination of a weak tissue-specific promoter with the Cre-loxP system could thus be used to enhance the strength of tissue-specific promoters in vitro and in vivo.
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Affiliation(s)
- Shingo Nakamura
- Department of Surgery, School of Medicine, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama, Japan
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Guo ZS, Thorne SH, Bartlett DL. Oncolytic virotherapy: molecular targets in tumor-selective replication and carrier cell-mediated delivery of oncolytic viruses. Biochim Biophys Acta Rev Cancer 2008; 1785:217-31. [PMID: 18328829 DOI: 10.1016/j.bbcan.2008.02.001] [Citation(s) in RCA: 90] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2007] [Revised: 02/01/2008] [Accepted: 02/04/2008] [Indexed: 01/13/2023]
Abstract
Tremendous advances have been made in developing oncolytic viruses (OVs) in the last few years. By taking advantage of current knowledge in cancer biology and virology, specific OVs have been genetically engineered to target specific molecules or signal transduction pathways in cancer cells in order to achieve efficient and selective replication. The viral infection and amplification eventually induce cancer cells into cell death pathways and elicit host antitumor immune responses to further help eliminate cancer cells. Specifically targeted molecules or signaling pathways (such as RB/E2F/p16, p53, IFN, PKR, EGFR, Ras, Wnt, anti-apoptosis or hypoxia) in cancer cells or tumor microenvironment have been studied and dissected with a variety of OVs such as adenovirus, herpes simplex virus, poxvirus, vesicular stomatitis virus, measles virus, Newcastle disease virus, influenza virus and reovirus, setting the molecular basis for further improvements in the near future. Another exciting new area of research has been the harnessing of naturally tumor-homing cells as carrier cells (or cellular vehicles) to deliver OVs to tumors. The trafficking of these tumor-homing cells (stem cells, immune cells and cancer cells), which support proliferation of the viruses, is mediated by specific chemokines and cell adhesion molecules and we are just beginning to understand the roles of these molecules. Finally, we will highlight some avenues deserving further study in order to achieve the ultimate goals of utilizing various OVs for effective cancer treatment.
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Affiliation(s)
- Z Sheng Guo
- University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
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Kang JH, Chung JK, Lee YJ, Kim KI, Jeong JM, Lee DS, Lee MC. Evaluation of transcriptional activity of the oestrogen receptor with sodium iodide symporter as an imaging reporter gene. Nucl Med Commun 2006; 27:773-7. [PMID: 16969258 DOI: 10.1097/01.mnm.0000230075.46694.8e] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
BACKGROUND Oestrogen receptors are ligand-dependent transcription factors whose activity is modulated either by oestrogens or by an alternative signalling pathway. Oestrogen receptors interact via a specific DNA-binding domain, the oestrogen responsive element (ERE), in the promoter region of sensitive genes. This binding leads to an initiation of gene expression and hormonal effects. OBJECTIVE To determine the transcriptional activity of the oestrogen receptor, we developed a molecular imaging system using sodium iodide symporter (NIS) as a reporter gene. METHODS The NIS reporter gene was placed under the control of an artificial ERE derived from pERE-TA-SEAP and named as pERE-NIS. pERE-NIS was transferred to MCF-7, human breast cancer cells, which highly expressed oestrogen receptor-alpha with lipofectamine. Stably expressing cells were generated by selection with G418 for 14 days. After treatment of 17beta-oestradiol and tamoxifen with serial doses, the (125)I uptake was measured for the determination of NIS expression. The inhibition of NIS activity was performed with 50 micromol x l(-1) potassium perchlorate. RESULTS The MCF7/pERE-NIS treated with 17beta-oestradiol accumulated (125)I up to 70-80% higher than did non-treated cells. NIS expression was increased according to increasing doses of 17beta-oestradiol. MCF7/pERE-NIS treated with tamoxifen also accumulated (125)I up to 50% higher than did non-treated cells. Potassium perchlorate completely inhibited (125)I uptake. When MDA-MB231 cells, the oestrogen receptor-negative breast cancer cells, were transfected with pERE-NIS, (125)I uptake of MDA-MB-231/pERE-NIS did not increase. CONCLUSION This pERE-NIS reporter system is sufficiently sensitive for monitoring transcriptional activity of the oestrogen receptor. Therefore, cis-enhancer reporter systems with ERE will be applicable to the development of a novel selective oestrogen receptor modulator with low toxicity and high efficacy.
