Amplified Muc1-specific gene expression in colon cancer cells utilizing a binary system in adenoviral vectors.
Mucin-1 is expressed in a variety of colon carcinomas and Muc-1/DF3 promoters have been utilized to reduce systemic toxicity through specific gene expression. To overcome weak expression, which is much lower than the widely used cytomegalovirus-promoter (CMV), new adenoviral vectors containing a binary system of transgene amplification have been developed. The Muc-1/DF3 promoter was used to control the expression of a Gal4VP16 fusion protein. This vector also contained Gal4 binding sites enabling the fusion protein to act as a transactivator, inducing transgene expression within the same construct. Mucin-1 expression was analyzed in a variety of colon cancer cell lines. After infection with recombinant adenoviruses, transgene expression was quantified using the luciferase system. Integration of the Gal4VP16-binary resulted in an up to 250-fold increase of Muc-1/DF3-specific gene expression. In mucin-positive cell lines utilizing this amplified Muc-1/DF3 promoter, expression was up to 590-fold higher as compared to the CMV-promoter. Western blot detected the presence of Gal4VP16 in infected muc-1-positive but not-negative cell lines. These new adenoviral vectors combing highly efficient and specific transgene expression and will contribute to the safety and efficacy of experimental approaches in cancer gene therapy.
Block
Andreas
A
Department of Medicine and Gastroenterology, University of Hamburg, University Hospital Hamburg-Eppendort, Martinistrasse 52, D-20246 Hamburg, Germany. block@uke.uni-hamburg.de
Milasinovic
Dragan
D
Mueller
Juergen
J
Schaefer
Peter
P
Schaefer
Hansjoerg
H
Greten
Heiner
H
D000256:Adenoviridae / Q000235:genetics; D001483:Base Sequence; D015153:Blotting, Western; D003110:Colonic Neoplasms / Q000235:genetics* / Q000378:metabolism; D003587:Cytomegalovirus / Q000235:genetics; D005784:Gene Amplification; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000096:biosynthesis / Q000235:genetics*; D011401:Promoter Regions, Genetic; D015534:Trans-Activators / Q000235:genetics; D015533:Transcriptional Activation; D019076:Transgenes / Q000235:genetics; D014407:Tumor Cells, Cultured
C070633:Gal-VP16; D018396:Mucin-1; D015534:Trans-Activators