Topotecan inhibits vascular endothelial growth factor production and angiogenic activity induced by hypoxia in human neuroblastoma by targeting hypoxia-inducible factor-1alpha and -2alpha.
Neuroblastoma produce angiogenic peptides, and the extent of angiogenesis correlates with tumor progression and poor clinical outcome. Hence, angiogenic factor inhibition represents an important therapeutic option. One of the major drives to tumor angiogenesis is hypoxia, a decrease in oxygen tension that characterizes the tumor microenvironment. We investigated the effects of the topoisomerase I inhibitor, topotecan, on vascular endothelial growth factor (VEGF) induction by hypoxia in advanced-stage human neuroblastoma cells. Topotecan counteracted hypoxic induction of VEGF and decreased angiogenic activity of conditioned medium from hypoxic cultures in vivo in the chick chorioallantoic membrane. Promoter-driven reporter studies showed the role of both hypoxia-inducible factor (HIF)-1alpha and -2alpha in VEGF transcription activation by hypoxia, because (a) overexpression of either protein by cotransfection with expression vectors resulted in VEGF promoter transactivation, which was abrogated by mutation in the HIF-binding site, and (b) targeted knockdown of HIF-1alpha/2alpha by RNA interference inhibited hypoxia-stimulated VEGF transcriptional activity and protein secretion. Topotecan-inhibitory effects on VEGF induction by hypoxia were mediated through suppression of both HIF-1alpha and HIF-2alpha protein accumulation and transactivation properties, which was specific and required ongoing RNA transcription. A similar pattern of results was obtained in cells treated with the hypoxia-mimetic agent, desferrioxamine. These data provide the first evidence that topotecan is a potent inhibitor of HIF-1alpha and HIF-2alpha subunits in hypoxic neuroblastoma cells, leading to decreased VEGF expression and angiogenic activity. An important clinical implication of these findings is that therapies targeted to the HIF pathway have the potential to inhibit neuroblastoma angiogenesis and growth.
Puppo
Maura
M
Laboratory of Molecular Biology, G. Gaslini Institute, L.go Gerolamo Gaslini 5, 16147 Genova Quarto, Italy.
Battaglia
Florinda
F
Ottaviano
Catherine
C
Delfino
Silvana
S
Ribatti
Domenico
D
Varesio
Luigi
L
Bosco
Maria Carla
MC
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D051792:Basic Helix-Loop-Helix Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D015687:Cell Hypoxia / Q000187:drug effects; D045744:Cell Line, Tumor; D002642:Chick Embryo; D003676:Deferoxamine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors* / Q000235:genetics; D009389:Neovascularization, Pathologic / Q000473:pathology*; D009447:Neuroblastoma / Q000098:blood supply* / Q000235:genetics / Q000473:pathology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D013997:Time Factors; D019772:Topotecan / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics; D000099162:Endothelial PAS Domain-Containing Protein 1
D000970:Antineoplastic Agents; D051792:Basic Helix-Loop-Helix Proteins; D003676:Deferoxamine; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D019772:Topotecan; D042461:Vascular Endothelial Growth Factor A; D000099162:Endothelial PAS Domain-Containing Protein 1