Aberrant activation of HGF/MET signaling as a therapeutic target in AML

HGF/MET 信号传导异常激活作为 AML 的治疗靶点

• 批准号: 8871431
• 负责人: Alex Kentsis
• 金额: $ 15.63万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8871431
• 关键词: Accounting; Acute Myelocytic Leukemia; Acute Promyelocytic Leukemia; Adult; Advisory Committees; Alternative Therapies; Antibodies; Award; Biochemistry; Biological; Biological Assay; Biophysics; Bone marrow biopsy; Candidate Disease Gene; Cell Line; Cell Survival; Cells; Cellular biology; Cessation of life; Chemicals; Child; Chromosome abnormality; Clinical; Clinical Trials; Combination Drug Therapy; Complex; Dana-Farber Cancer Institute; Dasatinib; Dependence; Development; Diagnosis; Diagnostic; Disease; Elements; FLT3 gene; Foundations; Future; Gene Expression Profiling; Genetic; Genetic Engineering; Genomic Instability; Genomics; Growth; Hematologic Neoplasms; Hematology; Hepatocyte Growth Factor; Immunohistochemistry; In Vitro; Investigation; Karyotype; Lead; Libraries; Ligands; Lymphoma; MET Oncogene; Mediating; Mentors; Mentorship; Methods; Modeling; Molecular; Multiple Myeloma; Mus; Mutate; Mutation; Oncogenes; Oncogenic; Pathway interactions; Patients; Pediatric Hematology; Pediatric Oncology; Phosphotransferases; Physicians; Play; Protein Tyrosine Kinase; Proteomics; RNA Interference; Receptor Protein-Tyrosine Kinases; Refractory; Relapse; Research Proposals; Resistance; Resistance development; Risk; Role; Scientist; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Signaling Molecule; Specimen; Surveys; System; Testing; Therapeutic; Therapeutic Intervention; Training; Transduction Gene; Treatment Efficacy; Tyrosine Kinase Inhibitor; autocrine; cancer cell; carcinogenesis; career; cell growth; clinically relevant; collaborative environment; combinatorial; functional genomics; genome-wide; high risk; improved; in vivo; in vivo Model; inhibitor/antagonist; innovation; insight; kinase inhibitor; leukemia; mouse model; nanoimmunoassay; new therapeutic target; oncogene addiction; oncology; preclinical study; receptor; small hairpin RNA; targeted treatment; therapeutic target; therapy resistant; tool

