Development of Angiogenesis Inhibitors

血管生成抑制剂的开发

• 批准号: 10262694
• 负责人: William Douglas Figg
• 金额: $ 46.81万
• 依托单位: DIVISION OF CLINICAL SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10262694
• 关键词: Affect; Alkaloids; Angiogenesis Inhibitors; Angiogenesis Pathway; Anti-Inflammatory Agents; Antineoplastic Agents; Aorta; Benzamides; Binding; Biological Assay; Biological Markers; Biological Models; C-terminal; Cell Proliferation; Chemical Structure; Chickens; Clinical; Collaborations; Complex; Cullin Proteins; Cysteine; Cytochrome P450; Development; Dexamethasone; Disease; Drug Design; EP300 gene; Elements; Embryo; Embryonic Development; Endothelial Cells; Energy Metabolism; FDA approved; Fluorescence; Genes; Genetic Polymorphism; Genotype; Hematologic Neoplasms; Histidine; Human; Hyperbilirubinemia; Hypoalbuminemia; Hypoxia; Hypoxia Inducible Factor; Immunomodulators; In Vitro; Inflammation; Isoenzymes; Laboratories; Legal patent; Libraries; Maintenance; Malignant Neoplasms; Malignant neoplasm of prostate; Mediating; Metabolic Biotransformation; Modeling; Molecular; Molecular Target; Multiple Myeloma; Mus; Natural Products; New Agents; Newly Diagnosed; Outcome; Pathway interactions; Patients; Pharmacology; Pharmacology Study; Pharmacology and Toxicology; Porifera; Process; Property; Prospective Studies; Pyrroloiminoquinones; Rattus; Regimen; Regulation; Reporting; Research; Role; Saphenous Vein; Series; Single Nucleotide Polymorphism; Small Interfering RNA; Solid Neoplasm; Structure-Activity Relationship; Thalidomide; Therapeutic; Toxic effect; Toxicology; Transactivation; Treatment Efficacy; Tube; Tumor Angiogenesis; Ubiquitin; United States National Institutes of Health; Work; Xenograft Model; Zebrafish; analog; angiogenesis; base; cancer therapy; cell motility; chemotherapy; cohort; design; drug development; high throughput screening; hypoxia inducible factor 1; immunoregulation; improved; in vivo; in vivo Model; inhibitor/antagonist; knock-down; lenalidomide; migration; model development; novel; patient population; phase 2 study; pleiotropism; potential biomarker; pre-clinical; preclinical study; prospective; protein expression; protein protein interaction; recruit; repository; research clinical testing; response; secondary analysis; side effect; small molecule inhibitor; targeted treatment; trend; tumor growth; ubiquitin ligase; ubiquitin-protein ligase

Development of immunomodulatory drugs. The antiangiogenic properties of thalidomide reported by D'Amato and colleagues prompted its clinical evaluation in various solid tumors, including prostate cancer. Thalidomide has demonstrated clinical activity in various malignancies affecting immunomodulatory and angiogenesis pathways. The development of novel thalidomide analogs with improved efficacy and decreased toxicity is an ongoing research effort in our laboratory. Previously, we showed that one of the products of cytochrome P450 2C19 isozyme biotransformation of thalidomide, 5'-OH-thalidomide, is responsible for the drug's antiangiogenic activity. Based on the chemical structure of this metabolite, we collaborate with Drs. Nigel Greig (NIA/NIH) and Michael Gutschow to synthesize novel thalidomide analogs, evaluate them using in vitro and in vivo models to assess activity, and characterize their structure-activity-relationships for further rational drug design. We have synthesized over 315 novel analogs of thalidomide and screened them for inhibition of inflammation and angiogenesis using various in vitro, ex vivo, and in vivo drug development models (e.g., rat aorta ring model, human saphenous vein model, cultured endothelial cells, migration and tube formation assays). In collaboration with Dr. Neil Vargesson, we conducted an in vivo screen of a library of new analogs to determine which agents demonstrate activity using the in vivo zebrafish and chicken embryo model systems. We identified the most potent of these agents and have patented them. We continue to develop these compounds, which appear to have minimal side effects in initial preclinical toxicology studies and may have improved pharmacology over the two FDA approved thalidomide analogs. We have optimized for both antiangiogenic and anti-inflammatory properties of these immunomodulatory drugs (IMiDS), which means the clinical indication can go beyond hematological malignancies and