Reduction of lung metastasis, cell invasion, and adhesion in mouse melanoma by statin-induced blockade of the Rho/Rho-associated coiled-coil-containing protein kinase pathway.
BACKGROUND
Melanomas are highly malignant and have high metastatic potential; hence, there is a need for new therapeutic strategies to prevent cell metastasis. In the present study, we investigated whether statins inhibit tumor cell migration, invasion, adhesion, and metastasis in the B16BL6 mouse melanoma cell line.


METHODS
The cytotoxicity of statins toward the B16BL6 cells were evaluated using a cell viability assay. As an experimental model, B16BL6 cells were intravenously injected into C57BL/6 mice. Cell migration and invasion were assessed using Boyden chamber assays. Cell adhesion analysis was performed using type I collagen-, type IV collagen-, fibronectin-, and laminin-coated plates. The mRNA levels, enzyme activities and protein levels of matrix metalloproteinases (MMPs) were determined using RT-PCR, activity assay kits, and Western blot analysis, respectively; the mRNA and protein levels of vary late antigens (VLAs) were also determined. The effects of statins on signal transduction molecules were determined by western blot analyses.


RESULTS
We found that statins significantly inhibited lung metastasis, cell migration, invasion, and adhesion at concentrations that did not have cytotoxic effects on B16BL6 cells. Statins also inhibited the mRNA expressions and enzymatic activities of matrix metalloproteinases (MMPs). Moreover, they suppressed the mRNA and protein expressions of integrin α2, integrin α4, and integrin α5 and decreased the membrane localization of Rho, and phosphorylated LIM kinase (LIMK) and myosin light chain (MLC).


CONCLUSIONS
The results indicated that statins suppressed the Rho/Rho-associated coiled-coil-containing protein kinase (ROCK) pathways, thereby inhibiting B16BL6 cell migration, invasion, adhesion, and metastasis. Furthermore, they markedly inhibited clinically evident metastasis. Thus, these findings suggest that statins have potential clinical applications for the treatment of tumor cell metastasis.
Kidera
Yasuhiro
Y
Division of Pharmacotherapy, Kinki University School of Pharmacy, Kowakae, Higashi-Osaka 577-8502, Japan.
Tsubaki
Masanobu
M
Yamazoe
Yuzuru
Y
Shoji
Kaori
K
Nakamura
Haruyuki
H
Ogaki
Mitsuhiko
M
Satou
Takao
T
Itoh
Tatsuki
T
Isozaki
Misako
M
Kaneko
Junichi
J
Tanimori
Yoshihiro
Y
Yanae
Masashi
M
Nishida
Shozo
S
D000284:Administration, Oral; D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000494:pharmacology*; D015153:Blotting, Western; D002448:Cell Adhesion / Q000187:drug effects*; D002465:Cell Movement / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology; D005260:Female; D000077340:Fluvastatin; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology; D016023:Integrins / Q000378:metabolism; D054461:Lim Kinases / Q000378:metabolism; D008175:Lung Neoplasms / Q000201:enzymology / Q000517:prevention & control* / Q000556:secondary; D020782:Matrix Metalloproteinases / Q000235:genetics / Q000378:metabolism; D008546:Melanoma, Experimental / Q000188:drug therapy* / Q000201:enzymology / Q000556:secondary; D051379:Mice; D008810:Mice, Inbred C57BL; D018994:Myosin Light Chains / Q000378:metabolism; D009361:Neoplasm Invasiveness; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects*; D019821:Simvastatin / Q000494:pharmacology; D013997:Time Factors; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000970:Antineoplastic Agents; D016326:Extracellular Matrix Proteins; D005229:Fatty Acids, Monounsaturated; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D007211:Indoles; D016023:Integrins; D018994:Myosin Light Chains; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D000077340:Fluvastatin; D019821:Simvastatin; D054461:Lim Kinases; D054460:rho-Associated Kinases; D020782:Matrix Metalloproteinases; D020741:rho GTP-Binding Proteins