Tubastatin A HCl (AG-CR-13900, TubA)

Alias: AG-CR-13900; TubA; Tubastatin A hydrochloride; Tubastatin A HCl; TSA HCl

Cat No.:V0281 Purity: =98.73%

COA Product Data Instructions for use SDS

Tubastatin A HCl (AG-CR-13900, TubA) Chemical Structure CAS No.: 1310693-92-5
Product category: HDAC

This product is for research use only, not for human use. We do not sell to patients.

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Other Forms of Tubastatin A HCl (AG-CR-13900, TubA):

Tubastatin A (TubA, AG-CR1-3900)

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Top Publications Citing lnvivochem Products

Nature 2021, 597(7874):119-125.

Cell 2020,183:1202-1218.e25.

Science Adv 2022: 8, eabm9427.

Nature 597, 119–125 (2021)

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Immunity 2023,56(3):620-634.e11.

J Am Chem Soc 2022,144:21831-21836.

Cell 2020,183:1202-1218.e25.

Science Adv 2022:8,eabm9427.

Nature 2021,597(7874):119-125.

Cell 2020,183:1202-1218.e25.

PNAS Nexus 2022,1(3):pgac063.

ACS Nano 2023,17(10):9110-9125.

Biomaterial 2023 Sep16:302:122333.

InvivoChem's Tubastatin A HCl (AG-CR-13900, TubA) has been cited by 1 publication

[1] Scientific Reports | (2020) 10:6064

Purity & Quality Control Documentation

Current batch：

Purity: =98.73%

COA

MSDS

NMR

HPLC

Instructions

Product Description
Bioactivity & Protocols
Physicochemical Properties
Solubility Data
Calculators
Biological Data

Product Description

Tubastatin A HCl, the hydrochloride salt of Tubastatin A (also known as TubA, AG-CR-13900), is a tubacin analog that acts as a potent and specific inhibitor of histone deacetylase 6 (HDAC6) with potential antitumor, neuroprotective and anti-inflammatory activities. Its selectivity (>1,000-fold) for inhibiting HDAC6 is greater than that of other HDAC isoforms, with the exception of HDAC8, whose IC50 is 0.9 μM.

Biological Activity I Assay Protocols (From Reference)

Targets

HDAC6 ( IC50 = 15 nM ); HDAC8 ( IC50 = 854 nM ); HDAC1 ( IC50 = 16400 nM )

ln Vitro

In vitro activity: Tubastatin A is largely selective for each of the 11 HDAC isoforms and retains over 1000-fold selectivity against all isoforms, with the exception of HDAC8, where selectivity is only about 57 layers. Tubastatin A initiates dose-dependent protection against homocysteic acid (HCA)-induced neuronal cell death as early as 5 μM and achieves near-complete protection at 10 μM in assays for HCA-induced neurodegeneration[1]. Tubastatin A suppresses T cell proliferation in vitro at 100 ng/mL by increasing Foxp³⁺ T-regulatory cells (Tregs)[2]. Alpha-tubulin hyperacetylation early in the myogenic process would impair myotube formation in CC12 cells treated with Tubastatin A; however, myotube elongation happens when alpha-tubulin is hyperacetylated in myotubes[3]. According to a recent study, treatment with tubastatin A increases cell elasticity as measured by atomic force microscopy (AFM) tests in mouse ovarian cancer cell lines MOSE-E and MOSE-L[4] without significantly altering the actin microfilament or microtubule networks.

ln Vivo

In mouse models of inflammation and autoimmunity, such as multiple forms of experimental colitis and fully major histocompatibility complex (MHC)-incompatible cardiac allograft rejection, daily treatment with Tubastatin A at 0.5 mg/kg inhibits HDAC6 to promote Tregs suppressive activity[2].

Enzyme Assay

The Reaction Biology HDAC Spectrum platform is utilized for the execution of enzyme inhibition experiments. Isolated recombinant human protein is utilized in the HDAC1, 2, 4, 5, 6, 7, 8, 9, 10, and 11 assays; the HDAC3/NcoR2 complex is utilized in the HDAC3 test. Fluorogenic peptide derived from p53 residues 379–382 (RHKKAc) serves as the substrate for HDAC1, 2, 3, 6, 10, and 11 assays; fluorogenic diacyl peptide derived from p53 residues 379–382 (RHKAcKAc) serves as the substrate for HDAC8. For the HDAC4, 5, 7, and 9 assays, acetyl-Lys (trifluoroacetyl)-AMC substrate is utilized. After dissolving tubastatin A in DMSO, it is tested in 10-dose IC50 mode using a 3-fold serial dilution regimen that begins at 30 μM. Trichostatin A (TSA), the control compound, is tested in a 10-dose IC50 using a 3-fold serial dilution that begins at 5 μM. Curve-fitting the dose/response slopes yields IC50 values.

Cell Assay

The cerebral cortex of fetal Sprague-Dawley rats (embryonic day 17) is used to cultivate primary cortical neurons. Twenty-four hours after plating, all experiments are started. Glutamate-mediated excitotoxicity cannot harm the cells in these circumstances. Cells are washed with warm PBS before being put in minimum essential medium with 5.5 g/L glucose, 10% fetal calf serum, 2 mM L-glutamine, and 100 μM cystine for cytotoxicity investigations. The glutamate analogue homocysteate (HCA; 5 mM) is added to the media to cause oxidative stress. HCA is prepared by diluting solutions that have been concentrated 100 times and adjusted to pH 7.5. Neurons are treated with Tubastatin A at the indicated concentrations in addition to HCA. The MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) is used to determine viability after a 24-hour period.

