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Philip N Tsichlis - One of the best experts on this subject based on the ideXlab platform.
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abstract 4777 a phosphoproteomics analysis reveals akt isoform specific signals that link rna splicing to non small cell lung cancer
Cancer Research, 2014Co-Authors: Ioannis Sanidas, Christos Polytarchou, Maria Hatziapostolou, Scott A Ezell, Michael J Comb, Dimitrios Iliopoulos, Filippos Kottakis, Ailan Guo, Jianxin Xie, Philip N TsichlisAbstract:Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Akt isoforms exhibit different functional properties. To address the signaling differences between Akt1, Akt2 and AKT3 we examined the phosphoproteomes of a set of isogenic mouse cell lines that express different Akt isoforms. Based on this screen, we identified a total of 606 Akt phosphorylation targets, of which many are phosphorylated in an isoform specific manner. Bioinformatics analyses of these data revealed that Akt isoforms regulate differentially multiple cellular functions. One of these functions was RNA metabolism which was represented by 25 proteins phosphorylated by at least one of the Akt isoforms. One of these proteins was IWS1, which is involved in the assembly of RNA Pol II transcriptional elongation complex, and which was found to be phosphorylated at the conserved site Ser720/Thr721 by AKT3 and Akt1. Here we show that this phosphorylation event is required for the recruitment of the histone methyltransferase SETD2 to the complex and the trimethylation of histone H3 at K36 in the body of the transcribed genes. H3K36me3 provides a docking site for MRG15 and its binding partner, the splicing suppressor PTB, and regulates PTB-dependent alternative splicing. One of the targets is FGFR-2 whose alternative splicing gives rise to two isoforms, IIIb, which is expressed in epithelial cells and IIIc, which is expressed in mesenchymal cells, promotes EMT and is associated with more aggressive tumors. IWS1 phosphorylation by AKT3/Akt1 shifts splicing toward the IIIc isoform and promotes tumor growth and invasiveness both in culture and in animals. Addressing the expression of FGFR-2 in a set of lung-derived normal and tumor samples revealed that whereas the overall expression was similar in both, there was a shift toward the IIIc isoform in the tumor samples. More important, the relative expression of the IIIc and IIIb isoforms in non-small-cell-lung-carcinomas (NSCLCs) correlated with the stoichiometry of IWS1 phosphorylation and the latter correlated with Akt phosphorylation and AKT3 expression. These findings combined, underpin the importance of this pathway in the pathogenesis of lung cancer. Overall, our data suggest that Akt isoform-dependent phosphorylation events are essential for RNA processing and provide novel insights into the role of Akt in carcinogenesis. Citation Format: Ioannis Sanidas, Christos Polytarchou, Maria Hatziapostolou, Scott A. Ezell, Filippos Kottakis, Lan Hu, Ailan Guo, Jianxin Xie, Michael J. Comb, Dimitrios Iliopoulos, Philip N. Tsichlis. A phosphoproteomics analysis reveals Akt isoform-specific signals that link RNA splicing to non-small cell lung cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4777. doi:10.1158/1538-7445.AM2014-4777
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The protein kinase Akt1 regulates the interferon response through phosphorylation of the transcriptional repressor EMSY.
Proceedings of the National Academy of Sciences of the United States of America, 2012Co-Authors: Scott A Ezell, Teeru Bihani, George Sourvinos, Ioannis Sanidas, Christos Polytarchou, Maria Hatziapostolou, Michael J Comb, Philip N TsichlisAbstract:The protein kinases Akt1, Akt2, and AKT3 possess nonredundant signaling properties, few of which have been investigated. Here, we present evidence for an Akt1-dependent pathway that controls interferon (IFN)-regulated gene expression and antiviral immunity. The target of this pathway is EMSY, an oncogenic interacting partner of BRCA2 that functions as a transcriptional repressor. Overexpression of EMSY in hTERT-immortalized mammary epithelial cells, and in breast and ovarian carcinoma cell lines, represses IFN-stimulated genes (ISGs) in a BRCA2-dependent manner, whereas its knockdown has the opposite effect. EMSY binds to the promoters of ISGs, suggesting that EMSY functions as a direct transcriptional repressor. Akt1, but not Akt2, phosphorylates EMSY at Ser209, relieving EMSY-mediated ISG repression. The Akt1/EMSY/ISG pathway is activated by both viral infection and IFN, and it inhibits the replication of HSV-1 and vesicular stomatitis virus (VSV). Collectively, these data define an Akt1-dependent pathway that contributes to the full activation of ISGs by relieving their repression by EMSY and BRCA2.
