Limited specificity of promoter constructs for gene therapy in osteosarcoma.
Osteosarcoma (OS), a malignant bone neoplasia in childhood, has poor prognosis if metastases appear in the lung. A novel therapeutic approach could consist in a gene therapeutic treatment of OS metastases. However, if promiscuous viral vectors are applied for the delivery of potentially toxic transgenes, their misdelivery into normal tissues could cause severe complications. This problem could be circumvented by application of OS-specific promoters for transgene expression control. We analysed the function of promoters described to be tumour-, osteosarcoma- or osteoblast-specific. Expression rates driven by osteoblast- specific fragments from the collagen1A1-promoter, the human Osteocalcin-promoter, the bone-sialoprotein promoter and the beta-catenin promoter depending on vitamin supplementation were analysed in five OS cell lines, in normal lung fibroblasts and in a non-osteoblastic prostate cancer cell line (LNCaP) by dual luciferase assays. In addition, an unspecific but doxycyclin-repressible promoter construct (pAd.3r-luc) was examined. We found that all constructs were active in OS cell lines to varying extents. The complete human Osteocalcin promoter and the bone-sialoprotein promoter were partially induced by vitamin D3 or C respectively while the pAd.3r-luc activity could be shut down by doxycyclin. In contrast, the human Osteocalcin-promoter was not activated by vitamin D3 in LNCaP cells; its action remained relatively low. Interestingly, excepting the beta-catenin promoter, we measured strong activities of all promoters in lung fibroblast cells. Our study demonstrates that promoter activity should be evaluated not only for the target cells of the gene therapeutic approaches, but also for neighbouring normal tissues. Unspecific but repressible promoters could represent an alternative.
Pollmann
Annika
A
Department of Paediatric Haematology and Oncology, Clinic for Children's Health, Martinistrasse 52, D-20246 Hamburg, Germany.
Kabisch
Hartmut
H
Block
Andreas
A
Müller
Jürgen
J
Hellwinkel
Olaf J C
OJ
D045744:Cell Line, Tumor; D003598:Cytoskeletal Proteins / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D008168:Lung / Q000378:metabolism; D009928:Organ Specificity; D012516:Osteosarcoma / Q000235:genetics* / Q000628:therapy*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015534:Trans-Activators / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics; D051176:beta Catenin
C495270:CTNNB1 protein, human; D003598:Cytoskeletal Proteins; D015534:Trans-Activators; D051176:beta Catenin