Development of first-in-class histone acetyltransferase (HAT) activators for precision targeting of epigenetic derangements in lymphoma

弥漫性大B细胞淋巴瘤(DLCBL)是最常见的淋巴瘤类型，影响约30,000名患者 每年一次。对DLBCL的分子发病机制的最新见解将该病分为两个分子 亚型：生发中心(GC)和ABC亚型。ABC-DLCBL的靶向分子特征研究 发展阶段，但对于更常见的GC-DLCBL还没有实现这样的成功。GC是一个 淋巴小室，负责通过体细胞超突变产生高亲和力抗体 和类切换重组。表观遗传修饰物，如EZH2、组蛋白乙酰转移酶(HATS)和 转录抑制因子Bcl6对于B细胞的发育是必不可少的，允许必要的突变和沉默 体细胞超突变所必需的肿瘤抑制因子。这种生理状态在一定程度上是通过 组蛋白的乙酰化减少和甲基化增加，从而使其处于转录抑制状态。 影响这些表观遗传和转录修饰因子HATS、EZH2和Bcl6的突变已被确认为 GC来源的淋巴瘤的驱动事件。 鉴于表观遗传功能障碍在GC来源的B细胞淋巴瘤发病机制中的关键重要性， 我们假设，如果HAT等位基因的失活突变对GC-DLBCL至关重要，那么药物激活HAT等位基因的突变对GC-DLBCL至关重要 野生型酶应该显示出治疗效果。此外，我们认为，联合瞄准 具有HAT激活剂和其他表观遗传修饰剂(HDAC和EZH2抑制剂)的表观遗传机制 可能会导致深刻的表观遗传修饰，从而协同诱导细胞程序性死亡。最后， 如果突变状态和EP300等特定基因的表达水平与HAT的反应相关 激活剂或联合表观遗传疗法，那么纳米串表达面板可能被开发为 反应的生物标志物。 本提案的目标将通过以下具体目标进行评估：(1)确定 HAT激活剂YF2与HAT酶的结合及其对细胞乙酰化的功能影响 免费化验。(2)确定HAT激活剂结合临床可用表观遗传学的效果 用于细胞系和小鼠的修饰剂。表观遗传修饰剂将包括HDAC和EZH2抑制剂。 HAT激活剂联合表观遗传修饰剂对翻译后效应的影响 将确定下游目标的修饰(甲基化、乙酰化)和基因表达。(3) 询问淋巴瘤细胞系EP300突变状态和表观遗传基因特征以了解精确度 用帽子激活剂瞄准。将确定候选人的基本基因突变和表达谱 淋巴瘤细胞系中的表观遗传和转录修饰因子，并将与HAT激活剂的IC50相关 以及联合表观遗传疗法的协同系数。如果我们接受这些假设，这将是 代表着一个直接靶向突变的机会，例如HAT和EZH2，它们驱动GC-DLBCL。

Diffuse large B-cell lymphoma (DLCBL) is the most common type of lymphoma affecting ~30,000 patients annually. Recent insights into the molecular pathogenesis of DLBCL has divided this disease into two molecular subtypes: germinal center (GC) and ABC subtypes. Targeting molecular features of ABC-DLCBL are in the final stages of development, but such success has not been realized for the more common GC-DLCBL. The GC is a compartment of the lymph node responsible for generating high affinity antibodies via somatic hyper-mutation and class switch recombination. Epigenetic modifiers such as EZH2, histone acetyltransferases (HATs), and the transcriptional repressor, Bcl6, are essential to B-cell development allowing for requisite mutagenesis and silencing of tumor suppressors necessary for somatic hypermutation. This physiologic state is partly achieved by decreased acetylation and increased methylation of histones enforcing a transcriptionally repressed state. Mutations affecting these epigenetic and transcriptional modifiers, HATs, EZH2 and Bcl6, have been identified as driving events in GC-derived lymphomas. Given the critical importance of epigenetic dysfunction in the pathogenesis of GC-derived B-cell lymphomas, we hypothesize that if inactivating mutations in HAT alleles are crucial to GC-DLBCL then drugs activating the wild-type enzyme should demonstrate therapeutic effect. In addition, we believe that combined targeting of epigenetic machinery with HAT activators and other epigenetic modifying agents (HDAC and EZH2 inhibitors) may induce profound epigenetic modification leading to synergistic induction of programmed cell death. Finally, if mutational status and expression levels of specific genes such as EP300 correlate with response to HAT activators or combined epigenetic therapy, then a NanoString expression panel may be developed as a biomarker for response. The objectives of this proposal will be evaluated by addressing the following specific aims: (1) Characterize the HAT activator YF2 by determining binding to HAT enzymes and its functional effects on acetylation in cell- free assays. (2) Determine the effects of HAT activators in combination with clinically available epigenetic modifying agents in cell lines and mice. Epigenetic modifying agents will include HDAC and EZH2 inhibitors. The effects of HAT activators in combination with epigenetic modifying agents on post-translational modifications (methylation, acetylation), and gene expression of downstream targets will be determined. (3) Interrogate the mutational status of EP300 and epigenetic gene signatures of lymphoma cell lines to inform precision targeting with HAT activators. Basal gene mutation and expression profiles will be determined for candidate epigenetic and transcriptional modifiers in lymphoma cell lines and will be correlated to the IC50 of HAT activators and the synergy coefficients of combined epigenetic therapy. Should we accept these hypotheses, this would represent an opportunity to directly target mutations, such as HAT and EZH2, which drive GC-DLBCL.