Inhibiting Multi-Functional ALDOA for Cancer Therapy

抑制多功能 ALDOA 用于癌症治疗

• 批准号: 10494262
• 负责人: GARTH POWIS
• 金额: $ 49.81万
• 依托单位: PHUSIS THERAPEUTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-12 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10494262
• 关键词: 1-Phosphatidylinositol 3-Kinase; Actins; Active Sites; Affinity; Aldolase A; Allosteric Regulation; Allosteric Site; Binding; Binding Proteins; Biological; Biomass; Blood; C-terminal; Cancer Cell Growth; Catalytic Domain; Cell Communication; Cell Nucleus; Cell Proliferation; Cell Survival; Cell physiology; Cells; Chemicals; Complex; Consensus; Crystallization; Cytoskeletal Proteins; Cytoskeleton; Cytosol; Development; Distal; E-Cadherin; EP300 gene; Elements; Energy Metabolism; Energy Supply; Energy-Generating Resources; Enzymes; Evolution; Fibrosis; Fructose; Fructosediphosphate Aldolase; Gene Family; Genes; Genetic Transcription; Glycolysis; Glycolysis Inhibition; Goals; Growth; Growth Factor; Human; Hypoxia; Interphase Cell; Lead; Link; Malignant Neoplasms; Malignant neoplasm of pancreas; Mediating; Mesenchymal; Molecular; Mus; Neoplasm Metastasis; Nuclear; Nuclear Proteins; Oncogenes; Pancreas; Pathway interactions; Patients; Pharmacology; Phenotype; Process; Prognosis; Proteins; Reporting; Repression; Resolution; Role; Solid Neoplasm; Stream; Structure; Tail; Therapeutic; Tight Junctions; Time; Transcription Coactivator; Transcriptional Activation; Vascular Endothelial Growth Factors; X-Ray Crystallography; Xenograft Model; aerobic glycolysis; alpha Tubulin; anaerobic glycolysis; angiogenesis; anti-cancer; base; cancer cell; cancer initiation; cancer therapy; checkpoint therapy; design; improved; in vivo; inhibitor; member; novel; novel drug class; novel therapeutics; pancreatic cancer cells; polymerization; prevent; response; small molecule; transcription factor; tumor; tumor growth; tumor xenograft

Fructose-bisphosphate aldolase A (ALDOA) is an ancient, highly expressed gene that has acquired three distinct cellular activities. The best understood activity is catalyzing a key step in aerobic glycolysis. ALDOA also has protein binding activities independent of its catalytic activity (“moonlighting”), that include binding to cytoskeletal proteins, which use an “EΦE” motif that fits into a “moonlighting pocket” of ALDO. Binding to the cytoskeleton actin holds ALDO in an inactive form until it is needed, when it is released by an increase in the levels of its substrate fructose-1, 6-bisphosphate and/or by growth-factor-induced PI-3-kinase activity. Another moonlighting function of ALDOA is its presence in the nucleus of cancer cells, where it is associated with increased proliferation. One possibility is that nuclear ALDOA binds to the EΦE motif on EID-1 in the nucleus to inhibit HIF-1's transcriptional co-activator, p300. ALDOA levels are increased in many cancers, particularly pancreatic cancer, where it has been linked to poor patient survival and an increase in metastasis. Pancreatic cancer cell has high levels of anaerobic glycolysis that is further increased by the hypoxia inducible transcription factor-1 (HIF-1). HIF-1 induces ALDOA and other glycolytic enzymes that in turn maintain high HIF-1 activity through an AMPK/p300-dependent feed-forward loop. HIF-1 activity leads to VEGF release and angiogenesis, and the induction of other cancer cell survival mechanisms. Increased glycolysis provides hypoxic cancer cells with an increased supply of energy (ATP) and essential metabolites for biomass synthesis. Elevated ALDOA is also associated with low E-cadherin, a component of cancer cell tight junctions (TJ) necessary for cell-cell interactions, giving a more mesenchymal phenotype and increased tumor metastasis. This most likely is due to binding of ALDOA to the EΦE motif of cytoskeleton actin causing its polymerization and disassembly, or other inactivation of the TJ with loss of E-cadherin. All this makes ALDOA an exceptional druggable anti-cancer target. Our X-ray crystallography studies have identified a new role for the C-terminal domain of ALDO in its catalytic activity, as well as a reactive Cys289 residue that allows allosteric regulation of catalytic activity. We have identified a novel lead probe allosteric inhibitor of ALDOA that forms a complex with Cys289 inhibiting glycolysis, HIF-1 activity and the proliferation of cancer cells, and in vivo inhibits glycolysis and tumor growth in a tumor xenograft model. Using this biologically stable allosteric inhibitor as a lead, one of our goals is to make more potent inhibitors that bind to the allosteric site Cys289, as well as high affinity bifunctional inhibitors that simultaneously engage the active site and moonlighting pocket. We will use X-ray crystallography to help us design inhibitors that modulate nuclear regulators of HIF-1 activity; and not least to design direct inhibitors of ALDOA catalytic activity. We will use these compounds as pharmacological probes to inhibit the various activities of ALDOA, and as potential leads for new therapies for the treatment of pancreatic, and other cancers.

