High dose acetaminophen with n-acetylcysteine rescue as a novel STAT3 inhibitor with anti-cancer stem cell properties

高剂量对乙酰氨基酚与 n-乙酰半胱氨酸救援作为具有抗癌干细胞特性的新型 STAT3 抑制剂

• 批准号: 10517287
• 负责人: Alexander Neuwelt
• 金额: --
• 依托单位: VA VETERANS ADMINISTRATION HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-10-01 至 2026-09-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10517287
• 关键词: Acetaminophen; Acetylcysteine; Analgesics; Antidotes; Award; Binding; Biological Assay; Cancer Model; Cancer Patient; Cancer Relapse; Caring; Cells; Chemoresistance; Cisplatin; Clinical; Clinical Research; Collaborations; Computer software; Data; Dedications; Disease; Doctor of Philosophy; Dominant-Negative Mutation; Dose; Drug Targeting; Drug usage; Education; Enrollment; Epidermal Growth Factor Receptor; Exhibits; Foundations; Free Radicals; Funding; Genes; Genetic Transcription; Glutathione; Goals; Head; Hepatoblastoma; Hepatotoxicity; Human; Hydrogen Bonding; In Vitro; In complete remission; Injury; Invaded; K-Series Research Career Programs; Knock-out; Knowledge; Laboratory Research; Length; Liver; Maintenance; Malignant Neoplasms; Malignant neoplasm of lung; Mediating; Mentors; Methods; Mitochondria; Modeling; Molecular; Mus; Natural regeneration; Non-Small-Cell Lung Carcinoma; Overdose; Patient Selection; Patients; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phenotype; Phosphorylation; Physicians; Physiology; Play; Population; Process; Prognosis; Proliferating; Property; Proteins; Regulation; Research; Research Personnel; Resistance; Resources; Role; STAT3 gene; Scientist; Series; Signal Transduction; Site-Directed Mutagenesis; Stat3 protein; Testing; Time; Toxic effect; Transfection; Tumorigenicity; Universities; Virginia; Work; Xenograft procedure; anti-cancer; anticancer activity; antitumor effect; cancer cell; cancer stem cell; career; chemotherapy; driver mutation; early phase clinical trial; experience; experimental study; in silico; in vivo; inhibitor; insight; lung cancer cell; neoplastic cell; novel; patient derived xenograft model; pre-clinical; predictive marker; response; self-renewal; skills; src Homology Region 2 Domain; stem cell biology; stem cell biomarkers; stem cell growth; stem cell population; symposium; synergism; targeted treatment; tumor; tumor growth; tumor microenvironment; vector

High dose acetaminophen (AAP) with delayed n-acetylcysteine (NAC) rescue has shown promise in early phase clinical trials, inducing disease shrinkage in 8/14 and an objective response in 3/14 assessable patients with diverse tumor types. However, the mechanism of anti-tumor activity of high dose AAP is not well understood. It was previously felt that high dose AAP has anti-tumor activity via similar processes as its mechanism of hepatotoxicity, namely free radical injury and glutathione depletion. However, pre-clinical and clinical evidence suggests that glutathione is not depleted within the tumor in response to high-dose AAP treatment. This suggests an alternate mechanism of anti-tumor activity of high dose AAP. In this proposal, we outline preliminary data suggesting that: 1.) High dose AAP has anti-cancer stem cell (CSC) activity and 2.) High dose AAP functions as a novel inhibitor of STAT3 phosphorylation. STAT3 is a protein that has previously been shown to play an important role in the maintenance and proliferation of a viable CSC population. We hypothesize that the anti-CSC activity in non-small cell lung cancer (NSCLC) of high dose AAP is via a STAT3 dependent mechanism. In specific aim #1, sub-aim A, we propose to evaluate the mechanism of STAT3 inhibition by AAP. We have obtained preliminary data suggesting that AAP binds STAT3 in the SH2 domain via a strong hydrogen bond formed at Lys626. We aim to validate these preliminary findings using orthogonal approaches such as microscale thermopheresis (MST). These studies will be performed in collaboration with co-mentor Umesh Desai, PhD, head of Medicinal Chemistry at Virginia Commonwealth University (VCU). In sub-aim B, we evaluate the mechanism of the anti-CSC activity of high dose AAP. We will generate constructs of human NSCLC with altered STAT3 proteins: empty vector (STAT3 KO), STAT3 K626K (WT) and STAT3 K626A (predicted to have decreased AAP-binding). We will determine the sensitivity of STAT3-altered constructs to the anti-CSC effects of AAP or AAP + NAC. If human NSCLC cells with STAT3 K626A have decreased responsiveness to the anti-CSC effects of high dose AAP, this would suggest that the anti-CSC effects of AAP are mediated by direct binding to STAT3. In specific aim #2, we will evaluate the mechanisms of synergy between chemotherapy (in NSCLC without targetable driver mutation) and targeted therapy (in NSCLC with EGFR or ALK driver alteration) with AAP, both in vitro and in vivo. We will investigate whether AAP-enhances chemotherapy/targeted therapy anti-tumor activity via a a STAT3 dependent mechanism. In this CDA application, Dr. Neuwelt proposes to work with an outstanding group of established scientists whom have the experience and expertise necessary to mentor Dr. Neuwelt on the proposed project. Dr. Neuwelt’s primary mentor will be Dr. Patel, a senior VA-merit funded researcher with expertise in cancer stem cell biology. Dr. Larner, a co-mentor on the project, discovered the role of STAT3 in mitochondrial function and has dedicated much of his career to understanding STAT3 physiology. If awarded a CDA, Dr. Neuwelt will enroll in relevant classes at local Universities, attend scientific conferences, take care of VA-based patients, and grow his research lab, all with the goal of becoming an independent VA-based physician scientist.

