Role Of Monocytes In AIDS And As Targets For Antiviral Therapy

对艾滋病毒的天然防御 HIV在消除或中和宿主细胞衍生的防御分子方面的熟练程度有助于HIV感染CD4+趋化因子共受体+靶标。在这些天然保护分子中，一类细胞内载脂蛋白B的mRNA编辑酶--催化多肽样胞苷脱氨酶是结构性表达的，但被HIV病毒感染性因子灭活。最近，我们证明了I型干扰素是一种在天然免疫和获得性免疫中具有多种功能的细胞因子，在体外和体内都是一种有效的HIV抑制因子，其抗病毒活性的部分机制是通过诱导内源性APOBEC3家族成员来实现的。干扰素增强胞苷脱氨酶的能力提供了一种可能性，即病毒和靶细胞之间的平衡可能被改变，有利于宿主。使用微阵列鉴定和分离逆转录病毒抑制物与相关的干扰素毒性，进一步表征了干扰素诱导的转录图谱，揭示了包括IL-27在内的多个疑似抗病毒活性的分子。为了确定是否可以通过使用下游的IL-27来对抗HIV来规避干扰素的毒性，我们检查了IL-27是否直接调节胞苷脱氨酶。尽管IL-27可以诱导APOBEC，但它是以延迟的方式完成的。对潜在调控事件的剖析揭示了最初依赖于IL-27的干扰素和/或干扰素的诱导，进而诱导APOBEC3，并被干扰素受体阻断所抑制。除了巨噬细胞，IL-27-干扰素的连接在CD4+T细胞中也起作用，这与依赖干扰素的途径一致，是宿主细胞防御艾滋病毒的基础。 尽管干扰素的抗艾滋病毒作用已经被证明，但在未经治疗的仅感染艾滋病毒的受试者中，其有效性和安全性尚不清楚。因此，未经治疗的无病毒性肝炎的HIV感染志愿者接受聚乙二醇化干扰素治疗12周，检测HIV RNA、CD4计数、药代动力学、2，5寡腺苷合成酶(OAS)活性的药效学测定、干扰素诱导基因(IFIG)诱导和淋巴细胞增殖反应。根据这些临床研究，聚乙二醇化干扰素2a在HIV单一感染患者中具有显著的抗病毒活性，这种抗HIV作用与OAS蛋白和IFIG诱导有关。在正在进行的研究中，我们正在通过评估特定的IFIG及其对抑制HIV的贡献来确定降低病毒负担的机制，包括APOBEC、BST/tetherin、TRIM22和其他在体外研究中涉及的因素。 干扰素治疗减少了艾滋病毒感染者的病毒负担，但这种细胞因子也会导致免疫功能障碍和毒性，限制了其用途。通过对干扰素受体(IFNAR)结合位点的详细定位和产生具有影响受体相互作用的螺旋C结构变化的干扰素杂交种和突变体，可以改变限制逆转录病毒复制的能力。我们的数据显示，与野生型干扰素分子相比，干扰素构建物具有阻止HIV复制的不同能力，并且HIV抑制的方向性大小与APOBEC3基因的表达水平相关。在与IFNAR结合后，突变体触发离散的或共享的细胞内信号通路(JAK/Stat/PI3K/NF&954；B)，导致抗病毒调节可能与潜在的毒性效应分离，例如吲哚乙胺2，3-双加氧酶(IDO)。通过探索干扰素的结构和功能，相对于其诱导APOBEC和其他抗病毒分子的能力，有可能设计出新的干扰素相关分子，在减少毒性的同时保持其抗病毒和抗肿瘤活性的有益作用。基于这些有希望的研究，我们计划探索灵长类研究，以评估选定的突变体在临床前研究中是否表现出类似的最佳APOBEC/IDO比率活性，这可能对人类HIV感染的干预具有潜在的意义。 在早期的研究中，我们证明了一种合成的三萜类化合物和PPAR配体，2-氰基-3，12-二氧代油酸-1，9-二烯-28-酸(CDDO)，已知可以影响p21激酶抑制剂的表达(与病毒生命周期有关)，抑制病毒在巨噬细胞和PBMC中的复制。CDDO的甲酯衍生物(CDDO-Me)是唯一的口服生物利用型药物，没有显著的毒性，由于这种甲酯衍生物针对的是组织，而不是血液，我们专注于确定它在扁桃体组织来源的CD4+HIV靶细胞中抑制HIV的能力。在CDDO-Me抑制扁桃体细胞中HIV感染的潜在机制中，通过诱导血红素氧合酶(HO-1)(最近被证明可以干扰HIV感染)以及诱导抗病毒I型干扰素。这些途径和其他途径将在我们的体外研究中以及在患者群体中进一步评估。由于抗逆转录病毒治疗的特点往往是高毒性，并经常导致耐药病毒株的出现，因此识别可单独使用或与现有抗病毒药物联合使用的针对宿主细胞分子的新的抗病毒药物，可能会在治疗艾滋病毒感染方面提供额外的资源。

Innate defense against HIV Infection of CD4+ chemokine co-receptor+ targets by HIV is aided by the proficiency of HIV in eliminating or neutralizing host cell-derived defensive molecules. Among these innate protective molecules, a family of intracellular apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like (APOBEC) cytidine deaminases is constitutively expressed, but inactivated by HIV viral infectivity factor. Recently, we demonstrated that type I Interferon (IFNα), a cytokine with multiple functions in innate and adaptive immunity and a potent inhibitor of HIV in vitro and in vivo, exerts its anti-viral activity, in part, by inducing endogenous APOBEC3 family members. The ability of IFNα to augment cytidine deaminases offered the possibility that the balance between virus and target cell might be altered in favor of the host. Further characterization of transcriptional profiles induced by IFNα using microarrays to identify and dissociate retroviral inhibitors from associated IFN toxicities, revealed multiple molecules with suspected anti-viral activity, including IL-27. To establish whether IFNα toxicity might be sidestepped through the use of downstream IL-27 against HIV, we examined whether IL-27 directly regulated cytidine deaminases. Although IL-27 induces APOBECs, it does so in a delayed fashion. Dissecting the underlying regulatory events uncovered an initial IL-27-dependent induction of IFNα and/or IFNβ, which in turn, induces APOBEC3, inhibited by IFNα/β receptor blockade. In addition to macrophages, the IL-27-IFNα connection is operative in CD4+ T cells, consistent with an IFNα-dependent pathway underlying host cell defense to HIV. Although the anti-HIV effects of IFNα have been demonstrated, efficacy