could have activity in solid tumors. This work in antiangiogenic/anti-cancer drug development serves not only to advance the field of antiangiogenic therapy but also to discover new treatment paradigms that focus on immunomodulation for advanced, metastatic disease. We have recently completed characterization of the antiangiogenic activity of polyfluorinated benzamides as well as adamantyl and noradamantyl phthalimidines. Efforts are ongoing to identify potential leads for in vivo toxicology and pharmacology studies in xenograft models. Cereblon (CRBN) is a substrate recruiter element of the E3 cullin 4-RING ubiquitin ligase complex, and a binding target of IMiDs. CRBN is responsible for the pleiotropic effects of IMiDs, yet its function in angiogenesis and in mediating the antiangiogenic effects of IMiDs remains unclear. We investigated the role of CRBN in the angiogenic process and in propagating the antiangiogenic effects of IMiDs in vitro. siRNA-mediated CRBN knock down in human endothelial cells (HUVEC and HMVEC-L), did not affect endothelial cell proliferation, migration, or tube formation. Using CRBN-deficient mice, we further demonstrated that microvessal formation can occur independently of cereblon in the ex vivo mouse aortic ring model. The cereblon E3 ubiquitin ligase complex can recruit endothelial cell-specific factors, AGO2 (associated with angiogenesis), and SALL4 (associated with embryogenesis/angiogenesis), for ubiquitin-mediated degradation. Knockdown of CRBN caused a corresponding increase in AGO2 and SALL4 protein expression and IMiD treatment was able to rescue the siCRBN effect to increase the CRBN expression. These findings suggest one potential mechanism of action that likely involves a tightly coordinated regulation of CRBN with endothelial cell targets and highlight the need to further elucidate the mechanism(s), which could include cereblon-independent pathways, through which IMiDs exert their antiangiogenic effects. Carfilzomib-lenalidomide-dexamethasone (KRd) therapy has yielded promising results in patients with newly diagnosed multiple myeloma (NDMM). CRBN is the direct molecular target of lenalidomide and genetic polymorphisms in CRBN have been associated with lenalidomide efficacy. In this study, we assessed the correlation of five single nucleotide variants (SNVs) in the CRBN gene with clinical response and outcomes in patients with NDMM administered KRd therapy with lenalidomide maintenance, achieving favorable trial endpoints in a prospective Phase II study (NCT01402284). Of the observed SNVs, no associations with KRd therapy response were found in this patient cohort, although strong trends in hypoalbuminemia grade and hyperbilirubinemia grade emerged across the CRBN rs1672753 genotype (P = 0.0008) and the rs1714327 genotype (P = 0.0010), respectively. Our results do not provide conclusive support for the predictive utility of CRBN gene polymorphisms as potential biomarkers of clinical response to lenalidomide-based therapy in our patient population. However, these findings remain to be validated in prospective studies using larger patient populations. Development of HIF-1alpha inhibitors. The hypoxia-inducible factor (HIF) is fundamentally involved in tumor angiogenesis, invasion, and energy metabolism. Inhibition of HIF-1 represents an attractive therapeutic strategy for targeting hypoxia, a hallmark of many solid tumors, and tumor angiogenesis. One promising approach for directly inhibiting HIF-1 activity is by disrupting the tight binding between HIF-1a and p300. Previously, our laboratory developed an in vitro fluorescence binding assay that can be used in a high-throughput screen to identify small-molecule inhibitors of HIF-1a through inhibiting the binding interaction between the C-terminal transactivation domain (CTAD) of HIF-1a and the cysteine/histidine-rich 1 (CH1) domain of p300. Using our HIF-1a/p300 assay, we performed high-throughput screen of NCI's Natural Products Repository in collaboration with Dr. Kirk Gustafson (Molecular Targets Laboratory/NCI). This effort led to the discovery of a series of pyrroloiminoquinone alkaloids including discorhabdin and makaluvamine alkaloids, originating from a Latrunculia sp. of marine sponge, as potential HIF-1a/p300 inhibitors. Preclinical studies are ongoing to understand the mechanisms of these novel compounds.

