CCT196969通过靶向HDAC5/RXRA/ASNS轴下调天冬酰胺合成抑制TNBC。

IF 12.8 1区 医学 Q1 ONCOLOGY
Qiong Yuan, Qi Wang, Jun Li, Liyang Yin, Shu Liu, Xuyu Zu, Yingying Shen
{"title":"CCT196969通过靶向HDAC5/RXRA/ASNS轴下调天冬酰胺合成抑制TNBC。","authors":"Qiong Yuan, Qi Wang, Jun Li, Liyang Yin, Shu Liu, Xuyu Zu, Yingying Shen","doi":"10.1186/s13046-025-03494-5","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Triple-negative breast cancer (TNBC) seriously threatens the health of patients, and new therapeutic targets and drugs need to be explored. Studies have shown that CCT196969 can inhibit melanoma and colorectal cancer. However, the role of CCT196969 in TNBC is unclear.</p><p><strong>Methods: </strong>CCT196969 inhibited TNBC via in vitro and in vivo experiments. Transcriptomic analysis, metabolomics analysis, proteomic analysis, and other experiments were used to determine that CCT196969 inhibited asparagine synthetase (ASNS) expression and downstream mTOR signaling pathway, and that Retinoid X Receptor Alpha (RXRA) was the upstream transcription factor that regulated ASNS. The binding sites of RXRA and ASNS promoter were determined by luciferase and Chromatin Immunoprecipitation (CHIP) assay. Histone Deacetylase 5 (HDAC5) was confirmed as the key target of CCT196969 by target capture assay, Cell thermal shift assay (CETSA), Surface plasmon resonance (SPR) and other experiments. qPCR, CHX tracer, MG132, immunofluorescence (IF) and Co-Immunoprecipitation (CO-IP) assay were used to detect the regulation of HDAC5 on RXRA transcription and post-translation level, and the key domains of interaction and binding between HDAC5 and RXRA. The binding sites of HDAC5 and RXRA were predicted by PyMOL software. The effect of HDAC5 on the acetylation and ubiquitination levels of RXRA was examined by CO-IP experiment. The deacetylation site of HDAC5 to RXRA was investigated by IP experiments and mass spectrometry.</p><p><strong>Results: </strong>This study reveals that CCT196969 can inhibit TNBC by down-regulating the expression of ASNS, inhibiting asparagine synthesis and downstream mTORC pathway. Mechanistically, CCT196969 targeted and inhibited HDAC5, reducing the interaction of its 1-291 region with RXRA's 1-98 region, and further resulting in an increase in RXRA acetylation (K410 and K412) and a decrease in ubiquitination levels. Together, these effects up-regulated the transcriptional and post-translational levels of RXRA. Finally, RXRA inhibited the expression of ASNS at the transcriptional level by binding to the - 1114/-1104 region on the ASNS promoter as a transcription suppressor.</p><p><strong>Conclusions: </strong>This study reveals a previously unrecognized anti-TNBC mechanism of CCT196969 through the HDAC5/RXRA/ASNS axis. This provides potential candidate targets for the treatment of TNBC and a theoretical basis for the clinical treatment of TNBC patients with CCT196969.</p>","PeriodicalId":50199,"journal":{"name":"Journal of Experimental & Clinical Cancer Research","volume":"44 1","pages":"231"},"PeriodicalIF":12.8000,"publicationDate":"2025-08-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12333201/pdf/","citationCount":"0","resultStr":"{\"title\":\"CCT196969 inhibits TNBC by targeting the HDAC5/RXRA/ASNS axis to down-regulate asparagine synthesis.\",\"authors\":\"Qiong Yuan, Qi Wang, Jun Li, Liyang Yin, Shu Liu, Xuyu Zu, Yingying Shen\",\"doi\":\"10.1186/s13046-025-03494-5\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Triple-negative breast cancer (TNBC) seriously threatens the health of patients, and new therapeutic targets and drugs need to be explored. Studies have shown that CCT196969 can inhibit melanoma and colorectal cancer. However, the role of CCT196969 in TNBC is unclear.</p><p><strong>Methods: </strong>CCT196969 inhibited TNBC via in vitro and in vivo experiments. Transcriptomic analysis, metabolomics analysis, proteomic analysis, and other experiments were used to determine that CCT196969 inhibited asparagine synthetase (ASNS) expression and downstream mTOR signaling pathway, and that Retinoid X Receptor Alpha (RXRA) was the upstream transcription factor that regulated ASNS. The binding sites of RXRA and ASNS promoter were determined by luciferase and Chromatin Immunoprecipitation (CHIP) assay. Histone Deacetylase 5 (HDAC5) was confirmed as the key target of CCT196969 by target capture assay, Cell thermal shift assay (CETSA), Surface plasmon resonance (SPR) and other experiments. qPCR, CHX tracer, MG132, immunofluorescence (IF) and Co-Immunoprecipitation (CO-IP) assay were used to detect the regulation of HDAC5 on RXRA transcription and post-translation level, and the key domains of interaction and binding between HDAC5 and RXRA. The binding sites of HDAC5 and RXRA were predicted by PyMOL software. The effect of HDAC5 on the acetylation and ubiquitination levels of RXRA was examined by CO-IP experiment. The deacetylation site of HDAC5 to RXRA was investigated by IP experiments and mass spectrometry.</p><p><strong>Results: </strong>This study reveals that CCT196969 can inhibit TNBC by down-regulating the expression of ASNS, inhibiting asparagine synthesis and downstream mTORC pathway. Mechanistically, CCT196969 targeted and inhibited HDAC5, reducing the interaction of its 1-291 region with RXRA's 1-98 region, and further resulting in an increase in RXRA acetylation (K410 and K412) and a decrease in ubiquitination levels. Together, these effects up-regulated the transcriptional and post-translational levels of RXRA. Finally, RXRA inhibited the expression of ASNS at the transcriptional level by binding to the - 1114/-1104 region on the ASNS promoter as a transcription suppressor.</p><p><strong>Conclusions: </strong>This study reveals a previously unrecognized anti-TNBC mechanism of CCT196969 through the HDAC5/RXRA/ASNS axis. This provides potential candidate targets for the treatment of TNBC and a theoretical basis for the clinical treatment of TNBC patients with CCT196969.</p>\",\"PeriodicalId\":50199,\"journal\":{\"name\":\"Journal of Experimental & Clinical Cancer Research\",\"volume\":\"44 1\",\"pages\":\"231\"},\"PeriodicalIF\":12.8000,\"publicationDate\":\"2025-08-08\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12333201/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of Experimental & Clinical Cancer Research\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1186/s13046-025-03494-5\",\"RegionNum\":1,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"ONCOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Experimental & Clinical Cancer Research","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1186/s13046-025-03494-5","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"ONCOLOGY","Score":null,"Total":0}
引用次数: 0

