目的:疾病复发和治疗耐药仍然是癌症治疗的严重障碍。白血病干细胞(LSC)具有治疗抗性和复发的原因。深度静止状态似乎使癌症干细胞(CSC)能够获得对疾病进展和治疗抵抗至关重要的新体细胞突变。正常造血干细胞(HSC)和LSC具有许多共同特征,从而使LSC的安全消除复杂化。最近的一项研究表明,长寿的正常卵母细胞存在没有线粒体复合物I(MC-1),以发育调节的方式表达,从而减轻他们对ROS的脆弱性。静态CSC依靠线粒体FAO,没有复杂的I表达式,从而避免了破坏性ROS的产生,类似于长寿的正常人干细胞。对治疗抗性生物学的更深入了解对于开发最佳策略以实现完全治愈白血病很重要。
方法:这里,使用scRNA测序和ATAC-seq对原发性慢性粒细胞白血病(CML)患者样本,结合生物信息学分析,我们进一步检查了先前表征的体外伊马替尼选择的CD34-CD38-CMLLSC群体的异质性.我们利用了一系列的功能分析,包括单细胞代谢组学和海马分析,以验证最深的静态白血病引发剂(LI)子集的存在。
结果:目前的研究揭示了CML患者治疗耐药LSC的异质性及其存在两种功能上不同的状态。最深层静止的LI抑制MC-1的表达,但高度依赖于脂肪酸氧化(FAO)的代谢需求,并且ATAC-seq显示该人群中染色质可及性增加。所有这些都符合极其原始的,静止的干性转录签名。重要的是,特异性CREB结合蛋白(CBP)/β-catenin拮抗剂ICG-001启动LSC的分化,包括LI,随着分化和MC-1,CD34,CD38和BCR-ABL1表达增加,从而降低染色质可及性,从而使它们对伊马替尼重新敏感。
结论:我们研究了CML患者与LSC异质性相关的生物学方面,并证明了特定小分子CBP/β-catenin拮抗剂安全消除深度静止治疗耐药CSC的能力。这些观察结果可能代表了一种有吸引力的可推广的治疗策略,可以帮助开发更好的方案来根除静止的LSC群体。
OBJECTIVE: Disease relapse and therapy resistance remain serious impediments to treating cancer. Leukemia stem cells (
LSC) are therapy resistant and the cause of relapse. A state of deep quiescence appears to enable cancer stem cells (CSC) to acquire new somatic mutations essential for disease progression and therapy resistance. Both normal hematopoietic stem cells (HSC) and
LSC share many common features, thereby complicating the safe elimination of
LSC. A recent study demonstrated that long lived normal oocytes exist without mitochondrial complex I (MC-1), expressing it in a developmentally regulated fashion, thereby mitigating their vulnerability to ROS. Quiescent CSC rely on mitochondrial FAO, without complex I expression, thereby avoiding the generation of damaging ROS, similar to long lived normal human stem cells. A deeper understanding of the biology of therapy resistance is important for the development of optimal strategies to attain complete leukemia cures.
METHODS: Here, using scRNA-sequencing and ATAC-seq on primary chronic myelogenous leukemia (CML) patient samples, combined with bioinformatics analyses, we further examine the heterogeneity of a previously characterized in vitro imatinib-selected CD34-CD38- CML LSC population. We utilized a series of functional analyses, including single-cell metabolomic and Seahorse analyses, to validate the existence of the deepest quiescent leukemia initiators (LI) subset.
RESULTS: Current study revealed heterogeneity of therapy resistant
LSC in CML patients and their existence of two functionally distinct states. The most deeply quiescent LI suppress the expression of MC-1, yet are highly dependent on fatty acid oxidation (FAO) for their metabolic requirements and ATAC-seq demonstrated increased chromatin accessibility in this population, all consistent with an extremely primitive, quiescent stemness transcriptional signature. Importantly, the specific CREB binding protein (CBP)/β-catenin antagonist ICG-001 initiates the differentiation of
LSC, including LI, decreases chromatin accessibility with differentiation and increasing expression of MC-1, CD34, CD38 and BCR-ABL1, thereby resensitizing them to imatinib.
CONCLUSIONS: We investigated the biological aspects related to LSC heterogeneity in CML patients and demonstrated the ability of specific small molecule CBP/β-catenin antagonists to safely eliminate deeply quiescent therapy resistant CSC. These observations may represent an attractive generalizable therapeutic strategy that could help develop better protocols to eradicate the quiescent LSC population.