Studies of Chemokine-Receptor Interactions with Chemokines and alarmins

趋化因子受体与趋化因子和警报素相互作用的研究

• 批准号: 7965166
• 负责人: JOOST J OPPENHEIM
• 金额: $ 115.53万
• 依托单位: DIVISION OF BASIC 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/7965166
• 关键词: Accounting; Adjuvant; Antibody Formation; Antigens; Antiviral Agents; Autoimmunity; B-Lymphocytes; C3H/HeN Mouse; CCR6 gene; Cell Line; Cells; Chimeric Proteins; Colon Carcinoma; Coupled; Cytoplasmic Granules; Cytotoxic T-Lymphocytes; Defensins; Dendritic Cells; Development; Endotoxins; Eosinophil-Derived Neurotoxin; Epithelial Cells; Family; G-Protein-Coupled Receptors; GTP-Binding Proteins; Gastrointestinal tract structure; Gi-alpha protein; Homologous Gene; Host Defense; Human; IgG1; Immune; Immune response; Immunity; Immunization; Immunologic Adjuvants; Infection; Inflammation; Ingestion; Injury; Interleukin-4; Interleukin-5; Laboratories; Lactoferrin; Leukocytes; Ligands; Malignant Neoplasms; Mononuclear; Mus; Myelogenous; Natural Killer Cells; Organism; Pain; Parasitic Diseases; Pattern; Pertussis Toxin; Preparation; Production; Property; Receptor Cell; Recruitment Activity; Signal Transduction; Source; System; T-Lymphocyte; TLR2 gene; TLR4 gene; Therapeutic; Tissues; Tracheobronchial; Trauma; Trees; Tumor Antigens; Vaccines; alpha-Defensins; aluminum sulfate; antimicrobial peptide; base; beta-Defensins; cathelicidin; cell injury; chemokine; chemokine receptor; cytokine; eosinophil; granulocyte; granulysin; in vivo; keratinocyte; macrophage; melanoma; member; microbial; monocyte; receptor; response; tumor; tumor growth

Overall, my laboratory investigates the interactions of cognate chemokine ligands and chemokine mimics with G-protein coupled chemotactic receptors and activating receptors with resultant effects on inflammation, immunity, autoimmunity, cancer and algesia. We have shown that a variety of antimicrobial peptides (AMPs) mimic chemokines and also have the capacity to rapidly activate host immune responses. We have proposed calling these early warning signals alarmins. Alarmins are characterized by having chemotactic activity for cells expressing GiPCR, together with the capacity to activate iDC to mature into antigen- presenting, T lymphocyte activating dendritic cells (mDC) with resultant in vivo immunoadjuvant effects. These activities of alarmins, if administered together with an antigen, result in considerable augmentation of both cellular and humoral in vivo immune responses to the antigen. We previously identified both alpha and beta types of defensins as alarmins with chemotactic and activating effects on immature dendritic cells (iDC) and in vivo immunoadjuvant effects. Some of the beta defensins interact with the CCR6 chemokine receptor, while alpha defensins interact with an as yet unknown G-Protein Coupled Receptors (GiPCR). Another antimicrobial peptide known as cathelicidin (LL37) and its murine homologue CRAMP are chemotactic for FPRL-1 receptors expressed on monocytes and precursors of iDC, induce the maturation of iDC and are equally as potent adjuvants in vivo as alum. Although alarmins are structurally distinct, they are rapidly released from granules of leukocytes or damaged cells. Alarmins can also be induced in response to proinflammatory stimulants by keratinocytes or epithelial cells lining the GI tract, GU tract and tracheobronchial tree. As such, alarmins probably represent an early warning system to alert the host defense to danger signals During the previous year we also investigated the immune activating and chemotactic effects of another leukocyte granule derived alarmin known as eosinophil derived neurotoxin (EDN), which is a member of the RNAse family, and has antiviral activity including anti-HIV activity. EDN based on its interactions with a pertussis toxin susceptible GiPCR is chemotactic for iDC, and mDC. In addition, EDN based on interactions with TLR2 activates iDC to produce multiple proinflammatory cytokines and to mature into mDC. EDN also has potent in vivo immunostimulating effects. However, EDN activated DC in a manner that preferentially promotes a Th2 pattern of polarization with greater induction of IgG1 antibody production by B-lymphocytes and the production of Interleukin (IL)-5 and 13, but not IL-4 or IFNgamma. Although EDN has the properties of an alarmin this suggests that the adjuvant effect of EDN may prove more useful in vaccines aimed at immunization against parasitic diseases rather than tumors. We have also shown that lactoferrin has alarmin activities. We have used GMP preparations of human lactoferrin (TLF) for our studies. TLF is being evaluated as an anti-tumor therapeutic at other centers. TLF is chemotactic for monocytic APCs, but uses an unidentified receptor to recruit and mobilize mononuclear cells. Since C3H/HeJ, unlike C3H/HeN, mice are not activated by TLF, this suggests TLR4 is involved in the immune adjuvant effects of TLF. TLF activity appears to survive transport through the gastrointestinal tract of mice. We are therefore investigating whether TLF can exacerbate IBD and whether ingestion of lactoferrin may thus predispose subjects to develop colon cancer. During the past year we identified granulysin, which is normally stored in the granules of natural killer (NK) cell and cytotoxic T cells (CTL), as an alarmin. Both 9kDa and 15kDa forms of granulysin are chemotactic for immature myeloid derived dendritic cells (iDC) and can activate such iDC to mature into antigen presenting dendritic cells (mDC). Granulysin interacts with an unidentified pertussis toxin sensitive Gi alpha protein cell receptor (GiPCR) to induce chemotactic responses. Like TLF, granulysin also fails to induce maturation of C3H/HeJ iDC, suggesting TLR4 is involved in the immune activating effect of granulysin. The possibility that endotoxin contamination accounts for the TLR4 stimulating effect of granulysin has been ruled out. Consequently, we can add NK and CTL cells to eosinophils, granulocytes, epithelial cells and macrophages as sources of alarmins. We are at present preparing fusion proteins consisting of some of the more potent alarmins fused to several different melanoma tumor antigens in order to obtain a potent antitumor vaccine.