摘要

背景:三阴性乳腺癌(triple negative breast cancer, TNBC)严重威胁着患者的健康,需要探索新的治疗靶点和药物。研究表明,CCT196969具有抑制黑色素瘤和结直肠癌的作用。然而,CCT196969在TNBC中的作用尚不清楚。方法:CCT196969通过体外和体内实验抑制TNBC。通过转录组学分析、代谢组学分析、蛋白质组学分析等实验,确定CCT196969抑制天冬酰胺合成酶(ASNS)表达和下游mTOR信号通路,而类视黄嘌呤X受体α (RXRA)是调控ASNS的上游转录因子。采用荧光素酶和染色质免疫沉淀(CHIP)法测定RXRA和ASNS启动子的结合位点。通过靶捕获实验、细胞热移实验(CETSA)、表面等离子体共振(SPR)等实验证实组蛋白去乙酰化酶5 (HDAC5)是CCT196969的关键靶点。采用qPCR、CHX示踪剂、MG132、免疫荧光(IF)和Co-Immunoprecipitation (CO-IP)检测HDAC5对RXRA转录和翻译后水平的调控,以及HDAC5与RXRA相互作用和结合的关键结构域。利用PyMOL软件预测HDAC5和RXRA的结合位点。通过CO-IP实验检测HDAC5对RXRA乙酰化和泛素化水平的影响。利用IP实验和质谱法研究了HDAC5对RXRA的去乙酰化位点。结果:本研究发现CCT196969通过下调ASNS表达、抑制天冬酰胺合成及下游mTORC通路抑制TNBC。机制上,CCT196969靶向并抑制HDAC5,减少其1-291区域与RXRA 1-98区域的相互作用,进一步导致RXRA乙酰化(K410和K412)的增加和泛素化水平的降低。总之,这些作用上调了RXRA的转录和翻译后水平。最后,RXRA通过结合ASNS启动子上的- 1114/-1104区域作为转录抑制因子,在转录水平上抑制ASNS的表达。结论:本研究揭示了CCT196969通过HDAC5/RXRA/ASNS轴抗tnbc的机制。这为治疗TNBC提供了潜在的候选靶点,为临床治疗CCT196969 TNBC患者提供了理论基础。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
CCT196969 inhibits TNBC by targeting the HDAC5/RXRA/ASNS axis to down-regulate asparagine synthesis.

