title: Reversal of multidrug resistance in gastric cancer cells by CDX2 downregulation.
abstract: OBJECTIVE
  To explore the role of CDX2 in the multi-drug resistance (MDR) process of gastric cancer in vitro and in vivo.
  
  
  METHODS
  A cisplatin-resistant gastric cancer cell line with stable downregulation of CDX2 was established. mRNA and protein expression levels of CDX2, survivin, cyclin D1, and c-Myc were detected by western blotting and semi-quantitative reverse-transcriptase polymerase chain reaction (RT-PCR). The influence of downregulation of CDX2 on MDR was assessed by measuring IC50 of SGC7901/DDP cells to cisplatin, doxorubicin, and 5-fluorouracil, rate of doxorubicin efflux, apoptosis, and cell cycle progression detected by flow cytometry. In addition, we determined the in vivo effects of CDX2 small interfering RNA (siRNA) on tumor size, and apoptotic cells in tumor tissues were detected by deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling and hematoxylin and eosin staining.
  
  
  RESULTS
  CDX2 siRNA led to downregulation of endogenous CDX2 mRNA (0.31 ± 0.05 vs 1.10 ± 0.51, 0.31 ± 0.05 vs 1.05 ± 0.21, P = 0.003) and protein (0.12 ± 0.08 vs 0.51 ± 0.07, 0.12 ± 0.08 vs 0.55 ± 0.16, P = 2.57 × 10(-4)) expression. It significantly promoted the sensitivity of SGC7901/DDP cells to cisplatin (0.12 ± 0.05 vs 0.33 ± 0.08, 0.12 ± 0.05 vs 0.39 ± 0.15, P = 0.001), doxorubicin (0.52 ± 0.13 vs 4.11 ± 1.25, 0.52 ± 0.13 vs 4.05 ± 1.44, P = 2.81 × 10(-4)), and 5-fluorouracil (0.82 ± 0.13 vs 2.81 ± 0.51, 0.82 ± 0.13 vs 3.28 ± 1.03, P = 1.71 × 10(-4)). Flow cytometry confirmed that the percentage of apoptotic cells increased after CDX2 downregulation (32.15% ± 2.15% vs 17.63% ± 3.16%, 32.15% ± 2.15% vs 19.3% ± 2.25%, P = 1.73 × 10(-6)). This notion was further supported by the observation that downregulation of CDX2 blocked entry into the S-phase of the cell cycle (31.53% ± 3.78% vs 65.05% ± 7.25%, 31.53% ± 3.78% vs 62.27% ± 5.02%, P = 7.55 × 10(-7)). Furthermore, downregulation of CDX2 significantly increased intracellular accumulation of doxorubicin (0.21 ± 0.06 vs 0.41 ± 0.11, 0.21 ± 0.06 vs 0.40 ± 0.08, P = 0.003). In molecular studies, semiquantitative RT-PCR and western blotting revealed that CDX2 downregulation could inhibit expression of c-Myc, survivin and cyclin D1.
  
  
  CONCLUSIONS
  CDX2 may be involved in regulating multiple signaling pathways in reversing MDR, suggesting that CDX2 may represent a novel target for gastric cancer therapy.
authors.affiliation: Department of Surgery, the First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China.
authors.forename: Lin-Hai
authors.initials: LH
authors.lastname: Yan
authors.forename: Xiao-Tong
authors.initials: XT
authors.lastname: Wang
authors.forename: Jie
authors.initials: J
authors.lastname: Yang
authors.forename: Chao
authors.initials: C
authors.lastname: Lian
authors.forename: Fan-Biao
authors.initials: FB
authors.lastname: Kong
authors.forename: Wei-Yuan
authors.initials: WY
authors.lastname: Wei
authors.forename: Wen
authors.initials: W
authors.lastname: Luo
authors.forename: Qiang
authors.initials: Q
authors.lastname: Xiao
authors.forename: Yu-Bo
authors.initials: YB
authors.lastname: Xie
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D000071616:CDX2 Transcription Factor; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000494:pharmacology; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D005472:Fluorouracil / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D013274:Stomach Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D000077022:Survivin; D013997:Time Factors; D014162:Transfection
chemical_list: D000970:Antineoplastic Agents; C107489:BIRC5 protein, human; C530104:CCND1 protein, human; D000071616:CDX2 Transcription Factor; C491270:CDX2 protein, human; D018398:Homeodomain Proteins; D051034:Inhibitor of Apoptosis Proteins; C489427:MYC protein, human; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D000077022:Survivin; D019938:Cyclin D1; D004317:Doxorubicin; D002945:Cisplatin; D005472:Fluorouracil
keywords: Drug resistance; Gastric cancer; Homeobox gene CDX2; Murine model; RNA interference