果糖二磷酸醛缩酶 A (ALDOA) 是一种古老的、高度表达的基因，已获得三个 不同的细胞活动。最容易理解的活性是催化有氧糖酵解的关键步骤。阿尔多阿 还具有独立于其催化活性（“兼职”）的蛋白质结合活性，包括与 细胞骨架蛋白，它使用“EΦE”基序，适合 ALDO 的“兼职口袋”。绑定到 细胞骨架肌动蛋白使 ALDO 保持非活性状态，直到需要时，通过增加 其底物果糖-1, 6-二磷酸水平和/或生长因子诱导的 PI-3-激酶活性。其他 ALDOA 的兼职功能是它存在于癌细胞的细胞核中，在那里它与 增殖增加。一种可能性是核 ALDOA 与细胞核中 EID-1 上的 EΦE 基序结合 抑制 HIF-1 的转录辅激活因子 p300。许多癌症中的 ALDOA 水平都会升高，特别是 胰腺癌，它与患者生存率低和转移增加有关。胰 癌细胞具有高水平的无氧糖酵解，缺氧诱导进一步增加了无氧糖酵解 转录因子-1 (HIF-1)。 HIF-1 诱导 ALDOA 和其他糖酵解酶，从而维持高水平 HIF-1 活性通过 AMPK/p300 依赖性前馈环路进行。 HIF-1 活性导致 VEGF 释放并 血管生成以及其他癌细胞存活机制的诱导。糖酵解增加提供 缺氧的癌细胞，能量 (ATP) 和生物量必需代谢物的供应增加 合成。 ALDOA 升高还与 E-钙粘蛋白降低有关，E-钙粘蛋白是癌细胞紧密连接的组成部分 (TJ) 是细胞间相互作用所必需的，提供更多的间充质表型并增加肿瘤 转移。这很可能是由于 ALDOA 与细胞骨架肌动蛋白的 EΦE 基序的结合导致其 聚合和分解，或 TJ 的其他失活，导致 E-钙粘蛋白损失。这一切都让ALDOA 一个特殊的药物抗癌靶点。我们的 X 射线晶体学研究确定了 ALDO 的 C 端结构域的催化活性，以及​​反应性 Cys289 残基，使 催化活性的变构调节。我们已经鉴定出一种新型的 ALDOA 先导探针变构抑制剂 与 Cys289 形成复合物，抑制糖酵解、HIF-1 活性和癌细胞增殖，以及 体内抑制肿瘤异种移植模型中的糖酵解和肿瘤生长。使用这种生物稳定的变构 作为先导抑制剂，我们的目标之一是制造更有效的抑制剂，与变构位点 Cys289 结合，如 以及同时参与活性位点和兼职口袋的高亲和力双功能抑制剂。 我们将使用X射线晶体学来帮助我们设计调节HIF-1活性核调节因子的抑制剂； 尤其是设计 ALDOA 催化活性的直接抑制剂。我们将使用这些化合物作为 抑制 ALDOA 各种活性的药理学探针，并作为新疗法的潜在线​​索 胰腺癌和其他癌症的治疗。