Novel monocyte effector function in CLL immune therapy

描述(由申请人提供)：慢性淋巴细胞白血病(CLL)是成人白血病最常见的形式，目前可用的治疗方法是无法治愈的。一种有希望的治疗形式是抗体疗法，即CLL细胞被抗CD20抗体如利妥昔单抗和ofatumab靶向。这导致了存活率的显著改善，特别是结合其他疗法，但完全缓解仍然相对罕见。单核细胞和巨噬细胞是抗体治疗的关键介质，这依赖于Fc？受体(Fc？R)活性。我们发现，这些Fc？R的激活导致sFlt-1的产生，sFlt-1是一种可溶性的、抑制形式的血管内皮生长因子受体。SFlt-1可抑制血管内皮生长因子信号转导，而血管内皮生长因子信号转导通路在CLL细胞存活中起重要作用。这些结果使我们假设，Fc？R介导的sFlt-1的产生可以抑制CLL细胞中的抗凋亡信号，这是抗体介导的抗肿瘤作用的重要部分。因此，在抗体治疗的背景下加强单核/巨噬细胞sFlt-1的产生可能是增强其有效性的有力手段。为了验证这一假设的预测，我们提出：目标1：分析sFlt-1对抗CD20的反应产生和功能。在这里，我们将a)确定sFlt-1与抗CD20抗体结合后主要负责产生sFlt-1的细胞，检测sFlt-1对CLL细胞存活的影响，并研究血管内皮生长因子介导的肿瘤细胞存活被抑制的机制(S)。我们还将研究单核细胞产生sFlt-1是否使CLL细胞更容易发生直接凋亡，以响应临床前和临床上用于治疗CLL的药物。目的：分析sFlt-1在体内抗体治疗中的产生和功能。我们将使用已建立的CLL小鼠模型来测试在抗体介导的B细胞耗竭过程中是否使用小鼠CD20抗体来检测sFlt-1的产生，b)确定负责sFlt-1产生的相关Fc？R效应细胞(S)，c)测试中和sFlt-1是否降低sFlt-1的产生 抗CD20抗体在小鼠CLL模型中的有效性，以及d)接受ofatumab单一治疗的CLL患者中sFlt-1水平是否与应答和无进展生存期相关。目的3：阐明Fc？R聚集性诱导sFlt-1的机制。在这里，我们将确定a)哪个激活的Fc？R负责sFlt-1的诱导，以及这种诱导是否受Fc？RIIb和SHIP的负调控，b)Fc？R诱导的sFlt-1的产生是直接的还是间接的影响，c)参与sFlt-1诱导的信号通路(S)。摘要：本研究完成后，我们将全面探索抗CD20抗体介导的单核巨噬细胞杀伤肿瘤细胞的全新机制。这些机制研究将为进一步提高抗CD20抗体的疗效提供信息，例如在CLL中使用ofatumumab，但可能在使用类似基于抗体的治疗的其他肿瘤中也是如此。

DESCRIPTION (provided by applicant): Chronic Lymphocytic Leukemia (CLL) is the most common form of adult leukemia and is incurable with currently available therapies. One promising form of treatment has been antibody therapy, where CLL cells are targeted by anti-CD20 antibodies such as rituximab and ofatumumab. This has led to significant improvements in survival, especially in combination with other therapies, but complete remissions are still relatively rare. Monocytes and macrophages are critical mediators of antibody therapy, and this depends upon Fc? receptor (Fc?R) activity. We have found that activation of these Fc?R leads to production of sFlt-1, a soluble, inhibitory form of the VEGF receptor. sFlt-1 can inhibit VEGF signaling, which has been shown to play a major role in CLL cell survival. These results led us to hypothesize that Fc?R-mediated sFlt-1 production can dampen anti- apoptotic signals in CLL cells, and that this accounts for a significant portion of antibody-mediated antitumor effects. Hence, strengthening monocyte / macrophage sFlt-1 production within the context of antibody therapy may be a powerful means of enhancing its effectiveness. To test the predictions of this hypothesis we propose: Aim 1: Analysis of sFlt-1 production and function in response to anti-CD20. Here, we will a) identify the cells primarily responsible for sFlt-1 production upon binding anti-CD20 antibodies, examine the effect of sFlt-1 on CLL cell survival, and study the mechanism(s) by which VEGF-mediated survival in tumor cells is inhibited. We will also b) examine whether sFlt-1 production by monocytes makes CLL cells more susceptible to direct apoptosis in response to agents in use preclinically and clinically for the treatment of CLL. Aim 2: Analysis of sFlt-1 production and function in antibody treatment in vivo. We will use established murine models of CLL to a) test whether sFlt-1 is produced during antibody-mediated B cell depletion using a murine CD20 antibody, b) identify relevant Fc?R-bearing effector cell(s) responsible for sFlt-1 production, c) test whether neutralizing sFlt-1 reduces the efficacy of anti-CD20 antibody in a murine CLL model, and d) Whether sFlt-1 levels in CLL patients receiving monotherapy with ofatumumab correlate with response and progression free survival. Aim 3: Elucidation of mechanism of sFlt-1 induction by Fc?R clustering. Here, we will determine a) which activating Fc?R is responsible for sFlt-1 induction and whether this induction is negatively regulated by Fc?RIIb and SHIP, b) whether Fc?R-induced sFlt-1 production is a direct or indirect effect, and c) the signaling pathway(s) involved in sFlt-1 induction. Summary: At completion of this study we will have fully explored an entire new mechanism of anti-CD20 mediated killing of tumor cells by monocytes and macrophages. These mechanistic studies will provide information to further enhance the efficacy of both anti-CD20 antibodies such as ofatumumab in CLL but potentially a wide variety of other tumors where similar antibody based treatments are utilized.