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| A 1ST STEP IN THE DEVELOPMENT OF GENE-THERAPY FOR COLORECTAL-CARCINOMA - CLONING, SEQUENCING, AND EXPRESSION OF ESCHERICHIA-COLI CYTOSINE DEAMINASE |
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| Author(s): AUSTIN EA, HUBER BE |
| Source: MOLECULAR PHARMACOLOGY Volume: 43 Issue: 3 Pages: 380-387 Published: MAR 1993 |
| Times Cited: 168 References: 28 |
| Abstract: We have developed a new approach involving gene therapy for the treatment of primary and metastatic tumors in the liver. As a first step toward the development of this gene therapy treatment for metastatic colorectal carcinoma, the Escherichia coli gene that encodes cytosine deaminase (CD) (EC 3.5.4.1) has been cloned. By using positive genetic selection, a plasmid carrying a 10.8-kilobase BamHI/EcoRI DNA insert was isolated that had CD enzymatic activity. Genetic screening, followed by enzymatic assays, identified a 3-kilobase DNA fragment that retained CD activity. Deamination of cytosine and 5-fluorocytosine (5-FC) by cloned CD was demonstrated. DNA and protein sequencing identified an open reading frame of 427 amino acids that encodes CD. To demonstrate that expression of CD in eukaryotic cells allows metabolism of the nontoxic prodrug 5-FC to the toxic metabolite 5-fluorouracil, CD was cloned into a eukaryotic expression vector and transfected into a human colorectal carcinoma cell line. Growth inhibition studies showed a shift in the IC50 for 5-FC from 17,000 muM in the parental cell line to 30 muM in cells expressing CD. |
| Document Type: Article |
| Language: English |
Addresses:
1. WELLCOME RES LABS, DIV CELL BIOL, 3030 CORNWALLIS RD, RES TRIANGLE PK, NC 27709 USA |
| Publisher: WILLIAMS & WILKINS, 351 WEST CAMDEN ST, BALTIMORE, MD 21201-2436 |
| Subject Category: Pharmacology & Pharmacy |
| IDS Number: KT629 |
| ISSN: 0026-895X |
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