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Affiliation(s)
- Joo Hyun Kang
- Laboratory of Nuclear Medicine, Korea Institute of Radiological and Medical Science, Korea
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13
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Liu BH, Yang Y, Paton JFR, Li F, Boulaire J, Kasparov S, Wang S. GAL4-NF-kappaB fusion protein augments transgene expression from neuronal promoters in the rat brain. Mol Ther 2006; 14:872-82. [PMID: 16904943 DOI: 10.1016/j.ymthe.2006.05.020] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/25/2005] [Revised: 04/10/2006] [Accepted: 05/21/2006] [Indexed: 11/27/2022] Open
Abstract
Targeted gene expression mediated by a mammalian cellular promoter is desirable for gene therapy in the brain, where there are a variety of different neuronal phenotypes, several types of supportive cells, and blood vessels. However, this approach can be hampered by weak activity of some cellular promoters. In view of the potency of the transcription factor NF-kappaB in regulating neuronal gene expression, we have assessed whether it can be used to enhance the strength of neuron-specific promoters. Our approach was to use a neuronal promoter to drive expression of a chimeric transactivator, which consisted of a part of the transcriptional activation domain of the NF-kappaB p65 protein fused to the DNA-binding domain of GAL4 protein from yeast. The second copy of the neuronal promoter was modified by introducing the unique GAL4 binding sequences at its 5' end and used to drive the expression of a transgene. Binding of the chimeric transcriptional activator upstream of the second promoter was expected to potentiate its transcriptional activity. In this study, the approach was applied to the platelet-derived growth factor beta chain and synapsin-1 neuron-specific promoters and tested in vitro and in vivo using plasmid, lentiviral, and baculoviral vectors. We observed up to a 100-fold improvement in reporter gene expression in cultured neurons and 20-fold improvement in the rat brain in vivo. Moreover, the cell-type specificity of the two tested promoters was well preserved and restricted to neurons. Finally, the expression driven by the new lentiviral vectors with the p65-potentiated synapsin-1 promoter showed no signs of decline or cell damage 4 weeks after injection. This approach should be suitable for constructing powerful and stable gene expression systems based on weak cell-specific promoters in neuronal phenotypes.
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Affiliation(s)
- B H Liu
- Institute of Bioengineering and Nanotechnology, 31 Biopolis Way, 138669, Singapore
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Hattori Y, Maitani Y. Two-step transcriptional amplification-lipid-based nanoparticles using PSMA or midkine promoter for suicide gene therapy in prostate cancer. Cancer Sci 2006; 97:787-98. [PMID: 16800821 PMCID: PMC11159223 DOI: 10.1111/j.1349-7006.2006.00243.x] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022] Open
Abstract
A two-step transcriptional amplification system (TSTA) was used to enhance the efficacy of suicide gene therapy for treatment of prostate cancer. We designed a TSTA system and constructed two types of plasmid: one containing GAL4-VP16 fusion protein under the control of a tumor-specific promoter, the other containing luciferase or herpes simplex virus thymidine kinase (HSV-tk) under the control of a synthetic promoter. The TSTA systems using nanoparticles based on lipids were evaluated by measuring the amount of induced luciferase activity as a function of prostate-specific membrane antigen (PSMA) and midkine (Mk) promoters, specific for LNCaP and PC-3 prostate cancer cells, respectively. In LNCaP cells that were PSMA-positive, the TSTA system featuring the PSMA enhancer and promoter exhibited activity that was 640-fold greater than a system consisting of one-step transcription with the PSMA promoter. In contrast, this difference in activity did not occur in PSMA-negative PC-3 cells. In Mk-positive PC-3 cells, the TSTA system with the Mk promoter exhibited a five-fold increase in activity over one-step transcription, but such activity was not induced in Mk-negative LNCaP cells. When using HSV-tk for suicide gene therapy, TSTA systems featuring the PSMA or Mk promoter inhibited in vitro cell growth in the presence of ganciclovir. Furthermore, the TSTA system featuring the Mk promoter suppressed in vivo growth of PC-3 tumor xenografts to a greater extent than one-step transcription. These findings show that TSTA systems can enhance PSMA and Mk promoter activities and selectively inhibit PC-3 cell growth in tumors. This suggests that TSTA systems featuring tumor-specific promoters are suitable for cancer treatment by gene therapy.
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Affiliation(s)
- Yoshiyuki Hattori
- Institute of Medicinal Chemistry, Hoshi University, Ebara 2-4-41, Shinagawa-ku, Tokyo 142-8501, Japan
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Chang JY, Zhang X, Komaki R, Cheung R, Fang B. Tumor-specific apoptotic gene targeting overcomes radiation resistance in esophageal adenocarcinoma. Int J Radiat Oncol Biol Phys 2006; 64:1482-94. [PMID: 16580499 PMCID: PMC1458528 DOI: 10.1016/j.ijrobp.2005.11.045] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2005] [Revised: 11/17/2005] [Accepted: 11/18/2005] [Indexed: 10/24/2022]
Abstract
PURPOSE To overcome radiation resistance in esophageal adenocarcinoma by tumor-specific apoptotic gene targeting using tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). METHODS AND MATERIALS Adenoviral vector Ad/TRAIL-F/RGD with a tumor-specific human telomerase reverse transcription promoter was used to transfer TRAIL gene to human esophageal adenocarcinoma and normal human lung fibroblastic cells (NHLF). Activation of apoptosis was analyzed by Western blot, fluorescent activated cell sorting, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate labeling (TUNEL) assay. A human esophageal adenocarcinoma mouse model was treated with intratumoral injections of Ad/TRAIL-F/RGD plus local radiotherapy. RESULTS The combination of Ad/TRAIL-F/RGD and radiotherapy increased the cell-killing effect in all esophageal adenocarcinoma cell lines but not in NHLF cells. This combination also significantly reduced clonogenic formation (p < 0.05) and increased sub-G1 deoxyribonucleic acid accumulation in cancer cells (p < 0.05). Activation of apoptosis by Ad/TRAIL-F/RGD plus radiotherapy was demonstrated by activation of caspase-9, caspase-8, and caspase-3 and cleaved poly (adenosine diphosphate-ribose) polymerase in vitro and TUNEL assay in vivo. Combined Ad/TRAIL-F/RGD and radiotherapy dramatically inhibited tumor growth and prolonged mean survival in the esophageal adenocarcinoma model to 31.6 days from 16.7 days for radiotherapy alone and 21.5 days for Ad/TRAIL-F/RGD alone (p < 0.05). CONCLUSIONS The combination of tumor-specific TRAIL gene targeting and radiotherapy enhances the effect of suppressing esophageal adenocarcinoma growth and prolonging survival.
