[FAK related non-kinase (FRNK) inhibits migration of a human breast carcinoma cell line MDA-MB-435].

BACKGROUND & OBJECTIVE: Over-expression or over-activation of focal adhesion kinase (FAK) correlates with cancer migration. FAK related non-kinase (FRNK) acts as an endogenous inhibitor of FAK, which can compete with FAK for focal adhesion binding sites. This study was designed to determine the inhibitory effects of FRNK gene on migration of a human breast carcinoma cell line MDA-MB-435 and explore the possible mechanisms.

METHODS

The functional fragment of FRNK cDNA was amplified by reverse transcription polymerase chain reaction (RT-PCR) and cloned into pcDNA3.1 vector. The recombinant pcDNA3.1-FRNK was transfected into MDA-MB-435 cells using lipofectamine 2000. The stably transfected cells were selected in a medium containing geneticin (G418). Expression of FRNK in stably transfected MDA-MB-435 cells and MMP-9 in both wild-type and transfected MDA-MB-435 cells was detected by Western blot. The effect of FRNK on cell migration was determined using cell wound healing and Boyden chamber assays, respectively, in vitro.

RESULTS

The recombinant plasmid pcDNA3.1-FRNK was successfully constructed and MDA-MB-435 cells stably transfected with pcDNA3.1-FRNK were obtained. MMP-9 protein expression was inhibited by 73.1% in MDA-MB-435 cells transfected with pcDNA3.1-FRNK compared to wild-type cells. In wound healing study, migrated cell count was significantly lower in MDA-MB-435/FRNK cells (0.35+/-0.02) than that in wild type cells (0.58+/-0.06, P<0.05). In Boyden chamber assay, the number of migrated MDA-MB-435/FRNK cells was (65.15+/-8.56), which was 66.57% of migrated wild type cells (97.86+/-5.44).

CONCLUSIONS

These findings suggest that FRNK may inhibit the migration of the human breast carcinoma cell line MDA-MB-435. And the suppressive effect may be due to the down-regulation of MMP-9 in MDA-MB-435 cells.