Inhibition of HDAC1/2 Along with TRAP1 Causes Synthetic Lethality in Glioblastoma Model Systems
Abstract
:1. Introduction
2. Material and Methods
2.1. Cell Cultures and Growth Conditions
2.2. Reagents
2.3. Cell Viability Assays
2.4. Flow Cytometry
2.5. Transfection of siRNAs
2.6. Extracellular Flux Analysis
2.7. Western Blot Analysis
2.8. Real-Time PCR Analysis
2.9. Chromatin Immunoprecipitation (CHIP) Sequencing
2.10. Subcutaneous Xenograft Model
2.11. TUNEL and Ki67 Staining
2.12. Statistical Analysis
2.13. Study Approval
3. Results
3.1. FDA Approved HDAC Inhibitors and the Mitochondrial Chaperone Inhibitor, Gamitrinib, Lead to a Synergistic Reduction of Cellular Viability in Glioblastoma Models
3.2. Dual Inhibition of TRAP1 and HDAC Elicits Enhanced Activation of a Cell Death with Apoptotic Features
3.3. The Combination Treatment Modulates the Expression of the Bcl-2 Protein Family Members and Pro-Apopotic Members of the Bcl-2 Family Members are Required for Cell Death Execution
3.4. The Combination Treatment of Gamitrinib and HDAC Inhibitor Mediates an Integrated Stress Response
3.5. Combined Inhibition of HDAC and TRAP1 Modulates Tumor Cell Metabolism
3.6. The Combination Treatment of Gamitrinib and Panobinostat Reduces Tumor Growth More Potently than Single Treatments in Glioblastoma PDX Models in Mice
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
- Neftel, C.; Laffy, J.; Filbin, M.G.; Hara, T.; Shore, M.E.; Rahme, G.J.; Richman, A.R.; Silverbush, D.; Shaw, M.L.; Hebert, C.M.; et al. An Integrative Model of Cellular States, Plasticity and Genetics for Glioblastoma. Cell 2019, 178, 835–849.e21. [Google Scholar] [CrossRef] [PubMed]
- Bunse, L.; Pusch, S.; Bunse, T.; Sahm, F.; Sanghvi, K.; Friedrich, M.; Alansary, D.; Sonner, J.K.; Green, E.; Deumelandt, K.; et al. Suppression of antitumor T cell immunity by the oncometabolite (R)-2-hydroxyglutarate. Nat. Med. 2018, 24, 1192–1203. [Google Scholar] [CrossRef]
- Gupta, S.K.; Kizilbash, S.H.; Carlson, B.L.; Mladek, A.C.; Boakye-Agyeman, F.; Bakken, K.K.; Pokorny, J.L.; Schroeder, M.A.; Decker, P.A.; Cen, L.; et al. Delineation of MGMT Hypermethylation as a Biomarker for Veliparib-Mediated Temozolomide-Sensitizing Therapy of Glioblastoma. J. Natl. Cancer Inst. 2016, 108. [Google Scholar] [CrossRef] [Green Version]
- Ghosh, J.C.; Siegelin, M.D.; Vaira, V.; Faversani, A.; Tavecchio, M.; Chae, Y.C.; Lisanti, S.; Rampini, P.; Giroda, M.; Caino, M.C.; et al. Adaptive mitochondrial reprogramming and resistance to PI3K therapy. J. Natl. Cancer Inst. 2015, 107. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Kang, B.H.; Siegelin, M.D.; Plescia, J.; Raskett, C.M.; Garlick, D.S.; Dohi, T.; Lian, J.B.; Stein, G.S.; Languino, L.R.; Altieri, D.C. Preclinical characterization of mitochondria-targeted small molecule hsp90 inhibitors, gamitrinibs, in advanced prostate cancer. Clin. Cancer Res. 2010, 16, 4779–4788. [Google Scholar] [CrossRef] [Green Version]
- Siegelin, M.D.; Dohi, T.; Raskett, C.M.; Orlowski, G.M.; Powers, C.M.; Gilbert, C.A.; Ross, A.H.; Plescia, J.; Altieri, D.C. Exploiting the mitochondrial unfolded protein response for cancer therapy in mice and human cells. J. Clin. Investig. 2011, 121, 1349–1360. [Google Scholar] [CrossRef] [Green Version]