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Affiliation(s)
- Joe Y Chang
- Department of Radiation Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
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16
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Abstract
Hepatitis C is a global problem with significantly associated morbidity and mortality. Although some recent therapeutic advances have shown rates of sustained virologic remission of 50% or higher, combination therapy with interferon and ribavirin is often not well tolerated and is giving rise to a growing number of nonresponders. As a result, a large number of experimental drugs for the treatment of chronic hepatitis C are in development. As the clinical trial reports are made available, physicians need to become familiar with issues related to the design of these studies and to develop strategies to interpret the evidence they yield. The articles in this supplement describe the issues in clinical trial design and the evaluation of evidence from clinical trials in patients with chronic hepatitis C.
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Affiliation(s)
- B R Bacon
- Division of Gastroenterology and Hepatology, Saint Louis University Liver Center, Saint Louis University School of Medicine, St Louis, MO 63110, USA.
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17
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Szymanski P, Anwer K, Sullivan SM. Development and characterization of a synthetic promoter for selective expression in proliferating endothelial cells. J Gene Med 2006; 8:514-23. [PMID: 16475217 DOI: 10.1002/jgm.875] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
BACKGROUND Systemic administration of non-viral gene therapy provides better access to tumors than local administration. Development of a promoter that restricts expression of cytotoxic proteins to the tumor vasculature will increase the safety of the system by minimizing expression in the non-dividing endothelial cells of the vasculature of non-target tissues. METHODS Cell cycle promoters were tested for selective expression in dividing cells vs. non-dividing cells in vitro and promoter strength was compared to the cytomegalovirus (CMV) promoter. Successful promoter candidates were tested in vivo using two proliferating endothelium mouse models. Ovarectomized mice were injected with estradiol prior to lipoplex administration and expression levels were measured in the lungs and uterus 4 days after administration. The second model was a subcutaneous tumor model and expression levels were measured in the lungs and tumors. For both animal models, expression levels from the proliferating endothelium promoter were compared to that obtained from a CMV promoter. RESULTS The results showed that the Cdc6 promoter yielded higher expression in proliferating vs. non-proliferating cells. Secondly, promoter strength could be selectively increased in endothelial cells by the addition of a multimerized endothelin enhancer (ET) to the Cdc6 promoter. Thirdly, comparison of expression levels in the lungs vs. uterus in the ovarectomized mouse model and lungs vs. tumor in the mouse tumor model showed expression was much higher in the uterus and the tumor than in the lungs for the ET/Cdc6 promoter, and expression levels were comparable to that of the CMV promoter in the hypervascularized tissues. CONCLUSIONS These results demonstrate that the combination of the endothelin enhancer with the Cdc6 promoter yields selective expression in proliferating endothelium and can be used to express cytotoxic proteins to treat vascularized tumors.
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Zhang X, Cheung RM, Komaki R, Fang B, Chang JY. Radiotherapy sensitization by tumor-specific TRAIL gene targeting improves survival of mice bearing human non-small cell lung cancer. Clin Cancer Res 2005; 11:6657-68. [PMID: 16166445 PMCID: PMC1351100 DOI: 10.1158/1078-0432.ccr-04-2699] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
PURPOSE To sensitize non-small cell lung cancer (NSCLC) to radiotherapy by tumor-specific delivery of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene. EXPERIMENTAL DESIGN The TRAIL was delivered to human NSCLC cell lines and normal human bronchial epithelial cells by the replication-defective adenoviral vector Ad/TRAIL-F/RGD using a tumor-specific human telomerase reverse transcriptase promoter. Cancer growth was studied using 2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide inner salt and clonogenic assays. Activation of the apoptosis pathway was analyzed in a Western blot and sub-G(1) DNA accumulation. A xenograft mouse lung cancer model was treated by intratumoral injections of Ad/TRAIL-F/RGD and local radiotherapy; the other groups received one of these treatments alone or a control agent. Apoptosis and TRAIL expression in tumors were also analyzed. RESULTS Ad/TRAIL-F/RGD specifically targets human NSCLC cells without significant effect in normal human bronchial epithelial cells. The combination of Ad/TRAIL-F/RGD and radiotherapy significantly improved cell-killing effect in all NSCLC cell lines tested (P < 0.05). Expression of TRAIL showed a dose-dependent relationship with Ad/TRAIL-F/RGD, and radiation seemed to increase TRAIL expression. Activation of the apoptosis by TRAIL and radiation was shown by activation of caspase-9, caspase-8, caspase-3, and poly(ADP-ribose) polymerase and increased DNA sub-G(1) accumulation. The combination of TRAIL and radiotherapy significantly increased apoptosis in vivo, inhibited tumor growth, and prolonged mean survival in mice bearing human NSCLC to 43.7 days compared with 23.7 days (TRAIL only) and 16.5 days (radiotherapy only; P < 0.05). CONCLUSIONS The combination of Ad/TRAIL-F/RGD and radiotherapy significantly improved therapeutic efficacy in suppressing NSCLC tumor growth and prolonging survival. Ad/TRAIL-F/RGD may improve the therapeutic ratio of radiotherapy in NSCLC.
