A soluble truncated cadherin induces breast cancer cell apoptosis and growth inhibition

摘要

Purpose To identify the characteristics and function of the truncated cadherin cDNA which encodes a soluble molecule containing the sequence of VE-cadherin extracellular domain repeats from repeat 1 to 4 (designated as CED1–4) and a secreting signal peptide at N terminal. Methods A pMSCV/CED1–4 vector was constructed. Recombinant retrovirus ReCED1–4 and ReEmpty were produced by 293 package cells and transfected into MDA-MB435 human breast cancer cells. The expression of CED1–4 in transfectants and their supernatant was analyzed by RT-PCR and Western blot, respectively. MDA-MB435 cell proliferation assays were performed in vitro and in vivo. CED-14-induced apoptosis was demonstrated using Annexin V binding, TUNEL and caspase 3 assays. The expression of integrin β1 and c-fos mRNA was detected by RT-PCR. Results The constructed soluble CED1–4 encoded 484 amino acids and a secreting signal peptide (27 amino acids). CED1–4 was expressed by MDA-MB435/CED1–4 cells, and detected in the supernatant of CED1–4 tranfectants. CED1–4 transfection significantly inhibited the growth of MDA-MB435 cells in vitro and in vivo. About 22-fold increase in the early apoptotic cells in MDA-MB435/CED1–4 cells was observed as compared with MDA-MB435/empty cells. Increased activity of caspase 3 in MDA-MB435/CED1–4 cells was more than two times as compared with that of the control cells. Interestingly, integrin β1 transcriptional level in MDA-MB435/CED1–4 cells was down-regulated as compared with control cells. The resistance of fibronectin to CED1–4 apoptotic inducibility was confirmed by detection of caspase 3. The blockage of c-fos transcriptional expression was detected in MDA-MB435/CED1–4 cells. Conclusions The soluble truncated cadherin may be considered an apoptotic inducer and growth inhibitor in the MDA-MB435 breast carcinoma cell line. Down-regulation of integrin β1 and blockage of c-fos expression may be related to CED1–4-induced apoptosis and growth inhibition.