- Kang, B.H.; Plescia, J.; Song, H.Y.; Meli, M.; Colombo, G.; Beebe, K.; Scroggins, B.; Neckers, L.; Altieri, D.C. Combinatorial drug design targeting multiple cancer signaling networks controlled by mitochondrial Hsp90. J. Clin. Investig. 2009, 119, 454–464. [Google Scholar] [CrossRef] [Green Version]
- Gyurkocza, B.; Plescia, J.; Raskett, C.M.; Garlick, D.S.; Lowry, P.A.; Carter, B.Z.; Andreeff, M.; Meli, M.; Colombo, G.; Altieri, D.C. Antileukemic activity of shepherdin and molecular diversity of hsp90 inhibitors. J. Natl. Cancer Inst. 2006, 98, 1068–1077. [Google Scholar] [CrossRef] [Green Version]
- Siegelin, M.D. Inhibition of the mitochondrial Hsp90 chaperone network: A novel, efficient treatment strategy for cancer? Cancer Lett. 2013, 333, 133–146. [Google Scholar] [CrossRef]
- Zhang, Y.; Nguyen, T.T.T.; Shang, E.; Mela, A.; Humala, N.; Mahajan, A.; Zhao, J.; Shu, C.; Torrini, C.; Sanchez-Quintero, M.J.; et al. MET Inhibition Elicits PGC1alpha-Dependent Metabolic Reprogramming in Glioblastoma. Cancer Res. 2020, 80, 30–43. [Google Scholar] [CrossRef] [Green Version]
- Eleutherakis-Papaiakovou, E.; Kanellias, N.; Kastritis, E.; Gavriatopoulou, M.; Terpos, E.; Dimopoulos, M.A. Efficacy of Panobinostat for the Treatment of Multiple Myeloma. J. Oncol. 2020, 2020, 7131802. [Google Scholar] [CrossRef] [PubMed]
- Zhang, Y.; Ishida, C.T.; Ishida, W.; Lo, S.L.; Zhao, J.; Shu, C.; Bianchetti, E.; Kleiner, G.; Sanchez-Quintero, M.J.; Quinzii, C.M.; et al. Combined HDAC and Bromodomain Protein Inhibition Reprograms Tumor Cell Metabolism and Elicits Synthetic Lethality in Glioblastoma. Clin. Cancer Res. 2018, 24, 3941–3954. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Nguyen, T.; Zhang, Y.; Shang, E.; Shu, C.; Torrini, C.; Zhao, J.; Bianchetti, E.; Mela, A.; Humala, N.; Mahajan, A.; et al. HDAC inhibitors elicit metabolic reprogramming by targeting super-enhancers in glioblastoma models. J. Clin. Investig. 2020. [Google Scholar] [CrossRef] [PubMed]
- Chou, T.C. Drug combination studies and their synergy quantification using the Chou-Talalay method. Cancer Res. 2010, 70, 440–446. [Google Scholar] [CrossRef] [Green Version]
- Li, A.; Walling, J.; Kotliarov, Y.; Center, A.; Steed, M.E.; Ahn, S.J.; Rosenblum, M.; Mikkelsen, T.; Zenklusen, J.C.; Fine, H.A. Genomic changes and gene expression profiles reveal that established glioma cell lines are poorly representative of primary human gliomas. Mol. Cancer Res. 2008, 6, 21–30. [Google Scholar] [CrossRef] [Green Version]
- Wang, J.; Jiang, J.; Chen, H.; Wang, L.; Guo, H.; Yang, L.; Xiao, D.; Qing, G.; Liu, H. FDA-approved drug screen identifies proteasome as a synthetic lethal target in MYC-driven neuroblastoma. Oncogene 2019, 38, 6737–6751. [Google Scholar] [CrossRef]
- Prabhu, V.V.; Talekar, M.K.; Lulla, A.R.; Kline, C.L.B.; Zhou, L.; Hall, J.; van den Heuvel, A.P.J.; Dicker, D.T.; Babar, J.; Grupp, S.A.; et al. Single agent and synergistic combinatorial efficacy of first-in-class small molecule imipridone ONC201 in hematological malignancies. Cell Cycle 2018, 17, 468–478. [Google Scholar] [CrossRef] [Green Version]
- Chae, Y.C.; Angelin, A.; Lisanti, S.; Kossenkov, A.V.; Speicher, K.D.; Wang, H.; Powers, J.F.; Tischler, A.S.; Pacak, K.; Fliedner, S.; et al. Landscape of the mitochondrial Hsp90 metabolome in tumours. Nat. Commun. 2013, 4, 2139. [Google Scholar] [CrossRef] [Green Version]