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Affiliation(s)
| | | | | | - Bingliang Fang
- Thoracic and Cardiovascular Surgery, The University of Texas M. D. Anderson Cancer Center, Houston, Texas
| | - Joe Y. Chang
- Radiation Oncology, and
- *Corresponding author: Joe Y. Chang, M.D., Ph.D., Department of Radiation Oncology, The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030. Telephone: (713) 563-2300; Fax: (713) 563-2331; E-mail:. Bingliang Fang: co-corresponding author
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Iyer M, Salazar FB, Lewis X, Zhang L, Wu L, Carey M, Gambhir SS. Non-invasive imaging of a transgenic mouse model using a prostate-specific two-step transcriptional amplification strategy. Transgenic Res 2005; 14:47-55. [PMID: 15865048 DOI: 10.1007/s11248-004-2836-1] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022]
Abstract
Non-invasive assessment of transgenic animals using bioluminescence imaging offers a rapid means of evaluating disease progression in animal models of disease. One of the challenges in the field is to develop models with robust expression to image repetitively live intact animals through solid tissues. The prostate-specific antigen (PSA) promoter is an attractive model for studying gene regulation due to its hormonal response and tissue-specificity permitting us to measure signaling events that occur within the native tissues. The use of the GAL4-VP16 activator offers a powerful means to augment gene expression levels driven by a weak promoter. We have used a two-step transcriptional amplification (TSTA) system to develop a transgenic mouse model to investigate the tissue-specificity and developmental regulation of firefly luciferase (fl) gene expression in living mice using bioluminescence imaging. We employed an enhanced prostate-specific promoter to drive the yeast transcriptional activator, GAL4-VP16 (effector). The reporter construct carries five Gal4 binding sites upstream of the fl gene. We generated a transgenic mouse model using a single vector carrying the effector and reporter constructs. The transgenic mice show prostate-specific expression as early as three weeks of age. The bioluminescence signal in the prostate is significantly higher than in other organs. We also demonstrate that blocking androgen availability can downregulate the fl expression in the prostate. The transgenic mice display normal physical characteristics and developmental behavior, indicating that the high level of GAL4 driven expression is well tolerated. These findings suggest that the GAL4-VP16 transactivator can be used to amplify reporter gene expression from a relatively weak promoter in a transgenic mouse model. The transgenic TSTA model in conjunction with other transgenic cancer models should also help to detect and track malignancies. The strategies developed will be useful for transgenic research in general by allowing for amplified tissue specific gene expression.
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Affiliation(s)
- M Iyer
- The Crump Institute for Molecular Imaging, Department of Molecular & Medical Pharmacology, Stanford University School of Medicine, 318 Campus Drive, IE, Stanford, CA, USA
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Jacob D, Schumacher G, Bahra M, Davis J, Zhu HB, Zhang LD, Teraishi F, Neuhaus P, Fang BL. Fiber-modified adenoviral vector expressing the tumor necrosis factor-related apoptosis-inducing ligand gene from the human telomerase reverse transcriptase promoter induces apoptosis in human hepatocellular carcinoma cells. World J Gastroenterol 2005; 11:2552-6. [PMID: 15849810 PMCID: PMC4305742 DOI: 10.3748/wjg.v11.i17.2552] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: Because of a major resistance to chemotherapy, prognosis of hepatocellular carcinoma (HCC) is still poor. New treatments are required and gene therapy may be an option. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in multiple malignant tumors, and using adenoviral vectors has shown a targeted tumor-specific therapy. However, repeated administration of adenoviral vectors can lead to cell resistance, which may be caused by the initial coxsackie-adenovirus receptor (CAR). One technique to overcome resistance is the use of modified adenoviral vectors containing an Arg-Gly-Asp (RGD) sequence. In this study we constructed an adenoviral vector (designated Ad/TRAIL-F/RGD) with RGD-modified fibers, expressing the TRAIL gene from the human telomerase reverse transcriptase (hTERT) promoter, and evaluated its antitumor activity in HCC cell lines.
METHODS: To investigate the effects of Ad/TRAIL-F/RGD in human HCC cell lines Hep G2 and Hep 3b, cells were infected with Ad/CMV-GFP (vector control), Ad/gTRAIL (positive control), and Ad/TRAIL-F/RGD. Phosphate-buffered saline (PBS) was used as control. Cell viability was determined by proliferation assay (XTT), and apoptosis induction by fluorescence activated cell sorting (FACS).
