HDAC8

2024年3月8日，杭州师范大学叶向阳/谢恬团队在Journal of Medicinal Chemistry在线发表了题为“Design, Synthesis, and Structure–Activity Relationship of Novel Pyridazinone-Based PARP7/HDACs Dual Inhibitors for Elucidating the Relationship between Antitumor Immunity and HDACs Inhibition”的文章。该团队开发了一系列基于哒嗪酮的PARP7/HDACs双重抑制剂，其中化合物9l具有强效的PARP7/HDACs双重抑制活性，在体内外均表现出优异的抗肿瘤能力。组蛋白去乙酰化酶（HDACs）在染色体结构和基因表达调控中发挥着重要作用，对肿瘤细胞分化、增殖和免疫调节等过程具有许多影响，已被验证为抗肿瘤的重要靶点，目前已有五种药物（1-5）获得上市批准（图1）。聚[二磷酸腺苷（ADP）-核糖]聚合酶7（PARP7）是PARPs家族中单PARP亚家族的成员之一，能使自身和其他底物蛋白发生MARylate反应，即催化单个ADP-核糖（MAR）从烟酰胺腺嘌呤二核苷酸（NAD）共价转移到其底物上。PARP7被证实通过抑制IFN-I信号转导参与细胞膜NA感应和多种病毒RNA感染。抑制PARP7能以免疫细胞依赖的方式诱导肿瘤细胞的IFN-I信号转导，从而导致肿瘤细胞死亡，使得PARP7成为一个极具吸引力的治疗靶点。Atamparib（RBN-2397，图1）是一种口服可用的PARP7选择性抑制剂（IC50< 3 nM），目前正在进行2期临床试验。BN-2397通过激活IFN-I信号通路抑制肿瘤免疫逃逸，从而抑制胰腺癌、卵巢癌、非小细胞肺等多种实体瘤的肿瘤生长。开发同时靶向HDACs和PARP7实现协同抗肿瘤作用具有重要意义。图1 HDACs抑制剂和PARP7抑制剂RBN-2397的结构。基于RBN-2397的双重抑制剂设计作者对临床候选药物RBN-2397与PARP7蛋白的共晶结构进行分析，发现其嘧啶环上的氮与ASP580形成氢键作用；哒嗪酮基团在NAD亚口袋中充当NAD模拟结合，并与PARP7蛋白的残基GLY565、GLN576和SER604形成氢键；哌嗪上的羰基和TYR596形成重要的氢键（图2A）。此外，嘧啶环上的CF3与GLY573形成额外的氢键作用，但其对PARP7活性并不重要。为了能容纳HDACs抑制剂的药效团（异羟肟酸），设计将异羟肟酸取代RBN-2397嘧啶环上的可修饰基团CF3，合成了化合物7具有HDAC1（IC50 = 34.0 nM）、HDAC6（IC50 = 83.7 nM）和PARP7（IC50 = 2.7 nM）的抑制作用（图2B）。图2 （A）RBN-2397与PARP7蛋白共晶结构中的相互作用；（B）第一种PARP7/HDACs双重抑制剂化合物7的结构；（C）化合物7与PARP7蛋白的相互作用；（D）化合物7与PARP7催化结构域结合的表面图；（E）BN-2397（绿色）和化合物7（蓝色）在PARP7中的叠加。随后，作者在此基础上进行异羟肟酸和哒嗪酮支架之间的片段Y和R1的结构改造（图3A），以进一步提升对PARP7、HDAC1和HDAC6的抑制活性。但化合物8a-8g的效力并没有显著提升。接着，作者保留了化合物7的Y和R1区域，在嘧啶环和异羟肟酸之间插入不同长度和类型的linker以提高对HDACs的抑制活性（图3B）。在将linker从具有酰胺官能团的直烷基链、吡唑基烷基链和苄氧基替换为烯丙基苄基后，得到活性最好的化合物9l（对PARP7、HDAC1和HDAC6的IC50分别为3.1 nM、35.0 nM和6.4 nM）。图3 PARP7/HDACs双重抑制剂8和9的设计。在体外细胞活性测试中，化合物9l对NCI-H1373和MV-4-11两种细胞系均显示出优异的抑制活性（≥ 99% at 10 μM，图4），比RBN-2397和SAHA效力更强。且与RBN-2397相比，化合物9l对MV-4-11和U937两种血液肿瘤细胞株显示出更强的抑制活性，与SAHA相比，活性略好。在对PARP7缺乏敏感性的NCI-2066细胞系中，化合物9l显示出比RBN-2397更强的抗细胞增殖活性。图4 PARP7/HDACs双重抑制剂的抗增殖活性测试。化合物9l可恢复IFN-β的产生和组蛋白H3的乙酰化研究表明，HDACs抑制剂，尤其是在用SAHA处理的细胞中，会降低磷酸化STAT1的水平和干扰素刺激基因的表达。作者以RBN-2397作为对照，在用固定浓度为1 μM的RBN-2397处理的NCI-H1373细胞中，加入不同浓度的SAHA显示，SAHA的增加会降低STAT1的磷酸化水平（在SAHA的最高浓度下，磷酸化可完全被抑制），而H3的乙酰化水平则不受影响（图5A）。化合物9l能显著提高p-STAT1和乙酰化H3的水平，而9h对p-STAT1水平没有影响（图 5B）。图5 （A）HDACi对PARP7i（RBN-2397）诱导p-STAT1水平的影响；（B）RBN-2397和SAHA组合以及不同浓度的9h和9l对Ac-H3和p-STAT1水平的影响。进一步的实验发现，随着9l浓度的增加，STAT1磷酸化水平、H3乙酰化水平和微管蛋白乙酰化水平逐渐升高（图6）。化合物9l通过同时抑制PARP7和HDACs可导致H3乙酰化水平升高，IFN-β和CXCL10的mRNA表达水平升高。这些发现意味着化合物9h只抑制组蛋白去乙酰化酶的活性，而化合物9l则具有同时抑制PARP7和HDAC的能力。图6 化合物对IFN-β和CXCL10表达水平的影响以及化合物对p-STAT1水平、Ac-H3和Ac-Tubulin的影响。化合物9l对PARPs和HDACs的选择性探究化合物9l具有强效的PARP7和HDAC6抑制活性，并且相比HDAC1，对HDAC6具有一定程度的选择性。化合物9l具有与SAHA相似的选择性，它对HDAC6的抑制活性（IC50 = 6.4 nM）略好于HDAC1-3（IC50 = 30~35 nM）；对HDAC8的活性较低，对HDAC11的活性最差（图7）。在对PARP同工酶的抑制活性中，化合物9l与RBN-2397具有相似的选择性。它对PARP7和PARP2的活性水平相同，但对PARP1、PARP5A和PARP5B的活性略高。图7 化合物9l对HDACs和PARPs的抑制活性。化合物9l的体内活性探究化合物9l以5 mg/kg的剂量通过静脉注射（iv）途径给药。结果显示，该化合物的清除率高达94.2 mL/min/kg，半衰期（t1/2）为13.3 h（表 8A）。口服10 mg/kg剂量的9l表现出极低的血浆暴露量，导致口服生物利用度较低（%F低至2.4%）。初步的PK评估表明，在异种移植小鼠模式的体内抗癌疗效概念验证（POC）研究中，口服给药是首选的给药途径。在BALB/c裸鼠CT26异种移植模型中，腹腔给药治疗17天，能明显减少肿瘤的生长（图8B、C）。当给药剂量为75 mg/kg时，9l的抗肿瘤效果最好，对肿瘤生长的抑制作用高于SAHA（50 mg/kg）和RBN-2397（50 mg/kg）。此外，实验小鼠均未出现体重明显减轻等副作用。图8 （A）化合物9l的PK表征；（B、C）在CT26荷瘤小鼠体内的抗肿瘤作用。总结在本研究中，作者首次设计并合成了一系列新型PARP7/HDACs双重抑制剂。在这些新化合物中，化合物9l对PARP7和HDACs在酶水平上都有很强的抑制活性。为了揭示同时在分子水平上靶向PARP7和HDACs能否产生协同药理作用，研究人员对化合物9l进行了一系列体外和体内实验。结果表明，同时靶向PARP7和HDACs可能无法实现协同抗癌效果，因为这两种途径的作用会相互抵消。不过，先导物9l是有史以来首次报道的PARP7/HDACs双重抑制剂，可作为进一步生物学研究的分子工具。原文链接：https://pubs.acs.org/doi/10.1021/acs.jmedchem.4c00090声明：发表/转载本文仅仅是出于传播信息的需要，并不意味着代表本公众号观点或证实其内容的真实性。据此内容作出的任何判断，后果自负。若有侵权，告知必删！长按关注本公众号   粉丝群/投稿/授权/广告等请联系公众号助手 觉得本文好看，请点这里↓

