Engineered HGF-NK1 antagonists for Met-targeted cancer imaging and therapy

用于 Met 靶向癌症成像和治疗的工程化 HGF-NK1 拮抗剂

• 批准号: 8657879
• 负责人: JENNIFER R COCHRAN
• 金额: $ 31.25万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8657879
• 关键词: 2-Fluoro-2-deoxyglucose; Affinity; Alanine; Animal Model; Apoptosis; Area; Binding; Biochemical; Biodistribution; Biological; Biological Assay; Blood capillaries; CD34 gene; Cancer Biology; Cancer Diagnostics; Cancer Etiology; Cell Culture Techniques; Cell Proliferation; Cell surface; Circular Dichroism; Clinical; Clinical Trials; Complement; Cysteine; Development; Dimerization; Disease Management; Drug Design; Drug Kinetics; Engineering; Exhibits; FDA approved; Fluorescent Dyes; Funding; Generations; Goals; Growth; Heating; Heparin; Hepatocyte Growth Factor; Homodimerization; Hot Spot; Human; Image; Imagery; Immunohistochemistry; In Vitro; Individual; Kringles; Label; Life; Ligands; Location; Luciferases; Lysine; Malignant Neoplasms; Mammalian Cell; Mammary Neoplasms; Measures; Mediating; Metabolism; Modeling; Molecular Sieve Chromatography; Monitor; Mus; Mutagenesis; Mutation; Mutation Analysis; N-terminal; Neoplasm Metastasis; Optics; Patients; Phase I Clinical Trials; Point Mutation; Positron-Emission Tomography; Pre-Clinical Model; Primary Neoplasm; Property; Prostate carcinoma; Proteins; Radioisotopes; Radiolabeled; Receptor Activation; Receptor Protein-Tyrosine Kinases; Recombinants; Research; Sampling; Scanning; Signal Pathway; Signal Transduction; Site; Staging; Staining method; Stains; Structure; Structure-Activity Relationship; Surface Plasmon Resonance; System; Temperature; Testing; Therapeutic; Therapeutic Agents; Therapeutic Intervention; Tissues; Translations; Treatment Efficacy; Tube; Tumor Cell Line; United States National Institutes of Health; Work; Xenograft Model; Xenograft procedure; abstracting; amino group; angiogenesis; base; bioluminescence imaging; biophysical properties; cancer diagnosis; cancer imaging; cancer therapy; capillary; chelation; density; design; directed evolution; flexibility; gain of function; high throughput screening; imaging probe; improved; in vivo; inhibitor/antagonist; insight; light scattering; loss of function; lung Carcinoma; meetings; migration; molecular imaging; mutant; neoplastic cell; non-invasive imaging; novel strategies; optical imaging; outcome forecast; pre-clinical; preclinical study; radiotracer; receptor; receptor binding; receptor expression; research study; response; therapeutic target; tumor; tumor growth; tumor metabolism; tumor progression; tumor xenograft; uptake; von Willebrand Factor

Project Summary/Abstract Dysregulation of cell signaling pathways that mediate proliferation, survival, and migration are an underlying cause of cancer, and result in invasion and metastasis of many human tumors. In particular, dysregulation and over-expression of the Met tyrosine kinase receptor correlates to poor prognosis in many human tumors, making it an attractive target for therapeutic intervention. There are currently no FDA-approved therapeutics targeting the Met receptor; however, a few candidate molecules have recently entered early stage clinical trials. Therefore, molecules that potently inhibit Met receptor activation would have a significant clinical impact on cancer therapy. In addition, studies to develop Met-targeted molecular imaging agents for non- invasive visualization of Met expression in vivo have been extremely limited. The availability of such imaging agents would aid in cancer diagnosis, staging, and disease management, as well as help identify patients who would be good candidates for Met-targeted therapies. To develop robust Met-targeting agents we used the N- terminal and first Kringle domain (NK1) of the natural Met activating ligand, hepatocyte growth factor (HGF), as a basis for engineering potent Met receptor antagonists. Using directed evolution, we engineered NK1 mutants with significant improvements in Met binding affinity and thermal stability compared to wild-type NK1. Rationally-designed, site-directed mutations introduced into these NK1 proteins transformed them into Met receptor antagonists. In Aim 1 of the proposal, we will perform pre-clinical studies on fluorescently-labeled and radiolabeled NK1 mutants to test their ability to non-invasively image Met expression in living subjects, with the goal of developing them as in vivo molecular imaging agents. In Aim 2, we will perform pre-clinical studies to measure the effects of NK1 mutants on tumor growth, metastasis, and angiogenesis in several mouse tumor models. During the course of treatment, non-invasive imaging will be used to monitor growth and progression of the primary tumor and metastases, and to monitor changes in Met expression and metabolism at the tumor site. In Aim 3, we will fully characterize the binding of a larger panel of engineered NK1 mutants to Met-expressing tumor cells, and will determine their ability to dimerize and subsequently inhibit Met receptor activation. In Aim 4, we will use these engineered NK1 proteins to probe sequence-structure-function relationships of ligand- receptor interactions in the Met receptor system, and provide biochemical and biophysical insight into their mechanism of action. In all four aims, results will be compared to wild-type NK1 to determine the effects of Met receptor binding affinity and protein stability on biological activity in cell culture and animal models. Upon completion of this proposal, we will have evaluated the potential of engineered NK1 proteins as molecular imaging and therapeutic agents in pre-clinical models, an important step on the path to clinical translation.