RESULTS: Cells treated with Ad/TRAIL-F/RGD and Ad/gTRAIL showed a significantly reduced cell viability in comparison to PBS and Ad/CMV-GFP treatment in both cell lines. Whereas, treatment with PBS and Ad/CMV-GFP had no cell-killing effect. The reduced cell viability was caused by induction of apoptosis as shown by FACS analysis. The amount of apoptotic cells was similar after incubation with Ad/gTRAIL and Ad/TRAIL-F/RGD.
CONCLUSION: The new RGD modified vector Ad/TRAIL-F/RGD could become a potent therapeutic agent for the treatment of HCC, adenovirus resistant tumors, and CAR low or negative cancer cells.
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Affiliation(s)
- Dietmar Jacob
- Department of General, Visceral and Transplantation Surgery, Humboldt University of Berlin, CharitA Virchow Clinic, Augustenburger Platz 1, 13353 Berlin, Germany.
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21
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Chan CHF, Stanners CP. Novel mouse model for carcinoembryonic antigen-based therapy. Mol Ther 2005; 9:775-85. [PMID: 15194045 DOI: 10.1016/j.ymthe.2004.03.009] [Citation(s) in RCA: 65] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2003] [Accepted: 03/11/2004] [Indexed: 12/22/2022] Open
Abstract
Many novel cancer therapies, including immunotherapy and gene therapy, are specifically targeted to tumor-associated molecules, among which carcinoembryonic antigen (CEA) represents a popular example. Discrepancies between preclinical experimental data in animal models and clinical outcome in terms of therapeutic response and toxicity, however, often arise. Preclinical testing can be compromised by the lack of CEA and other closely related human CEA family members in rodents, which lack analogous genes for most human CEA family members. Here, we report the construction of a transgenic mouse with a 187-kb human bacterial artificial chromosome (CEABAC) that contains part of the human CEA family gene cluster including complete human CEA (CEACAM5), CEACAM3, CEACAM6, and CEACAM7 genes. The spatiotemporal expression pattern of these genes in the CEABAC mice was found to be remarkably similar to that of humans. This novel mouse will ensure better assessment than previously utilized models for the preclinical testing of CEA-targeted therapies and perhaps allow the testing of CEACAM6, which is overexpressed in many solid tumors and leukemias, as a therapeutic target. Moreover, expression of CEA family genes in gastrointestinal, breast, hematopoietic, urogenital, and respiratory systems could facilitate other clinical applications, such as the development of therapeutic agents against Neisseria gonorrhoeae infections, which use CEA family members as major receptors.
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Affiliation(s)
- Carlos H F Chan
- Department of Biochemistry and McGill Cancer Centre, McGill University, Montreal, Quebec, Canada H3G 1Y6
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22
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Poulsen TT, Pedersen N, Poulsen HS. Replacement and Suicide Gene Therapy for Targeted Treatment of Lung Cancer. Clin Lung Cancer 2005; 6:227-36. [PMID: 15694015 DOI: 10.3816/clc.2005.n.002] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
Lung cancer is the leading cause of cancer-related death in the developed world; consequently, novel therapeutic strategies are in high demand. A major problem with the present treatment modalities is the lack of tumor specificity giving rise to dose-limiting toxicity and side effects. Gene therapy constitutes an experimental approach gaining increased attention as a putative future cancer therapeutic strategy. Using this strategy, cancer cytotoxicity can be obtained by replacing mutated genes with functional analogues or introducing a suicide gene into the malignant cells. Insight into the molecular biology of cancer cells has identified a number of regulatory gene sequences, which can be used to selectively activate the therapeutic gene specifically in cancer cells, thereby reducing nonspecific toxicity. Although further improvements are necessary, recent encouraging results have shown promise for future clinical application of gene therapy. This article presents an update on the experimental and clinical results obtained within the field of lung cancer gene therapy, concentrating on strategies to specifically activate expression of the therapeutic gene in cancer cells. Furthermore, status of the development of delivery vector constructs for lung cancer gene therapy will be presented.
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Affiliation(s)
- Thomas T Poulsen
- Department of Radiation Biology, National University Hospital, Copenhagen, Denmark
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23
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Jacob D, Davis J, Zhu H, Zhang L, Teraishi F, Wu S, Marini FC, Fang B. Suppressing orthotopic pancreatic tumor growth with a fiber-modified adenovector expressing the TRAIL gene from the human telomerase reverse transcriptase promoter. Clin Cancer Res 2004; 10:3535-41. [PMID: 15161713 DOI: 10.1158/1078-0432.ccr-03-0512] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
An adenoviral vector with RGD-modified fibers and expressing the human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene from the human telomerase reverse transcriptase (hTERT) promoter (designated Ad/TRAIL-F/RGD) was constructed, and its antitumor activity was evaluated in vitro and in vivo. An in vitro study showed that treatment with Ad/TRAIL-F/RGD elicited a high rate of apoptosis in human pancreatic and colon cancer cell lines that were either susceptible or resistant to conventional adenovectors. In vivo study showed that direct administration of Ad/TRAIL-F/RGD to an orthotopic implantation tumor model established in the pancreatic tails of nu/nu mice significantly suppressed tumor growth: tumors in the animals treated with Ad/TRAIL-F/RGD were approximately eight times smaller than those in animals treated with a control vector. We also evaluated hTERT promoter activity and the effect of Ad/TRAIL-F/RGD on mesenchymal stem cells. Our results showed that transgene expression from the hTERT promoter in human bone marrow mesenchymal stem cells was minimal. No adverse effect was observed in mesenchymal stem cells treated with Ad/TRAIL-F/RGD. Together, our results suggest that Ad/TRAIL-F/RGD could become a potent therapeutic agent for the management of pancreatic cancer.
