Loss function of PITX1 contributes to poor prognosis of gastric cancer patients by enhancing chemotherapy resistance to 5-fluorocytosine and cisplatin

【作者】 沈小慧； Fengchang Qiao； Pihai Gong； Zhujiang Zhao； 樊红；

【Author】 Xiaohui Shen;Fengchang Qiao;Pihai Gong;Zhujiang Zhao;Hong Fan;Department of Medical Genetics and Developmental Biology, Medical School of Southeast University, The Key Laboratory of Developmental Genes and Human Diseases, Ministry of Education, Southeast University;

【机构】 Department of Medical Genetics and Developmental Biology, Medical School of Southeast University, The Key Laboratory of Developmental Genes and Human Diseases, Ministry of Education, Southeast University；

【摘要】 Gastric cancer(GC) is the third leading cause of cancer-related death worldwide, and systemic chemotherapy is the major treatment strategy for advanced GC patients. Paired-like homeodomain transcription factor 1(PITX1) has been implicated as a tumour suppressor in various cancers. In the present study, we found that PITX1 expression was downregulated at the transcriptional level in 44%(14/32) of GC patients. The result of Kaplan-Meier curve indicated that GC patients with lower levels of PITX1 had a worse prognosis than those with higher levels of PITX1(*P = 0.027). Poor prognosis following chemotherapy is the general outcome owing to recurrent resistance. A Cell Counting kit-8 assay was performed to detect the impact of PITX1 expression on the sensitivity of GC cells to 5-fluorocytosine(5?FU) and cisplatin(CDDP). The results revealed that the overexpression of PIXT1 increased the sensitivity of GC cells AGS and BGC-823 to 5?FU/CDDP. Besides, PIXT1 knockdown decreased the sensitivity of GC cells MCG-803 and SGC-7901 to 5?FU/CDDP. To further assess the mechanism by which PITX1 contributes to chemotherapy insensitivity, a total of co-expressed genes, 1620 genes were screened by a KEGG analysis, the biological processes of which were primarily implicated in necroptosis and apoptosis. PDCD5, a cell apoptosis-related gene, was thus a candidate gene according to the results of CHIP-Seq and GO analysis. A luciferase reporter assay showed that the transcription activity of the motif was significantly higher in AGS cells transfected with pPITX1 than in cells transfected with pcDNA3.1 plasmids without the motif region. An EMSA was applied to certify the physical interaction of PITX1 with PDCD5. Furthermore, the expression level of PDCD5 in GC cells was regulated by PITX1 positively. Finally, we found that PDCD5 promoted cell apoptosis in GC cell lines by flow cytometry and Annexin V staining. Collectively, these data indicated that PITX1 enhanced the cytotoxicity of 5?FU and CDDP in GC cells partially by targeting the PDCD5 promoter to induce cell apoptosis.更多还原

【Abstract】 Gastric cancer(GC) is the third leading cause of cancer-related death worldwide, and systemic chemotherapy is the major treatment strategy for advanced GC patients. Paired-like homeodomain transcription factor 1(PITX1) has been implicated as a tumour suppressor in various cancers. In the present study, we found that PITX1 expression was downregulated at the transcriptional level in 44%(14/32) of GC patients. The result of Kaplan-Meier curve indicated that GC patients with lower levels of PITX1 had a worse prognosis than those with higher levels of PITX1(*P = 0.027). Poor prognosis following chemotherapy is the general outcome owing to recurrent resistance. A Cell Counting kit-8 assay was performed to detect the impact of PITX1 expression on the sensitivity of GC cells to 5-fluorocytosine(5?FU) and cisplatin(CDDP). The results revealed that the overexpression of PIXT1 increased the sensitivity of GC cells AGS and BGC-823 to 5?FU/CDDP. Besides, PIXT1 knockdown decreased the sensitivity of GC cells MCG-803 and SGC-7901 to 5?FU/CDDP. To further assess the mechanism by which PITX1 contributes to chemotherapy insensitivity, a total of co-expressed genes, 1620 genes were screened by a KEGG analysis, the biological processes of which were primarily implicated in necroptosis and apoptosis. PDCD5, a cell apoptosis-related gene, was thus a candidate gene according to the results of CHIP-Seq and GO analysis. A luciferase reporter assay showed that the transcription activity of the motif was significantly higher in AGS cells transfected with pPITX1 than in cells transfected with pcDNA3.1 plasmids without the motif region. An EMSA was applied to certify the physical interaction of PITX1 with PDCD5. Furthermore, the expression level of PDCD5 in GC cells was regulated by PITX1 positively. Finally, we found that PDCD5 promoted cell apoptosis in GC cell lines by flow cytometry and Annexin V staining. Collectively, these data indicated that PITX1 enhanced the cytotoxicity of 5?FU and CDDP in GC cells partially by targeting the PDCD5 promoter to induce cell apoptosis.更多还原