scholarly journals Therapeutic targeting of ARID1A and PI3K/AKT pathway alterations in cholangiocarcinoma

PeerJ ◽  
2022 ◽  
Vol 10 ◽  
pp. e12750
Supharada Tessiri ◽  
Anchalee Techasen ◽  
Sarinya Kongpetch ◽  
Achira Namjan ◽  
Watcharin Loilome ◽  

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 cytometry, and Western blots, respectively. Results The analysis of a total of 795 CCA samples revealed that ARID1A alterations significantly co-occurred with mutations of EPHA2 (p < 0.001), PIK3CA (p = 0.047), and LAMA1 (p = 0.024). Among the EPHA2 mutant CCA tumors, 82% of EPHA2 mutant tumors co-occurred with ARID1A truncating mutations. CCA tumors with ARID1A and EPHA2 mutations correlated with better survival compared to tumors with ARID1A mutations alone. We detected that 30% of patients with PIK3CA driver missense mutations harbored ARID1A-truncated mutations and 60% of LAMA1-mutated CCA co-occurred with truncating mutations of ARID1A. Interestingly, ARID1A-deficient CCA cell lines and ARID1A-knockdown CCA cells led to increased sensitivity to treatment with MK-2206 compared to the control. Treatment with MK-2206 induced apoptosis in ARID1A-knockdown KKU-213A and HUCCT1 cell lines and decreased the expression of pAKTS473 and mTOR. Conclusion These findings suggest a dependency of ARID1A-deficient CCA tumors with the activation of the PI3K/AKT-pathway, and that they may be more vulnerable to selective AKT pathway inhibitors which can be used therapeutically.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2830-2830
Eriko Suzuki ◽  
Ali R. Jazirehi ◽  
Benjamin Bonavida

Abstract Rituximab (chimeric anti-CD20 monoclonal antibody) has been used in the treatment of B-NHL. We have reported in vitro that rituximab treatment signals B-NHL cell lines Ramos and Daudi and inhibits both the ERK 1/2 MAPK and NF-κB signaling pathways leading to selective inhibition of Bclxl expression and sensitization to drug-induced apoptosis. The inhibition of the NF-κB signaling pathway by rituximab was shown to be due, in part, to the induction of the Raf Kinase Inhibitor Protein (RKIP) (Jazirehi, et al., 2005 Cancer Research 65:264–276). The PI3K-Akt signaling pathway is a key regulator of cell survival and aberrant activation of the PI3K-Akt pathway has been implicated in both drug resistance and resistance to apoptosis-inducing stimuli. Akt can promote cell survival by indirectly activating the proximal transcription factor NF-κB through the phosphorylation of I-kappa B kinase (I-κB) (Ozes et. al. Nature401:82–85, 1999). This study investigated whether NF-κB inhibition by rituximab and downregulation of Bclxl expression was also the result of rituximab-mediated inhibition of the PI3K-Akt pathway. Ramos and Daudi B-NHL cell lines were treated with rituximab (20 ug/ml) and cell lysates were prepared and both Akt and phospho-Akt (p-Akt) expression were examined by western blot. The findings demonstrate that both cell lines show constitutively activated p-Akt and treatment with rituximab significantly inhibited p-Akt but not Akt. Time kinetics analysis demonstrated that inhibition of p-Akt was first detected at 3–6 hours following rituximab treatment and inhibition was maintained up to 24 hours. Concomitantlly, a similar time kinetics revealed inhibition of NF-κB activity as assessed by EMSA. Since the inhibition of NF-kB activity resulted in significant downregulation of Bclxl expression, we also examined the role of the Akt pathway in the regulation of Bclxl expression. Tumor cells were treated with the Akt inhibitor LY294002 and analysis of cell lysates showed significant downregulation of Bclxl expression. Rituximab was previously shown to sensitize B-NHL cells to drug-induced apoptosis via inhibition of NF-κB activity and Bclxl expression. We examined if inhibition of the Akt pathway also chemosensitized the cells. Treatment of Ramos cells with the Akt inhibitor LY294002 significantly sensitized the cells to CDDP-induced apoptosis and synergy was achieved. Altogether, these findings demonstrate, for the first time, that rituximab inhibits the Akt pathway and that this pathway is involved in the regulation of tumor- cell resistance to chemotherapeutic drugs. This study also proposes that the Akt pathway is a potential targeting pathway for therapeutic intervention in the treatment of rituximab and drug-resistant B-NHL.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2488-2488 ◽  
Xavier Leleu ◽  
Xiaoying Jia ◽  
Anne-Sophie Moreau ◽  
Evdoxia Hatjiharisi ◽  
Hai Ngo ◽  