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Affiliation(s)
- Dietmar Jacob
- Department of Thoracic and Cardiovascular Surgery, The University of Texas M. D. Anderson Cancer Center, The University of Texas Graduate School of Biomedical Sciences, Houston, Texas 77030, USA
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He C, Lao WF, Hu XT, Xu XM, Xu J, Fang BL. Anti-liver cancer activity of TNF-related apoptosis-inducing ligand gene and its bystander effects. World J Gastroenterol 2004; 10:654-9. [PMID: 14991932 PMCID: PMC4716903 DOI: 10.3748/wjg.v10.i5.654] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
AIM: To observe the anti-liver cancer activity of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene and its bystander effects on hepatocellular carcinoma (HCC) cell line SMMC7721.
METHODS: Full-length cDNA of human TRAIL was transferred into SMMC7721 cells with a binary adenoviral vector system. Polymerase-chain reaction following reverse transcription (RT-PCR) was used to determine the expression of TRAIL gene. Effects of the transfected gene on proliferation of SMMC7721 cells were measured by MTT assay. Its influence on apoptosis was demonstrated by fluorescence-activated cell sorting (FACS). The bystander effect was observed by co-culturing the SMMC7721 cells with and without the transfected TRAIL gene at different ratios, and the culture medium supernatant from the transfected cells was also examined for its influence on SMMC7721 cells.
RESULTS: The growth-inhibition rate and apoptotic cell fraction in the cells transfected with the TRAIL gene, Bax gene or only LacZ gene were 91.2%, 48.0%, 28.8% and 29.1%, 12.5%, 6.6%, respectively. The growth-inhibition rate of transfection with these three sequences in normal human fibroblasts was 6.1%, 45.5% and 7.6%, respectively, indicating a discriminative inhibition of TRAIL transfection on the cancer cells. In the co-culturing test, addition of the transfected TRAIL to SMMC7721 cells in proportions of 5%, 25%, 50%, 75% and 100%, resulted in a growth-inhibition of 15.9%, 67%, 80.2%, 86.4% and 87.7%, respectively. We failed to observe a significant growth-inhibition effect of the culture medium supernatant on SMMC7721 cells.
CONCLUSION: TRAIL gene transferred by a binary adenoviral vector system can inhibit proliferation of SMMC7721 cells and induce their apoptosis. A bystander effect was observed, which seemed not to be mediated by soluble factors.
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Affiliation(s)
- Chao He
- Department of Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016, Zhejiang Province, China.
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Lin T, Zhang L, Davis J, Gu J, Nishizaki M, Ji L, Roth JA, Xiong M, Fang B. Combination of TRAIL gene therapy and chemotherapy enhances antitumor and antimetastasis effects in chemosensitive and chemoresistant breast cancers. Mol Ther 2003; 8:441-8. [PMID: 12946317 DOI: 10.1016/s1525-0016(03)00203-x] [Citation(s) in RCA: 46] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/27/2023] Open
Abstract
We recently found that breast cancer cell lines that are resistant to chemotherapy or to the recombinant TRAIL protein are susceptible to TRAIL gene therapy. However, it is unclear whether a combination of TRAIL gene therapy and chemotherapy will have enhanced antitumor activity or can be used for the treatment of metastasis. In this study, we investigated the combined effect of TRAIL gene therapy and chemotherapeutic agents, including doxorubicin, paclitaxel, vinorelbine, gemcitabine, irinotecan, and floxuridine, in different breast cancer cell lines. In all the cell lines tested, including a breast cancer cell line that is resistant to chemotherapy, the combination of TRAIL gene therapy and cytotoxic agents had either a synergistic or an additive effect. An in vivo study showed that aerosolized administration of an adenovector expressing the GFP-TRAIL fusion protein from the human telomerase reverse transcriptase promoter (designated Ad/gTRAIL) also decreased the number of lung metastases from both doxorubicin-sensitive and doxorubicin-resistant breast cancer cell lines. The combination of TRAIL gene therapy and chemotherapy resulted in a further reduction of lung metastatic nodules with minimal toxicity. These results suggest that a combination of TRAIL gene therapy and chemotherapy is effective in the treatment of metastatic diseases.
