Simultaneous pharmacological profiling of oncogenic gene fusion proteins in cancer

研究综述 人类基因组编码1600多个转录因子(TF)，以及其他 共同执行调控的辅因子、染色质调节剂和结构蛋白 编码在核DNA中的指令。这些蛋白质的功能失调，统称为 作为基因调控蛋白(GRP)，已知可导致多种疾病，如癌症， 与炎症相关的疾病和神经系统疾病。在癌症中，这些蛋白质经常 重新排列和融合以产生新的蛋白质，从而导致 各种类型的白血病、肉瘤等肿瘤。尽管这些蛋白质很重要， 由于在体外模拟其活性方面的挑战，GRP一直被认为是不可用药的。 我们已经通过实施细胞内功能蛋白质组学药物解决了这些缺点 量化小分子对GRPs丰度影响的发现平台 与多种细胞和组织类型的基因组结合。该平台基于染色质 盐析提取与数据独立分析质谱联用 (国际象棋-DIA)，这是最近报道的。在这份提案中，我们将把这项技术应用于 致癌融合蛋白的药物开发。首先，我们将使用混合血统白血病 (MLL)重排白血病进行致癌融合蛋白的技术开发 蛋白质组学战略。在AML和所有患者的子集中发现MLL重排， 通常发生在儿童中，用现有的治疗方案治疗仍然具有挑战性。几个 MLL重排白血病的候选药物目前正在进行临床试验，这些药物将 用于验证CHESS-DIA方法报告MLL靶向能力的准确性 破坏活细胞中MLL复合体的化合物。经过验证的MLL国际象棋-DIA测试， 然后，我们将进行试点筛查，以证明该分析在筛查环境中的实用性，使用 国家癌症研究所的机制多样性化合物集，并补充已知 MLL复合体的抑制剂。这些化合物含有各种各样的生物活性，许多 它们通过未知的机制起作用。这提供了一个寻找新的 能够破坏MLL复合体的化合物。最后，我们将使用开发的路线图 对于MLL来说，要为许多常见的致癌融合开发CHESS-DIA检测，然后 开发一种方法，将这些检测方法统一在一个单一的、统一的细胞内致癌检测方法中 融合蛋白。

RESEARCH SUMMARY The human genome encodes more than 1,600 transcription factors (TFs), along with additional cofactors, chromatin regulators, and structural proteins that collectively execute the regulatory instructions encoded within the nuclear DNA. Dysfunctions of these proteins, collectively known as Gene Regulatory Proteins (GRPs), are known to drive multiple diseases such as cancer, inflammation-related, and neurological conditions. In cancer, these proteins are frequently rearranged and fused to create new proteins which cause the initiation and progression of various types of leukemia, sarcoma and other tumors. Despite the importance of these proteins, GRPs have been considered undruggable due to challenges in modeling their activity in vitro. We have solved these shortcomings by implementing an in-cell functional proteomics drug discovery platform that quantifies the effects of small-molecules on the abundance of GRPs bound to the genome in a diversity of cell and tissue types. The platform is based on Chromatin Extraction by Salt Separation, coupled to Data Independent Analysis mass spectrometry (ChESS-DIA), which was recently reported. In this proposal, we will apply this technology for drug development of oncogenic fusion proteins. First, we will use Mixed Lineage Leukemia (MLL) rearranged leukemia to perform technology development of the oncogenic fusion protein proteomics strategy. MLL rearrangements are found in a subset of AML and ALL patients, commonly in children, and remain challenging to treat with existing therapeutic options. Several drug candidates for MLL-rearranged leukemia are currently in clinical trials, and these will be used to validate the accuracy of the ChESS-DIA assay for reporting the ability of MLL-targeting compounds to disrupt the MLL complex in live cells. With a validated MLL ChESS-DIA assay, we will then conduct a pilot screen to prove the assay’s utility in a screening setting, using the National Cancer Institute’s Mechanistic Diversity compound set supplemented with known inhibitors of the MLL complex. These compounds contain a diverse array of bioactivities, many of which act through unknown mechanisms. This provides an opportunity to find new compounds capable of disrupting the MLL complex. Lastly, we will use the roadmap developed for MLL to develop ChESS-DIA assays for many of the common oncogenic fusions, then develop a method to unify these assays together in a single, unified, in-cell assay for oncogenic fusion proteins.