and safety in untreated subjects infected with only HIV was unknown. Therefore, untreated HIV-infected volunteers without viral hepatitis were treated with pegylated IFNα2a for 12 weeks and changes in HIV RNA, CD4 counts, pharmacokinetics, pharmacodynamic measurements of 2,5 oligoadenylate synthetase (OAS) activity, induction of IFN-inducible genes (IFIG) and lymphoproliferative responses were measured. Based on these clinical studies, peg-IFNα2a has significant anti-viral activity in HIV-monoinfected patients and this anti-HIV effect correlated with OAS protein and IFIG induction. In ongoing studies, we are defining the mechanisms of reduced viral burden by evaluating specific IFIG and their contribution to inhibition of HIV, including APOBEC, BST/tetherin, TRIM22 and others implicated in in vitro studies. IFNα therapy reduces viral burden in HIV infected individuals, but this cytokine can also lead to immune dysfunction and toxicities, limiting its utility. Through detailed mapping of IFN receptor (IFNAR) binding sites and generation of IFNα hybrids and mutants with structural changes in helix C that influence receptor interactions, the ability to limit retroviral replication can be altered. Our data show a differential ability of IFNα constructs to block HIV replication when compared to the wildtype IFNα molecule, and the directional magnitude of HIV inhibition correlated with levels of APOBEC3 gene expression. Subsequent to binding with distinct affinities to IFNAR, the mutants trigger discreet or shared intracellular signaling pathways (Jak/Stat/PI3K/NFκB) leading to antiviral regulation that may be dissociated from underlying toxic effects, exemplified by indoleamine 2,3-dioxygenase (IDO). By exploring the structure and function of IFNα relative to its ability to induce APOBEC and other anti-viral molecules, it may become possible to design novel IFN-related molecules, which preserve its beneficial roles in anti-viral and anti-tumor activity while reducing toxicities. Based on these promising studies, we plan to explore primate studies to evaluate whether selected mutants exhibit similar optimal APOBEC to IDO ratio activity in pre-clinical studies, with potential implications for intervention in HIV infection of humans. In earlier studies, we demonstrated that a synthetic triterpenoid and PPARγ ligand, 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO), known to influence p21 kinase inhibitor expression (linked to viral life cycle), suppressed viral replication in macrophages and PBMC. A methyl ester derivative of CDDO (CDDO-Me), which is uniquely orally bioavailable, does not have significant toxicities, and since this methyl ester derivative targets tissues, rather than blood, we have focused on defining its ability to inhibit HIV in tonsil tissue-derived CD4+ HIV target cells. Among the potential mechanisms by which CDDO-Me inhibits HIV infection in tonsil cells is through induction of hemoxygenase (HO-1), recently shown to interfere with HIV infection, as well as induction of antiviral type I IFN. These and other pathways will be further evaluated in our in vitro studies as well as in patient populations. As anti-retroviral therapy is often characterized by high toxicity and frequently results in the emergence of drug resistant virus strains, the identification of new anti-viral agents targeting host cell molecules that can be used independently or in conjunction with current anti-viral drugs may provide additional resources in the treatment of HIV infection.