Project 2 - Joshua Hood, PhD

项目 2 - 约书亚·胡德博士

• 批准号: 10026256
• 负责人: Joshua L. Hood
• 金额: $ 21.84万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-10 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10026256
• 关键词: 3-Dimensional; Adjuvant; Antigen Presentation; Attenuated; Autologous; BAY 54-9085; Bee Venoms; Bees; Biological Assay; Blood; Cancer Etiology; Cancer Vaccines; Cell Line; Cells; Centers of Research Excellence; Cirrhosis; Coupled; Development; Doctor of Philosophy; Effector Cell; Enzyme-Linked Immunosorbent Assay; FDA approved; Growth; Harvest; HepG2; Hepatitis; Homing; Human; Immune; Immune Cell Suppression; Immune checkpoint inhibitor; Immune response; Immunity; Immunocompetent; Immunosuppression; Immunotherapeutic agent; Immunotherapy; Impairment; In Vitro; Inflammatory; Injections; Investigation; Kupffer Cells; Liver; Liver Fibrosis; Liver diseases; Malignant Epithelial Cell; Malignant Neoplasms; Malignant neoplasm of lung; Mediating; MicroRNAs; Modeling; Modernization; Mononuclear; Mus; Myeloid-derived suppressor cells; Neoplasm Metastasis; Nivolumab; Participant; Pathogenicity; Pathologic; Patients; Peptides; Phagocytes; Phenotype; Plasma; Population; Primary carcinoma of the liver cells; Process; Proteins; Quantitative Reverse Transcriptase PCR; Reporting; Research; Role; Source; Sting Injury; System; Testing; Therapeutic; Time; Toxic effect; Toxicology; Treatment Efficacy; Tyrosine Kinase Inhibitor; Vaccine Adjuvant; arginase; cancer therapy; checkpoint therapy; exosome; extracellular vesicles; hepatocellular carcinoma cell line; in vivo; light scattering; macrophage; melanoma; mortality; nanocarrier; nanomaterials; nanomedicine; nanovesicle; neoplastic cell; non-alcoholic; nonalcoholic steatohepatitis; polarized cell; problem drinker; recruit; response; tumor; tumor growth; zeta potential

Hepatocellular carcinoma (HCC) is the 3rd cause of cancer-related mortality. It is inherently difficult to treat given the often-times co-occurrence of underlying cirrhosis which exacerbates conventional chemotherapeutic toxicity. A sizeable number of patients remain non-responsive or “untreatable,” to cutting edge immune checkpoint inhibitor therapy necessitating the development of new or augmentative therapies. Given their role in removing foreign nanomaterials as participants of the mononuclear phagocyte system, coupled to their role in mediating HCC, Kupffer cells (KCs) are an ideal immunotherapeutic target for small extracellular vesicles (sEVs). Small EVs are enriched in exosome nanovesicles. Our previous studies demonstrate that melanoma and lung cancer sEVs can directly induce a pro-tumor M2-like Mφ phenotype or be modified with melittin peptide to induce anti-tumor M1-like Mφs. Theoretically, HCC sEVs might also be modified with melittin to induce anti-tumor M1 KC Mφs. The bee venom peptide melittin is a powerful adjuvant for activating M1 immunity. The FDA approves the use of bee venom injections for bee sting immunotherapy. In this proposal we hypothesize that (i.) HCC sEVs can be converted into stable melittin adjuvant nanocarriers, and (ii.) melittin-modified HCC sEVs associate with and induce KCs toward an anti-tumor M1-like phenotype in vivo. In aim 1, HCC sEVs will be converted into stable melittin nanocarriers. A fluorescent red, bioluminescent, 3D HepG2-Red-Fluc spheroid sEV source model will be developed. We will compare 2D versus 3D HepG2-Red-Fluc sourced natural and melittinized sEVs for their ability to influence primary KC polarization in vitro and determine differences in M1/M2 polarizing miRNA content using qRT-pcr. In aim 2, an orthotopic syngeneic model using bioluminescent Hepa1-6-Fluc-Neo cells in immunocompetent C57L/6 mice will be used. Natural vs. melittinized Hepa1-6-Fluc-Neo sEVs will be compared to determine whether they influence KC M1/M2 polarization. We will also assess whether pre- treatment with melittinized Hepa1-6-Fluc-Neo sEVs inhibits subsequent orthotopic HCC growth. Studies will be accomplished via utilization of all U of L Hepatobiology & Toxicology COBRE cores. The results of these studies will further our understanding of the relationship between natural HCC sEVs and KC tumor supportive functions, and a therapeutic means to antagonize this pathogenic relationship using melittin-modified KC sEVs will also be evaluated. Translationally, sEV populations might be harvested from the blood of HCC patients, modified into personalized immunotolerant nanomedicines using melittin, or other agents and re-administered. The proposed studies also serve as a platform to pursue pathologic and therapeutic sEV investigations concerning other Mφ driven liver diseases including hepatitis, alcoholic and non-alcoholic-SH, and liver fibrosis.

