A hammerhead ribozyme suppresses expression of hepatocyte growth factor/scatter factor receptor c-MET and reduces migration and invasiveness of breast cancer cells.
OBJECTIVE
Hepatocyte growth factor/scatter factor (HGF/SF), via its receptor c-MET, has been implicated to play a pivotal role in breast cancer development and progression. This study examined a transgene-consisting of a combination of U1snRNA, hammerhead ribozyme, and antisense, designed to inhibit c-met expression-and its impact on the migration and in vitro invasion of breast cancer cells.


METHODS
A hammerhead ribozyme targeting human c-MET was cloned into a modified pZeoU1EcoSpe vector and transfected into breast cancer cells MDA MB 231 and MCF-7 by electroporation. Expression of MET mRNA and protein was determined. Migration and in vitro invasiveness of transfected cells were also analyzed.


RESULTS
Breast cancer cells were transfected with the ribozyme-containing plasmids. Stable transfectants manifested an almost complete loss of MET mRNA and protein, as shown by reverse transcription-PCR, Northern blotting, and Western blotting, respectively, whereas the wild-type plasmid had no effects. Met-ribozyme transfected cells exhibited reduced migration and in vitro invasiveness through extracellular matrix (Matrigel), compared with the wild-type cells and cells transfected with empty plasmid.


CONCLUSIONS
These data show that targeting c-MET by way of a hammerhead ribozyme encoding antisense to c-MET is an effective approach in reducing the invasiveness of breast cancer cells.
Jiang
W G
WG
University Department of Surgery, University of Wales College of Medicine, Cardiff CF14 4XN, United Kingdom. jiangw@cf.ac.uk
Grimshaw
D
D
Lane
J
J
Martin
T A
TA
Abounader
R
R
Laterra
J
J
Mansel
R E
RE
Abounder
R
R
D001483:Base Sequence; D001943:Breast Neoplasms / Q000235:genetics / Q000473:pathology* / Q000628:therapy; D002465:Cell Movement / Q000235:genetics; D016373:DNA, Antisense / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008969:Molecular Sequence Data; D009361:Neoplasm Invasiveness / Q000235:genetics / Q000517:prevention & control; D010957:Plasmids / Q000235:genetics; D019859:Proto-Oncogene Proteins c-met / Q000235:genetics* / Q000378:metabolism; D016337:RNA, Catalytic / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured
D016373:DNA, Antisense; D016337:RNA, Catalytic; D012333:RNA, Messenger; D012342:RNA, Small Nuclear; C405483:U1 small nuclear RNA; D019859:Proto-Oncogene Proteins c-met