Department of Medicine V, Hematology, Oncology and Rheumatology, University Hospital Heidelberg, 69120, Heidelberg, Germany.
J Hematol Oncol. 2024 Sep 16;17(1):85. doi: 10.1186/s13045-024-01604-y.
Relapsed and refractory acute myeloid leukemia (AML) carries a dismal prognosis. CAR T cells have shown limited efficacy in AML, partially due to dysfunctional autologous T cells and the extended time for generation of patient specific CAR T cells. Allogeneic NK cell therapy is a promising alternative, but strategies to enhance efficacy and persistence may be necessary. Proteasome inhibitors (PI) induce changes in the surface proteome which may render malignant cells more vulnerable to NK mediated cytotoxicity. Here, we investigated the potential benefit of combining PIs with CAR-expressing allogeneic NK cells against AML.
We established the IC50 concentrations for Bortezomib and Carfilzomib against several AML cell lines. Surface expression of class-I HLA molecules and stress-associated proteins upon treatment with proteasome inhibitors was determined by multiparameter flow cytometry. Using functional in vitro assays, we explored the therapeutic synergy between pre-treatment with PIs and the anti-leukemic efficacy of NK cells with or without expression of AML-specific CAR constructs against AML cell lines and primary patient samples. Also, we investigated the tolerability and efficacy of a single PI application strategy followed by (CAR-) NK cell infusion in two different murine xenograft models of AML.
AML cell lines and primary AML patient samples were susceptible to Bortezomib and Carfilzomib mediated cytotoxicity. Conditioned resistance to Azacitidine/Venetoclax did not confer primary resistance to PIs. Treating AML cells with PIs reduced the surface expression of class-I HLA molecules on AML cells in a time-and-dose dependent manner. Stress-associated proteins were upregulated on the transcriptional level and on the cell surface. NK cell mediated killing of AML cells was enhanced in a synergistic manner. PI pre-treatment increased effector-target cell conjugate formation and Interferon-γ secretion, resulting in enhanced NK cell activity against AML cell lines and primary samples in vitro. Expression of CD33- and CD70-specific CARs further improved the antileukemic efficacy. In vivo, Bortezomib pre-treatment followed by CAR-NK cell infusion reduced AML growth, leading to prolonged overall survival.
PIs enhance the anti-leukemic efficacy of CAR-expressing allogeneic NK cells against AML in vitro and in vivo, warranting further exploration of this combinatorial treatment within early phase clinical trials.
复发和难治性急性髓系白血病（AML）预后不良。嵌合抗原受体 T 细胞（CAR T 细胞）在 AML 中的疗效有限，部分原因是自体 T 细胞功能障碍和生成患者特异性 CAR T 细胞的时间延长。异体自然杀伤（NK）细胞治疗是一种很有前途的替代方法，但可能需要增强疗效和持久性的策略。蛋白酶体抑制剂（PI）诱导细胞表面蛋白质组发生变化，这可能使恶性细胞更容易受到 NK 介导的细胞毒性作用。在这里，我们研究了将 PI 与表达嵌合抗原受体（CAR）的异体 NK 细胞联合应用于 AML 的潜在益处。
我们确定了硼替佐米（Bortezomib）和卡非佐米（Carfilzomib）对几种 AML 细胞系的 IC50 浓度。通过多参数流式细胞术测定用蛋白酶体抑制剂处理后细胞表面Ⅰ类 HLA 分子和应激相关蛋白的表达情况。使用功能体外测定，我们探索了 PI 预处理与 NK 细胞联合治疗的协同作用，NK 细胞未经或经 AML 特异性 CAR 构建物表达，对 AML 细胞系和原发性患者样本的抗白血病疗效。此外，我们还在两种不同的 AML 异种移植模型中研究了单一 PI 应用策略后输注（CAR-）NK 细胞的耐受性和疗效。
AML 细胞系和原发性 AML 患者样本对硼替佐米和卡非佐米介导的细胞毒性敏感。对阿扎胞苷/维奈托克的条件性耐药并未赋予对 PI 的原发性耐药性。PI 处理以时间和剂量依赖的方式降低 AML 细胞表面Ⅰ类 HLA 分子的表达。应激相关蛋白在转录水平和细胞表面上调。NK 细胞对 AML 细胞的杀伤以协同方式增强。PI 预处理增加效应细胞-靶细胞共轭形成和干扰素-γ分泌，导致体外 NK 细胞对 AML 细胞系和原发性样本的活性增强。表达 CD33 和 CD70 特异性 CAR 进一步提高了抗白血病疗效。在体内，硼替佐米预处理后输注 CAR-NK 细胞可减少 AML 生长，从而延长总生存期。
PI 增强了表达嵌合抗原受体的异体 NK 细胞在体外和体内对 AML 的抗白血病疗效，这需要在早期临床试验中进一步探索这种联合治疗方法。
