DISCOVERY OF FIRST-IN-CLASS NSD2 DEGRADERS FOR CANCER THERAPY

摘要 表观遗传失调是癌症的一个新的标志，也是与癌症相关的例子 表观基因组调节蛋白的过度表达、突变、易位或异常募集 正在迅速崛起。核受体结合集结构域2(NSD2)是表观遗传学中的关键分子 调节，已知其能力单甲基化和二甲基化组蛋白3(H3K36)的赖氨酸36。此标记已关联 H3K36me2转录活跃，H3K36me2水平升高导致正常沉默基因的异常激活。 因此，NSD2是一种有效的癌蛋白，并已被认为是多种肿瘤的治疗靶点。 癌症。研究发现，在1000名儿童癌症患者中，NSD2基因是突变频率最高的基因之一 代表21种不同儿科癌症亚型的基因组。儿童急性淋巴细胞白血病的临床研究 (所有)细胞系和患者样本显示功能突变(E1099K)的反复获得，导致 H3K36me2水平升高。此外，NSD2参与了多发性骨髓瘤(MM)的发病 是第二常见的血癌。15%-20%的多发性骨髓瘤患者存在染色体易位 4和14[t(4；14)(p16.3；q32)]，其中NSD2被认为是主要致癌驱动因素。 除了其催化SET结构域之外，NSD2还包含几个甲基赖氨酸(KME)阅读器结构域，包括 四个PHD指和两个PWWP结构域，被认为在传播H3K36me2和 将NSD2招募到其致癌靶基因。尽管NSD2清楚地显示出作为治疗靶点的希望 肿瘤学方面，到目前为止还没有小分子配体的报道，目前还不清楚哪些结构域是 “可用药”，在促进肿瘤发生方面起着最关键的作用。创造高品质的NSD2化学品 探针显然将提供急需的洞察力，并作为评估临床前目标的关键试剂 有效性，同时提供了立即过渡到药物发现努力的潜力。我们最近的努力是 靶向NSD2的N端PWWP结构域已成功地产生了有效的先导化合物。 随着新型NSD2抑制剂的问世，我们的目标是探索NSD2蛋白降解剂的发展 它利用了小分子抑制剂的优势，同时超越了传统的 受体药理学。这种新兴的药物发现策略特别适合NSD2，因为它 是一种大的多结构域蛋白质，我们缺乏最佳靶向结构域的知识，2)抑制单个 结构域可能不足以复制NDS2基因敲除结果，更广泛地防止 肿瘤发生和3)KME阅读器结构域通常被视为困难的药物靶点。总体目标 这项建议的目的是应用药物化学、化学生物学和癌症生物学方法来发现 一流的NSD2双功能降解器，以更好地了解NSD2肿瘤生物学，评估NSD2 临床前目标有效性，并作为潜在的治疗药物。

ABSTRACT Epigenetic dysregulation is an emerging hallmark of cancers, and examples of cancer associated overexpression, mutation, translocation, or aberrant recruitment of the proteins which regulate the epigenome are rapidly emerging. NSD2 (nuclear receptor-binding SET domain-containing 2) is a key player in epigenetic regulation, known for its ability to mono- and dimethylate lysine 36 of histone 3 (H3K36). This mark is associated with active transcription, and elevated levels of H3K36me2 lead to aberrant activation of normally silenced genes. Consequently, NSD2 is a potent oncoprotein and has been implicated as a therapeutic target for a variety of cancers. NSD2 was found to be among the most frequently mutated genes across 1,000 pediatric cancer genomes representing 21 different pediatric cancer subtypes. A study in pediatric acute lymphoblastic leukemia (ALL) cell lines and patient samples revealed a recurring gain of function mutation (E1099K) which resulted in elevated H3K36me2 levels. Furthermore, NSD2 is involved in the pathogenesis of multiple myeloma (MM) which is the second most common blood cancer. 15-20% of MM patients carry a translocation between chromosomes 4 and 14 [t(4;14)(p16.3;q32)] of which NSD2 is thought to be the primary oncogenic driver. In addition to its catalytic SET domain, NSD2 contains several methyl-lysine (Kme) reader domains including four PHD fingers and two PWWP domains which are thought to be critical in propagating H3K36me2 and recruiting NSD2 to its oncogenic target genes. Although NSD2 clearly shows promise as a therapeutic target in oncology, no small molecule ligands have been reported to date and it remains unclear as to which domains are `druggable' and function most critically in promoting tumorigenesis. The creation of high-quality NSD2 chemical probes will clearly provide much needed insight and serve as critical reagents in assessing preclinical target validity, while providing the potential for an immediate transition to a drug discovery effort. Our recent efforts to target the N-terminal PWWP domain of NSD2 have been successful in producing potent lead compounds. With novel NSD2 inhibitors in hand, we aim to explore the development of NSD2 protein degradation reagents which capitalize on advantages of small molecule inhibitors while moving beyond the limitations of traditional receptor pharmacology. This emerging drug discovery strategy is particularly well-suited for NSD2 because 1) it is a large multi-domain protein and we lack knowledge of the best domain to target, 2) inhibition of a single domain may not be sufficient to phenocopy NDS2 genetic knockdown results and more broadly prevent tumorigenesis, and 3) Kme reader domains are generally viewed as difficult drug targets. The overall objective of this proposal is to apply medicinal chemistry, chemical biology, and cancer biology approaches to discover first-in-class NSD2 bifunctional degraders in order to better understand NSD2 cancer biology, to assess NSD2 preclinical target validity, and as potential therapeutic agents.