MAT2A as key regulator and therapeutic target in MLL leukemogenesis

Epigenetic dysregulation plays a pivotal role in MLL pathogenesis, therefore serving as suitable therapeutic point of attack. SAM is the universal methyl donor in human cells and is synthesized by MAT2A. MAT2A is already known to be deregulated in different cancer types. Here, we used our human CRISPR/Cas9-MLLr leukemia model, faithfully mimicking MLL patients’ pathology with indefinite growth potential in in vitro culture, to evaluate the unknown role of MAT2A as therapeutic target. Comparable to publicly available patient data, we detected MAT2A to be significantly overexpressed in our CRISPR/Cas9-MLLr model compared to healthy controls. By using non-MLLr and MLLr cell lines and our model, we detected an MLLr specific enhanced response to PF-9366, a new MAT2A inhibitor, by alteration of proliferation, viability, differentiation, apoptosis and cell cycling. Moreover, the combinational treatment of PF-9366 with chemotherapy or targeted therapies against the SAM-dependent methyltransferases DOT1L and PRMT5, revealed even more pronounced effects. In summary, we uncovered MAT2A as a key regulator in MLL leukemogenesis and its inhibition led to significant anti-leukemic effects. Therefore, our study paves the avenue for clinical application of PF-9366 to improve the treatment of poor prognosis MLLr leukemia. huCB CD34+ cells were engineered to bear endogenous MLL-AF4 translocations. Generated cells were treated with 15 µM of PF-9366 (Carbosynth) or DMSO for 4 days. RNA sequencing was performed using NextSeq mRNA Stranded Sequencing (Illumina) of two biological replicates comparing treated and non-treated cells.

表观遗传失调在MLL致病过程中发挥关键作用，因此成为极具潜力的治疗靶点。S-腺苷甲硫氨酸（SAM）是人类细胞中通用的甲基供体，由MAT2A编码合成。已有研究表明，MAT2A在多种癌症类型中存在表达失调。本研究利用我们构建的人类CRISPR/Cas9-MLLr白血病模型——该模型可在体外培养中具备无限增殖潜能，能够忠实复现MLL患者的病理特征——来评估MAT2A作为治疗靶点的未知作用。与公开可用的患者数据集一致，我们发现相较于健康对照，CRISPR/Cas9-MLLr模型中的MAT2A呈显著高表达。通过使用非MLLr与MLLr细胞系及本研究构建的模型，我们观察到MLLr细胞对新型MAT2A抑制剂PF-9366存在特异性增强的应答反应，具体体现为增殖、细胞活力、分化、凋亡及细胞周期的改变。此外，将PF-9366与化疗药物或针对SAM依赖性甲基转移酶DOT1L、PRMT5的靶向疗法联合使用时，可观察到更为显著的抗白血病效应。综上，本研究揭示MAT2A是MLL白血病发生过程中的关键调控因子，抑制MAT2A可产生显著的抗白血病作用。因此，本研究为PF-9366的临床应用铺平了道路，有望改善预后不良的MLLr白血病的治疗效果。本研究对人类脐带血CD34+（huCB CD34+）细胞进行工程改造，使其携带内源性MLL-AF4染色体易位。将改造后的细胞用15 μM的PF-9366（购自Carbosynth公司）或二甲基亚砜（DMSO）处理4天。通过Illumina NextSeq mRNA链特异性测序平台对两组生物学重复样本（处理组与未处理组）进行RNA测序。