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akt2 regulates all akt isoforms and promotes resistance to hypoxia through induction of mir 21 upon oxygen deprivation
Cancer Research, 2011Co-Authors: Christos Polytarchou, Maria Hatziapostolou, Ioanna G Maroulakou, Dimitrios Iliopoulos, Filippos Kottakis, Kevin Struhl, Philip N TsichlisAbstract:The growth and survival of tumor cells in an unfavorable hypoxic environment depend upon their adaptibility. Here we show that both normal and tumor cells expressing the protein kinase Akt2 are more resistant to hypoxia than cells expressing Akt1 or AKT3. This is due to the differential regulation of miR-21, which is upregulated by hypoxia only in Akt2-expressing cells. By upregulating miR-21 upon oxygen deprivation, Akt2 downregulates PTEN and activates all three Akt isoforms. MiR-21 also targets PDCD4 and Spry1, and the combined downregulation of these proteins with PTEN is sufficient to confer resistance to hypoxia. Furthermore, the miR-21 induction by Akt2 during hypoxia depends upon the binding of NF-κB, CREB and CBP/p300 to the miR-21 promoter, in addition to the regional acetylation of histone H3K9, all of which are under the control of Akt2. Analysis of the Akt2-miR-21 pathway in hypoxic MMTV-PyMT-induced mouse mammary adenocarcinomas and human ovarian carcinomas confirmed the activity of the pathway in vivo. Taken together, this study identifies a novel Akt2-dependent pathway that is activated by hypoxia and promotes tumor resistance via induction of miR-21.
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micrornas differentially regulated by akt isoforms control emt and stem cell renewal in cancer cells
Science Signaling, 2009Co-Authors: Dimitrios Iliopoulos, Christos Polytarchou, Maria Hatziapostolou, Ioanna G Maroulakou, Filippos Kottakis, Kevin Struhl, Philip N TsichlisAbstract:Although Akt is known to play a role in human cancer, the relative contribution of its three isoforms to oncogenesis remains to be determined. We expressed each isoform individually in an Akt1 −/− / Akt2 −/− / AKT3 −/− cell line. MicroRNA profiling of growth factor–stimulated cells revealed unique microRNA signatures for cells with each isoform. Among the differentially regulated microRNAs, the abundance of the miR-200 family was decreased in cells bearing Akt2. Knockdown of Akt1 in transforming growth factor–β (TGFβ)–treated MCF10A cells also decreased the abundance of miR-200; however, knockdown of Akt2, or of both Akt1 and Akt2, did not. Furthermore, Akt1 knockdown in MCF10A cells promoted TGFβ-induced epithelial-mesenchymal transition (EMT) and a stem cell–like phenotype. Carcinomas developing in MMTV-cErbB2/ Akt1 −/− mice showed increased invasiveness because of miR-200 down-regulation. Finally, the ratio of Akt1 to Akt2 and the abundance of miR-200 and of the messenger RNA encoding E-cadherin in a set of primary and metastatic human breast cancers were consistent with the hypothesis that in many cases breast cancer metastasis may be under the control of the Akt–miR-200–E-cadherin axis. We conclude that induction of EMT is controlled by microRNAs whose abundance depends on the balance between Akt1 and Akt2 rather than on the overall activity of Akt.
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VEGF stimulation of mitochondrial biogenesis: requirement of AKT3 kinase
FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 2008Co-Authors: Gary L. Wright, Philip N Tsichlis, Ioanna G Maroulakou, Juanita Eldridge, Tiera L. Liby, Vijayalakshmi Sridharan, Robin C. Muise-helmericksAbstract:The growth factor, vascular endothelial growth factor (VEGF), induces angiogenesis and promotes endothelial cell (EC) proliferation. Affymetrix gene array analyses show that VEGF stimulates the expression of a cluster of nuclear-encoded mitochondrial genes, suggesting a role for VEGF in the regulation of mitochondrial biogenesis. We show that the serine threonine kinase AKT3 specifically links VEGF to mitochondrial biogenesis. A direct comparison of Akt1 vs. AKT3 gene silencing was performed in ECs and has uncovered a discrete role for AKT3 in the control of mitochondrial biogenesis. Silencing of AKT3, but not Akt1, results in a decrease in mitochondrial gene expression and mtDNA content. Nuclear-encoded mitochondrial gene transcripts are also found to decrease when AKT3 expression is silenced. Concurrent with these changes in mitochondrial gene expression, lower O2 consumption was observed. VEGF stimulation of the major mitochondrial import protein TOM70 is also blocked by AKT3 inhibition. In support of ...
