Immune-suppressive Myeloid Cells in the Glioma Microenvironment: Signaling Mechanisms and Novel Therapeutic Strategies

 描述(申请人提供)：多形性胶质母细胞瘤(GBM)是成人最常见的原发恶性脑肿瘤，确诊后中位生存期为15-21个月。抗GBM免疫策略构成了新的和令人兴奋的治疗佐剂，以提高手术、化疗和放射治疗的存活率。然而，开发有效的抗GBM免疫反应以提高患者的存活率一直是具有挑战性的。由于可以诱导针对GBM抗原的全身免疫反应，临床上的失败被认为是由于GBM诱导的免疫抑制。GBM患者的免疫抑制是由多种机制介导的，其中包括积聚在肿瘤微环境中的未成熟髓样细胞(IMCs)。未成熟髓系细胞的亚型有：(1)髓系抑制细胞(MDSCs)，(2)免疫抑制肿瘤相关巨噬细胞(TAMs)，(3)Tie2单核细胞(TEM)。基底膜将未成熟的髓样细胞招募到肿瘤微环境中，在那里它们抑制抗肿瘤免疫反应，例如，通过直接抑制T细胞效应器功能。其他免疫抑制机制包括：Treg、免疫抑制分子(即吲哚胺2，3-双加氧酶1)、细胞毒性T淋巴细胞抗原4(CTLA4)和程序性死亡1受体配体(PDL1)，以及细胞因子(如IL10、转化生长因子β)。为了确定吸引免疫抑制的IMC进入GBM微环境的分泌因子，我们对内源性和可移植的小鼠和人GBM细胞进行了DNA芯片分析，并确定CXCL12是一个可能的候选者。我们还在GBM微环境中的未成熟髓系细胞上发现了同源的CXCL12受体CXCR4，支持CXCL12/CXCR4在吸引IMC进入GBM微环境中发挥重要作用的假说。为了确定CXCL12-CXCR4信号在GBM进展和调节抗GBM免疫治疗中所起的作用，我们建议使用免疫活性的、基因工程的内源性小鼠GBM模型。颅内肿瘤是由睡美人(SB)介导的插入人类GBM中发现的基因改变引起的。初步数据显示，来自可移植和SB诱导的GBM的条件培养液在体外都能诱导IMCs的高水平扩张。在活体的基底膜模型中，我们观察到基底膜微环境和外周循环中IMCs的聚集。阻断CXCR4可显著延长内源性GBM小鼠的中位生存期。我们将使用CXCL12和/或CXCR4基因消融模型来检验这一假设，即CXCL12-CXCR4信号轴在决定GBM微环境的免疫谱中起主要作用，无论是定性还是定量，从而对疾病的进展具有深远的影响。我们进一步假设，阻断IMCs的积聚与抗GBM免疫刺激策略相结合将为治疗恶性脑癌提供一种强有力的辅助手段。

 DESCRIPTION (provided by applicant): Glioblastoma multiforme (GBM) is the most common primary malignant brain tumor in adults; median survival from diagnosis is ~15-21 months. Anti-GBM immune strategies constitute novel and exciting therapeutic adjuvants to improve survival due to surgery, chemo- and radiotherapy. However, it has been challenging to develop effective anti-GBM immune responses that translate into increased patients' survival. As systemic immune responses against GBM antigens can be induced, clinical failure is thought to be due to powerful GBM induced immune suppression. Immune suppression in GBM patients is mediated by various mechanisms that include immature myeloid cells (IMCs) that accumulate in the tumor microenvironment. Subtypes of immature myeloid cells are: (i) myeloid derived suppressor cells (MDSCs), (ii) immunosuppressive tumor associated macrophages (TAMs), and, (iii) Tie2+ monocytes (TEMs). GBMs recruit immature myeloid cells to the tumor microenvironment where they inhibit anti-tumor immune responses, for example, by directly inhibiting T-cell effector function. Additional immune suppressive mechanisms involve: accumulation of Tregs, immunosuppressive molecules (i.e., indoleamine2, 3-dioxygenase 1 (IDO), cytotoxic T-lymphocyte antigen 4 (CTLA4), and programmed death 1 receptor ligand (PDL1), and cytokines, (i.e., IL10, TGFβ). To identify secreted factors which attract immune-suppressive IMCs into the GBM microenvironment we performed DNA microarray analysis on endogenous and transplantable mouse and human GBM cells and identified CXCL12 as a possible candidate. We also identified CXCR4, the cognate CXCL12 receptor, on immature myeloid cells within the GBM microenvironment supporting the hypothesis that CXCL12/CXCR4 plays an important role in attracting IMCs to the GBM microenvironment. To ascertain the role played by CXCL12-CXCR4 signaling in GBM progression and in regulating anti-GBM immune therapies, we propose to use an immune competent, genetically engineered endogenous mouse GBM model. Intracranial tumors are induced by Sleeping Beauty (SB)-mediated insertion of genetic alterations found in human GBM. Preliminary data show that conditioned media from both transplantable and SB-induced GBM elicit a high level of IMCs' expansion in vitro. In GBM models in vivo, we observed accumulation of IMCs within the GBM microenvironment and in the peripheral circulation. CXCR4 blockade significantly prolonged median survival of mice bearing endogenous GBM. We will use CXCL12 and/or CXCR4 gene ablation models to test the hypothesis that CXCL12-CXCR4 signaling axis plays a major role in determining the immune profile, both qualitatively and quantitatively, of the GBM microenvironment and thus has profound effects on disease progression. We further hypothesize that blocking accumulation of IMCs in combination with anti-GBM immune stimulatory strategies will provide a powerful adjuvant approach to treat malignant brain cancer.