Tubastatin A, a tetrahydro-γ-carboline-capped selective HDAC6 inhibitor (HDAC6i), was rationally designed 10 years ago, and has become the best investigated HDAC6i to date. It shows efficacy in various neurological disease animal models, as HDAC6 plays a crucial regulatory role in axonal transport deficits, protein aggregation, as well as oxidative stress. In this work, we provide new insights into this HDAC6i by investigating the molecular basis of its interactions with HDAC6 through X-ray crystallography, determining its functional capability to elevate the levels of acetylated α-tubulin in vitro and in vivo, correlating PK/PD profiles to determine effective doses in plasma and brain, and finally assessing its therapeutic potential toward psychiatric diseases through use of the SmartCube screening platform.

Consortium for Fibrosis Research and Translation University of Colorado Anschutz Medical Campus Aurora Colorado 80045 United States

Department of Medicinal Chemistry and Pharmacognosy College of Pharmacy University of Illinois at Chicago Chicago Illinois 60612 United States

Department of Medicine Division of Cardiology University of Colorado Anschutz Medical Campus Aurora Colorado 80045 United States

Division of Experimental and Translational Neuroscience Krembil Research Institute University Health Network Toronto Ontario M5G 2C4 Canada

Laboratory of Structural Biology Institute of Biotechnology of the Czech Academy of Sciences Prumyslova 595 252 50 Vestec Czech Republic

StarWise Therapeutics LLC Chicago Illinois 60614 United States

Zobrazit více v PubMed

de Ruijter A. J.; van Gennip A. H.; Caron H. N.; Kemp S.; van Kuilenburg A. B. Histone deacetylases (HDACs): characterization of the classical HDAC family. Biochem. J. 2003, 370, 737–749. 10.1042/bj20021321. PubMed DOI PMC

Drummond D. C.; Noble C. O.; Kirpotin D. B.; Guo Z.; Scott G. K.; Benz C. C. Clinical development of histone deacetylase inhibitors as anticancer agents. Annu. Rev. Pharmacol. Toxicol. 2005, 45, 495–528. 10.1146/annurev.pharmtox.45.120403.095825. PubMed DOI

Boyault C.; Sadoul K.; Pabion M.; Khochbin S. HDAC6, at the crossroads between cytoskeleton and cell signaling by acetylation and ubiquitination. Oncogene 2007, 26, 5468–5476. 10.1038/sj.onc.1210614. PubMed DOI

Zhang Y.; Kwon S.; Yamaguchi T.; Cubizolles F.; Rousseaux S.; Kneissel M.; Cao C.; Li N.; Cheng H. L.; Chua K.; Lombard D.; Mizeracki A.; Matthias G.; Alt F. W.; Khochbin S.; Matthias P. Mice lacking histone deacetylase 6 have hyperacetylated tubulin but are viable and develop normally. Mol. Cell. Biol. 2008, 28, 1688–1701. 10.1128/MCB.01154-06. PubMed DOI PMC

Hammond J. W.; Huang C. F.; Kaech S.; Jacobson C.; Banker G.; Verhey K. J. Posttranslational modifications of tubulin and the polarized transport of kinesin-1 in neurons. Mol. Biol. Cell 2010, 21, 572–583. 10.1091/mbc.e09-01-0044. PubMed DOI PMC

Perdiz D.; Mackeh R.; Pous C.; Baillet A. The ins and outs of tubulin acetylation: more than just a post-translational modification?. Cell. Signalling 2011, 23, 763–771. 10.1016/j.cellsig.2010.10.014. PubMed DOI

North B. J.; Marshall B. L.; Borra M. T.; Denu J. M.; Verdin E. The human Sir2 ortholog, SIRT2, is an NAD+-dependent tubulin deacetylase. Mol. Cell 2003, 11, 437–444. 10.1016/S1097-2765(03)00038-8. PubMed DOI

Bobrowska A.; Donmez G.; Weiss A.; Guarente L.; Bates G. SIRT2 ablation has no effect on tubulin acetylation in brain, cholesterol biosynthesis or the progression of Huntington’s disease phenotypes in vivo. PLoS One 2012, 7, e3480510.1371/journal.pone.0034805. PubMed DOI PMC

Kozlov M. V.; Kleymenova A. A.; Konduktorov K. A.; Malikova A. Z.; Kochetkov S. N. Selective inhibitor of histone deacetylase 6 (tubastatin A) suppresses proliferation of hepatitis C virus replicon in culture of human hepatocytes. Biochemistry (Moscow) 2014, 79, 637–642. 10.1134/S0006297914070050. PubMed DOI