Nicole Grabinski - One of the best experts on this subject based on the ideXlab platform.
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downregulation of AKT3 increases migration and metastasis in triple negative breast cancer cells by upregulating s100a4
PLOS ONE, 2016Co-Authors: Astrid Grottke, Nicole Grabinski, Florian Ewald, Tobias Lange, Dominik Norz, Christiane Herzberger, Johanna Bach, Lareen Graser, Frank Hoppner, Bjorn NashanAbstract:Background Treatment of breast cancer patients with distant metastases represents one of the biggest challenges in today’s gynecological oncology. Therefore, a better understanding of mechanisms promoting the development of metastases is of paramount importance. The serine/threonine kinase AKT was shown to drive cancer progression and metastasis. However, there is emerging data that single AKT isoforms (i.e. AKT1, AKT2 and AKT3) have different or even opposing functions in the regulation of cancer cell migration in vitro, giving rise to the hypothesis that inhibition of distinct AKT isoforms might have undesirable effects on cancer dissemination in vivo.
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AKT3 regulates erbb2 erbb3 and estrogen receptor α expression and contributes to endocrine therapy resistance of erbb2 breast tumor cells from balb neut mice
Cellular Signalling, 2014Co-Authors: Nicole Grabinski, Katharina Mollmann, Karin Mildelangosch, Volkmar Muller, Udo Schumacher, Burkhard Brandt, Klaus Pantel, Manfred JuckerAbstract:Abstract ErbB2+ breast cancer is an aggressive breast cancer subtype generally associated with lower estrogen receptor alpha (ERα) expression and more aggressive tumor behavior compared to ERα+/ErbB2− breast cancer. The ErbB2+ phenotype is associated with resistance to endocrine therapy, e.g. the selective estrogen receptor modulator Tamoxifen. However, the mechanisms underlying endocrine resistance are not fully understood. Here, we investigated the impact of AKT signaling and distinct functional roles of AKT isoforms in ErbB2+ breast cancer from Balb-neuT mice. AKT isoform specific in vitro kinase assays revealed that AKT3 is activated in Balb-neuT breast tumors in comparison to normal murine breast tissue. Knock-down of AKT3, but not of AKT1 or AKT2, led to reduced expression and tyrosine-phosphorylation of ErbB2 and ErbB3 in Balb-neuT-derived mammary tumor cells. In contrast, expression of ERα was strongly up-regulated and phosphorylation of the AKT substrate Foxo3a which regulates ERα transcription was decreased in AKT3 knockdown cells. These data suggest that ERα expression is down regulated via AKT3/Foxo3a signaling in ErbB2+ breast cancer cells. Furthermore, up-regulation of ERα after depletion of AKT3 resulted in a significant increase in Tamoxifen responsiveness of Balb-neuT-derived mammary tumor cells. In addition, Tamoxifen resistant human breast cancer cell lines showed increased AKT3 expression and activity in comparison to Tamoxifen responsive MCF-7 cells. Finally, by AKT isoform specific in vitro kinase assays of human breast cancer samples, AKT3 activity was detected in ErbB2+ and triple negative tumors but not in ERα+ breast cancer. Our data indicate that AKT3 regulates the expression of ErbB2, ErbB3 and ERα and demonstrate that down-regulation of activated AKT3 can sensitize ErbB2+ breast cancer cells for treatment with Tamoxifen. Therefore, AKT3 targeting might be a new promising strategy for therapy of ErbB2+/ERα− breast cancer and might further increase the responsiveness to an endocrine therapy approach.
