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Co-activation of PIK3CA and Yap promotes development of hepatocellular and cholangiocellular tumors in mouse and human liver.

Li X, Tao J, Cigliano A, Sini M, Calderaro J, Azoulay D, Wang C, Liu Y, Jiang L, Evert K, Demartis MI, Ribback S, Utpatel K, Dombrowski F, Evert M, Calvisi DF, Chen X - Oncotarget (2015)

Bottom Line: Activation of the PI3K and Yes-associated protein (Yap) signaling pathways has been independently reported in human hepatocellular carcinoma (HCC).At the molecular level, PIK3CA/Yap tumors were characterized by activation of the mTORC1/2, ERK/MAPK, and Notch pathways.Simultaneous activation of PI3K and Yap pathways frequently occurred in human liver tumor specimens and their combined suppression was highly detrimental for the growth of HCC and CCA cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Hepatobiliary Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, P.R. China.

ABSTRACT
Activation of the PI3K and Yes-associated protein (Yap) signaling pathways has been independently reported in human hepatocellular carcinoma (HCC). However, the oncogenic interactions between these two cascades in hepatocarcinogenesis remain undetermined. To assess the consequences of the crosstalk between the PI3K and Yap pathways along liver carcinogenesis, we generated a mouse model characterized by combined overexpression of activated mutant forms of PIK3CA (PIK3CAH1047R) and Yap (YapS127A) in the mouse liver using hydrodynamic transfection (PIK3CA/Yap). In addition, suppression of PI3K and Yap pathways was conducted in human HCC and cholangiocarcinoma (CCA) cell lines. We found that concomitant activation of PI3K and Yap pathways triggered rapid liver tumor development in mice. Histologically, tumors were pure HCC, CCA, or mixed HCC/CCA. At the molecular level, PIK3CA/Yap tumors were characterized by activation of the mTORC1/2, ERK/MAPK, and Notch pathways. Simultaneous activation of PI3K and Yap pathways frequently occurred in human liver tumor specimens and their combined suppression was highly detrimental for the growth of HCC and CCA cell lines. In conclusion, our study demonstrates the oncogenic cooperation between PI3K and Yap pathways along liver carcinogenesis. The PIK3CA/Yap mouse represents an important preclinical liver tumor model for the development of novel therapeutics against this malignancy.

No MeSH data available.


Related in: MedlinePlus

Suppression of PIK3CA and Yap activity via specific inhibitors is highly detrimental for the growth of human HLF hepatocellular carcinoma (HCC) cell line and the human EGI1 cholangiocarcinoma (CCA) cell line(A) Treatment with the PIK3CA inhibitor, PIK75 (1 μM), or the Yap/TEAD disruptor, Verteporfin (Verte; 2 μM) decreased proliferation (left panel) and induced apoptosis (center panel) in the HLF HCC cell line when compared with control (untreated) and DMSO (solvent) treated cells. Of note, combined administration of PIK75 and Verteporfin further decreased the proliferation rate of HLF cells without further augmenting apoptosis. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in HLF cells were assessed by Western blot analysis (right panel). (B) A similar growth restraint patterns as those described in (A) was also detected when the EGI1 CCA cell line was subjected to the administration of the two inhibitors, either alone or in combination. Once again, the additive effects of the two drugs affected only the proliferation rate but not the apoptosis activity in EGI1 cells. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in EGI1 cells were assessed by Western blot analysis (right panel). Each bar represent mean ± SD of three independent experiments conducted in triplicate. Tukey-Kramer's test: P at least < 0.001 a, versus control (untreated cells); b, versus DMSO (solvent); c, versus PIK75; d, versus Verteporfin. Abbreviation: Verte, Verteporfin.
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Figure 8: Suppression of PIK3CA and Yap activity via specific inhibitors is highly detrimental for the growth of human HLF hepatocellular carcinoma (HCC) cell line and the human EGI1 cholangiocarcinoma (CCA) cell line(A) Treatment with the PIK3CA inhibitor, PIK75 (1 μM), or the Yap/TEAD disruptor, Verteporfin (Verte; 2 μM) decreased proliferation (left panel) and induced apoptosis (center panel) in the HLF HCC cell line when compared with control (untreated) and DMSO (solvent) treated cells. Of note, combined administration of PIK75 and Verteporfin further decreased the proliferation rate of HLF cells without further augmenting apoptosis. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in HLF cells were assessed by Western blot analysis (right panel). (B) A similar growth restraint patterns as those described in (A) was also detected when the EGI1 CCA cell line was subjected to the administration of the two inhibitors, either alone or in combination. Once again, the additive effects of the two drugs affected only the proliferation rate but not the apoptosis activity in EGI1 cells. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in EGI1 cells were assessed by Western blot analysis (right panel). Each bar represent mean ± SD of three independent experiments conducted in triplicate. Tukey-Kramer's test: P at least < 0.001 a, versus control (untreated cells); b, versus DMSO (solvent); c, versus PIK75; d, versus Verteporfin. Abbreviation: Verte, Verteporfin.

