Reprogramming the tumormicroenvironment to improve immunotherapy of glioblastoma

胶质母细胞瘤（GBM）是一种一致致命的恶性肿瘤，治疗选择有限。免疫检查点 阻断剂（ICB）已经彻底改变了几种恶性肿瘤的治疗，但在所有III期试验中均失败 在新诊断和复发的胶质母细胞瘤（GBM）患者中进行试验。ICB的这种有限功效是由于 GBM肿瘤微环境（TME）中由缺乏细胞毒性T细胞引起的严重免疫抑制 细胞，调节性T细胞，常驻巨噬细胞和小胶质细胞的丰度和骨髓浸润， 从骨髓中提取的细胞。Wnt信号传导通过帮助增殖、干性， 上皮细胞向间质细胞的转化和化学抗性。然而，Wnt信号转导在免疫中的作用， GBM中抑制是未知的。在我们的初步研究中，我们发现Wnt信号在细胞内升高， 鼠和人GBM。豪猪抑制剂WNT 974-阻断Wnt信号传导-与 抗PD-1抗体（α PD 1）可延长GBM荷瘤小鼠的存活时间。这种增加的存活率是 伴随着一种新的DC 3样树突状细胞状态的扩增和粒细胞髓样细胞的减少， 衍生的抑制细胞（gMDSC），其可以介导对应答肿瘤中的该组合的应答。 相比之下，反应不良的肿瘤显示单核细胞（m）MDSC增加，T细胞浸润不足， 和T细胞效应器功能，提示潜在的耐药机制。我们之前的研究表明， Wnt信号传导的缺失或药理学抑制破坏GBM脉管系统并使其渗漏。 此外，ICB本身会增加GBM患者的水肿，并需要使用高度耐受的类固醇。 免疫抑制我们的初步研究表明，氯沙坦，一种血管紧张素受体阻滞剂，可以减少， α PD 1诱导的水肿，并将免疫抑制性TME重新编程为免疫刺激性环境，以有利于T 细胞浸润和效应子功能。基于这些令人兴奋的发现，我们的总体假设是，Wnt 信号转导将GBM肿瘤微环境从免疫抑制重编程为免疫抑制 刺激，从而加强α PD 1治疗，并加入氯沙坦进一步增强结果， 克服治疗抵抗机制和减少水肿。我们将通过以下方式检验这一假设： 检查（a）抗原交叉呈递DC和（B）减少的gMDSC在介导免疫应答中的功能。 对WNT 974和α PD 1的反应（目的1）。我们还将检验对WNT 974 +α PD 1的抗性 由（a）mMDSC增加和（B）T细胞浸润和功能缺乏引起（目的2）。在目标3中，我们 检验氯沙坦与Wnt抑制剂和α PD 1联合使用将（a）减少mMDSC 和（B）减轻水肿并提供持久 WNT 974 +α PD 1难治性GBM的反应。如果成功，我们的结果将为设计提供信息， 未来的GBM临床试验，以改善ICB的结局，使用目前在非CNS临床试验中的药物 肿瘤（WNT 974：例如，NCT 01351103;和氯沙坦：NCT 03563248）。

Glioblastoma (GBM) is a uniformly fatal malignancy with limited treatment options. Immune checkpoint blockers (ICBs) have revolutionized the treatment of several malignancies, but have failed in all Phase III trials in newly diagnosed and recurrent glioblastoma (GBM) patients. This limited efficacy of ICBs is due to profound immunosuppression in the GBM tumor microenvironment (TME) caused by paucity of cytotoxic T cells, abundance of regulatory T cells, resident macrophages and microglia and infiltration of myeloid- derived cells from the bone marrow. Wnt signaling fuels GBM progression by aiding proliferation, stemness, epithelial-to-mesenchymal transition and chemoresistance. However, the role of Wnt signaling in immune suppression in GBM is not known. In our preliminary studies we found that Wnt signaling is elevated in murine and human GBMs. A porcupine inhibitor WNT974 -- that blocks Wnt signaling -- in combination with anti-PD-1 antibody (αPD1) prolonged the survival of GBM-bearing mice. This increased survival was accompanied by an expansion of a novel DC3-like dendritic cell state and decrease in granulocytic myeloid- derived suppressor cells (gMDSCs) that may mediate the response to this combination in responding tumors. By contrast, poorly-responding tumors showed an increase in monocytic (m) MDSCs, insufficient T cell infiltration and T cell effector function, suggesting potential resistance mechanisms. Our prior work shows that genetic deletion or pharmacological inhibition of Wnt signaling disrupts the GBM vasculature and makes it leaky. Moreover, ICBs themselves increase edema in GBM patients and require the use of steroids that are highly immunosuppressive. Our preliminary studies show that losartan, an angiotensin receptor blocker, can reduce αPD1-induced edema and reprogram the immunosuppressive TME to an immunostimulatory milieu to favor T cell infiltration and effector function. Building on these exciting findings, our overarching hypothesis is that Wnt signaling reprograms the GBM tumor microenvironment from immune suppressive to immune stimulatory, thus potentiating αPD1 therapy, and adding losartan further enhances the outcome by overcoming treatment resistance mechanisms, and reducing edema. We will test this hypothesis by examining the function of (a) antigen cross-presenting DCs and (b) decreased gMDSCs in mediating the response to WNT974 and αPD1 (Aim 1). We will also test the hypothesis that resistance to WNT974+αPD1 is caused by (a) increased mMDSCs and (b) lack of T cell infiltration and function (Aim 2). In Aim 3, we will test the hypothesis that the combination of losartan with Wnt-inhibition and αPD1 will (a) reduce mMDSCs infiltration and increase T cell infiltration and effector function and (b) alleviate edema and provide durable responses in GBMs that are refractory to WNT974+αPD1. If successful, our results will inform the design of future GBM clinical trials to improve the outcome of ICBs using agents currently in clinical trials for non-CNS tumors (WNT974: e.g., NCT01351103; and losartan: NCT03563248).