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AKT3 regulates erbb2 erbb3 and estrogen receptor α expression and contributes to endocrine therapy resistance of erbb2 breast tumor cells from balb neut mice
Cellular Signalling, 2014Co-Authors: Nicole Grabinski, Katharina Mollmann, Karin Mildelangosch, Volkmar Muller, Udo Schumacher, Burkhard Brandt, Klaus Pantel, Manfred JuckerAbstract:ErbB2(+) breast cancer is an aggressive breast cancer subtype generally associated with lower estrogen receptor alpha (ERα) expression and more aggressive tumor behavior compared to ERα(+)/ErbB2(-) breast cancer. The ErbB2(+) phenotype is associated with resistance to endocrine therapy, e.g. the selective estrogen receptor modulator Tamoxifen. However, the mechanisms underlying endocrine resistance are not fully understood. Here, we investigated the impact of AKT signaling and distinct functional roles of AKT isoforms in ErbB2(+) breast cancer from Balb-neuT mice. AKT isoform specific in vitro kinase assays revealed that AKT3 is activated in Balb-neuT breast tumors in comparison to normal murine breast tissue. Knock-down of AKT3, but not of AKT1 or AKT2, led to reduced expression and tyrosine-phosphorylation of ErbB2 and ErbB3 in Balb-neuT-derived mammary tumor cells. In contrast, expression of ERα was strongly up-regulated and phosphorylation of the AKT substrate Foxo3a which regulates ERα transcription was decreased in AKT3 knockdown cells. These data suggest that ERα expression is down regulated via AKT3/Foxo3a signaling in ErbB2(+) breast cancer cells. Furthermore, up-regulation of ERα after depletion of AKT3 resulted in a significant increase in Tamoxifen responsiveness of Balb-neuT-derived mammary tumor cells. In addition, Tamoxifen resistant human breast cancer cell lines showed increased AKT3 expression and activity in comparison to Tamoxifen responsive MCF-7 cells. Finally, by AKT isoform specific in vitro kinase assays of human breast cancer samples, AKT3 activity was detected in ErbB2(+) and triple negative tumors but not in ERα(+) breast cancer. Our data indicate that AKT3 regulates the expression of ErbB2, ErbB3 and ERα and demonstrate that down-regulation of activated AKT3 can sensitize ErbB2(+) breast cancer cells for treatment with Tamoxifen. Therefore, AKT3 targeting might be a new promising strategy for therapy of ErbB2(+)/ERα(-) breast cancer and might further increase the responsiveness to an endocrine therapy approach.
Manfred Jucker - One of the best experts on this subject based on the ideXlab platform.
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AKT3 regulates erbb2 erbb3 and estrogen receptor α expression and contributes to endocrine therapy resistance of erbb2 breast tumor cells from balb neut mice
Cellular Signalling, 2014Co-Authors: Nicole Grabinski, Katharina Mollmann, Karin Mildelangosch, Volkmar Muller, Udo Schumacher, Burkhard Brandt, Klaus Pantel, Manfred JuckerAbstract:Abstract ErbB2+ breast cancer is an aggressive breast cancer subtype generally associated with lower estrogen receptor alpha (ERα) expression and more aggressive tumor behavior compared to ERα+/ErbB2− breast cancer. The ErbB2+ phenotype is associated with resistance to endocrine therapy, e.g. the selective estrogen receptor modulator Tamoxifen. However, the mechanisms underlying endocrine resistance are not fully understood. Here, we investigated the impact of AKT signaling and distinct functional roles of AKT isoforms in ErbB2+ breast cancer from Balb-neuT mice. AKT isoform specific in vitro kinase assays revealed that AKT3 is activated in Balb-neuT breast tumors in comparison to normal murine breast tissue. Knock-down of AKT3, but not of AKT1 or AKT2, led to reduced expression and tyrosine-phosphorylation of ErbB2 and ErbB3 in Balb-neuT-derived mammary tumor cells. In contrast, expression of ERα was strongly up-regulated and phosphorylation of the AKT substrate Foxo3a which regulates ERα transcription was decreased in AKT3 knockdown cells. These data suggest that ERα expression is down regulated via AKT3/Foxo3a signaling in ErbB2+ breast cancer cells. Furthermore, up-regulation of ERα after depletion of AKT3 resulted in a significant increase in Tamoxifen responsiveness of Balb-neuT-derived mammary tumor cells. In addition, Tamoxifen resistant human breast cancer cell lines showed increased AKT3 expression and activity in comparison to Tamoxifen responsive MCF-7 cells. Finally, by AKT isoform specific in vitro kinase assays of human breast cancer samples, AKT3 activity was detected in ErbB2+ and triple negative tumors but not in ERα+ breast cancer. Our data indicate that AKT3 regulates the expression of ErbB2, ErbB3 and ERα and demonstrate that down-regulation of activated AKT3 can sensitize ErbB2+ breast cancer cells for treatment with Tamoxifen. Therefore, AKT3 targeting might be a new promising strategy for therapy of ErbB2+/ERα− breast cancer and might further increase the responsiveness to an endocrine therapy approach.
