PO.CH01.05 · 化学
色霉素A2通过整合的自噬和凋亡程序破坏白血病细胞活力
Chromomycin A2 disrupts leukemia cell viability via integrated autophagic and apoptotic programs
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
急性白血病由于复发率高且经常对标准治疗耐药，迫切需要新的治疗方法。分子分型的进展使得更具选择性、毒性更低的策略成为可能，尤其是对预后不良的患者。色霉素类作为潜在药物已崭露头角，因为它们结合富含GC的DNA区域、改变染色质结构、抑制RNA聚合酶并抑制白血病细胞存活所必需的转录程序。其中，色霉素A2尽管在某些实体瘤中有活性报道，但在血液系统肿瘤中仍鲜有探索。我们在一组广泛的白血病模型中研究了色霉素A2。分析了22株人类血液肿瘤细胞系（14株髓系，17株淋巴系），包括对venetoclax（n=2）、quizartinib（n=1）和ATRA（n=1）耐药的变异株。评估了细胞活力（MTT）、凋亡（annexin V/PI）以及在无细胞因子甲基纤维素中的自主克隆生长。离体实验采用原代AML（n=3）和ALL（n=3）样本。分子分析包括涵盖自噬、凋亡、DNA损伤和细胞周期调控相关基因的PCR阵列，以及Western印迹。统计学显著性定义为p <0.05。色霉素A2以剂量依赖方式降低所有模型的活力（IC50：0.40 - 4.42 nM）。耐药衍生株显示出略高的IC50值，但仍保持敏感性。该化合物诱导时间和剂量依赖性凋亡，并在≥5 nM时完全抑制自主克隆生长。早期反应涉及自噬标志物（LC3B-II积累、SQSTM1/p62减少），与转录输出减少和存活信号减弱一致。随暴露时间延长，凋亡标志物（切割的PARP1）和DNA损伤（gammaH2AX）占主导。基因表达数据表明存在时间序列：6小时时激活自噬和细胞周期阻滞，随后12小时时激活凋亡相关基因。富集分析突出了巨自噬、线粒体凋亡和G1 DNA损伤检查点通路。在离体实验中，IC50值范围为1.6至31.8 nM，在原代AML和ALL细胞中疗效为81-92%。色霉素A2在体外和离体模型（包括耐药亚型）中均显示出强烈的抗白血病活性。其对自噬、凋亡和DNA损伤的序贯诱导提示存在多靶点机制。对克隆生长的抑制表明其对白血病干/祖细胞具有潜在活性。这些结果支持进一步的临床前开发。由FAPESP、CAPES和CNPq资助。
查看英文原文 English abstract
Acute leukemias urgently require new therapeutic approaches due to high relapse rates and frequent resistance to standard treatments. Advances in molecular profiling have enabled more selective and less toxic strategies, especially for patients with poor prognosis. Chromomycins have emerged as potential agents because they bind GC-rich DNA regions, alter chromatin architecture, inhibit RNA polymerase, and suppress transcriptional programs essential for leukemic cell survival. Among them, chromomycin A2 remains largely unexplored in hematologic cancers despite reported activity in some solid tumors. We investigated chromomycin A2 in a broad panel of leukemia models. Twenty-two human blood cancer cell lines were analyzed (14 myeloid, 17 lymphoid), including resistant variants to venetoclax (n=2), quizartinib (n=1), and ATRA (n=1). Viability (MTT), apoptosis (annexin V/PI), and autonomous clonal growth in cytokine-free methylcellulose were evaluated. Ex vivo assays used primary AML (n=3) and ALL (n=3) samples. Molecular analyses included PCR arrays covering genes related to autophagy, apoptosis, DNA damage, and cell-cycle control, together with Western blotting. Statistical significance was defined as p <0.05. Chromomycin A2 reduced viability in all models in a dose-dependent manner (IC50: 0.40 - 4.42 nM). Resistant derivatives showed slightly higher IC 50 values but remained sensitive. The compound induced time- and dose-dependent apoptosis and fully suppressed autonomous clonal growth at ≥5 nM. Early responses involved autophagy markers (LC3B-II accumulation, SQSTM1/p62 reduction), compatible with decreased transcriptional output and weakened survival signaling. With longer exposure, apoptotic markers (cleaved PARP1) and DNA damage (gammaH2AX) predominated. Gene expression data indicated a temporal sequence: activation of autophagy and cell-cycle arrest at 6 h, followed by apoptosis-related genes at 12 h. Enrichment analyses highlighted macroautophagy, mitochondrial apoptosis, and G1 DNA damage checkpoint pathways. In ex vivo assays, IC 50 values ranged from 1.6 to 31.8 nM, with 81-92% efficacy in primary AML and ALL cells. Chromomycin A2 displayed strong antileukemic activity across in vitro and ex vivo models, including resistant subtypes. Its sequential induction of autophagy, apoptosis, and DNA damage suggests a multitarget mechanism. The inhibition of clonal growth indicates potential activity against leukemic stem/progenitor cells. These results support further preclinical development. Supported by FAPESP, CAPES, and CNPq.
利益披露 Disclosure
K. Lima, None..
E. O. Vieira, None..
R. C. Cavaglieri, None..
F. L. Nogueira, None..
G. M. Machado-Santelli, None..
L. V. Costa-Lotufo, None..
E. M. Rego, None..
J. A. Machado-Neto, None.