Abstract Background: Waldenstrom’s Macroglobulinemia (WM) is a low-grade lymphoplasmacytic lymphoma with limited options of therapy. The PI3k/Akt pathway is a critical regulator of cell survival. Our previous studies using proteomic analysis have demonstrated upregulation of members of the PI3k/Akt pathway in WM. We examined whether the new Akt inhibitor perifosine (NSC 639966; Keryx, NY) induces cytotoxicity in WM. Methods: WM cell lines (BCWM1 and WSU-WM) and IgM secreting low-grade lymphoma cell lines (MEC1, RL) were used. Primary CD19+ malignant cells were obtained from patients after informed consent. Inhibition of proliferation was measured using the MTT assay; DNA synthesis was measured using the thymidine uptake assay and apoptosis using Apo2.7 flow cytometry. Bone marrow stromal cells (BMSC) confer growth and resistance to conventional treatments. We therefore, tested the effect of perifosine on WM cells co-cultured with BMSC. Immunoblotting for signaling pathways was performed at different time (30 minutes to 16 hrs) and doses of therapy. In vivo activity of perifosine was assessed using a SCID-irradiated model with subcutaneous tumors in which perifosine was administered by oral gavage daily (35 mg/kg/day). A two-sided t-test was used to determine statistical differences. Results: Perifosine inhibited phosphorylation of Akt in a dose- and time- dependent fashion, as well as downstream GSK3a/b and ribosomal phospho-S6. Perifosine inhibited Akt activity as confirmed by Akt kinase assay. Perifosine induced significant cytotoxicity and inhibition of DNA synthesis with an IC50 of 5-20uM in all cell lines tested. Similar effects were observed in primary CD19+ patient WM cells. Perifosine induced apoptosis in WM cells as demonstrated by flow cytometry. The mechanism of apoptosis induced by perifosine was through activation of SAPK/JNK pathway, followed by caspase-8, -9 and PARP cleavage. The JNK inhibitor SP600125 abrogated perifosine-induced apoptosis. The growth inhibitory effects of perifosine were significant even in the presence of BMSC, IL-6 and IGF-1, which induce resistance to conventional therapies. Importantly, perifosine did not induce cytotoxicity in healthy donor peripheral blood mononuclear cells or in hematopietic stem cells in a methylcellulose colony forming cell assay, indicating lack of toxicity on normal cells. Interestingly, MAPK members such as MEK/ERK were activated by perifosine. The MEK inhibitor U0126 significantly enhanced perifosine-induced cytotoxicity in WM cells, indicating that this combination may be synergistic in vivo. Finally, perifosine induced significant reduction in WM tumor growth in vivo, as compared to control cohort treated with vehicle only at week 6 (p=0.05). Conclusion: Perifosine has significant antitumor activity in WM both in vitro and in vivo. These results provide the framework for clinical evaluation of perifosine in WM. Supported in part by the Leukemia and Lymphoma Society, the Lymphoma Research Foundation and an American Society of Hematology Scholar Award. * XL and XJ are co-first authors.

Cancers ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2323
Anita Thomas ◽  
Sascha Reetz ◽  
Philipp Stenzel ◽  
Katrin Tagscherer ◽  
Wilfried Roth ◽  

The PI3K/mTOR/AKT pathway might represent an intriguing option for treatment of penile cancer (PeCa). We aimed to assess whether members of this pathway might serve as biomarkers and targets for systemic therapy. Tissue of primary cancer from treatment-naïve PeCa patients was used for tissue microarray analysis. Immunohistochemical staining was performed with antibodies against AKT, pAKT, mTOR, pmTOR, pS6, pPRAS, p4EBP1, S6K1 and pp70S6K. Protein expression was correlated with clinicopathological characteristics as well as overall survival (OS), disease-specific survival (DSS), recurrence-free survival (RFS) and metastasis-free survival (MFS). AKT inhibition was tested in two primarily established, treatment-naïve PeCa cell lines by treatment with capivasertib and analysis of cell viability and chemotaxis. A total of 76 patients surgically treated for invasive PeCa were included. Higher expression of AKT was significantly more prevalent in high-grade tumors and predictive of DSS and OS in the Kaplan–Meier analysis, and an independent predictor of worse OS and DSS in the multivariate regression analysis. Treatment with pan-AKT inhibitor capivasertib in PeCa cell lines induced a significant downregulation of both total AKT and pAKT as well as decreased cell viability and chemotaxis. Selected protein candidates of the mTOR/AKT signaling pathway demonstrate association with histological and survival parameters of PeCa patients, whereas AKT appears to be the most promising one.

