Assessing resistance mechanisms in synNotch-CAR T cell-mediated glioblastoma therapy and evaluation of potential mitigation strategies

胶质母细胞瘤(GBM)是最常见的原发恶性脑肿瘤，中位总生存期不到2年，复发率接近100%。因此，迫切需要新的治疗策略。嵌合抗原受体T细胞(CAR T)治疗前景看好，但仍需解决一些挑战，如缺乏理想的靶抗原、T细胞耗竭和肿瘤诱导的免疫抑制。最近，人们发现，嵌合形式的Notch是一种1型跨膜蛋白，可以作为开发新的细胞-细胞接触信号通路(SynNotch)的通用平台。这使得T细胞的发展成为可能，这些细胞使用合成Notch受体展示定制的治疗反应程序。新一代创新的T细胞电路已经开发出来，它可以识别胶质母细胞瘤细胞，这是基于“先启动后杀死”的策略。在这一策略中，仅在脑或GBM细胞上表达的启动抗原被synNotch受体识别，从而诱导CAR(synNotch-car=sync)的表达。这提供了安全靶向低特异性GBM抗原的可能性，也与良好的T细胞在体内的持久性有关。尽管这一新方法看起来很有希望，但尚不清楚同步T细胞在完整的免疫系统中的工作效率。最近的研究表明，程序性死亡配体1(PD-L1)和转化生长因子1(TGF1)等反向免疫调节分子在肿瘤诱导的免疫抑制中起着关键作用。因此，现在需要研究将Sync-T细胞疗法与在完整的免疫系统中阻断这些分子相结合。本研究的目的是探讨同步T细胞治疗对免疫活性C57BL/6小鼠的潜在耐药机制，该C57BL/6小鼠携带低免疫原性小鼠胶质瘤细胞系的EGFRvIII表达的脑瘤。冈田实验室开发的B-sync T细胞将用于治疗。这些细胞被CNS特异性抗原Brivican激活，并在激活时表达抗EGFRvIII CAR。B-sync疗法单独和联合静脉输注针对PD-1或转化生长因子-β1的单抗的疗效将被研究。作为一个次要目标，我还将确定由同步T细胞本身直接表达的抗PD-1或转化生长因子-1是否可以克服潜在的局部免疫抑制，并可能优于抗体注射，当作为额外的有效载荷集成在synNotch电路中时。

Glioblastoma (GBM) is the most common primary malignant brain tumor with a median overall survival of less than 2 years and a recurrence rate of nearly 100%. Therefore, new therapeutic strategies are urgently needed. Chimeric antigen receptor T cell (CAR T) therapy is promising, but several challenges such as lack of ideal target antigens, T cell exhaustion, and tumor-induced immunosuppression must be addressed. Recently, it was discovered that chimeric forms of Notch, a type 1 transmembrane protein, can serve as a general platform for the development of new cell-cell contact signaling pathways (Synthetic Notch "synNotch"). This enables the development of T cells that exhibit customized therapeutic response programs using synthetic Notch receptors. A new generation of innovative T cell circuits that recognize glioblastoma cells based on the "prime-and-kill" strategy has been developed. In this strategy, priming antigen expressed exclusively on brain or GBM cells is recognized by the synNotch receptor, thereby inducing the expression of a CAR (synNotch-CAR=SYNC). This offers the possibility of safely targeting low-specificity GBM antigens and is also associated with excellent T cell persistence in vivo. Although this new approach appears promising, it is not yet known how efficiently SYNC T cells work in an intact immune system. Recent studies have shown that counter-active immunoregulatory molecules such as programmed death-ligand 1 (PD-L1) and transforming growth factor ß1 (TGF-β1) appear to play a key role in tumor-induced immunosuppression. Therefore, studies combining SYNC-T cell therapy with blockade of these molecules in an intact immune system are now needed. The aim of this study is to investigate potential resistance mechanisms of SYNC T cell therapy in immunocompetent C57BL/6 mice bearing EGFRvIII-expressing brain tumors of the low-immunogenic SB28 mouse glioma cell line. B-SYNC T cells developed by the Okada laboratory will be used as therapy. These cells are primed by the CNS-specific antigen Brevican and express an anti-EGFRvIII CAR upon activation. The efficacy of B-SYNC therapy alone and in combination with intravenous infusion of monoclonal antibodies directed against PD-1 or TGF-ß1 will be investigated. As a subordinate goal, I will also determine whether anti-PD-1 or TGF-ß1 expression directly by the SYNC T cell itself can overcome potential local immunosuppression and may be superior to infusion of antibodies, when integrated as an additional payload in the synNotch circuit.