Majid T.; Griffin D.; Criss Z. 2nd; Jarpe M.; Pautler R. G. Pharmocologic treatment with histone deacetylase 6 inhibitor (ACY-738) recovers Alzheimer’s disease phenotype in amyloid precursor protein/presenilin 1 (APP/PS1) mice. Alzheimers Dement. 2015, 1, 170–181. 10.1016/j.trci.2015.08.001. PubMed DOI PMC

d’Ydewalle C.; Krishnan J.; Chiheb D. M.; Van Damme P.; Irobi J.; Kozikowski A. P.; Vanden Berghe P.; Timmerman V.; Robberecht W.; Van Den Bosch L. HDAC6 inhibitors reverse axonal loss in a mouse model of mutant HSPB1-induced Charcot-Marie-Tooth disease. Nat. Med. 2011, 17, 968–974. 10.1038/nm.2396. PubMed DOI

Mo Z.; Zhao X.; Liu H.; Hu Q.; Chen X. Q.; Pham J.; Wei N.; Liu Z.; Zhou J.; Burgess R. W.; Pfaff S. L.; Caskey C. T.; Wu C.; Bai G.; Yang X. L. Aberrant GlyRS-HDAC6 interaction linked to axonal transport deficits in Charcot-Marie-Tooth neuropathy. Nat. Commun. 2018, 9, 1007.10.1038/s41467-018-03461-z. PubMed DOI PMC

Benoy V.; Van Helleputte L.; Prior R.; d’Ydewalle C.; Haeck W.; Geens N.; Scheveneels W.; Schevenels B.; Cader M. Z.; Talbot K.; Kozikowski A. P.; Vanden Berghe P.; Van Damme P.; Robberecht W.; Van Den Bosch L. HDAC6 is a therapeutic target in mutant GARS-induced Charcot-Marie-Tooth disease. Brain 2018, 141, 673–687. 10.1093/brain/awx375. PubMed DOI PMC

Guo W.; Naujock M.; Fumagalli L.; Vandoorne T.; Baatsen P.; Boon R.; Ordovas L.; Patel A.; Welters M.; Vanwelden T.; Geens N.; Tricot T.; Benoy V.; Steyaert J.; Lefebvre-Omar C.; Boesmans W.; Jarpe M.; Sterneckert J.; Wegner F.; Petri S.; Bohl D.; Vanden Berghe P.; Robberecht W.; Van Damme P.; Verfaillie C.; Van Den Bosch L.; et al. HDAC6 inhibition reverses axonal transport defects in motor neurons derived from FUS-ALS patients. Nat. Commun. 2017, 8, 861.10.1038/s41467-017-00911-y. PubMed DOI PMC

Rossaert E.; Pollari E.; Jaspers T.; Van Helleputte L.; Jarpe M.; Van Damme P.; De Bock K.; Moisse M.; Van Den Bosch L. Restoration of histone acetylation ameliorates disease and metabolic abnormalities in a FUS mouse model. Acta Neuropathol. Commun. 2019, 7, 107.10.1186/s40478-019-0750-2. PubMed DOI PMC

Xu X.; Kozikowski A. P.; Pozzo-Miller L. A selective histone deacetylase-6 inhibitor improves BDNF trafficking in hippocampal neurons from Mecp2 knockout mice: implications for Rett syndrome. Front. Cell. Neurosci. 2014, 8, 68.10.3389/fncel.2014.00068. PubMed DOI PMC

Gold W. A.; Lacina T. A.; Cantrill L. C.; Christodoulou J. MeCP2 deficiency is associated with reduced levels of tubulin acetylation and can be restored using HDAC6 inhibitors. J. Mol. Med. 2015, 93, 63–72. 10.1007/s00109-014-1202-x. PubMed DOI

Butler K. V.; Kalin J.; Brochier C.; Vistoli G.; Langley B.; Kozikowski A. P. Rational design and simple chemistry yield a superior, neuroprotective HDAC6 inhibitor, tubastatin A. J. Am. Chem. Soc. 2010, 132, 10842–10846. 10.1021/ja102758v. PubMed DOI PMC

Wang X. X.; Wan R. Z.; Liu Z. P. Recent advances in the discovery of potent and selective HDAC6 inhibitors. Eur. J. Med. Chem. 2018, 143, 1406–1418. 10.1016/j.ejmech.2017.10.040. PubMed DOI

Geraldy M.; Morgen M.; Sehr P.; Steimbach R. R.; Moi D.; Ridinger J.; Oehme I.; Witt O.; Malz M.; Nogueira M. S.; Koch O.; Gunkel N.; Miller A. K. Selective inhibition of histone deacetylase 10: hydrogen bonding to the gatekeeper residue is implicated. J. Med. Chem. 2019, 62, 4426–4443. 10.1021/acs.jmedchem.8b01936. PubMed DOI