2020 ◽  
Vol 2020 ◽  
pp. 1-11
Ruyue Tian ◽  
Hailun Jiang ◽  
Linlin Shao ◽  
Yang Yu ◽  
Qingdong Guo ◽  

Gastric cancer (GC) is one of the most common and fatal malignancies worldwide. MicroRNAs (miRNAs) play a critical role in tumor initiation, proliferation, and metastasis of gastric cancer. miR193b has been identified as a tumor suppressor in a variety of tumor types; however, its role in gastric cancer is yet to be determined. Here, we found a significant downregulation of miR193b expression in both human gastric cancer tissues (p<0.05) and human gastric cancer cell lines (p<0.01). Furthermore, the expression level of miR193b correlated with the tumor type, tumor size, and clinical stage (p<0.05). In vitro, miR193b overexpression inhibited cell survival and induced apoptosis in GC cell lines, indicating that miR193b plays a role in the development of gastric cancer. KRAS was verified as the target of miR193b, and KRAS overexpression attenuated miR193b-induced apoptosis (p<0.05). Moreover, we found that the Akt pathway negatively regulated miR193b, also affecting apoptosis. Further analyses indicated that PIK3CA mutation and KRAS amplification are two mutually exclusive pathways (p<0.01), and we hypothesize that both two pathways could result in the carcinogenic overactivation of KRAS. Thus, our results suggest that the Akt-miR193b-KRAS axis may act as a mechanism affecting apoptosis in gastric cancer cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4438-4438 ◽  
Lubing Gu ◽  
Ningxi Zhu ◽  
Harry W. Findley ◽  
Muxiang Zhou

Abstract PTEN is a tumor suppressor gene responsible for downregulating the phosphoinositide 3-kinase (PI3k)/Akt pathway. Loss of PTEN expression frequently occurs in human cancer leading to high Akt activation, which consequently confers neoplastic cell survival and resistance to chemotherapy-induced apoptosis. Here we report a mechanism by which loss of PTEN expression activates the transcription factor NF-kB through the PI3k/Akt pathway that induces activation of the IkBa kinase (IKK). Activation of NF-kB by loss of PTEN expression results in resistance to doxorubicin in acute lymphoblastic leukemia (ALL) cells. Initially, we examined 27 leukemia cell lines derived from children with ALL for the expression of PTEN and constitutive activation of NF-kB to evaluate whether there is a correlation between these two events. We found that 12 of the 27 lines lacked PTEN expression (PTEN-). Of 12 PTEN- ALL lines, 10 lines expressed constitutive NF-kB activation. In contrast, 11 of the 15 PTEN positive (PTEN+) lines were defect of NF-kB activation. Treatment of PTEN- line with PI3k kinase inhibitor Ly294002 caused downregulation of Akt activity accompanied by reduced activation of IKK and inhibition of constitutive NF-kB activation, resulting in increased sensitivity to doxorubicin-induced apoptosis. Similar to treatment with Ly294002, transfection of the PTEN expression plasmid into the PTEN- lines attenuated constitutive activation of both Akt and NF-kB, thereby sensitizing these cells to doxorubicin. These results suggest that both constitutive and inducible activation of NF-kB play an important role in chemotherapy resistance, and that loss of PTEN expression is at least one reason for the constitutive activation of NF-kB in ALL cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4473-4473
Eriko Suzuki ◽  
Kazuo Umezawa ◽  
Gustavo Helguera ◽  
Tracy R Daniels ◽  
Gary Schiller ◽  

