Tubastatin A Hydrochloride (TSA HCl) is an effective and specific HDAC6 inhibitor (IC50: 15 nM). It has selectivity (>1000-fold) against all other isozymes except HDAC8 (>57-fold).

HDAC6:15 nM

Tubastatin A 对所有11种HDAC异构体具有较高选择性，除HDAC8外对其他异构体的选择性超过1000倍，在HDAC8上大约有57倍的选择性。在同型半胱氨酸(HCA)诱导的神经退行性测定中，Tubastatin A 以剂量依赖的方式保护神经细胞免受HCA诱导的细胞死亡，从5μM开始显示保护效果，在10μM时几乎完全保护。[1] 在体外实验中，Tubastatin A 在100 ng/mL的剂量下增加Foxp3+ 调节性T细胞（Tregs）对T细胞增殖的抑制。[2] 在C2C12细胞中，Tubastatin A 的处理会导致在肌肉生成过程早期时，由于α-微管蛋白过度乙酰化而妨碍肌管形成；然而，当在肌管中α-微管蛋白过度乙酰化时，则会发生肌管伸长。[3] 最近的研究表明，Tubastatin A 处理通过原子力显微镜（AFM）测试表明增加了细胞弹性，而不对小鼠卵巢癌细胞系MOSE-E和MOSE-L中的微丝或微管网络造成严重变化。[4]

每日处理0.5 mg/kg Tubastatin A 通过抑制HDAC6来促进Tregs的抑制活性，在小鼠的炎症与自身免疫模型中表现出效果，这些模型包括多种实验性结肠炎和完全主要组织相容性复合体（MHC）-不相容心脏异体移植的拒绝反应。[2]

Enzyme Inhibition Assays: Enzyme inhibition assays are performed by the Reaction Biology Corporation, Malvern, PA, using the Reaction Biology HDAC Spectrum platform. (www.reactionbiology.com) The HDAC1, 2, 4, 5, 6, 7, 8, 9, 10, and 11 assays use isolated recombinant human protein; HDAC3/NcoR2 complex is used for the HDAC3 assay. Substrate for HDAC1, 2, 3, 6, 10, and 11 assays is a fluorogenic peptide from p53 residues 379-382 (RHKKAc); substrate for HDAC8 is fluorogenic diacyl peptide based on residues 379-382 of p53 (RHKAcKAc). Acetyl-Lys (trifluoroacetyl)-AMC substrate is used for HDAC4, 5, 7, and 9 assays. Tubastatin A is dissolved in DMSO and tested in 10-dose IC50 mode with 3-fold serial dilution starting at 30 μM. Control Compound Trichostatin A (TSA) is tested in a 10-dose IC50 with 3-fold serial dilution starting at 5 μM. IC50 values are extracted by curve-fitting the dose/response slopes.

Primary cortical neuron cultures are obtained from the cerebral cortex of fetal Sprague-Dawley rats (embryonic day 17) as described previously. All experiments are initiated 24 hours after plating. Under these conditions, the cells are not susceptible to glutamate-mediated excitotoxicity. For cytotoxicity studies, cells are rinsed with warm PBS and then placed in minimum essential medium (Invitrogen) containing 5.5 g/L glucose, 10% fetal calf serum, 2 mM L-glutamine, and 100 μM cystine. Oxidative stress is induced by the addition of the glutamate analogue homocysteate (HCA; 5 mM) to the media. HCA is diluted from 100-fold concentrated solutions that are adjusted to pH 7.5. In combination with HCA, neurons are treated with Tubastatin A at the indicated concentrations. Viability is assessed after 24 hours by MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method.(Only for Reference)