BRD4 x BRD7 x BRD9

近期，密歇根大学王少萌团队在Journal of Medicinal Chemistry杂志上发表了题为“Discovery of CW-3308 as a Potent, Selective, and Orally Efficacious PROTAC Degrader of BRD9”的研究论文。该研究报道了一个高效的靶向降解BRD9的PROTAC分子CW-3308，该分子在小鼠体内实现了91%的口服生物利用度，单次口服剂量有效地将滑膜肉瘤HS-SY-II异种移植肿瘤组织中的BRD9蛋白降低了>90%，并有效抑制小鼠HS-SY-II异种移植瘤的生长。 背景 BRD9在非小细胞肺癌，肝细胞癌，小儿横纹肌瘤，前列腺癌，宫颈癌等多种癌症中过表达，是一个潜在的癌症治疗靶点。研究表明，SMARCB1缺失的滑膜肉瘤和恶性横纹肌样瘤细胞的存活能力对BRD9的依赖性提高。目前针对滑膜肉瘤和恶性横纹肌样肿瘤的治疗方案是有限的，通常涉及蒽环类药物和帕唑帕尼，迫切需要新的治疗方法。因此，靶向BRD9是治疗滑膜肉瘤和恶性横纹肌瘤的一种有前景的治疗策略。 目前已有多类靶向BRD9溴结构域的高亲和力小分子抑制剂被研发出来，然而，这种抑制剂不能够完全消除由BRD9调控的基因转录功能并且不能完全抑制滑膜肉瘤细胞的生长。由于序列同源性紧密，BRD9的溴域抑制剂也会与BRD7和BRD4表现出结合亲和力。为了克服这些缺点，多种靶向降解BRD9的PROTAC被报道，如：dBRD9、VZ185、QA-68、CFT8634、FHD-609、C6。其中CFT8634（口服，NCT05355753）和FHD-609（静脉注射，NCT04965753）进入临床研究阶段用于研究滑膜肉瘤和SMARCB1缺失的实体瘤患者。然而，Ⅰ期临床研究中发现，FHD-609引发了四级QTc延长，并在2023年4月被提出暂停临床研究。CFT8634作为单药治疗药物因在Ⅰ期临床阶段缺乏足够的效力也处于暂停状态。因此，目前需要开发新的BRD9降解剂。 设计优化思路 考虑到目前进入临床研究阶段的可口服的PROTAC分子选用的是CRBN作为E3连接酶，为了实现开发口服可利用的小分子，作者依旧采用CRBN配体作为E3连接酶配体设计PROTAC。因此，作者对其之前开发的两种新的CRBN配体TX-11-Me和TX-16-Me进行评估，发现其保持了与传统的CRBN配体沙利度胺和来那度胺相似的亲和力，其细胞渗透性良好，并且表现出优秀的药代动力学特性。于是，作者选择TX-11-Me和TX-16-Me作为CRBN配体进一步开发可口服的BRD9降解剂。 作者选用抑制剂BI-7273作为BRD9降解剂的另一个靶蛋白招募配体，通过共晶结构确定了BI-7273的二甲基胺基基团为溶剂暴露区，是一个合适的连接位点。 作者首先利用TX-11-Me和BI-7273作为靶蛋白招募配体设计了化合物15-18。其中，化合物18和降解剂dBRD9相比提升了近三倍的降解活性。但是18在口服给药条件下的血浆暴露量极低（< 5 ng/mL）。这可能由于化合物18由两种碱性氨组成，这导致其大体积分布和高清除率，从而限制其口服血浆暴露。 因此，作者将其中一个氨基转化成酰胺，并设计合成了19-22。酰胺键的引入在化合物18的基础上进一步提升了化合物的降解效果。但是，以化合物20为例，其血浆暴露量仍然很低并且微粒体稳定性差。 考虑到相对于TX-11-Me的中等外排率，TX-16-Me则没有外排问题，因此，作者将TX-16-Me作为新的CRBN招募配体设计合成了化合物23-26。虽然化合物的降解效力有所降低，但是以23和26为代表的化合物表现出了较好的微粒体稳定性和半衰期。并且在剂量为10 mg/kg的小鼠中，化合物26表现出良好的口服血浆暴露，而化合物23表现出极低的口服血浆暴露。 综上，作者选用化合物26，对其BRD9蛋白质招募结构BI-7273进行了进一步结构优化。共晶结构显示BI-7273的N-甲基周围还有额外的空间可供进行结构修饰。因此作者设计合成了化合物27-42。 根据化合物的降解效力，作者挑选了27，29，37进行了口服暴露量的评估。其中化合物27（CW-3308）达到了更优的口服血浆暴露量。 进一步的微粒体和血浆稳定性以及体内PK测定表明，化合物CW-3308可作为候选化合物进行后续研究。 CW-3308的生物活性评估 CW-3308在横纹肌样肿瘤细胞系G401和滑膜肉瘤细胞系HS-SY-II中以浓度依赖的方式实现了对BRD9的选择性降解。 CW-3308能够以时间依赖的方式实现对BRD9的快速降解，在1 h内实现> 90%的降解效果，并且是通过PROTAC作用机制实现的降解效果。 作者测试了CW-3308对BRD9的亲和力：CW-3308与抑制剂BI-7273对BRD9表现出相似的亲和力，并且在10 微摩尔浓度条件下对BRD4没有表现出明显的亲和力。 CW-3308在细胞培养基中的稳定性较低。这表明CW-3308的降解效果是在极短的暴露时间下实现的，体现了CW-3308作为PROTAC的高效性。 相较于抑制剂BI-7273和来那度胺，CW-3308能够有效抑制G401细胞和HS-SY-Ⅱ细胞的生长。即使CW-3308在细胞培养基中的稳定性低，也能实现对肿瘤细胞有效的生长抑制。 药效学研究表明，CW-3308以50 mg/kg口服给药条件下在3 h实现了92.5%的降解，而在24h 只实现了45%的降解。药物浓度分析显示，CW-3308在3 h的血浆和肿瘤暴露量较高，而在24 h较低，与肿瘤组织中BRD9降解数据一致。 CW-3308以25 mg/kg和50 mg/kg的给药剂量下分别实现了57%和60%的肿瘤生长抑制，并且表现出良好的耐受性。 最后作者预测了CW-3308与BRD9和CRBN形成的三元复合物之间的相互作用。其中，BRD9蛋白上ZA环的ILE53的主链羰基与CRBN蛋白上TYR357的羟基之间距离为1.91 Å，揭示了强烈的蛋白质-蛋白质相互作用。预测得到的CW-3308:BRD9:CRBN三元复合物为新型BRD9降解剂的设计提供了结构模型。 综上所述，CW-3308是一种有前景的靶向降解BRD9的先导化合物，可用于广泛评估和优化BRD9降解物的开发，用于治疗滑膜肉瘤和横纹肌样肿瘤，以及其他BRD9依赖性人类疾病。 原文链接：https://doi-org.libproxy1.nus.edu.sg/10.1021/acs.jmedchem.4c00971 药物筛选中心（暨南大学）对外提供新药筛选服务 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↓