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Affiliation(s)
- Tongyu Lin
- Department of Thoracic and Cardiovascular Surgery, The University of Texas M. D. Anderson Cancer Center, 77030, Houston, Texas 77030, USA
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Abstract
Cancer gene therapy has been one of the most exciting areas of therapeutic research in the past decade. In this review, we discuss strategies to restrict transcription of transgenes to tumour cells. A range of promoters which are tissue-specific, tumour-specific, or inducible by exogenous agents are presented. Transcriptional targeting should prevent normal tissue toxicities associated with other cancer treatments, such as radiation and chemotherapy. In addition, the specificity of these strategies should provide improved targeting of metastatic tumours following systemic gene delivery. Rapid progress in the ability to specifically control transgenes will allow systemic gene delivery for cancer therapy to become a real possibility in the near future.
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Affiliation(s)
- Tracy Robson
- School of Biomedical Sciences, University of Ulster, Newtownabbey, Co. Antrim, BT37 0QB, Northern Ireland, UK
| | - David G. Hirst
- School of Biomedical Sciences, University of Ulster, Newtownabbey, Co. Antrim, BT37 0QB, Northern Ireland, UK
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Abstract
Cancer gene therapy is the transfer of genetic material to the cells of an individual with the goal of eradicating cancer cells, both in the primary tumor and metastases. Cancer gene therapy strategies exploit our expanding knowledge of the genetic basis of cancer, thereby allowing rationally targeted interventions at the molecular level. The successful implementation of cancer gene therapy in the clinic awaits the development of vectors capable of specific and efficient gene delivery to cancer cells. The first clinical applications of cancer gene therapy are likely to be in combination with conventional therapies, such as radiotherapy and immunotherapy.
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Affiliation(s)
- Joanne T Douglas
- Division of Human Gene Therapy, Department of Medicine, The Gene Therapy Center, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.
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Lin T, Huang X, Gu J, Zhang L, Roth JA, Xiong M, Curley SA, Yu Y, Hunt KK, Fang B. Long-term tumor-free survival from treatment with the GFP-TRAIL fusion gene expressed from the hTERT promoter in breast cancer cells. Oncogene 2002; 21:8020-8. [PMID: 12439752 DOI: 10.1038/sj.onc.1205926] [Citation(s) in RCA: 87] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/06/2002] [Revised: 07/31/2002] [Accepted: 08/01/2002] [Indexed: 12/28/2022]
Abstract
We evaluated anti-tumor activity and toxic effect of an adenoviral vector expressing the GFP/TRAIL fusion gene from the hTERT promoter (designated Ad/gTRAIL) on human breast cancer cell lines and on normal human breast cells. Treatment with Ad/gTRAIL elicited high levels of transgene expression and apoptosis in a variety of breast cancer cell lines. Furthermore, treatment with Ad/gTRAIL was effective in killing breast cancer lines resistant to doxorubicin or soluble TRAIL protein. In contrast, only minimal transgene expression and toxicity was detected in normal human primary mammary epithelial cells after treatment with this vector. An in vivo study further showed that the intralesional administration of Ad/gTRAIL effectively suppressed the growth of human tumor xenografts derived from both doxorubicin-sensitive and doxorubicin-resistant breast cancer lines. Specifically, about 50% of animals bearing doxorubicin-sensitive and doxorubicin-resistant breast cancer xenografts showed complete tumor regression and remained tumor-free for over 5 months. These results suggest that the adenovirus encoding the GFP/TRAIL gene driven by the hTERT promoter has potential application in cancer therapy.
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Affiliation(s)
- Tongyu Lin
- Department of Thoracic and Cardiovascular Surgery, Section of Thoracic Molecular Oncology, The University of Texas M. D. Anderson Cancer Center, Houston 77030, USA
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Huang X, Lin T, Gu J, Zhang L, Roth JA, Stephens LC, Yu Y, Liu J, Fang B. Combined TRAIL and Bax gene therapy prolonged survival in mice with ovarian cancer xenograft. Gene Ther 2002; 9:1379-86. [PMID: 12365003 DOI: 10.1038/sj.gt.3301810] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/01/2002] [Accepted: 05/07/2002] [Indexed: 11/08/2022]
Abstract
We evaluated the antitumor activity of the Bax gene and green fluorescent protein/tumor necrosis factor-related apoptosis-inducing ligand (GFP/TRAIL) fusion gene driven by the human telomerase reverse transcriptase promoter both separately and combined in the human ovarian cancer lines SKOV3ip and DOV13 and human lung cancer line H1299. In vitro study showed that both TRAIL- and Bax-expressing vectors elicited significant cell killing in H1299 and SKOV3ip cells, but only the GFP/TRAIL gene elicited significant cell killing in DOV13 cells. Combined TRAIL and Bax therapy also produced more profound cell killing in SKOV3ip and H1299 cells, but not DOV13 cells without escalation of the vector doses. To further evaluate the combined effects of Bax and TRAIL, abdominally spread tumors were established in nude mice via intraperitoneal inoculation of SKOV3ip cells followed by that of adenoviral vectors. Tumor growth, ascites formation, survival duration and toxicity were evaluated after treatment. We found that treatment using the Bax- or TRAIL-expressing vector alone significantly suppressed tumor growth and ascites formation, and prolonged animal survival when compared with that of using PBS or a control vector. Combined TRAIL and Bax therapy further prolonged survival significantly when compared with therapy using the TRAIL or Bax gene alone. Transgene expression and apoptosis induction were not detected in normal human ovarian epithelial cells in vitro or normal mouse tissues in vivo after intraperitoneal vector administration. Also, liver toxicity was not detected after either treatment. Thus, combined TRAIL and Bax gene therapy may be useful for treatment of abdominally spread tumors.