免疫调节药物的发展。D'Amato及其同事报道的沙利度胺的抗血管生成特性促使其在包括前列腺癌在内的各种实体肿瘤中的临床评估。沙利度胺在多种影响免疫调节和血管生成途径的恶性肿瘤中显示出临床活性。开发新的沙利度胺类似物，提高疗效和降低毒性是我们实验室正在进行的研究工作。先前，我们发现细胞色素P450 2C19同工酶生物转化沙利度胺的产物之一，5'- oh -沙利度胺，是该药物抗血管生成活性的原因。基于这种代谢物的化学结构，我们与dr。Nigel Greig （NIA/NIH）和Michael Gutschow将合成新的沙利度胺类似物，利用体外和体内模型评估其活性，并表征其结构-活性-关系，为进一步合理的药物设计提供依据。我们已经合成了超过315种新的沙利度胺类似物，并通过各种体外，离体和体内药物开发模型（例如，大鼠主动脉环模型，人隐静脉模型，培养内皮细胞，迁移和管形成实验）筛选它们对炎症和血管生成的抑制作用。与Neil Vargesson博士合作，我们对一个新的类似物库进行了体内筛选，以确定哪些药物在体内斑马鱼和鸡胚胎模型系统中表现出活性。我们确定了这些药物中最有效的药物，并申请了专利。我们继续开发这些化合物，这些化合物在最初的临床前毒理学研究中似乎副作用最小，并且可能比FDA批准的两种沙利度胺类似物具有更好的药理学效果。我们已经优化了这些免疫调节药物（IMiDS）的抗血管生成和抗炎特性，这意味着临床适应症可以超越血液系统恶性肿瘤，并且可能在实体肿瘤中具有活性。这项在抗血管生成/抗癌药物开发方面的工作不仅有助于推进抗血管生成治疗领域，而且还有助于发现新的治疗范式，重点关注晚期转移性疾病的免疫调节。我们最近完成了对多氟苯酰胺以及金刚烷酰和去甲金刚烷酰邻苯二胺抗血管生成活性的表征。目前正在努力确定异种移植模型体内毒理学和药理学研究的潜在线索。Cereblon （CRBN）是E3 cullin 4-RING泛素连接酶复合物的底物招募元件，也是IMiDs的结合靶点。CRBN对IMiDs的多效作用负责，但其在血管生成和介导IMiDs的抗血管生成作用中的功能尚不清楚。我们在体外研究了CRBN在血管生成过程中的作用，以及在IMiDs抗血管生成作用中的作用。在人内皮细胞（HUVEC和HMVEC-L）中，sirna介导的CRBN敲低不影响内皮细胞的增殖、迁移或管的形成。利用crbn缺陷小鼠，我们进一步证明了在离体小鼠主动脉环模型中，微血管形成可以独立于小脑发生。小脑E3泛素连接酶复合物可以招募内皮细胞特异性因子AGO2（与血管生成相关）和SALL4（与胚胎发生/血管生成相关），用于泛素介导的降解。CRBN的敲低引起AGO2和SALL4蛋白表达的相应增加，IMiD处理能够挽救siCRBN的作用，增加CRBN的表达。这些发现提示了一种潜在的作用机制，可能涉及到CRBN与内皮细胞靶点的紧密协调调节，并强调需要进一步阐明其机制，其中可能包括不依赖小脑的途径，IMiDs通过该途径发挥其抗血管生成作用。卡非佐米-来那度胺-地塞米松（KRd）治疗新诊断的多发性骨髓瘤（NDMM）患者取得了令人鼓舞的结果。CRBN是来那度胺的直接分子靶点，CRBN的遗传多态性与来那度胺的疗效有关。在这项研究中，我们评估了CRBN基因中5个单核苷酸变异（snv）与来那度胺维持治疗的NDMM患者的临床反应和结局的相关性，在一项前瞻性II期研究（NCT01402284）中获得了有利的试验终点。在观察到的snv中，在该患者队列中未发现与KRd治疗反应相关，尽管在CRBN rs1672753基因型（P = 0.0008）和rs1714327基因型中分别出现了低白蛋白血症等级和高胆红素血症等级的强烈趋势（P = 0.0010）。我们的研究结果并不能为CRBN基因多态性作为来那度胺治疗临床反应的潜在生物标志物的预测效用提供确凿的支持。然而，这些发现仍需在更大患者群体的前瞻性研究中得到验证。hif -1 α抑制剂的发展。缺氧诱导因子（HIF）从根本上参与肿瘤血管生成、侵袭和能量代谢。抑制HIF-1是针对缺氧的一种有吸引力的治疗策略，这是许多实体瘤和肿瘤血管生成的标志。一种有希望的直接抑制HIF-1活性的方法是破坏HIF-1a和p300之间的紧密结合。此前，我们的实验室开发了一种体外荧光结合试验，可用于高通量筛选，通过抑制HIF-1a的c端反活化结构域（CTAD）与p300的半胱氨酸/组氨酸富1 （CH1）结构域之间的结合相互作用来识别HIF-1a的小分子抑制剂。通过HIF-1a/p300检测，我们与Kirk Gustafson博士（Molecular Targets Laboratory/NCI）合作，对NCI的天然产物库进行了高通量筛选。这一努力导致了一系列吡咯亚胺醌类生物碱的发现，包括discorhabdin和makaluvamine生物碱，原产于海绵Latrunculia sp.，作为潜在的HIF-1a/p300抑制剂。临床前研究正在进行中，以了解这些新化合物的机制。