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AKT3 regulates erbb2 erbb3 and estrogen receptor α expression and contributes to endocrine therapy resistance of erbb2 breast tumor cells from balb neut mice
Cellular Signalling, 2014Co-Authors: Nicole Grabinski, Katharina Mollmann, Karin Mildelangosch, Volkmar Muller, Udo Schumacher, Burkhard Brandt, Klaus Pantel, Manfred JuckerAbstract:ErbB2(+) breast cancer is an aggressive breast cancer subtype generally associated with lower estrogen receptor alpha (ERα) expression and more aggressive tumor behavior compared to ERα(+)/ErbB2(-) breast cancer. The ErbB2(+) phenotype is associated with resistance to endocrine therapy, e.g. the selective estrogen receptor modulator Tamoxifen. However, the mechanisms underlying endocrine resistance are not fully understood. Here, we investigated the impact of AKT signaling and distinct functional roles of AKT isoforms in ErbB2(+) breast cancer from Balb-neuT mice. AKT isoform specific in vitro kinase assays revealed that AKT3 is activated in Balb-neuT breast tumors in comparison to normal murine breast tissue. Knock-down of AKT3, but not of AKT1 or AKT2, led to reduced expression and tyrosine-phosphorylation of ErbB2 and ErbB3 in Balb-neuT-derived mammary tumor cells. In contrast, expression of ERα was strongly up-regulated and phosphorylation of the AKT substrate Foxo3a which regulates ERα transcription was decreased in AKT3 knockdown cells. These data suggest that ERα expression is down regulated via AKT3/Foxo3a signaling in ErbB2(+) breast cancer cells. Furthermore, up-regulation of ERα after depletion of AKT3 resulted in a significant increase in Tamoxifen responsiveness of Balb-neuT-derived mammary tumor cells. In addition, Tamoxifen resistant human breast cancer cell lines showed increased AKT3 expression and activity in comparison to Tamoxifen responsive MCF-7 cells. Finally, by AKT isoform specific in vitro kinase assays of human breast cancer samples, AKT3 activity was detected in ErbB2(+) and triple negative tumors but not in ERα(+) breast cancer. Our data indicate that AKT3 regulates the expression of ErbB2, ErbB3 and ERα and demonstrate that down-regulation of activated AKT3 can sensitize ErbB2(+) breast cancer cells for treatment with Tamoxifen. Therefore, AKT3 targeting might be a new promising strategy for therapy of ErbB2(+)/ERα(-) breast cancer and might further increase the responsiveness to an endocrine therapy approach.
Y. Rebecca Chin - One of the best experts on this subject based on the ideXlab platform.
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Upregulation of AKT3 Confers Resistance to the AKT Inhibitor MK2206 in Breast Cancer
Molecular cancer therapeutics, 2016Co-Authors: Casey Stottrup, Tiffany Tsang, Y. Rebecca ChinAbstract:Acquired resistance to molecular targeted therapy represents a major challenge for the effective treatment of cancer. Hyperactivation of the PI3K/AKT pathway is frequently observed in virtually all human malignancies, and numerous PI3K and AKT inhibitors are currently under clinical evaluation. However, mechanisms of acquired resistance to AKT inhibitors have yet to be described. Here, we use a breast cancer preclinical model to identify resistance mechanisms to a small molecule allosteric AKT inhibitor, MK2206. Using a step-wise and chronic high-dose exposure, breast cancer cell lines harboring oncogenic PI3K resistant to MK2206 were established. Using this model, we reveal that AKT3 expression is markedly upregulated in AKT inhibitor-resistant cells. Induction of AKT3 is regulated epigenetically by the bromodomain and extra terminal domain proteins. Importantly, knockdown of AKT3, but not AKT1 or AKT2, in resistant cells restores sensitivity to MK2206. AKT inhibitor-resistant cells also display an epithelial to mesenchymal transition phenotype as assessed by alterations in the levels of E-Cadherin, N-Cadherin, and vimentin, as well as enhanced invasiveness of tumor spheroids. Notably, the invasive morphology of resistant spheroids is diminished upon AKT3 depletion. We also show that resistance to MK2206 is reversible because upon drug removal resistant cells regain sensitivity to AKT inhibition, accompanied by reexpression of epithelial markers and reduction of AKT3 expression, implying that epigenetic reprogramming contributes to acquisition of resistance. These findings provide a rationale for developing therapeutics targeting AKT3 to circumvent acquired resistance in breast cancer. Mol Cancer Ther; 15(8); 1964-74. ©2016 AACR.
