Dual Inhibition of PI3K-AKT-mTOR- and RAF-MEK-ERK-signaling is synergistic in cholangiocarcinoma and reverses acquired resistance to MEK-inhibitors (original) (raw)
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Journal of Cancer Research and Clinical Oncology, 2010
Purpose Hilar cholangiocarcinoma (Klatskin tumor) is one of the most difficult cancers to treat. We demonstrate activation of phosphoinositide-3-kinase (PI3K)/Akt signaling, which is a critical pathway for cell survival, in hilar cholangiocarcinoma cells. However, inhibition of PI3K has little effect on hilar cholangiocarcinoma cell survival. In this study, we investigated the mechanism by which hilar cholangiocarcinoma cells resist PI3K inhibitors. Methods Human hilar cholangiocarcinoma cells KKU-100 were treated with PI3K inhibitors, and cell viability and apoptosis assays were performed. The expression of a MAPK phosphatase (MKP-1) that contributes to cancer cell survival in response to multiple stress stimuli was assayed by quantitative real-time RT-PCR and western blotting. In addition, the effects of the MKP-1 inhibitor were studied in KKU-100 cells treated with PI3K inhibitors. Results Incubation of KKU-100 cells with PI3K inhibitors resulted in increased expression of MKP-1. Furthermore, we found that inhibition of MKP-1 using siRNA silencing sensitized KKU-100 cells to PI3K inhibitor-induced apoptosis via increased phosphorylation of p38 MAPK. Conclusions These results indicate that concurrent inhibition of PI3K and MKP-1 induces apoptosis in KKU-100 cells. Simultaneous targeting of the PI3K pathway and MKP-1 may be a useful approach to improve therapies directed against hilar cholangiocarcinoma.
Journal of Cancer, 2015
Hepatocellular carcinoma (HCC) is the sixth most common cancer, and the third most common cause of cancer related death worldwide. The multi-kinase inhibitor Sorafenib represents the only systemic treatment option until today, and results from clinical trials with allosteric mTOR inhibitors were sobering. Since the PI3K/AKT/mTOR and RAF/MEK/ERK signaling pathways are frequently upregulated in HCC, we have analyzed the effects of AKT inhibitor MK-2206, MEK inhibitor AZD6244 (ARRY 142886) and mTOR kinase inhibitor AZD8055, given as single drugs or in combination, on proliferation and apoptosis of three HCC cell lines in vitro. We show that all three inhibitor combinations synergistically inhibit proliferation of the three HCC cell lines, with the strongest synergistic effect observed after vertical inhibition of AKT and mTORC1/2. We demonstrate that AKT kinase activity is restored 24h after blockade of mTORC1/2 by increased phosphorylation of T308, providing a rationale for combined targeting of AKT and mTOR inhibition in HCC. Our data suggest that a combination of inhibitors targeting those respective pathways may be a viable approach for future application in patients with hepatocellular carcinoma.
Disease models & mechanisms, 2018
Although gemcitabine plus cisplatin is the gold standard chemotherapy regimen for advanced cholangiocarcinoma, the response rate has been disappointing. This study aims to investigate a novel therapeutic regimen [gemcitabine plus everolimus (rad001), an mTOR inhibitor] for cholangiocarcinoma. Gemcitabine, oxaliplatin, cetuximab and rad001 in various combinations were first evaluated using six cholangiocarcinoma cell lines. therapeutic efficacies of gemcitabine and rad001 alone and their combination were further evaluated using a xenograft mouse model and a chemically induced orthotopic cholangiocarcinoma rat model. In the study, gemcitabine plus rad001 exerted a synergistic therapeutic effect on the cholangiocarcinoma cells, irrespective of the mutation status. In the xenograft study, gemcitabine plus rad001 showed the best therapeutic effect on tumor volume change, and was associated with increased caspase-3 expression, decreased eIF4E expression, as well as overexpression of both ...
Therapeutic targeting of ARID1A and PI3K/AKT pathway alterations in cholangiocarcinoma
PeerJ
Background Genetic alterations in ARID1A were detected at a high frequency in cholangiocarcinoma (CCA). Growing evidence indicates that the loss of ARID1A expression leads to activation of the PI3K/AKT pathway and increasing sensitivity of ARID1A-deficient cells for treatment with the PI3K/AKT inhibitor. Therefore, we investigated the association between genetic alterations of ARID1A and the PI3K/AKT pathway and evaluated the effect of AKT inhibition on ARID1A-deficient CCA cells. Methods Alterations of ARID1A, PI3K/AKT pathway-related genes, clinicopathological data and overall survival of 795 CCA patients were retrieved from cBio Cancer Genomics Portal (cBioPortal) databases. The association between genetic alterations and clinical data were analyzed. The effect of the AKT inhibitor (MK-2206) on ARID1A-deficient CCA cell lines and stable ARID1A-knockdown cell lines was investigated. Cell viability, apoptosis, and expression of AKT signaling were analyzed using an MTT assay, flow c...
