Synthesis and pharmacological characterization of novel histone deacetylase 6 inhibitors

组蛋白脱乙酰酶(HDAC)是一种表观遗传调节因子，催化蛋白质中赖氨酸残基的脱乙酰基。HDAC通常在癌细胞中过度表达，在癌细胞中它们对肿瘤的发生起着决定性的作用。组蛋白去乙酰酶抑制剂(HDACi)抑制蛋白质去乙酰化，改变基因表达和信号转导。因此，癌细胞的生长就会受到影响。HDACi对通常无法治愈的白血病特别有效。广谱HDACi(PAN-HDACi)抑制几种HDAC同工酶，并产生普遍的细胞毒性。同工酶特异性的HDACi产生的副作用更少。在拟议的项目中，我们计划用高级化学方法合成HDAC6抑制剂(Mahboobi教授)，并在体外和基于细胞的生物检测(PD)中对它们进行测试。HDAC6有两个催化域和几个特定的功能和底物。目前还不清楚HDAC6对某些底物的催化活性是否取决于这两个HDAC结构域。细胞骨架蛋白微管蛋白-α和伴侣蛋白HSP90是HDAC6的靶标。去乙酰化的HSP90有助于癌蛋白的折叠，包括白血病融合蛋白BCR-ABL、AML1-ETO和PML-RARpha、突变体Flt3-ITD、泛白血病标记蛋白WT1和致癌p53。这些和HDAC6的癌症相关底物暗示了HDAC6抑制剂的用处。然而，HDAC6的生物学作用仍不完全清楚，也存在一些争议。部分原因是使用了被错误地认为是HDAC6特有的化合物。在初步研究中，我们合成了新型有效的HDAC6抑制剂。它们在几种癌细胞系中表现出很强的细胞毒性和细胞抑制活性。我们还首次证明了Survivin的表达依赖于HDAC6，Survivin是肿瘤发生和化疗耐药的启动子。此外，我们的新抑制剂显示出与伊马替尼联合抑制bcr-abl转化的白血病细胞的细胞毒活性。在分子水平上，这种效应与bcr-abl和WT1的耗尽以及乙酰化微管蛋白-α的积累有关。基于这些数据以及与晶体结构和同源模型的对接研究，我们将合成和表征更有效和特异的HDAC6抑制剂。此外，我们的目标是开发HDAC6的不可逆抑制剂。这类药物是抑制HDAC6的一种新的药理学原理。我们预计，较低的药物剂量将充分阻断HDAC6，因为较少的脱离HDAC6。我们针对HDAC6的新化合物的初步生物学特征将主要考察它们作为治疗急性和髓系白血病的药理药物的潜力。我们将用我们的新型抑制剂收集的数据通常也可以增加我们对HDAC6在肿瘤细胞中的分子效应和生物学功能的理解。

Histone deacetylases (HDACs) are epigenetic regulators catalyzing the deacetylation of lysine residues in proteins. HDACs are often overexpressed in cancer cells, in which they causally contribute to tumorigenesis. Histone deacetylase inhibitors (HDACi) inhibit protein deacetylation and alter gene expression and signaling. Consequently, cancer cell growth becomes compromised. HDACi are particularly effective against leukemia which is often not curable. Broad-range HDACi (pan-HDACi) inhibit several HDAC isoenzymes and produce general cytotoxicity. Isoenzyme-specific HDACi generate fewer side effects. Within the proposed project we plan to synthesize HDAC6 inhibitors with advanced chemistry (Prof. Mahboobi) and to test them in vitro and in cell based biological assays (PD. Dr. Krämer).HDAC6 has two catalytic domains and several specific functions and substrates. It is yet often unclear whether the catalytic activity of HDAC6 against certain substrates depends on both HDAC domains. The cytoskeletal protein tubulin-alpha and the chaperone HSP90 are targets of HDAC6. Deacetylated HSP90 assists the folding of oncoproteins including the leukemia fusion proteins BCR-ABL, AML1-ETO, and PML-RARalpha, mutant FLT3-ITD, the pan-leukemic marker protein WT1, and oncogenic p53. These and further cancer-relevant substrates of HDAC6 suggest the usefulness of HDAC6 inhibitors. The biological role of HDAC6 is though still incompletely understood and there are some controversies. These partly originate from the use of compounds that were erroneously assumed to be specific for HDAC6. In preliminary studies, we have synthesized novel and potent HDAC6 inhibitors. They exhibit potent cytotoxic and cytostatic activity in several cancer cell lines. We also show for the first time that the expression of survivin, a promoter of tumorigenesis and chemoresistance, depends on HDAC6. Moreover, our new inhibitors demonstrate combined cytotoxic activity with Imatinib against BCR-ABL transformed leukemic cells. At the molecular level this effect is linked to the depletion of BCR-ABL and WT1 and an accumulation of acetylated tubulin-alpha.Based on these data as well as on docking studies with crystal structures and homology models, we will synthesize and characterize even more potent and specific HDAC6 inhibitors. Furthermore, we aim to develop irreversible inhibitors for HDAC6. Such agents are a new pharmacological principle of HDAC6 inhibition. We expect that lower drug doses will sufficiently block HDAC6 due to a lesser detachment from HDAC6. The initial biological characterization of our new compounds targeting HDAC6 will mainly investigate their potential as pharmacological agents for the treatment of acute and myeloid leukemia. Data that we will collect with our novel inhibitors can also generally increase our understanding of the molecular effects and biological functions of HDAC6 in tumor cells.