- Hilf, N.; Kuttruff-Coqui, S.; Frenzel, K.; Bukur, V.; Stevanovic, S.; Gouttefangeas, C.; Platten, M.; Tabatabai, G.; Dutoit, V.; van der Burg, S.H.; et al. Actively personalized vaccination trial for newly diagnosed glioblastoma. Nature 2019, 565, 240–245. [Google Scholar] [CrossRef]
- Cruickshanks, N.; Zhang, Y.; Hine, S.; Gibert, M.; Yuan, F.; Oxford, M.; Grello, C.; Pahuski, M.; Dube, C.; Guessous, F.; et al. Discovery and Therapeutic Exploitation of Mechanisms of Resistance to MET Inhibitors in Glioblastoma. Clin. Cancer Res. 2019, 25, 663–673. [Google Scholar] [CrossRef] [Green Version]
- Darmanis, S.; Sloan, S.A.; Croote, D.; Mignardi, M.; Chernikova, S.; Samghababi, P.; Zhang, Y.; Neff, N.; Kowarsky, M.; Caneda, C.; et al. Single-Cell RNA-Seq Analysis of Infiltrating Neoplastic Cells at the Migrating Front of Human Glioblastoma. Cell Rep. 2017, 21, 1399–1410. [Google Scholar] [CrossRef] [Green Version]
- Mashimo, T.; Pichumani, K.; Vemireddy, V.; Hatanpaa, K.J.; Singh, D.K.; Sirasanagandla, S.; Nannepaga, S.; Piccirillo, S.G.; Kovacs, Z.; Foong, C.; et al. Acetate is a bioenergetic substrate for human glioblastoma and brain metastases. Cell 2014, 159, 1603–1614. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Sarkaria, J.N.; Carlson, B.L.; Schroeder, M.A.; Grogan, P.; Brown, P.D.; Giannini, C.; Ballman, K.V.; Kitange, G.J.; Guha, A.; Pandita, A.; et al. Use of an orthotopic xenograft model for assessing the effect of epidermal growth factor receptor amplification on glioblastoma radiation response. Clin. Cancer Res. 2006, 12, 2264–2271. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Lee, J.V.; Berry, C.T.; Kim, K.; Sen, P.; Kim, T.; Carrer, A.; Trefely, S.; Zhao, S.; Fernandez, S.; Barney, L.E.; et al. Acetyl-CoA promotes glioblastoma cell adhesion and migration through Ca(2+)-NFAT signaling. Genes Dev. 2018, 32, 497–511. [Google Scholar] [CrossRef] [Green Version]
- Guo, D.; Reinitz, F.; Youssef, M.; Hong, C.; Nathanson, D.; Akhavan, D.; Kuga, D.; Amzajerdi, A.N.; Soto, H.; Zhu, S.; et al. An LXR agonist promotes glioblastoma cell death through inhibition of an EGFR/AKT/SREBP-1/LDLR-dependent pathway. Cancer Discov. 2011, 1, 442–456. [Google Scholar] [CrossRef] [Green Version]
- Wang, X.; Yang, K.; Wu, Q.; Kim, L.J.Y.; Morton, A.R.; Gimple, R.C.; Prager, B.C.; Shi, Y.; Zhou, W.; Bhargava, S.; et al. Targeting pyrimidine synthesis accentuates molecular therapy response in glioblastoma stem cells. Sci. Transl. Med. 2019, 11. [Google Scholar] [CrossRef]
- Vander Heiden, M.G.; Cantley, L.C.; Thompson, C.B. Understanding the Warburg effect: The metabolic requirements of cell proliferation. Science 2009, 324, 1029–1033. [Google Scholar] [CrossRef] [Green Version]
- Hernandez-Munoz, I.; Taghavi, P.; Kuijl, C.; Neefjes, J.; van Lohuizen, M. Association of BMI1 with polycomb bodies is dynamic and requires PRC2/EZH2 and the maintenance DNA methyltransferase DNMT1. Mol. Cell Biol. 2005, 25, 11047–11058. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Vire, E.; Brenner, C.; Deplus, R.; Blanchon, L.; Fraga, M.; Didelot, C.; Morey, L.; Van Eynde, A.; Bernard, D.; Vanderwinden, J.M.; et al. The Polycomb group protein EZH2 directly controls DNA methylation. Nature 2006, 439, 871–874. [Google Scholar] [CrossRef]
