Hu Wentao, Cui Xiaoteng, Liu Hongyu, Li Ze, Chen Xu, Wang Qixue, Zhang Guolu, Wen Er, Lan Jinxin, Chen Junyi, Liu Jialin, Kang Chunsheng, Chen Ling
School of Medicine, Chinese PLA General Hospital, Nankai University, Beijing, China.
Department of Neurosurgery, Institute of Neurosurgery of Chinese PLA, Medical School of Chinese PLA, Chinese PLA General Hospital, Beijing, China.
J Exp Clin Cancer Res. 2025 Jan 3;44(1):3. doi: 10.1186/s13046-024-03254-x.
Glioblastoma multiforme (GBM) exhibits a cellular hierarchy with a subpopulation of stem-like cells known as glioblastoma stem cells (GSCs) that drive tumor growth and contribute to treatment resistance. NAD(H) emerges as a crucial factor influencing GSC maintenance through its involvement in diverse biological processes, including mitochondrial fitness and DNA damage repair. However, how GSCs leverage metabolic adaptation to obtain survival advantage remains elusive.
A multi-step process of machine learning algorithms was implemented to construct the glioma stemness-related score (GScore). Further in silico and patient tissue analyses validated the predictive ability of the GScore and identified a potential target, CYP3A5. Loss-of-function or gain-of-function genetic experiments were performed to assess the impact of CYP3A5 on the self-renewal and chemoresistance of GSCs both in vitro and in vivo. Mechanistic studies were conducted using nontargeted metabolomics, RNA-seq, seahorse, transmission electron microscopy, immunofluorescence, flow cytometry, ChIP‒qPCR, RT‒qPCR, western blotting, etc. The efficacy of pharmacological inhibitors of CYP3A5 was assessed in vivo.
Based on the proposed GScore, we identify a GSC target CYP3A5, which is highly expressed in GSCs and temozolomide (TMZ)-resistant GBM patients. This elevated expression of CYP3A5 is attributed to transcription factor STAT3 activated by EGFR signaling or TMZ treatment. Depletion of CYP3A5 impairs self-renewal and TMZ resistance of GSCs. Mechanistically, CYP3A5 maintains mitochondrial fitness to promote GSC metabolic adaption through the NAD⁺/NADH-SIRT1-PGC1α axis. Additionally, CYP3A5 enhances the activity of NAD-dependent enzyme PARP to augment DNA damage repair. Treatment with CYP3A5 inhibitor alone or together with TMZ effectively suppresses tumor growth in vivo.
Together, this study suggests that GSCs activate STAT3 to upregulate CYP3A5 to fine-tune NAD⁺/NADH for the enhancement of mitochondrial functions and DNA damage repair, thereby fueling tumor stemness and conferring TMZ resistance, respectively. Thus, CYP3A5 represents a promising target for GBM treatment.
多形性胶质母细胞瘤（GBM）呈现出一种细胞层级结构，其中存在一类称为胶质母细胞瘤干细胞（GSCs）的干细胞亚群，它们驱动肿瘤生长并导致治疗耐药。NAD（H）通过参与包括线粒体健康和DNA损伤修复在内的多种生物学过程，成为影响GSC维持的关键因素。然而，GSCs如何利用代谢适应来获得生存优势仍不清楚。
实施了一个多步骤的机器学习算法过程来构建胶质瘤干性相关评分（GScore）。进一步的计算机模拟和患者组织分析验证了GScore的预测能力，并确定了一个潜在靶点CYP3A5。进行了功能丧失或功能获得的基因实验，以评估CYP3A5对GSCs体外和体内自我更新及化疗耐药性的影响。使用非靶向代谢组学、RNA测序、海马实验、透射电子显微镜、免疫荧光、流式细胞术、染色质免疫沉淀-定量聚合酶链反应（ChIP-qPCR）、逆转录-定量聚合酶链反应（RT-qPCR）、蛋白质免疫印迹等进行机制研究。评估了CYP3A5药理抑制剂在体内的疗效。
基于提出的GScore，我们确定了一个GSC靶点CYP3A5，其在GSCs和对替莫唑胺（TMZ）耐药的GBM患者中高表达。CYP3A5的这种表达升高归因于由表皮生长因子受体（EGFR）信号或TMZ治疗激活的转录因子信号转导和转录激活因子3（STAT3）。CYP3A5的缺失损害了GSCs的自我更新和TMZ耐药性。机制上，CYP3A5通过NAD⁺/NADH-沉默信息调节因子1（SIRT1）-过氧化物酶体增殖物激活受体γ共激活因子1α（PGC1α）轴维持线粒体健康，以促进GSC代谢适应。此外，CYP3A5增强了NAD依赖酶聚（ADP-核糖）聚合酶（PARP）的活性，以增强DNA损伤修复。单独使用CYP3A5抑制剂或与TMZ联合治疗可有效抑制体内肿瘤生长。
总之，本研究表明GSCs激活STAT3以上调CYP3A5，从而微调NAD⁺/NADH，分别增强线粒体功能和DNA损伤修复，进而促进肿瘤干性并赋予TMZ耐药性。因此，CYP3A5是GBM治疗的一个有前景的靶点。