PRESS RELEASE Amphista Therapeutics unveils new differentiated mechanism of action for BRD9 degradation at 2024 Protein Degradation in Focus Symposium First presentation of a novel mechanism of action for the degradation of BRD9 – completely differentiated from cereblon- or VHL-based PROTACs Amphista’s bifunctional BRD9 degraders selectively induce BRD9 to DCAF16 proximity and serve as molecular glues, leading to strong and rapid degradation of BRD9 Discovery further underscores Amphista’s proprietary chemistry and high-quality science which is being applied to the discovery and development of additional targeted glues beyond BRD9 Cambridge, UK, May 23, 2024 – Amphista Therapeutics (“Amphista”), a leader in next generation targeted protein degradation (TPD) approaches, today announces the unveiling of a new mechanism of action for the degradation of BRD9, an emerging target in oncology, that is differentiated from cereblon- or VHL-based PROTACs, during an oral presentation at the 2024 Protein Degradation in Focus Symposium held in Dundee, UK. This significant news for the TPD field builds upon an earlier announcement by Amphista of two compelling data sets demonstrating in vivo efficacy and central nervous system (CNS) activity of its mechanistically differentiated protein degraders. At today’s presentation titled “Degradation of BRD9 by a novel targeted glue”, Dr Andrea Testa, Scientific Co-Founder and Senior Director, Discovery Chemistry showed data from proteomic and genetic validation studies which demonstrate that Amphista’s bifunctional degraders induced degradation of BRD9 by target-assisted E3 ligase recruitment. This novel mechanism selectively induces the proximity of BRD9 to DCAF16 and serves as a molecular glue. DCAF16 is a cullin-RING E3 ligase which can facilitate degradation of proteins of interest via ternary complex formation. Induction of this complex leads to deep and rapid degradation of BRD9 in vivo. Louise Modis, Chief Scientific Officer of Amphista Therapeutics, said: “The discovery of a novel mechanism for BRD9 degradation, differentiated from cereblon- or VHL-based PROTACs, is a testament to Amphista’s proprietary chemistry and know-how. Our ability to translate this novel approach into high-quality, drug-like compounds with oral bioavailability and activity in vivo is