- Lue, J.K.; Prabhu, S.A.; Liu, Y.; Gonzalez, Y.; Verma, A.; Mundi, P.S.; Abshiru, N.; Camarillo, J.M.; Mehta, S.; Chen, E.I.; et al. Precision Targeting with EZH2 and HDAC Inhibitors in Epigenetically Dysregulated Lymphomas. Clin. Cancer Res. 2019, 25, 5271–5283. [Google Scholar] [CrossRef] [Green Version]
- Zha, L.; Li, F.; Wu, R.; Artinian, L.; Rehder, V.; Yu, L.; Liang, H.; Xue, B.; Shi, H. The Histone Demethylase UTX Promotes Brown Adipocyte Thermogenic Program Via Coordinated Regulation of H3K27 Demethylation and Acetylation. J. Biol. Chem. 2015, 290, 25151–25163. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Luo, C.; Balsa, E.; Perry, E.A.; Liang, J.; Tavares, C.D.; Vazquez, F.; Widlund, H.R.; Puigserver, P. H3K27me3-mediated PGC1alpha gene silencing promotes melanoma invasion through WNT5A and YAP. J. Clin. Investig. 2020, 130, 853–862. [Google Scholar] [CrossRef] [PubMed]
- Wang, Q.; Mora-Jensen, H.; Weniger, M.A.; Perez-Galan, P.; Wolford, C.; Hai, T.; Ron, D.; Chen, W.; Trenkle, W.; Wiestner, A.; et al. ERAD inhibitors integrate ER stress with an epigenetic mechanism to activate BH3-only protein NOXA in cancer cells. Proc. Natl. Acad. Sci. USA 2009, 106, 2200–2205. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Albershardt, T.C.; Salerni, B.L.; Soderquist, R.S.; Bates, D.J.; Pletnev, A.A.; Kisselev, A.F.; Eastman, A. Multiple BH3 mimetics antagonize antiapoptotic MCL1 protein by inducing the endoplasmic reticulum stress response and up-regulating BH3-only protein NOXA. J. Biol. Chem. 2011, 286, 24882–24895. [Google Scholar] [CrossRef] [Green Version]
- Ramirez-Peinado, S.; Alcazar-Limones, F.; Lagares-Tena, L.; El Mjiyad, N.; Caro-Maldonado, A.; Tirado, O.M.; Munoz-Pinedo, C. 2-deoxyglucose induces Noxa-dependent apoptosis in alveolar rhabdomyosarcoma. Cancer Res. 2011, 71, 6796–6806. [Google Scholar] [CrossRef] [Green Version]
- Molina, J.R.; Sun, Y.; Protopopova, M.; Gera, S.; Bandi, M.; Bristow, C.; McAfoos, T.; Morlacchi, P.; Ackroyd, J.; Agip, A.A.; et al. An inhibitor of oxidative phosphorylation exploits cancer vulnerability. Nat. Med. 2018, 24, 1036–1046. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Nguyen, T.T.T.; Ishida, C.T.; Shang, E.; Shu, C.; Torrini, C.; Zhang, Y.; Bianchetti, E.; Sanchez-Quintero, M.J.; Kleiner, G.; Quinzii, C.M.; et al. Activation of LXRbeta inhibits tumor respiration and is synthetically lethal with Bcl-xL inhibition. EMBO Mol. Med. 2019, 11, e10769. [Google Scholar] [CrossRef]
- Ehrhardt, H.; Hofig, I.; Wachter, F.; Obexer, P.; Fulda, S.; Terziyska, N.; Jeremias, I. NOXA as critical mediator for drug combinations in polychemotherapy. Cell Death Dis. 2012, 3, e327. [Google Scholar] [CrossRef]
- Unterkircher, T.; Cristofanon, S.; Vellanki, S.H.; Nonnenmacher, L.; Karpel-Massler, G.; Wirtz, C.R.; Debatin, K.M.; Fulda, S. Bortezomib primes glioblastoma, including glioblastoma stem cells, for TRAIL by increasing tBid stability and mitochondrial apoptosis. Clin. Cancer Res. 2011, 17, 4019–4030. [Google Scholar] [CrossRef] [Green Version]
- Boedicker, C.; Hussong, M.; Grimm, C.; Dolgikh, N.; Meister, M.T.; Enssle, J.C.; Wanior, M.; Knapp, S.; Schweiger, M.R.; Fulda, S. Co-inhibition of BET proteins and PI3Kalpha triggers mitochondrial apoptosis in rhabdomyosarcoma cells. Oncogene 2020, 39, 3837–3852. [Google Scholar] [CrossRef]