项目摘要/摘要 介导增殖，存活和迁移的细胞信号通路失调是一种 癌症的根本原因，并导致许多人类肿瘤的侵袭和转移。尤其， MET酪氨酸激酶受体的失调和过表达与许多人的预后不良相关 人类肿瘤，使其成为治疗干预的有吸引力的靶标。目前没有FDA批准 针对MET受体的治疗剂；但是，一些候选分子最近进入了早期阶段 临床试验。因此，有效抑制MET受体激活的分子将具有明显的临床 对癌症治疗的影响。此外，为非 - 体内MET表达的侵入性可视化非常有限。这种成像的可用性 代理人将帮助癌症诊断，分期和疾病管理，并帮助识别患者 将是符合目标疗法的好候选人。为了开发强大的MET靶向剂，我们使用了N- 天然的末端和第一克林尔结构域（NK1）MET激活配体，肝细胞生长因子（HGF），AS 工程有效的MET受体拮抗剂的基础。使用定向进化，我们设计了NK1突变体 与野生型NK1相比，MET结合亲和力和热稳定性的显着改善。 引入这些NK1蛋白引入的理性设计的，定向的突变将它们转化为MET 受体拮抗剂。 在该提案的AIM 1中，我们将对荧光标记和放射标记的NK1进行临床前研究 突变体测试其非侵入图像在生物中表达表达的能力，目的是 将它们作为体内分子成像剂发展。在AIM 2中，我们将进行临床前研究以衡量 NK1突变体对几种小鼠肿瘤模型中肿瘤生长，转移和血管生成的影响。 在治疗过程中，非侵入性成像将用于监测 原发性肿瘤和转移，并监测肿瘤部位的MET表达和代谢的变化。在 AIM 3，我们将充分表征较大的工程NK1突变体对表达的结合 肿瘤细胞，并将确定其二聚和随后抑制MET受体激活的能力。目标 4，我们将使用这些工程化的NK1蛋白来探测配体 - 配体官方函数的关系 MET受体系统中的受体相互作用，并为其提供生化和生物物理洞察力 作用机理。在所有四个目标中，将将结果与野生型NK1进行比较，以确定Met的影响 受体结合亲和力和蛋白质稳定性在细胞培养和动物模型中生物学活性。之上 该提案的完成，我们将评估工程NK1蛋白作为分子的潜力 临床前模型中的成像和治疗剂，这是临床翻译道路的重要步骤。

项目成果

Dual display of proteins on the yeast cell surface simplifies quantification of binding interactions and enzymatic bioconjugation reactions.

• DOI: 10.1002/biot.201600696
• 发表时间: 2017-05
• 期刊: Biotechnology journal
• 影响因子: 4.7
• 作者: Lim S;Glasgow JE;Filsinger Interrante M;Storm EM;Cochran JR
• 通讯作者: Cochran JR

Delivery of an engineered HGF fragment in an extracellular matrix-derived hydrogel prevents negative LV remodeling post-myocardial infarction.

• DOI: 10.1016/j.biomaterials.2014.12.021
• 发表时间: 2015-03
• 期刊: BIOMATERIALS
• 影响因子: 14
• 作者: Sonnenberg, Sonya B.;Rane, Aboli A.;Liu, Cassie J.;Rao, Nikhil;Agmon, Gillie;Suarez, Sophia;Wang, Raymond;Munoz, Adam;Bajaj, Vaibhav;Zhang, Shirley;Braden, Rebecca;Schup-Magoffin, Pamela J.;Kwan, Oi Ling;De Maria, Anthony N.;Cochran, Jennifer R.;Christman, Karen L.
• 通讯作者: Christman, Karen L.