Abstract Multiple Myeloma (MM) is an incurable disease of monoclonal malignant plasma cells. Treatment of MM with conventional chemotherapeutic drugs has resulted in improved response rates, however, with no sufficient improvement in overall survival. Bortezomib has been recently used and results in significant clinical responses in refractory MM. However, many patients relapse and become refractory to cytotoxic therapies and, hence, the need for new therapies. We have generated an Anti-TfR-IgG3-Avidin Fusion Protein (Anti-TfR-IgG3-Av) that can bind MM which express high levels of transferrin receptor and can deliver biotinylated molecules into cancer cells (Ng et al PNAS2002; 79:10706). We have reported that treatment of MM with Anti-TfR-IgG3-Av results in inhibition of cell proliferation and direct cytotoxicity in few cell lines. Further, we have also found that Anti-TfR-IgG3-Av can sensitize resistant MM cells to drugs (eg. CDDP)-induced apoptosis. Sensitization by Anti-TfR-IgG3-Av resulted in the inhibition of several anti-apoptotic gene products like XIAP, Bid, Bcl-2 and BclXL. Since these gene products are regulated by the NF-κB pathway, we hypothesized that Anti-TfR-IgG3-Av may inhibit the AKT pathway in MM cell lines. The AKT signaling inactivates several pro-apoptotic factors, such as Bad, which is phosphorylated and inhibits its binding and inactivation of BclXL. AKT also activates IκB kinase (IKK) to phosphorylate IκB (inhibitor of NF-κB ) and leading to its proteasomal degradation and NF-κB nuclear localization. The AKT and NF-κB pathways result in the transcription of many anti-apoptotic gene products like XIAP, Bcl-2, survivin and BclXL. Treatment of MM cell lines with Anti-TfR-IgG3-Av resulted in inhibition of phospho-AKT and inhibition of NF-κB activity and downstream inhibition of above anti-apoptotic gene products. We then examined the roles of AKT and NF-κB in Anti-TfR-IgG3-Av-induced sensitization of MM to CDDP-apoptosis. Treatment of IM-9 cells with siRNA AKT, not control siRNA, resulted in inhibition of AKT concomitantly with inhibition of Bcl-2 and survive in. The cells treated with si-RNA AKT were sensitized to CDDP-induced apoptosis. These findings suggested that Anti-Anti-TfR-IgG3-Av-induced sensitization to CDDP may be due, in part, to inhibition of AKT. Likewise, the role of NF-κB inhibition by Anti-TfR-IgG3-Av in the sensitization to CDDP was demonstrated by the use of the specific NF-κB inhibitor, DHMEQ. Thus, both inhibition of AKT and NF-κB pathways by Anti-TfR-IgG3-Av play a major role in Anti-TfR-IgG3-Av-induced sensitization to CDDP. The apoptosis achieved by the combination of Anti-TfR-IgG3-Av and CDDP resulted from the complementation of several gene products modified by each agent alone and resulting in the activation of caspases 9, 8 and 3 and apoptosis. The above findings provide an underlying mechanism of Anti-TfR-IgG3-Av-induced cell signaling modification that renders drug-resistant MM cells sensitive to apoptosis by drugs.

2020 ◽  
Vol 9 (12) ◽  
pp. 4019
Jiunn-Chang Lin ◽  
Tsang-Pai Liu ◽  
Pei-Ming Yang

The mutation of cyclin dependent kinase inhibitor 2A (CDKN2A) is frequently found in pancreatic ductal adenocarcinoma (PDAC). However, its prognostic and therapeutic roles in PDAC have not been extensively investigated yet. In this study, we mined and integrated the cancer genomics and chemogenomics data to investigate the roles of CDKN2A genetic alterations in PDAC patients’ prognosis and treatment. We found that functional CDKN2A inactivation caused by mutations and deep deletions predicted poor prognosis in PDAC patients. CDKN2A inactivation was associated with the upregulation of genes related to estrogen response, which can be overcome by CDKN2A restoration. Chemosensitivity profiling of PDAC cell lines and patient-derived organoids found that CDKN2A inactivation was associated with the increased sensitivity to paclitaxel and SN-38 (the active metabolite of irinotecan). However, only paclitaxel can mimic the effect of CDKN2A restoration, and its drug sensitivity was correlated with genes related to estrogen response. Therefore, our study suggested that CDKN2A-inactivated PDAC patients could benefit from the precision treatment with paclitaxel, whose albumin-stabilized nanoparticle formulation (nab-paclitaxel) has been approved for treating PDAC.

Blood ◽  
2006 ◽  
Vol 108 (13) ◽  
pp. 4178-4186 ◽  
Shahab Uddin ◽  
Azhar R. Hussain ◽  
Abdul K. Siraj ◽  
Pulicat S. Manogaran ◽  
Naif A. Al-Jomah ◽  

Abstract Phosphatidylinositol 3′-kinase (PI3K) is a key player in cell-growth signaling in a number of lymphoid malignancies, but its role in diffuse large B-cell lymphoma (DLBCL) has not been fully elucidated. Therefore, we investigated the role of the PI3K/AKT pathway in a panel of 5 DLBCL cell lines and 100 clinical samples. Inhibition of PI3K by a specific inhibitor, LY294002, induced apoptosis in SUDHL4, SUDHL5, and SUDHL10 (LY-sensitive) cells, whereas SUDHL8 and OCI-LY19 (LY-resistant) cells were refractory to LY294002-induced apoptosis. AKT was phosphorylated in 5 of 5 DLBCL cell lines and inhibition of PI3K caused dephosphorylation/inactivation of constitutively active AKT, FOXO transcription factor, and GSK3 in LY-sensitive cell lines. In addition, there was a decrease in the expression level of inhibitory apoptotic protein, XIAP, in the DLBCL cell lines sensitive to LY294002 after treatment. However, no effect was observed in XIAP protein levels in the resistant DLBCL cell lines following LY294002 treatment. Finally, using immunohistochemistry, p-AKT was detected in 52% of DLBCL tumors tested. Furthermore, in univariate analysis, high p-AKT expression was associated with short survival. In multivariate analysis, this correlation was no longer significant. Altogether, these results suggest that the PI3K/AKT pathway may be a potential target for therapeutic intervention in DLBCL.

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