Ive activity, weaker apoptosis, and richer neovascularization than AFP-negative gastric cancers. On the other hand, high levels of AFP in fully developed hepatocarcinoma or in serum of the host are associated with more aggressive behavior, and increased 22948146 anaplasia [37,38]. Studies of AFP knockdown by siRNA found inhibited cell proliferation in hepatomas [39]. Therefore, AFP may function in a fundamental step in the progression of AFP-positive cancer. Downregulation of AFP expression may represent a relevant therapeutic strategy. We found that As2O3 could downregulate AFP mRNA and protein expression. Also, downregulation of AFP by As2O3 could inhibit cell proliferation and induce cell apoptosis in AFPGC FU97 cells. Moreover, AFP secretion in As2O3-treated cells was dose- and time-dependently decreased in the supernatant. Downregulation of AFP expression might contribute to As2O3-induced inhibition of cell growth and apoptosis. Thus, these data indicated that AFP expression is downregulated in response to As2O3 treatment. AFP may play an important role in the proliferation and apoptosis of AFPGC. In addition to being a point of convergence for numerous oncogenic signaling pathways, STAT3 also participates 
in cell growth and survival. In leukemia cells, As2O3 activates numerousNovel Therapy for AFP-Producing Gastric CancersFigure 6. Kaplan eier survival curves and Dimethylenastron custom synthesis log-rank test for patients with AFP-positive gastric cancer (AFPGC) stratified by AFP and STAT3 expression. (A) AFP positivity alone. (B) STAT3 positivity alone. (C) AFP and STAT3 double positivity compared with AFP positivity. (D) AFP and STAT3 double positivity compared with STAT3 positivity(all P,0.05). doi:10.1371/journal.pone.0054774.gFigure 7. Schematic illustration of As2O3-induced growth inhibition and apoptosis of FU97 cells. Inactivation of the ATBF1 gene in AFPGC, through mutation or reduced expression, may allow AFPGC cells to produce AFP protein and overexpress STAT3, which contributes to aggressive behavior and poor prognosis of AFPGC. As2O3 can inhibit AFPGC cell growth and induce cell apoptosis. The underlying mechanisms may involve downregulation of AFP and STAT3 expression and STAT3 downregulating the expression of anti-apoptotic Bcl-2 and upregulating that of the tumor suppressor Bax. Furthermore, AFP can dimerize with other proteins such as nuclear receptors, transcription factors and caspases, all of which can promote growth of tumor cells. AFP may dimerize with the transcription factor STAT3 to promote AFPGC growth. Therefore, AFP may interact with STAT3 in the signal pathway for chemotherapeutic efficiency of agents on AFPGC. doi:10.1371/journal.pone.0054774.gNovel Therapy for AFP-Producing Gastric Cancersintracellular signal 
transduction pathways, thus resulting in induction of apoptosis [40]. As2O3 inhibition of STAT3, before inhibition of cellular proliferation, has been described in multiple myeloma cells [41]. Also, As2O3 inhibits protein tyrosine kinase, thereby indirectly decreasing activation of STAT proteins [42].Therefore, downregulation of STAT3 has been purchase TBHQ considered one of the mechanisms of action of As2O3 in acute promyelocytic leukemia(APL).We found STAT3 activated in AFPGC cells, and As2O3 could downregulate STAT3 mRNA expression and STAT3 and pSTAT3 protein expression. Especially, downregulated expression of STAT3 and pSTAT3 was consistent with downregulated expression of AFP by As2O3. STAT3 might be inhibited by some factor during its activati.Ive activity, weaker apoptosis, and richer neovascularization than AFP-negative gastric cancers. On the other hand, high levels of AFP in fully developed hepatocarcinoma or in serum of the host are associated with more aggressive behavior, and increased 22948146 anaplasia [37,38]. Studies of AFP knockdown by siRNA found inhibited cell proliferation in hepatomas [39]. Therefore, AFP may function in a fundamental step in the progression of AFP-positive cancer. Downregulation of AFP expression may represent a relevant therapeutic strategy. We found that As2O3 could downregulate AFP mRNA and protein expression. Also, downregulation of AFP by As2O3 could inhibit cell proliferation and induce cell apoptosis in AFPGC FU97 cells. Moreover, AFP secretion in As2O3-treated cells was dose- and time-dependently decreased in the supernatant. Downregulation of AFP expression might contribute to As2O3-induced inhibition of cell growth and apoptosis. Thus, these data indicated that AFP expression is downregulated in response to As2O3 treatment. AFP may play an important role in the proliferation and apoptosis of AFPGC. In addition to being a point of convergence for numerous oncogenic signaling pathways, STAT3 also participates in cell growth and survival. In leukemia cells, As2O3 activates numerousNovel Therapy for AFP-Producing Gastric CancersFigure 6. Kaplan eier survival curves and log-rank test for patients with AFP-positive gastric cancer (AFPGC) stratified by AFP and STAT3 expression. (A) AFP positivity alone. (B) STAT3 positivity alone. (C) AFP and STAT3 double positivity compared with AFP positivity. (D) AFP and STAT3 double positivity compared with STAT3 positivity(all P,0.05). doi:10.1371/journal.pone.0054774.gFigure 7. Schematic illustration of As2O3-induced growth inhibition and apoptosis of FU97 cells. Inactivation of the ATBF1 gene in AFPGC, through mutation or reduced expression, may allow AFPGC cells to produce AFP protein and overexpress STAT3, which contributes to aggressive behavior and poor prognosis of AFPGC. As2O3 can inhibit AFPGC cell growth and induce cell apoptosis. The underlying mechanisms may involve downregulation of AFP and STAT3 expression and STAT3 downregulating the expression of anti-apoptotic Bcl-2 and upregulating that of the tumor suppressor Bax. Furthermore, AFP can dimerize with other proteins such as nuclear receptors, transcription factors and caspases, all of which can promote growth of tumor cells. AFP may dimerize with the transcription factor STAT3 to promote AFPGC growth. Therefore, AFP may interact with STAT3 in the signal pathway for chemotherapeutic efficiency of agents on AFPGC. doi:10.1371/journal.pone.0054774.gNovel Therapy for AFP-Producing Gastric Cancersintracellular signal transduction pathways, thus resulting in induction of apoptosis [40]. As2O3 inhibition of STAT3, before inhibition of cellular proliferation, has been described in multiple myeloma cells [41]. Also, As2O3 inhibits protein tyrosine kinase, thereby indirectly decreasing activation of STAT proteins [42].Therefore, downregulation of STAT3 has been considered one of the mechanisms of action of As2O3 in acute promyelocytic leukemia(APL).We found STAT3 activated in AFPGC cells, and As2O3 could downregulate STAT3 mRNA expression and STAT3 and pSTAT3 protein expression. Especially, downregulated expression of STAT3 and pSTAT3 was consistent with downregulated expression of AFP by As2O3. STAT3 might be inhibited by some factor during its activati.
