Effect of antiangiogenic therapy on luciferase activity in a cytomegalovirus- or HSP70-promoter-transfected M21 tumor model.
We investigated the effect of targeted gene therapy on heat shock protein 70 expression (Hsp70) and protein production (HSP70) in a melanoma tumor model (M21; M21-L). M21 and M21-L cells transfected with a plasmid containing the Hsp70 (Hspa1b) or the cytomegalovirus (CMV) promoter and the luciferase reporter gene were injected into mice; the resulting tumors grew to a size of 650 mm(3). Mice (five per group) were intravenously treated with an Arg-Gly-Asp peptide-nanoparticle/Raf-1 kinase inhibitor protein complex [RGD-NP/RAF(-)] or with a nanoparticle control. Bioluminescence imaging (IVIS®, Xenogen, USA) was performed at 12, 24, 48, and 72 h after the treatment cycle. Western blot analysis of HSP70 protein was performed to monitor protein expression. The size of the treated M21 tumors remained fairly constant (647.8 ± 103.4 mm(2) at the beginning versus 704.8 ± 94.4 mm(3) at the end of the experiment). The size of the M21-L tumors increased, similar to the untreated control tumors. Bioluminescent imaging demonstrated that when transcription was controlled by the CMV promoter, luciferase activity decreased to 17.9% ± 4.3% of baseline values in the treated M21 tumors. When transcription was controlled by the Hsp70 promoter, the highest luciferase activity (4.5 ± 0.7-fold increase over base-line values) was seen 24 h after injection in the M21 tumors; however, no luciferase activity was seen in the M21-L tumors. In accordance with bioluminescent imaging, western blot analysis showed a peak in HSP70 production at 24 h after the injection of the RGD-NP/RAF(-) complex in the M21 tumors; however, no HSP70 protein induction was seen in the M21-L tumors. Thus, targeted antiangiogenic therapy can induce Hsp70 expression and HSP70 protein in melanoma tumors.
Stanford School of Medicine, Lucas MRS Research Center, Department of Radiology, Stanford, California, USA. hundt@med.uni-marburg.de
Walter
W
Hundt
Christian
C
Schink
Silke
S
Steinbach
Caitlin E
CE
O'Connell-Rodwell
Andreas
A
Kiessling
Damiano
D
Librizzi
Mykhaylo
M
Burbelko
Samira
S
Guccione
D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D001483:Base Sequence; D045744:Cell Line, Tumor; D003587:Cytomegalovirus / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D015316:Genetic Therapy / Q000379:methods; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008156:Luciferases / Q000378:metabolism*; D049449:Luminescence; D051379:Mice; D008819:Mice, Nude; D008969:Molecular Sequence Data; D009368:Neoplasm Transplantation; D011401:Promoter Regions, Genetic; D013997:Time Factors; D014162:Transfection
D020533:Angiogenesis Inhibitors; D018840:HSP70 Heat-Shock Proteins; D008156:Luciferases