Mentions: Finally, we assessed the importance of the PI3K and Yap on the in vitro growth of human HCC and CCA cell lines. For this purpose, the PIK3CA specific inhibitor, PIK75 [27], and the disruptor of Yap-TEAD interaction, Verteporfin [28], were applied either alone or in combination in HLF and SK/Hep1 HCC cell lines and the EGI1 CCA cell line (Figure 8A and B, Supplementary Figure 5). Treatment with the two inhibitors alone resulted in a strong decrease of proliferation and induction of apoptosis in the three cell lines. A further reduction of proliferation was detected in the three cell lines when the two drugs were administered combinatorially, whereas no additive effects on apoptosis were observed (Figure 8A and B, Supplementary Figure 5). At the molecular level, inhibition of PIK3CA activity by PIK75 resulted, as expected, in the downregulation of PIK3CA canonical targets, such as phosphorylated NDRG1 and phosphorylated/inactivated 4EBP1 in HLF and EGI1 cell lines (Figure 8A and B). Of note, PIK75 administration was followed by decreased levels of Yap and connective tissue growth factor (CTGF), a Yap target, in both HLF and EGI1 cells (Figure 8A and 8B). Treatment with Yap/TEAD disruptor, Verteporfin, led instead to the reduction of Yap and CTGF levels in both HLF and EGI1 cell lines, whereas Verteporfin administration triggered downregulation of the PIK3CA targets, namely phosphorylated NDRG1 and phosphorylated/inactivated 4EBP1, only in HLF cells (Figure 8A and B). Altogether, the present data indicate that simultaneous inhibition of the PIK3CA and Yap cascades is extremely harmful for the in vitro growth of HCC and CCA cells.


Co-activation of PIK3CA and Yap promotes development of hepatocellular and cholangiocellular tumors in mouse and human liver.

Li X, Tao J, Cigliano A, Sini M, Calderaro J, Azoulay D, Wang C, Liu Y, Jiang L, Evert K, Demartis MI, Ribback S, Utpatel K, Dombrowski F, Evert M, Calvisi DF, Chen X - Oncotarget (2015)