Animal Protocol

In adoptive transfer and dextran sodium sulfate (DSS) models of colitis, the effects of HDAC6 targeting are assessed in groups of ten mice each. For five days, WT B6 mice's pH-balanced tap water is supplemented with freshly made 4% (wt/vol) DSS every day. Colitis is evaluated by daily monitoring of body weight, stool consistency, and fecal blood. Mice are treated daily for 7 days with either tubacin or niltubacin (0.5 mg/kg of body weight/day, i.p.). Hemoloccult feces are graded as 0 (absent), 2 (occult), or 4 (gross). Stool consistency is graded as 0 (hard), 2 (soft), or 4 (diarrhea). In order to evaluate the prevention of colitis in a T cell-dependent model, B6/Rag1−/− mice receive an intraperitoneal injection of CD⁴⁺ CD45RBhi T cells (1×10⁶) isolated from WT mice using magnetic beads (>95% cell purity, flow cytometry) along with CD4+ CD²⁵⁺ Tregs (1.25×10⁵) isolated from HDAC6^−/− or WT mice using magnetic beads (>90% Treg purity, flow cytometry). The mice are then observed every two weeks for signs of colitis. In order to evaluate treatment for established T cell-dependent colitis, CD⁴⁺ CD45RBhi cells (1×10⁶) are intraperitoneally injected into B6/Rag1^−/− mice. After colitis manifests, mice are also given treatment with HDAC6i (tubastatin A) or HSP90i (17-AAG) or CD⁴⁺ CD²⁵⁺ Tregs (5×10⁵ cells), which were isolated from HDAC6^−/− or WT mice as previously described. The mice's continued weight loss and the consistency of their feces are observed. When the study comes to an end, paraffin sections of colons stained with hematoxylin and eosin or Alcian Blue are either immunoperoxidase stained for Foxp³⁺ Treg infiltration or graded histologically.

References

[1]. Rational Design and Simple Chemistry Yield a Superior, Neuroprotective HDAC6 Inhibitor, Tubastatin A J. Am. Chem. Soc., 2010, 132 (31), pp 10842-10846.

[2]. Histone deacetylase 6 and heat shock protein 90 control the functions of Foxp3(+) T-regulatory cells. Mol Cell Biol. 2011 May;31(10):2066-78.

[3]. Dysferlin interacts with histone deacetylase 6 and increases alpha-tubulin acetylation. PLoS One. 2011;6(12):e28563.

[4]. Actin filaments play a primary role for structural integrity and viscoelastic response in cells. Integr Biol (Camb). 2012 May;4(5):540-9.

[5]. HDAC6 Inhibition Promotes Transcription Factor EB Activation and Is Protective in Experimental Kidney Disease. Front Pharmacol. 2018 Feb 1;9:34.

[6]. Target deconvolution of HDAC pharmacopoeia reveals MBLAC2 as common off-target. Nat Chem Biol. 2022 Apr 28.

These protocols are for reference only. InvivoChem does not independently validate these methods.

Physicochemical Properties

Molecular Formula	C20H21N3O2.HCL
Molecular Weight	371.86
Exact Mass	371.14
Elemental Analysis	C, 64.60; H, 5.96; Cl, 9.53; N, 11.30; O, 8.60
CAS #	1310693-92-5
Related CAS #	1310693-92-5 (HCl); 1252003-15-8
Appearance	Solid powder
SMILES	CN1CCC2=C(C1)C3=CC=CC=C3N2CC4=CC=C(C=C4)C(=O)NO.Cl
InChi Key	LJTSJTWIMOGKRJ-UHFFFAOYSA-N
InChi Code	InChI=1S/C20H21N3O2.ClH/c1-22-11-10-19-17(13-22)16-4-2-3-5-18(16)23(19)12-14-6-8-15(9-7-14)20(24)21-25;/h2-9,25H,10-13H2,1H3,(H,21,24);1H
Chemical Name	N-hydroxy-4-[(2-methyl-3,4-dihydro-1H-pyrido[4,3-b]indol-5-yl)methyl]benzamide;hydrochloride
Synonyms	AG-CR-13900; TubA; Tubastatin A hydrochloride; Tubastatin A HCl; TSA HCl
HS Tariff Code	2934.99.9001
Storage	Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility Data

Solubility (In Vitro)

DMSO: 10.8~74 mg/mL (29.0~199.0 mM)

Water: <1 mg/mL

Ethanol: <1 mg/mL

Solubility (In Vivo)

1% DMSO+30% polyethylene glycol+1% Tween 80: 30 mg/mL

(Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.6892 mL	13.4459 mL	26.8918 mL
	5 mM	0.5378 mL	2.6892 mL	5.3784 mL
	10 mM	0.2689 mL	1.3446 mL	2.6892 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.

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Molecular Weight Calculator allows you to calculate the molar mass and elemental composition of a compound, as detailed below:

Note: Chemical formula is case sensitive: C12H18N3O4 c12h18n3o4
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In vivo Formulation Calculator (Clear solution)

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Calculation results

Working concentration： mg/mL；

Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.

Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O，mix and clarify.

Note： (1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.

Biological Data