Inhibition of PI3K increases oxaliplatin sensitivity in cholangiocarcinoma cells
Cancer Cell International, 2009
Background Resistance of cholangiocarcinoma to chemotherapy is a major problem in cancer treatment. The mechanism of resistance is believed to involve phosphoinositide-3- kinase (PI3K)/Akt activation. Although the platinum-containing compound oxaliplatin has been extensively used in the treatment of several solid tumors, recent data regarding its use to treat cholangiocarcinoma are ambiguous. Oxaliplatin resistance in this disease could potentially involve PI3K pathways. We, therefore, examined the effects of PI3K pathways in cholangiocarcinoma cells in modulating oxaliplatin resistance. Results After exposing the cholangiocarcinoma cell lines RMCCA1 and KKU100 to oxaliplatin, the levels of Akt and mTOR phosphorylation increased, as shown by western blot analysis. The WST-1 cell proliferation assay showed increased inhibition of cell growth under high concentrations of oxaliplatin. The combination of oxaliplatin with LY294002, an inhibitor of PI3K, resulted in a remarkable arrest of cell proliferation. Deactivation of mTOR by RAD001 was also synergistic with oxaliplatin, although to a lesser extent. The combination of oxaliplatin and a PI3K inhibitor also resulted in a significant induction of apoptosis, as demonstrated by the TUNEL assay. Conclusion Activation of PI3K might protect cholangiocarcinoma cells from oxaliplatininduced cytotoxicity. Although the inhibition of PI3K and the inhibition of mTOR both enhance oxaliplatin-induced cytotoxicity, PI3K inhibition has a greater effect. Targeting the PI3K pathway may be a useful approach to improve the chemotherapeutic sensitivity of cholangiocarcinoma.
Combined CDK4/6 and pan-mTOR inhibition is synergistic against intrahepatic cholangiocarcinoma
Clinical cancer research : an official journal of the American Association for Cancer Research, 2018
Intrahepatic cholangiocarcinoma (ICC) is an aggressive cancer type, lacking effective therapies and associated with a dismal prognosis. Palbociclib is a selective CDK4/6 inhibitor, which has been shown to suppress cell proliferation in many experimental cancer models. Recently, we demonstrated that pan-mTOR inhibitors, such as MLN0128, effectively induce apoptosis, while having limited efficacy in restraining proliferation of ICC cells. Here, we tested the hypothesis that Palbociclib, due to its ant-proliferative properties in many cancer types, might synergize with MLN0128 to impair ICC growth. Human ICC cell lines and the AKT/YapS127A ICC mouse model were used to test the therapeutic efficacy of Palbociclib and MLN0128, either alone or in combination. Administration of Palbociclib suppressed ICC cell growth by inhibiting cell cycle progression. Concomitant administration of Palbociclib and MLN0128 led to a pronounced, synergistic growth constraint of ICC cell lines. Furthermore, w...
International Journal of Cancer, 2009
Mutations in KRAS or BRAF frequently manifest in constitutive activation of the MEK1/2-ERK1/2 signalling pathway. The MEK1/2-selective inhibitor, AZD6244 (ARRY-142886), blocks ERK1/2 activation and is currently undergoing clinical evaluation. Tumour cells can vary markedly in their response to MAPK or ERK kinase (MEK) inhibitors, and the presence of a BRAF mutation is thought to predict sensitivity, with the RAS mutations being associated with intrinsic resistance. We analysed cell proliferation in a panel of 19 colorectal cancer cell lines and found no simple correlation between BRAF or KRAS mutation and sensitivity to AZD6244, though cells that harbour neither mutation tended to be resistant. Cells that were sensitive arrested in G 1 and/or underwent apoptosis and the presence of BRAF or KRAS mutation was not sufficient to predict either fate. Cell lines that were resistant to AZD6244 exhibited low or no ERK1/2 activation or exhibited coincident activation of ERK1/2 and protein kinase B (PKB), the latter indicative of activation of the PI3K pathway. In cell lines with coincident ERK1/2 and PKB activation, sensitivity to AZD6244 could be re-imposed by any of the 3 distinct PI3K/ mTOR inhibitors. We conclude that AZD6244 is effective in colorectal cancer cell lines with BRAF or KRAS mutations. Sensitivity to MEK1/2 inhibition correlates with a biochemical signature; those cells with high ERK1/2 activity (whether mutant for BRAF or KRAS) evolve a dependency upon that pathway and tend to be sensitive to AZD6244 but this can be offset by high PI3K-dependent signalling. This may have implications for the use of MEK inhibitors in combination with PI3K inhibitors. ' 2009 UICC Mutant RAS proteins found in human cancer 1 can interact with a variety of effector signalling pathways to control cell fate, including the RAF-MEK1/2-ERK1/2 pathway. 2,3 Activated mutants of BRAF are found in a number of tumour types, including 70% of melanomas, 30% of papillary thyroid tumours and 10% of colorectal tumours. 