Downregulation of matrix metalloproteinase-2 (MMP-2) utilizing adenovirus-mediated transfer of small interfering RNA (siRNA) in a novel spinal metastatic melanoma model.
Matrix metalloproteinases (MMPs) comprise a class of secreted zinc-dependent endopeptidases implicated in the metastatic potential of tumor cells due to their ability to degrade the extracellular matrix (ECM) and basement membrane. Matrix metalloproteinase-2 (MMP-2) has been detected in high levels and correlates with invasiveness in human melanoma. We have studied the effect of adenovirus-mediated transfer of small interfering RNA (siRNA) against MMP-2 in the human melanoma cell line A2058. The delivery of these double-stranded RNA molecules represents an efficient technology in silencing disease-causing genes with known sequences at the post-transcriptional level. siRNA against MMP-2 mRNA (Ad-MMP-2) was found to decrease MMP-2 protein expression and activity in melanoma cells as demonstrated by western blotting and gelatin zymography. Furthermore, infection of cells with Ad-MMP-2 inhibited cellular migration and invasion as indicated by spheroid and matrigel assays. We also observed dose-dependent suppression of vascular network formation in an angiogenesis assay. Finally, we developed a nude mouse spinal metastatic model to investigate the local effects of tumor metastasis. Intravenous tail vein injection with Ad-MMP-2 on days 5, 9 and 11 after tumor implantation resulted in complete retention of neurological function as compared to control and scrambled vector (Ad-SV)-treated groups that showed complete paraplegia by day 14+/-2 days. Hematoxylin and eosin staining revealed decreased tumor size in the Ad-MMP-2-treated animals. This novel experimental model revealed that adenoviral-mediated transfer of RNA interference against MMP-2 results in the retention of neurological function and significantly inhibited tumor growth.
Tsung
Andrew J
AJ
Department of Neurosurgery, University of Illinois College of Medicine, Peoria, IL 61605, USA.
Kargiotis
Odysseas
O
Chetty
Chandramu
C
Lakka
Sajani S
SS
Gujrati
Meena
M
Spomar
Daniel G
DG
Dinh
Dzung H
DH
Rao
Jasti S
JS
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D045744:Cell Line, Tumor; D002465:Cell Movement; D015536:Down-Regulation / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000235:genetics / Q000378:metabolism; D061965:Matrix Metalloproteinase Inhibitors; D008545:Melanoma / Q000235:genetics / Q000473:pathology* / Q000628:therapy; D051379:Mice; D008819:Mice, Nude; D008954:Models, Biological; D009389:Neovascularization, Pathologic / Q000628:therapy; D009474:Neurons / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics* / Q000494:pharmacology*; D013125:Spinal Neoplasms / Q000235:genetics / Q000556:secondary* / Q000628:therapy*; D023041:Xenograft Model Antitumor Assays
D061965:Matrix Metalloproteinase Inhibitors; D034741:RNA, Small Interfering; D020778:Matrix Metalloproteinase 2