总的来说，我的实验室研究了同源趋化因子配体和趋化因子模拟物与g蛋白偶联趋化受体和激活受体的相互作用，从而对炎症、免疫、自身免疫、癌症和痛觉产生影响。我们已经证明，多种抗菌肽（amp）模拟趋化因子，也具有快速激活宿主免疫反应的能力。我们建议把这些早期预警信号称为警报。警报器的特点是对表达GiPCR的细胞具有趋化活性，同时具有激活iDC成熟为抗原呈递、T淋巴细胞激活树突状细胞（mDC）的能力，从而产生体内免疫佐剂作用。如果与抗原一起使用，这些警报器的活性会导致细胞和体液对抗原的体内免疫反应显著增强。我们之前发现α和β两种类型的防御素都是对未成熟树突状细胞（iDC）具有趋化和激活作用的警报器，并具有体内免疫佐剂作用。一些β -防御素与CCR6趋化因子受体相互作用，而α -防御素与一种未知的g蛋白偶联受体（GiPCR）相互作用。另一种抗菌肽被称为cathelicidin （LL37）及其小鼠同源物CRAMP，对单核细胞和iDC前体上表达的FPRL-1受体具有趋化作用，诱导iDC成熟，并且在体内与明矾一样是有效的佐剂。虽然警报器在结构上是不同的，但它们能从白细胞颗粒或受损细胞中迅速释放出来。在促炎刺激物的作用下，胃肠道、胃肠道和气管支气管内的角质形成细胞或上皮细胞也可诱发警报器。在过去的一年里，我们还研究了另一种白细胞颗粒来源的警报蛋白的免疫激活和趋化作用，称为嗜酸性神经毒素（EDN），它是RNAse家族的成员，具有抗病毒活性，包括抗hiv活性。EDN基于其与百日咳毒素敏感基因pcr的相互作用，对iDC和mDC具有趋化作用。此外，EDN基于与TLR2的相互作用，激活iDC产生多种促炎细胞因子，成熟为mDC。EDN还具有强大的体内免疫刺激作用。然而，EDN激活DC的方式优先促进Th2极化模式，更大程度地诱导b淋巴细胞产生IgG1抗体和白细胞介素(IL)-5和13的产生，但不诱导IL-4或IFNgamma。虽然EDN具有警报的特性，但这表明EDN的佐剂作用可能在针对寄生虫病而不是肿瘤的免疫疫苗中更有用。我们也证明了乳铁蛋白具有警示活性。我们采用GMP制备的人乳铁蛋白（TLF）进行研究。其他中心正在评估TLF作为抗肿瘤治疗药物。TLF对单核细胞APCs具有趋化作用，但使用一种未知的受体来招募和动员单核细胞。由于C3H/HeJ与C3H/HeN不同，小鼠不被TLF激活，这表明TLR4参与了TLF的免疫佐剂作用。TLF活性似乎在小鼠胃肠道运输中存活下来。因此，我们正在研究TLF是否会加剧IBD，以及摄入乳铁蛋白是否会因此使受试者易患结肠癌。在过去的一年里，我们发现颗粒蛋白是一种警报蛋白，它通常储存在自然杀伤细胞（NK）和细胞毒性T细胞（CTL）的颗粒中。9kDa和15kDa形式的颗粒蛋白对未成熟的髓源性树突状细胞（iDC）具有趋化作用，并能激活这些iDC成熟为抗原呈递树突状细胞（mDC）。颗粒酶与一种未知的百日咳毒素敏感Gi α蛋白细胞受体（GiPCR）相互作用，诱导趋化反应。与TLF一样，颗粒素也不能诱导C3H/HeJ iDC成熟，说明TLR4参与了颗粒素的免疫激活作用。排除了内毒素污染导致颗粒蛋白刺激TLR4的可能性。因此，我们可以将NK和CTL细胞添加到嗜酸性粒细胞、粒细胞、上皮细胞和巨噬细胞中作为警报来源。目前，我们正在制备融合蛋白，将一些更有效的警报器与几种不同的黑色素瘤肿瘤抗原融合，以获得一种有效的抗肿瘤疫苗。