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Affiliation(s)
- X Huang
- Department of Thoracic and Cardiovascular Surgery, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
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Barnett BG, Tillman BW, Curiel DT, Douglas JT. Dual targeting of adenoviral vectors at the levels of transduction and transcription enhances the specificity of gene expression in cancer cells. Mol Ther 2002; 6:377-85. [PMID: 12231174 DOI: 10.1006/mthe.2002.0670] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Adenoviral (Ad) vector-mediated strategies for cancer gene therapy mandate a vector that is capable of efficient expression of the therapeutic gene specifically within the target tumor cells. In one approach to the development of cancer cell-specific vectors, Ad vectors have been targeted at the level of transduction to achieve the selective delivery of the therapeutic gene. In an alternative approach to the derivation of cancer cell-specific vectors, Ad vectors have been targeted at the level of transcription by placing the therapeutic gene under the control of transcriptional regulatory sequences that are activated in tumor cells, but not in normal cells, and therefore target expression selectively to the tumor cell. In this report, we demonstrate that a higher degree of specificity for cancer cells can be achieved by combining the complementary approaches of transductional and transcriptional targeting, each of which is imperfect or "leaky" by itself.
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Affiliation(s)
- Brian G Barnett
- Division of Human Gene Therapy, Department of Medicine and the Gene Therapy Center, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA
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Zhang L, Gu J, Lin T, Huang X, Roth JA, Fang B. Mechanisms involved in development of resistance to adenovirus-mediated proapoptotic gene therapy in DLD1 human colon cancer cell line. Gene Ther 2002; 9:1262-70. [PMID: 12215894 DOI: 10.1038/sj.gt.3301797] [Citation(s) in RCA: 39] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2002] [Accepted: 04/29/2002] [Indexed: 01/20/2023]
Abstract
To evaluate resistance that develops in cancer cells during treatment with adenoviral vectors expressing proapoptotic genes, we repeatedly treated the human colon cancer cell line DLD1 with adenoviral vectors expressing the human Bax gene and the human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene. DLD1 cells resistant to the Bax- or TRAIL-expressing adenoviral vectors were then selected and designated as DLD1/Bax-R or DLD1/TRAIL-R cells, respectively. Further study showed that resistance in DLD1/Bax-R cells was caused by resistance to adenoviral infection, which can be overcome by dose escalation of the adenoviral vectors. However, resistance in DLD1/TRAIL-R cells was caused by resistance to the TRAIL gene. Therefore, different mechanisms are involved in the development of resistance during adenovirus-mediated proapoptotic gene therapy. A survey of molecules involved in TRAIL- or Bax-mediated apoptotic pathways showed no significant change in expression of death receptors, death decoy receptors; FLIP; Bcl-2; Bcl-xS; Bax; Bak; XIAP or caspase-2, -7, -8, or -9 in either DLD1/Bax-R or DLD1/TRAIL-R cells. Bcl-xL expression detected in both mRNA and protein level assays was three times higher in DLD1/TRAIL-R cells than in parental or DLD1/Bax-R cells. However, transfection of DLD1 cells with the Bcl-xL gene showed that overexpression of Bcl-xL is not sufficient for the resistance. Moreover, DLD1/Bax-R cells were sensitive to adenoviral vectors that expressed the TRAIL gene, but resistant to adenoviral vectors that expressed the Bak gene. In contrast, DLD1/TRAIL-R cells were sensitive to adenoviral vectors that expressed either Bax or Bak gene. Thus, alternative application of adenoviral vectors that expressed proapoptotic genes in different pathways or different cell killing models may delay or prevent development of resistance in adenovirus-mediated proapoptotic gene therapy.
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Affiliation(s)
- L Zhang
- Department of Thoracic and Cardiovascular Surgery, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
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Abstract
Current treatment of solid tumors is limited by severe adverse effects, resulting in a narrow therapeutic index. Therefore, cancer gene therapy has emerged as a targeted approach that would significantly reduce undesired side effects in normal tissues. This approach requires a clear understanding of the molecular biology of both the malignant clone and the biological vectors that serve as vehicles to target cancer cells. In this review we discuss novel approaches for conditional gene expression in cancer cells. Targeting transgene expression to malignant tissues requires the use of specific regulatory elements including promoters based on tumor biology, tissue-specific promoters and inducible regulatory elements. We also discuss the regulation of both replication and transgene expression by conditionally-replicative viruses. These approaches have the potential to restrict the expression of transgenes exclusively to tissues of interest and thereby to increase the therapeutic index of future vectors for cancer gene therapy.
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Affiliation(s)
- Y S Haviv
- Division of Human Gene Therapy, Departments of Medicine, Surgery and Pathology, University of Alabama at Birmingham, 1824 6th Avenue South, Birmingham, AL 35294, USA
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