Researchers have been working to better understand what drives melanoma brain metastasis. They now report on the identification of a cell signaling pathway that regulates the metastatic spread of melanoma cells to the brain. Melanoma is the deadliest form of skin cancer because of its ability to quickly grow and spread throughout the body. More than half of those with advanced melanoma will see the disease spread to the brain, where it rapidly progresses, often leading to death in only three to four months. Researchers in Moffitt Cancer Center's Donald A. Adam Melanoma and Skin Cancer Center of Excellence have been working to better understand what drives melanoma brain metastasis. In a new study published in Nature Communications, they report on the identification of a cell signaling pathway that regulates the metastatic spread of melanoma cells to the brain. Melanoma tumors are composed of subgroups of cells with different gene expression patterns with varied abilities to invade surrounding tissues and survive anticancer treatments. It is unclear how these different melanoma subgroups contribute to tumor development and progression. In previous studies, Moffitt researchers determined that the protein HDAC8 regulated resistance to BRAF and MEK inhibitors commonly used to treat melanoma. HDAC8 removes chemical modifications called acetyl groups from other proteins, leading to alterations in gene expression patterns. The Moffitt team hypothesized that HDAC8 may also be involved in the regulation of gene expression patterns of melanoma cell subgroups. The researchers performed laboratory experiments and demonstrated that HDAC8 activity increased melanoma cell survival under stress conditions, including low oxygen, UV radiation, and BRAF/MEK inhibitor treatment. HDAC8 activity also changed the gene expression pattern of melanoma cells and caused the cells to develop characteristics associated with cell subgroups that were able to migrate into and invade surrounding sites. Their pre-clinical experiments found that increased HDAC8 expression and activity enhanced the ability of melanoma cells to metastasize to the brain, while no significant impact was observed in the number of metastatic tumors to other organs, such as the liver or lung. The researchers further investigated the molecular pathways of HDAC8-mediated brain metastasis and discovered that HDAC8 chemically modified the protein EP300, which subsequently caused cells to develop invasive characteristics. The researchers confirmed the importance of EP300 to melanoma brain metastases by showing that increased expression of EP300 decreased cell invasion and caused melanoma cells to be more sensitive to cell death. "These data show the importance of HDAC8 and EP300 activity to melanoma cell invasion to the brain and suggest that agents that target these pathways may inhibit brain metastasis," said Keiran Smalley, Ph.D., lead study author and director of Moffitt's Melanoma and Skin Cancer Center of Excellence. "Our work provides the first evidence that stress induced HDAC8 is a regulator of an invasive melanoma cell state that leads to increased brain metastasis." This study was supported by the National Institutes of Health (R01CA262483, R21CA267141, P30CA076292, P30CA247796) and by state appropriations provided in Florida Statute §381.915.