Suppression of PIK3CA and Yap activity via specific inhibitors is highly detrimental for the growth of human HLF hepatocellular carcinoma (HCC) cell line and the human EGI1 cholangiocarcinoma (CCA) cell line(A) Treatment with the PIK3CA inhibitor, PIK75 (1 μM), or the Yap/TEAD disruptor, Verteporfin (Verte; 2 μM) decreased proliferation (left panel) and induced apoptosis (center panel) in the HLF HCC cell line when compared with control (untreated) and DMSO (solvent) treated cells. Of note, combined administration of PIK75 and Verteporfin further decreased the proliferation rate of HLF cells without further augmenting apoptosis. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in HLF cells were assessed by Western blot analysis (right panel). (B) A similar growth restraint patterns as those described in (A) was also detected when the EGI1 CCA cell line was subjected to the administration of the two inhibitors, either alone or in combination. Once again, the additive effects of the two drugs affected only the proliferation rate but not the apoptosis activity in EGI1 cells. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in EGI1 cells were assessed by Western blot analysis (right panel). Each bar represent mean ± SD of three independent experiments conducted in triplicate. Tukey-Kramer's test: P at least < 0.001 a, versus control (untreated cells); b, versus DMSO (solvent); c, versus PIK75; d, versus Verteporfin. Abbreviation: Verte, Verteporfin.
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Figure 8: Suppression of PIK3CA and Yap activity via specific inhibitors is highly detrimental for the growth of human HLF hepatocellular carcinoma (HCC) cell line and the human EGI1 cholangiocarcinoma (CCA) cell line(A) Treatment with the PIK3CA inhibitor, PIK75 (1 μM), or the Yap/TEAD disruptor, Verteporfin (Verte; 2 μM) decreased proliferation (left panel) and induced apoptosis (center panel) in the HLF HCC cell line when compared with control (untreated) and DMSO (solvent) treated cells. Of note, combined administration of PIK75 and Verteporfin further decreased the proliferation rate of HLF cells without further augmenting apoptosis. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in HLF cells were assessed by Western blot analysis (right panel). (B) A similar growth restraint patterns as those described in (A) was also detected when the EGI1 CCA cell line was subjected to the administration of the two inhibitors, either alone or in combination. Once again, the additive effects of the two drugs affected only the proliferation rate but not the apoptosis activity in EGI1 cells. The effects of PIK75 and Verteporfin treatment on PIK3CA targets (phosphorylated-NDRG1 and phosphorylated/inactivated 4EBP1) as wells as on Yap and its effector, CTGF, in EGI1 cells were assessed by Western blot analysis (right panel). Each bar represent mean ± SD of three independent experiments conducted in triplicate. Tukey-Kramer's test: P at least < 0.001 a, versus control (untreated cells); b, versus DMSO (solvent); c, versus PIK75; d, versus Verteporfin. Abbreviation: Verte, Verteporfin.
Mentions: Finally, we assessed the importance of the PI3K and Yap on the in vitro growth of human HCC and CCA cell lines. For this purpose, the PIK3CA specific inhibitor, PIK75 [27], and the disruptor of Yap-TEAD interaction, Verteporfin [28], were applied either alone or in combination in HLF and SK/Hep1 HCC cell lines and the EGI1 CCA cell line (Figure 8A and B, Supplementary Figure 5). Treatment with the two inhibitors alone resulted in a strong decrease of proliferation and induction of apoptosis in the three cell lines. A further reduction of proliferation was detected in the three cell lines when the two drugs were administered combinatorially, whereas no additive effects on apoptosis were observed (Figure 8A and B, Supplementary Figure 5). At the molecular level, inhibition of PIK3CA activity by PIK75 resulted, as expected, in the downregulation of PIK3CA canonical targets, such as phosphorylated NDRG1 and phosphorylated/inactivated 4EBP1 in HLF and EGI1 cell lines (Figure 8A and B). Of note, PIK75 administration was followed by decreased levels of Yap and connective tissue growth factor (CTGF), a Yap target, in both HLF and EGI1 cells (Figure 8A and 8B). Treatment with Yap/TEAD disruptor, Verteporfin, led instead to the reduction of Yap and CTGF levels in both HLF and EGI1 cell lines, whereas Verteporfin administration triggered downregulation of the PIK3CA targets, namely phosphorylated NDRG1 and phosphorylated/inactivated 4EBP1, only in HLF cells (Figure 8A and B). Altogether, the present data indicate that simultaneous inhibition of the PIK3CA and Yap cascades is extremely harmful for the in vitro growth of HCC and CCA cells.

Bottom Line: Activation of the PI3K and Yes-associated protein (Yap) signaling pathways has been independently reported in human hepatocellular carcinoma (HCC).At the molecular level, PIK3CA/Yap tumors were characterized by activation of the mTORC1/2, ERK/MAPK, and Notch pathways.Simultaneous activation of PI3K and Yap pathways frequently occurred in human liver tumor specimens and their combined suppression was highly detrimental for the growth of HCC and CCA cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Hepatobiliary Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, P.R. China.

ABSTRACT
Activation of the PI3K and Yes-associated protein (Yap) signaling pathways has been independently reported in human hepatocellular carcinoma (HCC). However, the oncogenic interactions between these two cascades in hepatocarcinogenesis remain undetermined. To assess the consequences of the crosstalk between the PI3K and Yap pathways along liver carcinogenesis, we generated a mouse model characterized by combined overexpression of activated mutant forms of PIK3CA (PIK3CAH1047R) and Yap (YapS127A) in the mouse liver using hydrodynamic transfection (PIK3CA/Yap). In addition, suppression of PI3K and Yap pathways was conducted in human HCC and cholangiocarcinoma (CCA) cell lines. We found that concomitant activation of PI3K and Yap pathways triggered rapid liver tumor development in mice. Histologically, tumors were pure HCC, CCA, or mixed HCC/CCA. At the molecular level, PIK3CA/Yap tumors were characterized by activation of the mTORC1/2, ERK/MAPK, and Notch pathways. Simultaneous activation of PI3K and Yap pathways frequently occurred in human liver tumor specimens and their combined suppression was highly detrimental for the growth of HCC and CCA cell lines. In conclusion, our study demonstrates the oncogenic cooperation between PI3K and Yap pathways along liver carcinogenesis. The PIK3CA/Yap mouse represents an important preclinical liver tumor model for the development of novel therapeutics against this malignancy.

No MeSH data available.


Related in: MedlinePlus