- Haydn, T.; Metzger, E.; Schuele, R.; Fulda, S. Concomitant epigenetic targeting of LSD1 and HDAC synergistically induces mitochondrial apoptosis in rhabdomyosarcoma cells. Cell Death Dis. 2017, 8, e2879. [Google Scholar] [CrossRef] [PubMed]
- Engert, F.; Schneider, C.; Weibeta, L.M.; Probst, M.; Fulda, S. PARP Inhibitors Sensitize Ewing Sarcoma Cells to Temozolomide-Induced Apoptosis via the Mitochondrial Pathway. Mol. Cancer Ther. 2015, 14, 2818–2830. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Tse, C.; Shoemaker, A.R.; Adickes, J.; Anderson, M.G.; Chen, J.; Jin, S.; Johnson, E.F.; Marsh, K.C.; Mitten, M.J.; Nimmer, P.; et al. ABT-263: A potent and orally bioavailable Bcl-2 family inhibitor. Cancer Res. 2008, 68, 3421–3428. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Souers, A.J.; Leverson, J.D.; Boghaert, E.R.; Ackler, S.L.; Catron, N.D.; Chen, J.; Dayton, B.D.; Ding, H.; Enschede, S.H.; Fairbrother, W.J.; et al. ABT-199, a potent and selective BCL-2 inhibitor, achieves antitumor activity while sparing platelets. Nat. Med. 2013, 19, 202–208. [Google Scholar] [CrossRef]
- Karpel-Massler, G.; Ishida, C.T.; Bianchetti, E.; Shu, C.; Perez-Lorenzo, R.; Horst, B.; Banu, M.; Roth, K.A.; Bruce, J.N.; Canoll, P.; et al. Inhibition of Mitochondrial Matrix Chaperones and Antiapoptotic Bcl-2 Family Proteins Empower Antitumor Therapeutic Responses. Cancer Res. 2017, 77, 3513–3526. [Google Scholar] [CrossRef] [Green Version]
- Nguyen, T.T.T.; Ishida, C.T.; Shang, E.; Shu, C.; Bianchetti, E.; Karpel-Massler, G.; Siegelin, M.D. Activation of LXR Receptors and Inhibition of TRAP1 Causes Synthetic Lethality in Solid Tumors. Cancers 2019, 11, 788. [Google Scholar] [CrossRef] [Green Version]
- Zhang, G.; Frederick, D.T.; Wu, L.; Wei, Z.; Krepler, C.; Srinivasan, S.; Chae, Y.C.; Xu, X.; Choi, H.; Dimwamwa, E.; et al. Targeting mitochondrial biogenesis to overcome drug resistance to MAPK inhibitors. J. Clin. Investig. 2016, 126, 1834–1856. [Google Scholar] [CrossRef]
Gene | Forward Sequence | Reserve Sequence |
---|---|---|
GRP78 (HSPA5) | CTGTCCAGGCTGGTGTGCTCT | CTTGGTAGGCACCACTGTGTTC |
Noxa (PMAIP1) | CTGGAAGTCGAGTGTGCTACTC | TGAAGGAGTCCCCTCATGCAAG |
XBP1 | CTGCCAGAGATCGAAAGAAGGC | CTCCTGGTTCTCAACTACAAGGC |
CHOP (DDIT3) | GGTATGAGGACCTGCAAGAGGT | CTTGTGACCTCTGCTGGTTCTG |
CEBPB | AGAAGACCGTGGACAAGCACAG | CTCCAGGACCTTGTGCTGCGT |
ATF3 | CGCTGGAATCAGTCACTGTCAG | CTTGTTTCGGCACTTTGCAGCTG |
ATF4 | TTCTCCAGCGACAAGGCTAAGG | CTCCAACATCCAATCTGTCCCG |
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Nguyen, T.T.T.; Zhang, Y.; Shang, E.; Shu, C.; Quinzii, C.M.; Westhoff, M.-A.; Karpel-Massler, G.; Siegelin, M.D. Inhibition of HDAC1/2 Along with TRAP1 Causes Synthetic Lethality in Glioblastoma Model Systems. Cells 2020, 9, 1661. https://doi.org/10.3390/cells9071661
Nguyen TTT, Zhang Y, Shang E, Shu C, Quinzii CM, Westhoff M-A, Karpel-Massler G, Siegelin MD. Inhibition of HDAC1/2 Along with TRAP1 Causes Synthetic Lethality in Glioblastoma Model Systems. Cells. 2020; 9(7):1661. https://doi.org/10.3390/cells9071661
Chicago/Turabian StyleNguyen, Trang T. T., Yiru Zhang, Enyuan Shang, Chang Shu, Catarina M. Quinzii, Mike-Andrew Westhoff, Georg Karpel-Massler, and Markus D. Siegelin. 2020. "Inhibition of HDAC1/2 Along with TRAP1 Causes Synthetic Lethality in Glioblastoma Model Systems" Cells 9, no. 7: 1661. https://doi.org/10.3390/cells9071661