Heat-inducible in vivo gene therapy of colon carcinoma by human mdr1 promoter-regulated tumor necrosis factor-alpha expression.
The promoter of the human multidrug resistance gene (mdr1) harbors defined heat-responsive elements, which could be exploited for construction of heat-inducible expression vectors. To analyze the hyperthermia inducibility of the mdr1 promoter in vitro and in vivo, we used the pcDNA3-mdrp-hTNF vector construct for heat-induced tumor necrosis factor alpha (TNF-alpha) expression in transfected HCT116 human colon carcinoma cells at mRNA level by quantitative real-time reverse transcription-PCR and at protein level by TNF-alpha ELISA. For the in vitro studies, the pcDNA3-mdrp-hTNF-transfected tumor cells were treated with hyperthermia at 43 degrees C for 2 h. In the animal studies, stably transfected or in vivo jet-injected tumor-bearing Ncr:nu/nu mice were treated for 60 min at 42 degrees C to induce TNF-alpha expression. Both the in vitro and in vivo experiments show that hyperthermia activates the mdr1 promoter in a temperature- and time-dependent manner, leading to an up to 4-fold increase in mdr1 promoter-driven TNF-alpha expression at mRNA and an up to 3-fold increase at protein level. The in vivo heat-induced TNF-alpha expression combined with Adriamycin (8 mg/kg) treatment leads to the inhibition of tumor growth in the animals. These experiments support the idea that heat-induced mdr1 promoter-driven expression of therapeutic genes is efficient and feasible for combined cancer gene therapy approaches.
Walther
Wolfgang
W
Max-Delbrück-Center for Molecular Medicine, Robert-Rössle-Strasse 10, 13092 Berlin, Germany. wowalt@mdc-berlin.de
Arlt
Franziska
F
Fichtner
Iduna
I
Aumann
Jutta
J
Stein
Ulrike
U
Schlag
Peter M
PM
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D003110:Colonic Neoplasms / Q000235:genetics* / Q000473:pathology / Q000628:therapy*; D003131:Combined Modality Therapy; D004317:Doxorubicin / Q000494:pharmacology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D015316:Genetic Therapy / Q000379:methods*; D045325:HCT116 Cells; D006801:Humans; D006979:Hyperthermia, Induced; D007277:Injections, Jet; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D016896:Treatment Outcome; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics* / Q000378:metabolism
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D004317:Doxorubicin