Kozikowski A. P.; Shen S.; Pardo M.; Tavares M. T.; Szarics D.; Benoy V.; Zimprich C. A.; Kutil Z.; Zhang G.; Barinka C.; Robers M. B.; Van Den Bosch L.; Eubanks J. H.; Jope R. S. Brain penetrable histone deacetylase 6 inhibitor SW-100 ameliorates memory and learning impairments in a mouse model of Fragile X Syndrome. ACS Chem. Neurosci. 2019, 10, 1679–1695. 10.1021/acschemneuro.8b00600. PubMed DOI PMC

Hai Y.; Christianson D. W. Histone deacetylase 6 structure and molecular basis of catalysis and inhibition. Nat. Chem. Biol. 2016, 12, 741–747. 10.1038/nchembio.2134. PubMed DOI PMC

Miyake Y.; Keusch J. J.; Wang L.; Saito M.; Hess D.; Wang X.; Melancon B. J.; Helquist P.; Gut H.; Matthias P. Structural insights into HDAC6 tubulin deacetylation and its selective inhibition. Nat. Chem. Biol. 2016, 12, 748–754. 10.1038/nchembio.2140. PubMed DOI

Porter N. J.; Mahendran A.; Breslow R.; Christianson D. W. Unusual zinc-binding mode of HDAC6-selective hydroxamate inhibitors. Proc. Natl. Acad. Sci. U. S. A. 2017, 114, 13459–13464. 10.1073/pnas.1718823114. PubMed DOI PMC

Porter N. J.; Osko J. D.; Diedrich D.; Kurz T.; Hooker J. M.; Hansen F. K.; Christianson D. W. Histone deacetylase 6-selective inhibitors and the influence of capping groups on hydroxamate-zinc denticity. J. Med. Chem. 2018, 61, 8054–8060. 10.1021/acs.jmedchem.8b01013. PubMed DOI PMC

Mackwitz M. K. W.; Hamacher A.; Osko J. D.; Held J.; Scholer A.; Christianson D. W.; Kassack M. U.; Hansen F. K. Multicomponent synthesis and binding mode of imidazo[1,2- a]pyridine-capped selective HDAC6 inhibitors. Org. Lett. 2018, 20, 3255–3258. 10.1021/acs.orglett.8b01118. PubMed DOI PMC

Vogerl K.; Ong N.; Senger J.; Herp D.; Schmidtkunz K.; Marek M.; Muller M.; Bartel K.; Shaik T. B.; Porter N. J.; Robaa D.; Christianson D. W.; Romier C.; Sippl W.; Jung M.; Bracher F. Synthesis and biological investigation of phenothiazine-based benzhydroxamic acids as selective histone deacetylase 6 inhibitors. J. Med. Chem. 2019, 62, 1138–1166. 10.1021/acs.jmedchem.8b01090. PubMed DOI PMC

Bhatia S.; Krieger V.; Groll M.; Osko J. D.; Ressing N.; Ahlert H.; Borkhardt A.; Kurz T.; Christianson D. W.; Hauer J.; Hansen F. K. Discovery of the first-in-class dual histone deacetylase-proteasome inhibitor. J. Med. Chem. 2018, 61, 10299–10309. 10.1021/acs.jmedchem.8b01487. PubMed DOI PMC

Jochems J.; Boulden J.; Lee B. G.; Blendy J. A.; Jarpe M.; Mazitschek R.; Van Duzer J. H.; Jones S.; Berton O. Antidepressant-like properties of novel HDAC6-selective inhibitors with improved brain bioavailability. Neuropsychopharmacology 2014, 39, 389–400. 10.1038/npp.2013.207. PubMed DOI PMC

Zhang L.; Liu C.; Wu J.; Tao J. J.; Sui X. L.; Yao Z. G.; Xu Y. F.; Huang L.; Zhu H.; Sheng S. L.; Qin C. Tubastatin A/ACY-1215 improves cognition in Alzheimer’s disease transgenic mice. J. Alzheimer's Dis. 2014, 41, 1193–1205. 10.3233/JAD-140066. PubMed DOI

Selenica M. L.; Benner L.; Housley S. B.; Manchec B.; Lee D. C.; Nash K. R.; Kalin J.; Bergman J. A.; Kozikowski A.; Gordon M. N.; Morgan D. Histone deacetylase 6 inhibition improves memory and reduces total tau levels in a mouse model of tau deposition. Alzheimer's Res. Ther. 2014, 6, 12.10.1186/alzrt241. PubMed DOI PMC