高剂量对乙酰氨基酚（AAP）与延迟的N-乙酰半胱氨酸（NAC）救援已显示出在早期 阶段临床试验，诱导8/14例患者的疾病缩小，3/14例可评估患者的客观缓解 不同类型的肿瘤然而，高剂量AAP的抗肿瘤作用机制尚不清楚 明白以前认为高剂量AAP通过与其类似的过程具有抗肿瘤活性。 肝毒性机制，即自由基损伤和谷胱甘肽耗竭。然而，临床前和 临床证据表明，谷胱甘肽在肿瘤内不会因高剂量AAP而耗尽， 治疗这表明高剂量AAP的抗肿瘤活性的替代机制。在本提案中，我们 初步数据显示：（1）高剂量AAP具有抗癌干细胞（CSC）活性和2.） 高剂量AAP是一种新型的STAT 3磷酸化抑制剂。STAT 3是一种蛋白质， 以前已经证明在维持和增殖有活力的CSC中起重要作用 人口我们推测，高剂量AAP对非小细胞肺癌（NSCLC）的抗CSC活性 是通过STAT 3依赖性机制。在具体目标#1，子目标A中，我们建议评估该机制 AAP对STAT 3的抑制作用。我们已经获得的初步数据表明，AAP结合STAT 3在SH 2 结构域通过在Lys 626处形成的强氢键。我们的目标是验证这些初步发现， 正交方法，如微量热分离法（MST）。这些研究将在 与共同导师Umesh Desai，博士，弗吉尼亚联邦药物化学负责人合作 大学（VCU）。在子目标B中，我们评估了高剂量AAP抗CSC活性的机制。我们 将产生具有改变的STAT 3蛋白的人NSCLC的构建体：空载体（STAT 3 KO），STAT 3 K626 K（WT）和STAT 3 K626 A（预测具有降低的AAP结合）。我们将确定敏感性 STAT 3-改变的构建体对AAP或AAP + NAC的抗CSC作用的影响。如果具有STAT 3的人NSCLC细胞 K626 A对高剂量AAP的抗CSC作用的反应性降低，这表明K626 A对高剂量AAP的抗CSC作用的反应性降低。 AAP的抗CSC作用是通过直接结合STAT 3介导的。在具体目标#2中，我们将评估 化疗（在无靶向驱动突变的NSCLC中）与靶向 AAP治疗（EGFR或ALK驱动基因改变的NSCLC），包括体外和体内研究。我们将调查 AAP是否通过STAT 3依赖性增强化疗/靶向治疗的抗肿瘤活性 机制在这个CDA应用程序中，Neuwelt博士建议与一个杰出的团队合作， 科学家谁拥有必要的经验和专业知识，以指导博士Neuwelt对拟议的项目。 博士Neuwelt的主要导师将是Patel博士，他是一位资深的退伍军人管理局奖学金资助的研究人员，具有癌症方面的专业知识 干细胞生物学Larner博士是该项目的共同导师，他发现了STAT 3在线粒体中的作用。 他的大部分职业生涯都致力于了解STAT 3的生理学。如果获得CDA，博士。 Neuwelt将参加当地大学的相关课程，参加科学会议，照顾VA为基础的 病人，并扩大他的研究实验室，所有的目标是成为一个独立的VA为基础的医生 科学家