Background: Triple-negative breast cancer (TNBC) seriously threatens the health of patients, and new therapeutic targets and drugs need to be explored. Studies have shown that CCT196969 can inhibit melanoma and colorectal cancer. However, the role of CCT196969 in TNBC is unclear.

Methods: CCT196969 inhibited TNBC via in vitro and in vivo experiments. Transcriptomic analysis, metabolomics analysis, proteomic analysis, and other experiments were used to determine that CCT196969 inhibited asparagine synthetase (ASNS) expression and downstream mTOR signaling pathway, and that Retinoid X Receptor Alpha (RXRA) was the upstream transcription factor that regulated ASNS. The binding sites of RXRA and ASNS promoter were determined by luciferase and Chromatin Immunoprecipitation (CHIP) assay. Histone Deacetylase 5 (HDAC5) was confirmed as the key target of CCT196969 by target capture assay, Cell thermal shift assay (CETSA), Surface plasmon resonance (SPR) and other experiments. qPCR, CHX tracer, MG132, immunofluorescence (IF) and Co-Immunoprecipitation (CO-IP) assay were used to detect the regulation of HDAC5 on RXRA transcription and post-translation level, and the key domains of interaction and binding between HDAC5 and RXRA. The binding sites of HDAC5 and RXRA were predicted by PyMOL software. The effect of HDAC5 on the acetylation and ubiquitination levels of RXRA was examined by CO-IP experiment. The deacetylation site of HDAC5 to RXRA was investigated by IP experiments and mass spectrometry.

Results: This study reveals that CCT196969 can inhibit TNBC by down-regulating the expression of ASNS, inhibiting asparagine synthesis and downstream mTORC pathway. Mechanistically, CCT196969 targeted and inhibited HDAC5, reducing the interaction of its 1-291 region with RXRA's 1-98 region, and further resulting in an increase in RXRA acetylation (K410 and K412) and a decrease in ubiquitination levels. Together, these effects up-regulated the transcriptional and post-translational levels of RXRA. Finally, RXRA inhibited the expression of ASNS at the transcriptional level by binding to the - 1114/-1104 region on the ASNS promoter as a transcription suppressor.

Conclusions: This study reveals a previously unrecognized anti-TNBC mechanism of CCT196969 through the HDAC5/RXRA/ASNS axis. This provides potential candidate targets for the treatment of TNBC and a theoretical basis for the clinical treatment of TNBC patients with CCT196969.

求助全文
通过发布文献求助,成功后即可免费获取论文全文。 去求助
来源期刊
CiteScore
18.20
自引率
1.80%
发文量
333
审稿时长
1 months
期刊介绍: The Journal of Experimental & Clinical Cancer Research is an esteemed peer-reviewed publication that focuses on cancer research, encompassing everything from fundamental discoveries to practical applications. We welcome submissions that showcase groundbreaking advancements in the field of cancer research, especially those that bridge the gap between laboratory findings and clinical implementation. Our goal is to foster a deeper understanding of cancer, improve prevention and detection strategies, facilitate accurate diagnosis, and enhance treatment options. We are particularly interested in manuscripts that shed light on the mechanisms behind the development and progression of cancer, including metastasis. Additionally, we encourage submissions that explore molecular alterations or biomarkers that can help predict the efficacy of different treatments or identify drug resistance. Translational research related to targeted therapies, personalized medicine, tumor immunotherapy, and innovative approaches applicable to clinical investigations are also of great interest to us. We provide a platform for the dissemination of large-scale molecular characterizations of human tumors and encourage researchers to share their insights, discoveries, and methodologies with the wider scientific community. By publishing high-quality research articles, reviews, and commentaries, the Journal of Experimental & Clinical Cancer Research strives to contribute to the continuous improvement of cancer care and make a meaningful impact on patients' lives.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:604180095
Book学术官方微信