truly exciting. As we continue to spearhead our efforts to advance new TPD approaches, our goal remains clear – to expand the offering of TPD medicines beyond CRBN and VHL-based agents and bring effective treatments to patients in areas of high unmet need.” In the results presented, Amphista’s sub-nanomolar BRD9 degraders showed activity in both solid and liquid cancer cell lines and demonstrated a high degree of selectivity over 8,000 proteins quantified by proteomics, including the closely related proteins BRD7 and BRD4. Notably, these compounds induced BRD9 degradation in a mouse xenograft model illustrating Amphista’s BRD9 degraders are orally bioavailable and active in vivo. This is the first-time in vivo degradation of BRD9 has been demonstrated via DCAF16, underscoring the potential therapeutic application of Amphista’s bifunctional degraders. Building on this knowledge, Amphista is applying its proprietary scientific know-how and expertise to the development of targeted glues which induce degradation of different targets, expanding the opportunity beyond BRD9. Future announcements will provide details of additional pipeline targets as part of Amphista’s ambitions to develop a first- and/or best-in-class portfolio of degraders with leading physicochemical properties. About Amphista Therapeutics Amphista Therapeutics is focused on transforming the lives of patients with severe diseases, including cancer and neurodegenerative disease, through the advancement of next generation targeted protein degradation (TPD) medicines. Amphista is applying its proprietary warhead chemistry and mechanistic know-how to generate bifunctional targeted glues with a differentiated mechanism and leading physicochemical properties. Its portfolio offers the potential for first- and/or best-in-class assets and expanding the offering of TPD medicines beyond CRBN and VHL-based agents. Founded by Advent Life Sciences, Amphista is a spin-out of TPD expert Professor Alessio Ciulli’s laboratories at the University of Dundee. Amphista is funded by leading life science investors including Forbion, Gilde Healthcare, Novartis Venture Fund, Advent Life Sciences, Eli Lilly & Company and The Dementia Discovery Fund, and also has strategic collaborations with Bristol Myers Squibb and Merck. For more information, please visit: Amphista and the Amphista logo are all trademarks or registered trademarks of Amphista Therapeutics Limited. For more information please contact: ICR Consilium Amber Fennell, Namrata Taak Email: Amphista@consilium-comms.com Tel: +44 (0)20 3709 5813