4,5 The BRAF 600E mutant can replicate many of the effects of mutant RAS; indeed, expression of endogenous BRAF 600E promotes anchorage-independent growth, morphologic transformation, cell proliferation and cell survival in primary mouse embryo fibroblasts. 6-11 As the only physiologically defined substrate of RAF, MEK1/2 plays a key role in mediating the effects of RAS and RAF. 12,13 Since genetic inhibition of the ERK1/2 pathway reverses RAS and RAF transformation, and pharmacological inhibition of MEK1/2 is feasible, 14 this pathway has attracted much attention in the search for new chemotherapeutics and selective small molecule MEK1/2 inhibitors have been identified, including PD184352 (CI-1040), PD0325901 and AZD6244 (ARRY-142886). Tumour cells can vary markedly in their response to MEK1/2 inhibition, and RAS mutation status alone is not sufficient as a prognostic marker for responses. This likely reflects the fact that RAS can activate multiple effectors including the class I PI3Kinases, which activate the phosphoinositide dependent kinase-1 (PDK1)-protein kinase B (PKB) pathway. 2,20 For example, both the RAF and PI3K pathways can promote expression of cyclin D1 and down-regulation of p27 KIP1 , 21,22 albeit by different mechanisms, and both pathways can promote cell survival. Consequently, tumour cells with RAS mutations might be less sensitive to mitogen-activated protein kinase (MAPK) or ERK kinase (MEK) inhibitors because of compensatory signalling by parallel non-extracellular signal-regulated kinase (ERK) effector pathways.
Liver International, 2019
Cholangiocarcinoma (CCA) is a deadly disease. While surgery may attain cure in a minor fraction of cases, therapeutic options in either the adjuvant or advanced setting are limited. The possibility of advancing the efficacy of therapeutic approaches to CCA relies on understanding its molecular pathogenesis and developing rational therapies aimed at interfering with oncogenic signaling networks that drive and sustain cholangiocarcinogenesis. These efforts are complicated by the intricate biology of CCA, which integrates not only the driving force of tumor-cell-intrinsic alterations at the genetic and epigenetic level, but also pro-tumorigenic cues conveyed to CCA cells by different cell types present in the rich tumor stroma. Herein, we review our current understanding of the Accepted Article mechanistic bases underpinning the activation of major oncogenic pathways causative of CCA pathogenesis. We subsequently discuss how this knowledge is being exploited to implement rationale-based and genotype-matched therapeutic approaches that predictably will radically transform CCA clinical management in the next decade. We conclude by highlighting mechanisms of therapeutic resistance in CCA and reviewing innovative approaches to combat resistance at the pre-clinical and clinical level. KEY POINTS Cholangiocarcinoma (CCA) is a deadly cancer worldwide as a result of limited therapeutic options and chemoresistance. CCA pathogenesis is associated with genetic and epigenetic alterations in tumor cells as well as important changes in the tumor microenvironment, which, collectively, lead to the activation of multiple signaling pathways responsible for driving tumor onset and progression. These pathways are linked to the control of cell proliferation, cell survival/death, metabolism, tissue morphogenesis and inflammation. A better characterization of the molecular mechanisms involved in CCA pathogenesis and chemoresistance is predicted to pave the way to the rational design of innovative therapies and to the prevention/bypass of chemoresistance. INTRODUCTORY STATEMENT Cholangiocarcinoma (CCA), the second most frequent primary liver cancer, is characterized by high mortality, clinical silence at early stages and rapid disease development and progression 1. The unfavorable clinical history of the disease is largely caused by the aggressive biology of the malignancy, the nature and mechanisms of which are still largely
Evolution of Treatment in Advanced Cholangiocarcinoma: Old and New towards Precision Oncology
International Journal of Molecular Sciences
Cholangiocarcinoma (CCA) is a malignant neoplasm arising in the epithelium of the biliary tract. It represents the second most common primary liver cancer in the world, after hepatocellular carcinoma, and it constitutes 10–15% of hepatobiliary neoplasms and 3% of all gastrointestinal tumors. As in other types of cancers, recent studies have revealed genetic alterations underlying the establishment and progression of CCA. The most frequently involved genes are APC, ARID1A, AXIN1, BAP1, EGFR, FGFRs, IDH1/2, RAS, SMAD4, and TP53. Actionable targets include alterations of FGFRs, IDH1/2, BRAF, NTRK, and HER2. “Precision oncology” is emerging as a promising approach for CCA, and it is possible to inhibit the altered function of these genes with molecularly oriented drugs (pemigatinib, ivosidenib, vemurafenib, larotrectinib, and trastuzumab). In this review, we provide an overview of new biologic drugs (their structures, mechanisms of action, and toxicities) to treat metastatic CCA, provid...