DESCRIPTION (provided by applicant): Despite improvement in treatment of acute myeloid leukemia (AML), high-risk disease such as complex karyotype AML remains largely refractory to current therapy, and is mostly fatal. Identification of effective therapeutic targets by using candidate gene approaches has been limited by the number and variety of genetic defects associated with AML. To identify new therapeutic targets, I carried out a genome-wide functional screen by using a retroviral library of short hairpin RNAs (shRNAs) in complex karyotype AML cells. I discovered that shRNA mediated depletion of hepatocyte growth factor (HGF), ligand of the receptor tyrosine kinase MET, specifically inhibits growth of AML but not other hematologic cancer cells. MET is a potent oncogene, whose aberrant activation is widely implicated in carcinogenesis, causing enhanced growth, survival, and genomic instability of cancer cells. However, mechanisms of carcinogenic MET signaling are currently not well understood, and HGF/MET signaling is not thought to play a role in AML. To validate this observation in patient specimens, I carried out immunohistochemistry of diagnostic bone marrow biopsies. I observed that HGF is aberrantly expressed and associated with activation of MET in about 15% of patients with AML, including most patients with complex karyotype disease. Analysis of cell lines derived from such patients showed that HGF expression was associated with autocrine activation of its receptor MET. Depletion of HGF or MET using shRNA or inhibition of MET using tyrosine kinase inhibitors and neutralizing anti-HGF antibody profoundly reduced proliferation and induced death of AML cells lines that express HGF but not those that lack HGF expression. This indicates the functional dependence or "oncogene addiction" to this pathway, and suggests that therapeutic inhibition of HGF/MET signaling may be used to improve the treatment of AML. However, detailed understanding of the molecular mechanisms by which this pathway promotes AML cell growth and survival is currently lacking. By genetically engineering AML cell lines to deplete and express specific signaling molecules, and isolating cell lines that are resistant to HGF/MET inhibition, I will identify signaling components that mediate HGF/MET "oncogene addiction," and strategies to overcome resistance to therapeutic inhibition of HGF/MET signaling. These studies will be combined with the investigation of antileukemic efficacy of HGF/MET inhibition in murine models of AML in vivo using functional nanoimmunoassay, phosphoproteomic and genomic methods to identify not only the optimal strategy to target this pathway clinically, but also how to optimally integrate it with other targeted inhibitors of AML signaling. These technical advances will circumvent limitations associated with empiric discovery of novel therapeutic targets in AML, and will identify the principal signaling pathways required for AML cell growth and survival. The cell line systems and mouse models will also allow for detailed analysis of gene expression and proteomic changes that accompany signaling by receptor tyrosine kinases, thus providing key mechanistic insights that will be important to a wide variety of biological and disease phenomena. Results of the in vitro and in vivo studies will be further investigated using primary patient specimens, and will lay the foundation for future preclinical studies and clinical trials of targeted therapies of AML. The specific aims are: Specific Aim 1: Dissect the molecular signaling pathways responsible for the HGF/MET dependence of AML cell growth and survival, and identify mechanisms that account for resistance to HGF/MET inhibition (years 1-3). Specific Aim 2: Assess the antileukemic efficacy of HGF/MET inhibition in murine AML models in vivo, both by itself and in concert with the inhibition of other leukemogenic tyrosine kinases, including FLT3 and KIT (years 3-5). The applicant, Dr. Alex Kentsis, a pediatric hematology/oncology fellow at the Dana-Farber Cancer Institute (DFCI) has outlined a 5-year career plan that will build upon his background in biophysics and clinical hematology/oncology. Under the mentorship of Dr. Thomas Look, a recognized leader in cancer cell biology and translational investigations of leukemia, Dr. Kentsis seeks to utilize powerful functional genomic and proteomic approaches using a combination of in vitro systems and murine models in vivo to study the role of aberrant HGF/MET signaling in AML. Dr. Kentsis will be mentored by an Advisory Committee of internationally recognized experts in the field. Finally, the plan is ideally carried out in the Department of Pediatric Oncology at DFCI, given its distinguished record for training physician-scientists in a rich and collaborative environment. With the support provided by the K08 award, Dr. Kentsis' project will lead to the development of clinically effective HGF/MET targeted therapy for AML.

