Use of a modified ornithine decarboxylase promoter to achieve efficient c-MYC- or N-MYC-regulated protein expression.
One of the advantages of viral-directed enzyme prodrug therapy (VDEPT) is its potential for tumor-specific cytotoxicity. However, the viruses used to deliver cDNAs encoding prodrug-activating enzymes transduce normal cells as well as tumor cells, and several approaches to achieve tumor-specific expression of the delivered cDNAs are being investigated. One such approach is to regulate transcription of the prodrug-activating enzyme with a promoter that is preferentially activated by tumor cells. Published data suggest that the most promising transcription factor/promoter/enhancer combinations are those activated by a tumor-specific transcription factor to retain tumor cell specificity but that are equal in strength to nonspecific viral promoters in their ability to up-regulate target cDNAs. This report identifies MYC-responsive, modified ornithine decarboxylase (ODC) promoter/enhancer sequences that up-regulate target protein expression in tumor cells overexpressing either N-MYC or c-MYC protein. The most efficient of the four constructs assessed contained six additional CACGTG MYC binding sites 5' to the endogenous ODC promoter (R6ODC). Reporter assays with this chimeric promoter/enhancer regulating expression of chloramphenicol acetyltransferase demonstrated 50-250-fold more activity in MYC-expressing cells compared with similar assays with promoterless plasmids. The R6ODC regulatory sequence was approximately equivalent to the CMV promoter in inducing expression of the neomycin resistance gene in c-MYC-expressing SW480 and HT-29 colon carcinoma cells and in N-MYC-expressing NB-1691 neuroblastoma cells. The modified ODC promoter may, therefore, be useful in achieving tissue-specific expression of target proteins in tumor cells that overexpress c- or N-MYC.
Iyengar
R V
RV
Department of Molecular Pharmacology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Pawlik
C A
CA
Krull
E J
EJ
Phelps
D A
DA
Burger
R A
RA
Harris
L C
LC
Potter
P M
PM
Danks
M K
MK
D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000493:pharmacokinetics / Q000494:pharmacology*; D001711:Biotransformation; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000493:pharmacokinetics / Q000494:pharmacology*; D043182:Carboxylesterase; D002265:Carboxylic Ester Hydrolases / Q000096:biosynthesis / Q000235:genetics; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D016259:Genes, myc / Q000235:genetics*; D005909:Glioblastoma / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D000077146:Irinotecan; D017570:MyoD Protein / Q000096:biosynthesis / Q000235:genetics; D009447:Neuroblastoma / Q000235:genetics / Q000378:metabolism; D009955:Ornithine Decarboxylase / Q000235:genetics*; D011401:Promoter Regions, Genetic; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis* / Q000235:genetics; D011817:Rabbits; D012208:Rhabdomyosarcoma / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D019076:Transgenes; D014407:Tumor Cells, Cultured
D000972:Antineoplastic Agents, Phytogenic; D017570:MyoD Protein; D016271:Proto-Oncogene Proteins c-myc; D000077146:Irinotecan; D015500:Chloramphenicol O-Acetyltransferase; D002265:Carboxylic Ester Hydrolases; D043182:Carboxylesterase; D009955:Ornithine Decarboxylase; D002166:Camptothecin