肝细胞癌（HCC）是癌症相关死亡率的第三个原因。它是天生的 鉴于潜在的肝硬化的经常同时存在，这很难治疗，这会加剧 常规的化学治疗毒性。大量患者仍然无反应或 “不可治疗”，要开发新的 或增强疗法。鉴于他们在去除外国纳米材料作为参与者中的作用 单核吞噬细胞系统，与它们在介导HCC中的作用相结合，Kupffer细胞（KC）是理想的 小细胞外蔬菜（SEV）的免疫治疗靶标。小型电动汽车富含外泌体 纳米层。我们以前的研究表明，黑色素瘤和肺癌SEV可以直接诱导 亲肿瘤的M2样Mφ表型，或用蜂毒蛋白胡椒进行修饰，以诱导抗肿瘤M1样Mφ。 理论上的HCC SEV也可以用蜂蜜素修饰，以诱导抗肿瘤M1 KCMφs。 蜜蜂毒液胡椒粉是激活M1免疫力的有力调整。 FDA 批准将蜜蜂毒液注射用于蜂刺免疫疗法。在这个建议中，我们假设 （i。）HCC SEV可以转换为稳定的蜂素调节纳米载体，以及（ii。）修饰的（ii。） HCC SEV与体内的抗肿瘤M1样表型相关并诱导KC。 在AIM 1中，HCC SEV将转换为稳定的蜂胶纳米载体。荧光红， 将开发生物发光的3D HEPG2-RED-FLUC球体SEV源模型。我们将比较2D 对3D HEPG2-红色液体的自然和融化SEV的影响能够影响原发性KC 在体外极化，并使用QRT-PCR确定M1/M2极化miRNA含量的差异。 在AIM 2中，使用生物发光Hepa1-6-纤维蛋白细胞的原位合理模型 将使用免疫功能的C57L/6小鼠。自然与融合的HEPA1-6-FLUC-NEO SEV将是 比较确定它们是否影响KC M1/M2极化。我们还将评估是否预先 用融化的HEPA1-6-FLUC-NEO SEV抑制后续原位HCC生长。研究 将通过利用所有肝病学和毒理学圆锥核心的利用来实现。 这些研究的结果将进一步了解我们对天然HCC之间关系的理解 SEV和KC肿瘤支持功能以及拮抗这种致病性的治疗手段 也将评估使用融化蛋白修饰的KC SEV的关系。在翻译上，人口种群 可以从HCC患者的血液中收获，并将其修改为个性化的不透明剂 纳米医学使用蜂胶或其他剂并重新管理。拟议的研究也是 购买有关其他Mφ驱动肝的病理和治疗性SEV调查的平台 包括肝炎，酒精和非酒精-SH以及肝纤维化在内的疾病。