Jian W.; Wei X.; Chen L.; Wang Z.; Sun Y.; Zhu S.; Lou H.; Yan S.; Li X.; Zhou J.; Zhang B. Inhibition of HDAC6 increases acetylation of peroxiredoxin1/2 and ameliorates 6-OHDA induced dopaminergic injury. Neurosci. Lett. 2017, 658, 114–120. 10.1016/j.neulet.2017.08.029. PubMed DOI

Wang Z.; Leng Y.; Wang J.; Liao H. M.; Bergman J.; Leeds P.; Kozikowski A.; Chuang D. M. Tubastatin A, an HDAC6 inhibitor, alleviates stroke-induced brain infarction and functional deficits: potential roles of alpha-tubulin acetylation and FGF-21 up-regulation. Sci. Rep. 2016, 6, 19626.10.1038/srep19626. PubMed DOI PMC

Shen S.; Benoy V.; Bergman J. A.; Kalin J. H.; Frojuello M.; Vistoli G.; Haeck W.; Van Den Bosch L.; Kozikowski A. P. Bicyclic-capped histone deacetylase 6 inhibitors with improved activity in a model of axonal Charcot-Marie-Tooth Disease. ACS Chem. Neurosci. 2016, 7, 240–258. 10.1021/acschemneuro.5b00286. PubMed DOI PMC

Vogl D. T.; Raje N.; Jagannath S.; Richardson P.; Hari P.; Orlowski R.; Supko J. G.; Tamang D.; Yang M.; Jones S. S.; Wheeler C.; Markelewicz R. J.; Lonial S. Ricolinostat, the first selective histone deacetylase 6 inhibitor, in combination with bortezomib and dexamethasone for relapsed or refractory multiple myeloma. Clin. Cancer Res. 2017, 23, 3307–3315. 10.1158/1078-0432.CCR-16-2526. PubMed DOI PMC

Sweeney M. D.; Sagare A. P.; Zlokovic B. V. Blood-brain barrier breakdown in Alzheimer disease and other neurodegenerative disorders. Nat. Rev. Neurol. 2018, 14, 133–150. 10.1038/nrneurol.2017.188. PubMed DOI PMC

Gertz M.; Kilford P. J.; Houston J. B.; Galetin A. Drug lipophilicity and microsomal protein concentration as determinants in the prediction of the fraction unbound in microsomal incubations. Drug Metab. Dispos. 2008, 36, 535–542. 10.1124/dmd.107.018713. PubMed DOI

Kim C.; Choi H.; Jung E. S.; Lee W.; Oh S.; Jeon N. L.; Mook-Jung I. HDAC6 inhibitor blocks amyloid beta-induced impairment of mitochondrial transport in hippocampal neurons. PLoS One 2012, 7, e4298310.1371/journal.pone.0042983. PubMed DOI PMC

Fukada M.; Hanai A.; Nakayama A.; Suzuki T.; Miyata N.; Rodriguiz R. M.; Wetsel W. C.; Yao T. P.; Kawaguchi Y. Loss of deacetylation activity of Hdac6 affects emotional behavior in mice. PLoS One 2012, 7, e3092410.1371/journal.pone.0030924. PubMed DOI PMC

Roberds S. L.; Filippov I.; Alexandrov V.; Hanania T.; Brunner D. Rapid, computer vision-enabled murine screening system identifies neuropharmacological potential of two new mechanisms. Front. Neurosci. 2011, 5, 103.10.3389/fnins.2011.00103. PubMed DOI PMC

Zhang H. K.; Eaton J. B.; Yu L. F.; Nys M.; Mazzolari A.; van Elk R.; Smit A. B.; Alexandrov V.; Hanania T.; Sabath E.; Fedolak A.; Brunner D.; Lukas R. J.; Vistoli G.; Ulens C.; Kozikowski A. P. Insights into the structural determinants required for high-affinity binding of chiral cyclopropane-containing ligands to alpha4beta2-nicotinic acetylcholine receptors: an integrated approach to behaviorally active nicotinic ligands. J. Med. Chem. 2012, 55, 8028–8037. 10.1021/jm3008739. PubMed DOI PMC

Gunosewoyo H.; Midzak A.; Gaisina I. N.; Sabath E. V.; Fedolak A.; Hanania T.; Brunner D.; Papadopoulos V.; Kozikowski A. P. Characterization of maleimide-based glycogen synthase kinase-3 (GSK-3) inhibitors as stimulators of steroidogenesis. J. Med. Chem. 2013, 56, 5115–5129. 10.1021/jm400511s. PubMed DOI PMC

Comprehensive Mechanistic View of the Hydrolysis of Oxadiazole-Based Inhibitors by Histone Deacetylase 6 (HDAC6)