Purpose:To clone and construct an eukaryotic expression vector of NOEY2 gene, and to observe the effects of NOEY2 transfection on the growth of human breast cancer cell line MDA MB 231. Methods:The coding region of NOEY2 was obtained with reverse transcription PCR, and then the PCR product was first cloned into pGEM T vector and further directionally subcloned into pcDNA3. Transfection reagent and selective antibiotic were lipofect AMINE and G418 respectively. The expression level of NOEY2 protein was detected by Western Blotting. The growth cures of the transfected and non transfected cells were recorded. The changes in cell cycle were analyzed by flow cytometry. Results:An eukaryotic expression vector of NOEY2 gene was constructed successfully. The cell transfected with NOEY2 showed definite expression of NOEY2, and the controls were negative. The growth of NOEY2 transfected cells was inhibited by 46.3%, compared with the parental cells on the seventh day after seeding. An obvious decrease in S phase and G2/M phase fraction and an increase in the percentage of G0/G1 phase cells were found in NOEY2 transfected cells. Conclusions:NOEY2 Transfection can inhibit the growth rate of MDA MB 231 cells in vitro, probably via the mechanism of G1 arrest. These data support the suggestion that NOEY2 is a tumor suppressive gene, which merits further investigation for its value as a therapy gene.

OBJECTIVETo investigate the expression of NOEY2 gene in breast cancer tissue and its relation to clinicopathological and other molecular parameters.

METHODSThe mRNA expression of NOEY2 gene was monitored in benign and malignant breast lesions by RT-PCR and in situ hybridization (ISH) with transcripted antisense RNA probes. The protein expression of estrogen receptor (ER), Ki67, p27 and p21(WAF1) in the 60 breast cancer lesions was detected by immunohistochemical method.

RESULTSAll 6 benign lesions, and 13 (72.2%) of the 18 breast cancers were NOEY2 positive by RT-PCR. By ISH, positive NOEY2 was obtained in all 10 benign lesions but only in 31 (51.7%) of the 60 breast cancer lesions. The difference was statistically significant (P = 0.025). NOEY2 positive rate tended to decrease with the increase of histological grade. However, NOEY2 expression was negatively correlated with the status of axillary lymph nodes. The positive NOEY2 rate was 75% in those without lymph node metastasis but only 26.7% in those with metastasis (P < 0.001). No correlation with other clinicopathological parameters including ER, Ki67, p27 or p21(WAF1) were found.

CONCLUSIONNOEY2 gene may be related with the pathogenesis of breast cancer. A link between NOEY2 loss expression and the spreading mechanism of breast cancer may possibly exist.

Breast Neoplasms ; metabolism ; Female ; Gene Expression ; Humans ; In Situ Hybridization ; RNA, Messenger ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; rho GTP-Binding Proteins ; biosynthesis ; genetics