Leveraging doxorubicin immune-modulation, blood-brain barrier opening, and personalized medicine for effective immunotherapy in glioblastoma. A mechanistic approach and pharmacokinetic trial.

摘要 尽管临床前有希望，并且偶尔会在部分患者中长期（呃）长期控制肿瘤生长，但抗 PD1 药物 检查点阻断（aPD1）未能改善胶质母细胞瘤（GBM）的预后。未能证明 抗 PD1 抗体 (aPD1) 的益处可能是由于 1) 免疫抑制微环境中几乎没有 免疫原性肿瘤细胞； 2）aPD1穿过血脑屏障（BBB）的能力不足，无法 到达瘤周脑部的浸润性肿瘤细胞； 3）GBM的分子异质性。在这里我们 系统地应对这些挑战。我们的目标是引起对 Balstilimab 的强烈抗肿瘤反应 (Agenus Bio)，一种新型 aPD1 抗体，通过低剂量阿霉素调节免疫反应， 之前已经展示过。为了让药物充分渗透到组织中，我们将暂时破坏血脑 颅骨植入式超声设备 (SonoCloud-9) 与静脉微泡相结合的屏障 （美国/MB）。在我们正在进行的研究和临床试验中，我们已经证明了这种方法的可行性，包括 对增强和非增强组织进行靶向活检并测量药物组织分布。最后，我们的目标是 识别分子特征，从而预测 aPD1 免疫疗法的益处 我们之前的工作已将 MAPK 激活（pERK 染色）确定为假定的生物标志物。我们首先会 完善临床前模型中 DOX 和 aPD1 的给药顺序。二、为了核实和 优化 我们将在试验的导入阶段治疗一些试点患者。三、优化方案 在一项多队列研究中对接受 DOX 和 aPD1 Balstilimab 治疗的复发性 GBM 患者进行了测试 （DOX aPD1）。主要终点是切除标本和/或外周血中的肿瘤免疫反应 免疫反应、增强和非增强脑组织中的药物浓度（目标活检） 切除时间）和安全性。临床结果和生物标志物是次要终点。四组患者 将检查 DOX/aPD1 治疗在不同时间点开始的位置： 诱导（新辅助治疗，之前 切除术）；或术中切除时，分别有或没有 US/MB。非干预标准 护理队列将作为对照。我们的提案有 3 个具体目标： SA1：表征和优化效果 DOX aPD1 对 GBM 抗肿瘤免疫的影响。 SA2：确定 US/MB 对浓度的影响 人脑中的 DOX aPD1 和 GBM 中的抗肿瘤免疫反应。 SA3：调查是否有肿瘤 pERK/MAPK 激活可预测 GBM 对 DOX aPD1 的反应。成功完成将决定 i) DOX和aPD1的免疫调节价值； ii) 诱导（切除前）治疗和体内治疗的价值 抗肿瘤免疫反应评估； iii) BBB开放对于aPD1功效的价值（或不存在）； iv) pERK 是否可以预测反应并识别将从治疗中受益的患者。

ABSTRACT Despite preclinical promise and occasional long(er) term tumor growth control in subsets of patients, anti-PD1 checkpoint blockade (aPD1) has failed to improve outcome in glioblastoma (GBM). The failure to demonstrate a benefit to an anti-PD1 antibody (aPD1) may be due to 1) an immunosuppressive microenvironment with few immunogenic tumor cells; 2) insufficient aPD1 penetration across the blood-brain barrier (BBB), which fail to reach the infiltrating tumor cells in the peritumoral brain; and 3) molecular heterogeneity of GBM. Here we systematically address each of these challenges. We aim to elicit robust anti-tumoral response to Balstilimab (Agenus Bio), a novel aPD1 antibody by modulating the immune response by low doses of doxorubicin as has been shown previously. To allow for adequate drug tissue penetration, we will transiently disrupt the blood-brain barrier by a skull implantable ultrasound device (SonoCloud-9) in association with intravenous microbubbles (US/MB). In our ongoing research and clinical trials we have shown the feasibility of this approach including targeted biopsies of enhancing and non-enhancing tissue and measure drug tissue distribution. Lastly, we aim at identifying molecular characteristics that will allow to predict a benefit from aPD1 immunotherapy based on our prior work where we have identified MAPK activation (pERK staining) as a putative biomarker. We will firstly refine the sequence of DOX & aPD1 administration in preclinical models. Second, for verification and optimization we will treat a few pilot patients in the lead-in phase of our trial. Third, the optimized regimen will be tested in a multi-cohort study in patients with recurrent GBM who will be treated by DOX and the aPD1 Balstilimab (DOX+aPD1). Primary endpoints are tumor immune response in the resection specimen and/or peripheral immune response, drug concentrations in enhancing and non-enhancing brain tissue (targeted biopsies at the time of resection) and safety. Clinical outcome and biomarkers are secondary endpoints. Four cohorts of patient will be examined where DOX/aPD1 treatment starts at different time points: Induction (neoadjuvant, prior to resection); or intraoperatively upon resection, each with and without US/MB. A non-interventional standard of care cohort will serve as control. Our proposal has 3 specific aims: SA1: Characterize & optimize the effect of DOX + aPD1 on anti-tumoral immunity in GBM. SA2: Determine the effect of US/MB on the concentration of DOX + aPD1 in the human brain and anti-tumoral immune response in GBM. SA3: Investigate whether tumor pERK/MAPK activation predicts GBM response to DOX + aPD1. Successful completion will determine the i) value of immune modulation by DOX and aPD1; ii) the value of induction (pre-resection) treatment and in vivo evaluation of anti-tumor immune response; iii) the value (or absence thereof) of BBB-opening for aPD1 efficacy; and iv) whether pERK predicts response and identifies patients that will benefit from treatment.