SPAG6, which is a novel cancer-testis antigen, is overexpressed in myeloid malignancies. Previously, SPAG6 was found in UPD (uniparental disomy) region of myeloid cell DNA from MDS patients and reported that SPAG6 may be a predictive marker of minimal residual disease in pediatric acute myeloid, but the biological role of SPAG6 in myeloid malignancies remains unclear. The present study was undertaken to determine the expression and functional significance of SPAG6 in malignant myeloid hematologic cell lines. A short hairpin RNA (shRNA) targeting SPAG6 was designed that could specifically inhibit SPAG6 expression at the mRNA and protein levels when introduced into the malignant myeloid hematologic cell lines SKM-1 and K562. The results from flow cytometry and CCK-8 assays showed that SPAG6 silencing inhibited the proliferation of SKM-1/K562 by inducing apoptosis. Furthermore, SPAG6 silencing resulted in activation of caspase-3, -9 and -8 and upregulated the mRNA and protein expression of p53 and PTEN. Then, we subcutaneously inoculated the monoclonal cells into NOD/SCID mice to establish xenograft models, and we found that the SPAG6-shRNA lentivirus dramatically inhibited tumor growth and increased apoptosis in vivo. These findings demonstrate that SPAG6 might have a role in malignant myeloid hematologic cell proliferation and apoptosis by regulating caspase proteins and p53, suggesting that SPAG6 may be a potential therapeutic target.

Myelodysplastic syndromes (MDSs) are a group of malignant clone hematopoietic stem-cell diseases, and the evolution and progression of MDS depend on the abnormal apoptosis of bone marrow cells. Our previous studies have indicated that sperm-associated antigen 6 (SPAG6), located in the uniparental disomy regions of myeloid cells, is overexpressed in patients with MDS as compared to controls, and SPAG6 can inhibit apoptosis of SKM-1. However, the concrete mechanism is still unclear. In the present study, it was found that the TNF-related apoptosis-inducing ligand (TRAIL)signal pathway was activated when the expression of SPAG6 was inhibited by SPAG6-shRNA lentivirus in SKM-1 cells. Additionally, the results of flow cytometry, Cell Counting Kit-8 assay and western blot analysis implied that the TRAIL signal pathway could be inhibited by a high expression of SPAG6. However, SPAG6 cannot influence the expression of TRAIL death receptors, except for FADD. Additionally the interaction between FADD and TRAIL death receptors also increased in SKM-1 cells infected with SPAG6-shRNA lentivirus. Thus, our study demonstrates that SPAG6 may regulate apoptosis in SKM-1 through the TRAIL signal pathway, indicating that SPAG6 could be a potential therapeutic target.

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