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Abstract A42: Increased AKT3 expression as a resistance mechanism to targeted therapy
Resistance Mechanisms, 2015Co-Authors: Y. Rebecca Chin, Alex TokerAbstract:Resistance to targeted therapy poses a major challenge for successful treatment of cancers. Despite a number of drugs targeting the PI 3-K/Akt pathway are in clinical development, mechanisms of resistance to drugs targeting this pathway have not been studied extensively. To determine how breast cancer cells develop resistance to Akt inhibitors (Akti), we generated Akti-resistant lines using dose escalation regimens with pan-Akt inhibitors (ATP-competitive: GDC0068, GSK690693; Allosteric: MK2206). Interestingly, in multiple Akti-resistant breast cancer lines, we observed a robust increase in the protein expression of AKT3, but not Akt1 or Akt2. We were intrigued by this observation, as we have recently demonstrated that AKT3 has a predominant function in regulating growth of triple negative breast cancer cells in three-dimensional spheroids and in xenograft models. The increased expression of AKT3 in Akti-resistant cells occurs via a genetic event, since the levels of AKT3 remain elevated after 3 weeks of Akt inhibitor removal. Moreover, upon Akt inhibitor removal in resistant lines, cells remain resistant to Akt inhibition. The enhanced AKT3 expression is observed in both luminal and basal-like triple-negative breast cancer resistant lines. Importantly, knockdown of AKT3, but not Akt1 or Akt2, results in re-sensitization of resistant cells to Akti. Mechanistically, using anti-phosphotyrosine receptor antibody array (RTK array), we observed increased phosphorylation of several RTKs in resistant cells, including EGFR, Her2, and ROR1. AKT3 knockdown in resistant cells results in reduction of pEGFR, pHer3 and pHer2, suggesting that AKT3 plays a role in the regulation of multiple RTKs in resistant cells. Epithelial-mesenchymal transition (EMT) is implicated in therapeutic resistance of cancers. Interestingly, several of our resistant lines display EMT properties including downregulation of E-Cadherin, and upregulation of N-Cadherin as well as Vimentin. We propose that enhanced AKT3 expression is a general mechanism by which breast cancer cells acquire resistance to Akti targeted therapy. Current efforts are aimed at investigating the mechanistic basis for AKT3-mediated resistance, and demonstrating the physiological relevance of these findings in clinical samples as well as in mouse models. The role of AKT3 in regulating EMT during establishment of resistance will also be explored. These studies will provide insights into molecular mechanisms of therapeutic resistance to drugs targeting the PI 3-K/Akt pathway, as well as information for designing combination strategies and next generation therapy. Citation Format: Y Rebecca Chin, Alex Toker. Increased AKT3 expression as a resistance mechanism to targeted therapy. [abstract]. In: Proceedings of the AACR Special Conference: Targeting the PI3K-mTOR Network in Cancer; Sep 14-17, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(7 Suppl):Abstract nr A42.
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Targeting AKT3 Signaling in Triple-Negative Breast Cancer
Cancer research, 2013Co-Authors: Y. Rebecca Chin, Taku Yoshida, Andriy Marusyk, Andrew H. Beck, Kornelia Polyak, Alex TokerAbstract:Triple negative breast cancer (TNBC) is currently the only major breast tumor subtype without effective targeted therapy and as a consequence in general has poor outcome. To identify new therapeutic targets in TNBC we performed an shRNA screen for protein kinases commonly amplified and overexpressed in breast cancer. Using this approach, we identified AKT3 as a gene preferentially required for the growth of TNBCs. Downregulation of AKT3 significantly inhibits the growth of TNBC lines in 3D spheroid cultures and in mouse xenograft models whereas loss of Akt1 or Akt2 have more modest effects. AKT3 silencing markedly upregulates the p27 cell cycle inhibitor and this is critical for the ability of AKT3 to inhibit spheroid growth. In contrast to Akt1, AKT3 silencing results in only a minor enhancement of migration and does not promote invasion. Depletion of AKT3 in TNBC sensitizes cells to the pan-Akt inhibitor GSK690693. These results imply that AKT3 has a specific function in TNBCs, thus, its therapeutic targeting may provide a new treatment option for this tumor subtype.