Isaac Hanson Francis deSouza was forced to step down as CEO following the company’s legal troubles. Photo: Victor J. Blue/Bloomberg via Getty Images US-based sequencing and diagnostics company Illumina reported in its Q2 filing that it is under investigation by the Securities and Exchange Commission (SEC) over its $8bn acquisition of Grail, the manufacturer of a proprietary multi-cancer early detection (MCED) test. This is the latest in a line of lawsuits faced by the firm over its decision. Grail began as a branch of Illumina and was then spun out as an independent start-up in 2016, though the parent retained a 12% stake. In 2020, Illumina opted to reacquire the company and announced a deal for $7.1bn. Recommended Reports Reports Histone Deacetylase 8 (Histone Deacetylase Like 1 or HDAC8 or EC 3.5.1.98) Drugs in Development b... GlobalData Reports Pharmaceuticals Industry Deals and Trends in April 2021 - Partnerships, Licensing, Investments, M... GlobalData View all The saga is long, complicated and neatly outlined here , but by June of this year Illumina had lost its CEO, been fined the maximum 10% of annual turnover by the EU (with an indication that a greater figure would have been sought if possible) and been ordered to divest Grail by the Federal Trade Commission (FTC). The issue at fault is twofold: at a base level Illumina stands accused of anticompetitive practices through the purchase, as the MCED test that Grail produces – and others like it by competitors – can only be processed by technology of which Illumina is the largest provider. The second problem is that, despite being told in July 2021 that the merger was under investigation by the European Commission, Illumina completed the deal a month later. The company has argued that its acquisition is a moral one that will accelerate the production of lifesaving tests and that it had to be completed within its initial deal period – ending in November 2021 – to avoid having to renegotiate, thereby slowing down the expansion. Regulators see it differently. The European Commission described the acquisition as “an unprecedented and very serious infringement undermining the effective functioning of the EU merger control system.” It found that the company had made a strategic decision to continue with the merger despite knowing it was possible it would be disallowed as the break-up fee for failing the takeover would outweigh the loss of profit from the divestment. This appears to be an accelerating trend in the medical field, following the news of UnitedHealth’s legal troubles . Legislation has not yet caught up to the "tidal wave" of mergers that the FTC reported in 2021, meaning investigations are either being suspended or cut short as regulators scramble to get on top. Companies may be taking advantage of this situation to pursue a more aggressive mergers and acquisitions (M&A) strategy than they would normally, knowing that the profits outweigh the costs of potential future fines. When companies beg forgiveness and not permission, there is a danger that anti-competitive practices will go unnoticed. Our signals coverage is powered by GlobalData’s Thematic Engine, which tags millions of data items across six alternative datasets - patents, jobs, deals, company filings, social media mentions and news — to themes, sectors and companies. These signals enhance our predictive capabilities, helping us to identify the most disruptive threats across each of the sectors we cover and the companies best placed to succeed.