Abstract
Background
The objective of this study was to determine whether rat insulin promoter (RIP) could, in a mouse model, direct expression of an adenovirus-mediated reporter gene specifically into pancreatic islets via systemic delivery.
Methods
Five hundred and eight base pairs of the RIP DNA sequence were constructed into an adenoviral vector containing a lacZ reporter gene (Adeno-RIP-lacZ). The cytomegalovirus (CMV) promoter was constructed to drive lacZ reporter-gene expression (Adeno-CMV-lacZ) and used as controls. In vitro transient transfection assays were performed to determine levels of reporter-gene expression and compared with that of liposome-mediated plasmid transfection. SCID mice were bred and housed in the barrier BL-4 animal facility. At 2 months of age, the human pancreatic cancer cell PANC-1 was intraperitoneally injected into male mice. Two months after the tumor cell inoculation, mice were injected with 107 adenoviral particles via tail veins. After gene delivery, mice were sacrificed at different time points to determine transgene expression levels. Complete necropsies were performed. Morphological alterations were determined using hematoxylin and eosin (H&E) staining, and distribution of the reporter lacZ gene was determined by immunohistochemistry analyses.
Results
Adenoviral-driven reporter-gene expression resulted in more than 5 times higher transgene expression compared with conventional plasmid transfections. In Adeno-RIP-lacZ-injected mice, lacZ expression was specifically detected in pancreatic islets. By contrast, in Adeno-CMV-lacZ-injected mice, lacZ gene expression was observed in multiple organs and tissues. Mononuclear cell infiltration and liver cell inflammation were found in Adeno-CMV-lacZ-treated mice. Similar phenomena were observed in islet cells of Adeno-RIP-lacZ-treated mice. A significantly higher level of reporter-gene expression was also found at the edge of in-vivo-inoculated human pancreatic tumors.
Conclusion
These results demonstrate that RIP-directed reporter-gene expression was found specifically in mouse pancreatic islets and implanted human pancreatic cancer cells. These data thus demonstrate that the combination of an adenoviral vector and a tissue-specific promoter could lead to an enhanced and more specific transgene expression in vivo.
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References
Ginsberg GG. New developments in pancreatic cancer. Semin Gastrointest Dis 2000;11:162
Brennan MF. Pancreatic cancer. J Gastroenterol Hepatol 2000;15:G13
Yeo CJ, Cameron JL. Neoplasms of the pancreas exocrine tumors, 5th edn. Philadelphia, PA, W. B. Saunders Company, 1997
Olson L, Qian J, Poitout V. Glucose rapidly and reversibly decreases INS-1 cell insulin gene transcription via decrements in STF-1 and C1 activator transcription factor activity. Mol Endocrinol 1998;12:207
Al-Hendy A, Auersperg N. Applying the Herpes simplex virus thymidine Kinase/Ganciclovir approach to Ovarian Cancer: an effective in vitro drug-sensitization system. Gynecol Obstet Invest 1997;42:268
Bonnekoh B, Greenhalgh D, Bundman D, et al. Inhibition of melanoma growth by adenoviral mediated HSV thymidine kinase gene transfer in vivo. J Invest Derm 1995;104:313
Fussenegger M. The impact of mammalian gene regulation concepts on functional genomic research, metabolic engineering, and advanced gene therapies. Biotechnol Prog 2001;17:1
Zhang R, Straus FH, DeGroot LJ. Adenoviral-mediated gene therapy for thyroid carcinoma using thymidine kinase controlled by thyroglobulin promoter demonstrated high specificity and low toxicity. Thyroid 2001;11:115
Piva MG, Navaglia F, Fogar DB, et al. CEA mRNA identification in peripheral blood is feasible for colorectal, but not for gastric or pancreatic cancer staging. Oncology 2000;59:323
Okai T, Kawashima A, Watanabe H, et al. Nonfunctioning islet cell carcinoma of the pancreas with high serum CEA CA19-9, K-ras codon 12 mutation and microsatellite instability. J Clin Gastroenterol 2000;30:307
DiMaio JM, Clary BM, Via DF, et al. Directed enzyme pro-drug gene therapy for pancreatic cancer in vivo. Surgery 1994;116:205
Marrelli D, Pintp E, Stefano AD, et al. Clinical utility of CEA, CA 19-9, and CA 72-4 in the follow-up of patients with resectable gastric cancer. Am J Surg 2001;181:16
Osaki T, Tanio Y, Tachibana I, et al. Gene therapy for carcinoembryonic antigen-producing human lung cancer cells by cell type-specific expression of herpes simplex virus thymidine kinase gene. Cancer Res 1994;54:5258
Minami S, Furui J, Kanamatsu T. Role of carcinoembryonic antigen in the progression of colon cancer cells that express carbohydrate antigen. Cancer Res 2001;61:2732
Tirone TA, Fagan SP, Templeton NS, et al. Insulinoma-induced hypoglycemic death in mice is prevented with beta cell-specific gene therapy. Ann Surg 2001;233:603
Tirone TA, Wang XP, Templeton NS, et al. Cell-specific cytotoxicity of human pancreatic adenocarcinoma cells using rat insulin promoter thymidine kinase directed gene therapy. World J Surg 2004;28:826
Aoki K, Yoshida T, Matsumoto N, et al. Gene therapy for peritoneal dissemination of pancreatic cancer by liposome-mediated transfer of herpes simplex virus thymidine kinase gene. Hum Gene Ther 1997;8:1105
