Cell signaling in granulocyte transfusion

项目摘要 粒细胞输注(GTX)已被用作治疗危及生命的疾病的一种方法。 严重中性粒细胞减少患者的细菌和真菌感染。然而，它的临床结果往往是 由于对炎症部位的重新募集效率低、体内死亡迅速和病原体杀灭能力差而受阻 移植的中性粒细胞的能力。拟议研究的最终目标是确定和 描述在粒细胞过程中可以改善中性粒细胞性能的细胞和分子事件 输血。在上一个资助阶段，我们证明了提升细胞内PtdIns(3，4，5)P3信号 通过破坏PTEN途径增强中性粒细胞功能，增加细菌清除，并降低 中性粒细胞减少相关性肺炎小鼠模型的死亡率。我们进一步揭示了PTEN 破坏输注的中性粒细胞显著提高了粒细胞输注的效率。这些 结果证实了PtdIns(3，4，5)P3通路作为改善临床症状的治疗靶点的有效性 粒细胞输注的结局。尽管如此，PTEN是一种众所周知的肿瘤抑制因子及其干扰 与许多实体和血液系统癌症的发生有关，这使得它不适合 作为治疗的靶点。最近，我们报道了中性粒细胞中的PtdIns(3，4，5)P3信号也可以升高 通过破坏InsP6K1，一种负责合成InsP7的酶，InsP7是一种胞质分子，它对 调节PtdIns(3，4，5)P3信号。InsP6K1缺陷的中性粒细胞具有增强的杀菌作用 能力和他们的招募到炎症部位是增强的。重要的是，纯合子InsP6K1 KO小鼠是可存活的，没有表现出任何严重的身体或行为异常。任何类型的肿瘤都不是 在这些老鼠身上发现的。基于这些有趣的结果，我们假设InsP6K1的中断应该 是一种更安全，但同样有效的改善粒细胞输注的治疗策略。在本建议中 研究中，我们将使用小鼠中性粒细胞减少相关肺炎模型来验证这一假说。首先，我们将 研究干扰InsP6K1是否能上调PtdIns(3，4，5)P3信号并增强其蓄积 (包括中性粒细胞减少相关的大肠杆菌肺炎输注中性粒细胞的招募和存活)(AIM I)。 此外，我们将确定在输血的中性粒细胞中干扰InsP6K1是否最终可以增强 宿主防御(AIM II)，并减轻(AIM III)中性粒细胞减少相关细菌和真菌的严重程度 肺炎。最后，对于未来的临床干预，InsP6K1的抑制很可能通过使用 化合物。因此，我们将检查InsP6K抑制剂TNP的预处理是否也可以 提高粒细胞输注治疗中性粒细胞减少相关性肺炎的疗效(目标IV)。因此，在 根据PPG的总主题，这项研究将提供新的治疗靶点和/或 改善中性粒细胞在粒细胞输注中的性能从而缓解 移植围手术期中性粒细胞减少相关感染的严重程度。

Project Summary Granulocyte transfusion (GTX) has been utilized as a therapeutic approach for the treatment of life-threatening bacterial and fungal infections in severely neutropenic patients. However, its clinical outcome is often hampered by inefficiency of recruitment to sites of inflammation, rapid in vivo death, and poor pathogen killing capability of transplanted neutrophils. The ultimate goal of the proposed research is to identify and characterize cellular and molecular events that can improve neutrophil performance during granulocyte transfusion. In the last funding period, we demonstrated that elevating intracellular PtdIns(3,4,5)P3 signaling pathway by disrupting PTEN enhances neutrophil function, augments bacterial clearance, and reduces mortality rate in a murine model of neutropenia-associated pneumonia. We further revealed that PTEN disruption in transfused neutrophils significantly improves the efficacy of granulocyte transfusion. These findings confirm the validity of the PtdIns(3,4,5)P3 pathway as a therapeutic target for improving the clinical outcome of granulocyte transfusion. Nevertheless, PTEN is a well-known tumor suppressor and its disruption has been implicated in tumorigenesis of numerous solid and hematologic cancers, which renders it unsuitable as a therapeutic target. Recently, we reported that PtdIns(3,4,5)P3 signal in neutrophils can also be elevated by disrupting InsP6K1, an enzyme responsible for the synthesis of InsP7, a cytosolic molecule that negatively regulates PtdIns(3,4,5)P3 signaling. InsP6K1 deficient neutrophils possess an enhanced bacteria killing capability and their recruitment to the site of inflammation is augmented. Importantly, homozygous InsP6K1 KO mice are viable and do not display any gross physical or behavioral abnormalities. No tumors of any kind is discovered in these mice. Based on these intriguing results, we hypothesize that disruption of InsP6K1 should be a safer, yet equally effective therapeutic strategy for improving granulocyte transfusion. In this proposed study, we will use a mouse neutropenia-related pneumonia model to test this hypothesis. First, we will investigate whether disrupting InsP6K1 can elevate PtdIns(3,4,5)P3 signal and enhance the accumulation (including recruitment and survival) of transfused neutrophils in neutropenia-related E.coli pneumonia (Aim I). In addition, we will determine whether disrupting InsP6K1 in transfused neutrophils can ultimately enhance the host defense in (Aim II), and alleviate the severity of (Aim III) neutropenia-related bacterial and fungal pneumonia. Finally, for future clinical intervention, inhibition of InsP6K1 will most likely be achieved by using chemical compounds. Thus we will examine whether pre-treatment with an InsP6K inhibitor, TNP, can also augment the efficacy of granulocyte transfusion in neutropenia-related pneumonia (Aim IV). Thus, in accordance with the general theme of the PPG, this study will provide novel therapeutic targets and/or strategies for improving the performance of neutrophils in granulocyte transfusion and therefore alleviating the severity of neutropenia-related infection in the peri-transplant setting.