Alex Toker - One of the best experts on this subject based on the ideXlab platform.
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Abstract A42: Increased AKT3 expression as a resistance mechanism to targeted therapy
Resistance Mechanisms, 2015Co-Authors: Y. Rebecca Chin, Alex TokerAbstract:Resistance to targeted therapy poses a major challenge for successful treatment of cancers. Despite a number of drugs targeting the PI 3-K/Akt pathway are in clinical development, mechanisms of resistance to drugs targeting this pathway have not been studied extensively. To determine how breast cancer cells develop resistance to Akt inhibitors (Akti), we generated Akti-resistant lines using dose escalation regimens with pan-Akt inhibitors (ATP-competitive: GDC0068, GSK690693; Allosteric: MK2206). Interestingly, in multiple Akti-resistant breast cancer lines, we observed a robust increase in the protein expression of AKT3, but not Akt1 or Akt2. We were intrigued by this observation, as we have recently demonstrated that AKT3 has a predominant function in regulating growth of triple negative breast cancer cells in three-dimensional spheroids and in xenograft models. The increased expression of AKT3 in Akti-resistant cells occurs via a genetic event, since the levels of AKT3 remain elevated after 3 weeks of Akt inhibitor removal. Moreover, upon Akt inhibitor removal in resistant lines, cells remain resistant to Akt inhibition. The enhanced AKT3 expression is observed in both luminal and basal-like triple-negative breast cancer resistant lines. Importantly, knockdown of AKT3, but not Akt1 or Akt2, results in re-sensitization of resistant cells to Akti. Mechanistically, using anti-phosphotyrosine receptor antibody array (RTK array), we observed increased phosphorylation of several RTKs in resistant cells, including EGFR, Her2, and ROR1. AKT3 knockdown in resistant cells results in reduction of pEGFR, pHer3 and pHer2, suggesting that AKT3 plays a role in the regulation of multiple RTKs in resistant cells. Epithelial-mesenchymal transition (EMT) is implicated in therapeutic resistance of cancers. Interestingly, several of our resistant lines display EMT properties including downregulation of E-Cadherin, and upregulation of N-Cadherin as well as Vimentin. We propose that enhanced AKT3 expression is a general mechanism by which breast cancer cells acquire resistance to Akti targeted therapy. Current efforts are aimed at investigating the mechanistic basis for AKT3-mediated resistance, and demonstrating the physiological relevance of these findings in clinical samples as well as in mouse models. The role of AKT3 in regulating EMT during establishment of resistance will also be explored. These studies will provide insights into molecular mechanisms of therapeutic resistance to drugs targeting the PI 3-K/Akt pathway, as well as information for designing combination strategies and next generation therapy. Citation Format: Y Rebecca Chin, Alex Toker. Increased AKT3 expression as a resistance mechanism to targeted therapy. [abstract]. In: Proceedings of the AACR Special Conference: Targeting the PI3K-mTOR Network in Cancer; Sep 14-17, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(7 Suppl):Abstract nr A42.
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Targeting AKT3 Signaling in Triple-Negative Breast Cancer
Cancer research, 2013Co-Authors: Y. Rebecca Chin, Taku Yoshida, Andriy Marusyk, Andrew H. Beck, Kornelia Polyak, Alex TokerAbstract:Triple negative breast cancer (TNBC) is currently the only major breast tumor subtype without effective targeted therapy and as a consequence in general has poor outcome. To identify new therapeutic targets in TNBC we performed an shRNA screen for protein kinases commonly amplified and overexpressed in breast cancer. Using this approach, we identified AKT3 as a gene preferentially required for the growth of TNBCs. Downregulation of AKT3 significantly inhibits the growth of TNBC lines in 3D spheroid cultures and in mouse xenograft models whereas loss of Akt1 or Akt2 have more modest effects. AKT3 silencing markedly upregulates the p27 cell cycle inhibitor and this is critical for the ability of AKT3 to inhibit spheroid growth. In contrast to Akt1, AKT3 silencing results in only a minor enhancement of migration and does not promote invasion. Depletion of AKT3 in TNBC sensitizes cells to the pan-Akt inhibitor GSK690693. These results imply that AKT3 has a specific function in TNBCs, thus, its therapeutic targeting may provide a new treatment option for this tumor subtype.
