ACSL4通过FBW7调控c-Myc稳定性在促进肝癌增殖中的作用及机制研究

Metabolic reprogramming and cell cycle regulation are two hallmarks of cancer. Their interactions and the resulting promotion on cancer cell proliferation and carcinogenesis are not well understood yet. Our previous work has identified through hepatocellular carcinoma (HCC) transcriptome profiling that long chain acyl coenzyme A synthase 4 (ACSL4), an upregulated lipid metabolic reprogramming enzyme, was highly expressed in HCC and closely related to poor prognosis. ACSL4 activated c-Myc to promote G1/S phase transition in cancer cells, but the molecular mechanism remains unclear. This study aims to examine the expression patterns of ACSL4/c-Myc in HCC cells and analyze the correlation of the abnormal expressions with clinico-pathological characteristics and recurrence of HCC, as well as the prognosis of the patients. We also plan to determine the effects of ACSL4 expression on HCC cell proliferation and demonstrate the molecular mechanisms of ACSL4 regulation on c-Myc stability via FBW7, using methods such as bioinformatics, co-immunoprecipitatation, ubiquitylation, and truncation mutant assay. In addition, in vivo models such as nude mice model, tail vein injection model, and PDX model are utilized to investigate into the ability of tumorigenesis and metastasis of HCC and the effects of rosiglitazone, an ACSL4 inhibitor, on targeted therapies for HCC. This study may discover a novel mechanism of interactions between lipid metabolism reprogramming and cycle regulation on promoting cancer cell proliferation, and provide new approaches for clinical management of HCC.

代谢重编程和细胞周期调控异常是肿瘤的两个重要特征，两者之间如何交互作用以促进肿瘤发生发展目前尚不清楚。我们前期研究发现，脂代谢重编程酶—长链脂酰辅酶Ａ合成酶4(ACSL4)在肝癌中显著高表达，并与不良预后密切相关。抑制ACSL4表达能下调cMyc水平并诱导肝癌细胞发生G1/S期阻滞，然而作用机制尚不清楚。本研究拟分析肝癌组织中ACSL4/cMyc的表达水平与临床病理特征和预后的相关性；明确ACSL4的表达变化对肝癌细胞增殖能力的影响，应用生物信息学、免疫共沉淀、泛素化及截短等技术阐明ACSL4通过FBW7调控cMyc稳定性促肝癌增殖的分子机制；采用裸鼠肝癌模型、尾静脉注射模型、PDX模型从体内研究ACSL4对肝癌细胞成瘤及转移能力的影响，证明ACSL4抑制剂罗格列酮对肝癌的靶向治疗作用。本研究将揭示脂代谢重编程与周期调控交互作用促进肝癌细胞快速增殖的新机制，为临床肝癌的转化治疗提供新思路。

项目的背景：肝细胞癌（HCC）是高度异质性，多基因驱动的恶性肿瘤。长链脂酰辅酶A合成酶4（ACSL4）是酰基辅酶A合成酶（ACS）家族的成员之一，在花生四烯酸（AA）代谢中起关键作用，已被证明与多种癌症的恶性表型相关。然而，在肝癌中其功能及其潜在的分子机制仍未完全阐明。.主要研究内容：采用转录组微阵列分析以鉴定AFP高表达肝细胞癌的新型分子标志物。通过实时荧光定量逆转录聚合酶链反应（qRT-PCR）、蛋白质免疫印迹（WB）和免疫组化（IHC）检测肝癌和癌旁组织中ACSL4的表达。应用功能学实验，CHX和MG132干预实验，泛素化实验，拯救实验和裸鼠异种移植模型来确定ACSL4在体内外促进肿瘤发展的潜在机制。通过IHC染色证明肝癌中ACSL4和c-Myc的相关性。.重要结果、关键数据：通过转录组分析将ACSL4鉴定为AFP高表达HCC亚型的新型分子标志物。ACSL4在HCC样本中表达显著上调，并与不良预后相关。功能上，ACSL4敲低导致细胞生长速率减慢，而外源性ACSL4过表达则促进了体内外的肿瘤生长。机制上，ACSL4通过泛素-蛋白酶体系统以ERK/FBW7依赖性的方式稳定原癌蛋白c-Myc。ACSL4升高介导的促进细胞生长能力可被c-MycsiRNA或其抑制剂10058-F4导致的c-Myc耗竭所抑制。相反，通过FBW7敲低导致的c-Myc上调部分逆转了ACSL4沉默带来的细胞表型的影响。临床肝癌样本中，ACSL4的表达与c-Myc呈正相关。.结论及科学意义：ACSL4是AFP高表达HCC亚型的新型分子标志物。我们的研究揭示了ACSL4促进HCC肿瘤进展的新机制：以依赖ERK/FBW7通路的方式在转录后水平稳定c-Myc表达。我们的发现表明ACSL4可能是HCC中有价值的预后生物标志物和潜在的治疗靶标。

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