Bouvet M, Ellis LM, Nishizaki M, et al. Adenovirus-mediated wild-type p53 tumor suppressor gene therapy induces apoptosis and suppresses growth of human pancreatic cancer. Ann Surg Oncol 1998;5:681
Chen SH, Shine D, Goodman J, et al. Gene Therapy for brain tumors: regression of experimental gliomas by adenovirus-mediated gene transfer in vivo. Proc Natl Acd Sci USA 1994;91:3054
Makinen K, Loimas S, Wahlfors J, et al. Evaluation of herpes simplex thymidine kinase mediated gene therapy in experimental pancreatic cancer. J Gene Med 2000;2:361
Qian C, Bilbao R, Bruna O, et al. Induction of sensitivity to ganciclovir in human hepatocellular carcinoma cells by adenovirus-mediated gene transfer of herpes simples virus thymidine kinase. Hepatology 1995;22:118
Ramesh R, Saeke T, Templeton NS, et al. Successful treatment of primary and disseminated human lung cancers by systemic delivery of tumor suppressor genes using an improved liposome vector. Mol Ther 2001;3:337
Yang L, Hwang R, Chiang Y, et al. Mechanisms for ganciclovir resistance in gastrointestinal tumor cells transduced with a retroviral vector containing the herpes simplex virus thymidine kinase gene. Clin Cancer Res 1998;4:731
Davis AR, Meyers K, Wilson JM. High throughput method for creating and screening recombinant adenoviruses. Gene Ther 1998;5:1148
Wang XP, Yazawa K, Yang J, et al. Specific gene expression and therapy for pancreatic cancer using the cytosine deaminase gene directed by the Rat Insulin Promoter. J Gastrointestinal Surg 2004;8:98
Yang L, Chiang Y, Lenz HJ, et al. Intercellular communication mediates the bystander effect during herpes simplex thymidine kinase/ganciclovir-based gene therapy of human gastrointestinal tumor cells. Hum Gene Ther 1998;9:719
Sander M, German M. The beta cell transcription factors and development of the pancreas. J Mol Med 1997;75:327
Jonsson J, Carlsson L, Edlund T, et al. Insulin promoter factor 1 is required for pancreas development in mice. Nature 1994;371:606
Ahlgren U, Jonsson J, Edlund H. The morphogenesis of the pancreatic mesenchyme is uncoupled from that of the pancreatic epithelium in IPF1/PDX1-deficient mice. Development 1996;122:1409
Dutta S, Gannon M, Peers B, et al. PDX:PBX complexes are required for normal proliferation of pancreatic cells during development. Proc Natl Acad Sci USA 2001;98:1065
Peshavaria M, Cissell MA., Henderson E, et al. The PDX-1 activation domain provides specific functions necessary for transcriptional stimulation in pancreatic beta-cells. Mol Endocrinol 2000;14:1907
Rooman I, Heremans Y, Heimberg H, et al. Modulation of rat pancreatic acinoductal transdifferentiation and expression of PDX-1 in vitro. Diabetologia 2000;43:907
Macfarlane WM, Shepherd RM, Cosgrove KE, et al. Glucose modulation of insulin mRNA levels is dependent on transcription factor PDX-1 and occurs independently of changes of intracellular Ca2+. Diabetes 2000;49:418
Ohneda K, Mirmira RG, Wang J, et al. The homeodomain of PDX-1 mediates multiple proteins-protein interactions in the formation of a transcriptional activation complex on the insulin promoter. Mol Cell Biol 2000;20:900
Marshak S, Benshushan E, Shoshkes M, et al. Beta-cell-specific expression of insulin and PDX-1 genes. Diabetes 2001;50:S131
Ashizawa S, Brunicardi FC, Wang XP. PDX-1 and the pancreas. Pancreas 2004;28:109
Dufayet D, Halvorsen T, Demeterco C, et al. Beta-cell differentiation from a human pancreatic cell line in vitro and in vivo. Mol Endocrino 2001;15:476
Grapin-Botton A, Majithia AR, Melton DA. Key events of pancreas formation are triggered in gut endoderm by ectopic expression of pancreatic regulatory genes. Genes Dev 2001;15:444
Kritzik MR, Krahl T, Good A, et al. Transcription factor expression during pancreatic islet regeneration. Mol Cell Endocrinol 2000;164:99
Gernan MS. Wang H. The insulin gene contains multiple transcriptional elements that respond to glucose. Mol Cell Biol 1994;14:4067
Karlsson O, Edlund T, Moss JB, et al. A mutational analysis of the insulin gene transcription control region: expression in beta cells is dependent on two related sequences within the enhancer. Proc Natl Acad Sci USA 1987;84:8819
Petersen HV, Serup P, Leonard J, et al. Transcriptional regulation of the human insulin gene is dependent on the homeodomain protein STF1/IPF1 acting through the CT boxes. Proc Natl Acad Sci USA 1994;91:10465
Acknowledgement
This project is supported in part by an Advances in Technology Transfer Grant (ATP1176) from the State of Texas Department of Education, and an NIH R01 grant (NCI-CA95731). We would like to thank our colleagues working in Dr. Brunicardi’s laboratory for their support and suggestions. Our gratitude also extends to Ms. Katie Elsbury for her continued assistance and editorial support.
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Wang, X., Olmsted-Davis, E., Davis, A. et al. Specific Targeting of Pancreatic Islet Cells In Vivo by Insulin-Promoter-Driven Adenoviral Conjugated Reporter Genes. World J. Surg. 30, 1543–1552 (2006). https://doi.org/10.1007/s00268-005-0688-3
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DOI: https://doi.org/10.1007/s00268-005-0688-3