ANTISENSE DOWN-REGULATION OF METALLOTHIONEIN INDUCES GROWTH ARREST AND APOPTOSIS IN HUMAN BREAST-CARCINOMA CELLS

Citation
Ab. Abdelmageed et Kc. Agrawal, ANTISENSE DOWN-REGULATION OF METALLOTHIONEIN INDUCES GROWTH ARREST AND APOPTOSIS IN HUMAN BREAST-CARCINOMA CELLS, Cancer gene therapy, 4(3), 1997, pp. 199-207
Citations number
55
Categorie Soggetti
Oncology,"Biothechnology & Applied Migrobiology
Journal title
ISSN journal
09291903
Volume
4
Issue
3
Year of publication
1997
Pages
199 - 207
Database
ISI
SICI code
0929-1903(1997)4:3<199:ADOMIG>2.0.ZU;2-7
Abstract
The association of increased metallothionein (MT) gene expression in b reast cancer with metastasis and poor prognosis has led us to investig ate the hypothesis that inhibition of MT gene expression may elicit an tiproliferative effects in breast carcinoma MCF7 cells. To monitor the effect of downregulation of MT protein on growth, MCF7 cells were tra nsiently transfected by electroporation with an 18-mer MT antisense ph osphorothioate oligomer (AO) or an 18-mer random oligomer (RO). The MT -AO is complementary to the region 7 bases downstream from the AUG tra nslational start site of the hMT-IIA gene. Transfection of MCF7 cells with the AO inhibited cell growth by 50-60% at 72 hours when compared to control cells or the cells transfected with RO. The AO-induced grow th inhibition was associated with alterations in morphology suggestive of apoptotic cell death. This was further confirmed by DNA linker cle avage into oligonucleosomal fragments and decreased bcl-2 protein leve ls in AO-transfected cells as opposed to the RO-transfected cells. Rev erse transcriptase polymerase chain reaction analysis showed that AO i nduced a 2-fold increase in the levels of c-fos and p53 transcripts in comparison to RO which had no significant effect. Conversely, c-myc t ranscripts were decreased by 2.5-fold in the AO-transfected cells when compared to the controls. Furthermore, MCF7 cells transfected with an expression plasmid pBAcNEO-sMT-IIA encompassing human MT-IIA cDNA, co nstitutively driven by beta-actin promotor, caused a 2.5-fold increase in intracellular levels of MT, as judged by PCR and western blot anal ysis, in comparison to the cells transfected with pBAcNEO plasmid. In contrast to the AO-induced growth inhibition, overexpression of cytopl asmic MT increased the cell multiplication by 2-fold compared with con trol cells or the cells transfected with the control plasmid 72 hours post-transfection. Moreover, the effects of AO on oncogene expression were reversed on increased expression of MT. These data suggest that o verexpression of MT potentiates the growth of MCF7 cells, whereas down regulation of MT elicits antiproliferative effects.