描述（由申请人提供）：尽管急性髓样白血病（AML）的治疗有所改善，但高风险疾病（例如复杂的核型AML）在很大程度上仍然对当前疗法难治性，并且大多是致命的。通过使用候选基因方法鉴定有效的治疗靶标已受到与AML相关的遗传缺陷的数量和种类的限制。为了识别新的治疗靶标，我通过在复杂的核型AML细胞中使用短发夹RNA（SHRNA）的逆转录病毒库进行了全基因组功能筛选。我发现SHRNA介导的肝细胞生长因子（HGF）的耗竭，受体酪氨酸激酶的配体Met，特别抑制了AML的生长，但没有抑制其他血液学癌细胞的生长。 MET是一种有效的致癌基因，其异常激活广泛与癌变有关，导致癌细胞的生长，存活和基因组不稳定。但是，目前尚不清楚致癌MET信号传导的机制，并且HGF/MET信号传导也不在AML中发挥作用。为了验证患者标本中的观察结果，我进行了诊断骨髓活检的免疫组织化学。我观察到HGF异常表达，并与大约15％的AML患者（包括大多数患有复杂核型疾病的患者）激活MET有关。对此类患者得出的细胞系的分析表明，HGF表达与其受体的自分泌激活有关。使用shRNA或使用酪氨酸激酶抑制剂对HGF或MET的耗竭或抑制Met对抗HGF抗体的中和抗HGF抗体可大大降低增殖和诱导HGF的AML细胞系的诱导死亡，而不是那些缺乏HGF表达的AML细胞。这表明功能依赖性或“癌基因成瘾”对这一途径，并表明可以使用HGF/MET信号的治疗性抑制来改善AML的处理。但是，目前缺乏该途径促进AML细胞生长和存活的分子机制的详细理解。通过遗传工程AML细胞系来耗尽并表达特定的信号分子，并隔离对HGF/MET抑制具有抗性的细胞系，我将确定介导HGF/MET的信号成分“癌基因成瘾”，以及克服对HGF/MET信号疗法抑制HGF/MET信号的抗性的策略。这些研究将与使用功能性纳米免疫测定，磷酸蛋白质组和基因组方法的AML体内鼠模型中HGF/MET抑制作用的抗白血病功效的研究结合使用，不仅鉴定出最佳的策略以临床上靶向这一途径，还可以将其与其他靶向启用的Aml抑制剂相结合。这些技术进步将规避与AML中新型治疗靶标的经验发现相关的局限性，并将确定AML细胞生长和生存所需的主要信号通路。细胞系系统和小鼠模型还将允许对受体酪氨酸激酶信号传导的基因表达和蛋白质组学变化进行详细分析，从而提供关键的机械见解，这些见解对多种生物学和疾病现象至关重要。体外和体内研究的结果将使用原发性患者标本进一步研究，并将为未来的临床前研究和AML靶向疗法的临床试验奠定基础。具体目的是：特定目标1：剖析负责HGF/MET依赖AML细胞生长和存活的分子信号传导途径，并确定对HGF/MET抑制的抗性的机制（1-3年）。具体目的2：评估HGF/MET抑制在体内鼠AML模型中的抗白血病疗效，无论是本身，以及与其他白血病酪氨酸激酶（包括FLT3和试剂盒）的抑制（3-5年）。申请人Alex Kentsis博士是Dana-Farber癌症研究所（DFCI）的儿科血液学/肿瘤学研究员Alex Kentsis博士，概述了一项为期5年的职业计划，该计划将基于他在生物物理学和临床血液学/肿瘤学方面的背景。在托马斯·莱克（Thomas Look）博士的指导下，托马斯·莱克（Thomas Look）是癌细胞生物学的公认领导者，以及对白血病的转化研究，肯蒂斯博士试图利用体外系统和鼠模型在体内使用强大的功能性基因组和蛋白质组学方法来研究AML中异常HGF/Met Metsign的作用。 Kentsis博士将由该领域的国际认可专家咨询委员会指导。最后，该计划是在DFCI的儿科肿瘤学系中进行的，鉴于其在丰富且协作的环境中培训医师科学家的杰出记录。在K08奖提供的支持下，Kentsis博士的项目将导致开发具有AML的临床有效HGF/MET目标疗法。

项目成果

Osteosarcoma With Apparent Ewing Sarcoma Gene Rearrangement.

具有明显尤文肉瘤基因重排的骨肉瘤。

• DOI: 10.1097/mph.0000000000000553
• 发表时间: 2016
• 期刊: Journal of pediatric hematology/oncology
• 作者: Mathias,MelissaD;Chou,AlexanderJ;Meyers,Paul;Shukla,Neerav;Hameed,Meera;Agaram,Narasimhan;Wang,Lu;Berger,MichaelF;Walsh,Michael;Kentsis,Alex
• 通讯作者: Kentsis,Alex

Autocrine activation of the MET receptor tyrosine kinase in acute myeloid leukemia.

• DOI: 10.1038/nm.2819
• 发表时间: 2012-07
• 期刊: Nature medicine
• 影响因子: 82.9

Advancing Urinary Protein Biomarker Discovery by Data-Independent Acquisition on a Quadrupole-Orbitrap Mass Spectrometer.

• DOI: 10.1021/acs.jproteome.5b00826
• 发表时间: 2015-11-06
• 期刊: Journal of proteome research
• 影响因子: 4.4
• 作者: Muntel J;Xuan Y;Berger ST;Reiter L;Bachur R;Kentsis A;Steen H
• 通讯作者: Steen H

Venous thrombosis associated with gene deletion of tissue factor pathway inhibitor.

与组织因子途径抑制剂基因缺失相关的静脉血栓形成。

• DOI: 10.1002/ajh.21542
• 发表时间: 2009
• 期刊: American journal of hematology
• 影响因子: 12.8
• 作者: Kentsis,Alex;Bradwin,Gary;Miller,DavidT;Trenor3rd,CameronC
• 通讯作者: Trenor3rd,CameronC

Forward genetic screen of human transposase genomic rearrangements.

• DOI: 10.1186/s12864-016-2877-x
• 发表时间: 2016-08-04
• 期刊: BMC genomics
• 影响因子: 4.4
• 作者: Henssen AG;Jiang E;Zhuang J;Pinello L;Socci ND;Koche R;Gonen M;Villasante CM;Armstrong SA;Bauer DE;Weng Z;Kentsis A
• 通讯作者: Kentsis A