Laboratory of Animal Genetics, Breeding and Reproduction in the Plateau Mountainous Region (Ministry of Education), College of Life Sciences, Guizhou University, Guiyang, 550025, Guizhou, China.
College of Animal Science, Guizhou University, Guiyang, 550025, Guizhou, China.
J Transl Med. 2023 Jul 6;21(1):445. doi: 10.1186/s12967-023-04288-z.
Prostate cancer (PCa) is a prevalent malignant disease affecting a significant number of males globally. Elevated expression of the Bloom's syndrome protein (BLM) helicase has emerged as a promising cancer biomarker, being associated with the onset and progression of PCa. Nevertheless, the precise molecular mechanisms governing BLM regulation in PCa remain elusive.
The expression of BLM in human specimens was analyzed using immnohistochemistry (IHC). A 5'-biotin-labeled DNA probe containing the promoter region of BLM was synthesized to pull down BLM promoter-binding proteins. Functional studies were conducted using a range of assays, including CCK-8, EdU incorporation, clone formation, wound scratch, transwell migration, alkaline comet assay, xenograft mouse model, and H&E staining. Mechanistic studies were carried out using various techniques, including streptavidin-agarose-mediated DNA pull-down, mass spectrometry (MS), immunofluorescence (IF), dual luciferase reporter assay system, RT-qPCR, ChIP-qPCR, co-immunoprecipitation (co-IP), and western blot.
The results revealed significant upregulation of BLM in human PCa tissues, and its overexpression was associated with an unfavorable prognosis in PCa patients. Increased BLM expression showed significant correlations with advanced clinical stage (P = 0.022) and Gleason grade (P = 0.006). In vitro experiments demonstrated that BLM knockdown exerted inhibitory effects on cell proliferation, clone formation, invasion, and migration. Furthermore, PARP1 (poly (ADP-ribose) polymerase 1) was identified as a BLM promoter-binding protein. Further investigations revealed that the downregulation of PARP1 led to increased BLM promoter activity and expression, while the overexpression of PARP1 exerted opposite effects. Through mechanistic studies, we elucidated that the interaction between PARP1 and HSP90AB1 (heat shock protein alpha family class B) enhanced the transcriptional regulation of BLM by counteracting the inhibitory influence of PARP1 on BLM. Furthermore, the combination treatment of olaparib with ML216 demonstrated enhanced inhibitory effects on cell proliferation, clone formation, invasion, and migration. It also induced more severe DNA damage in vitro and exhibited superior inhibitory effects on the proliferation of PC3 xenograft tumors in vivo.
The results of this study underscore the significance of BLM overexpression as a prognostic biomarker for PCa, while also demonstrating the negative regulatory impact of PARP1 on BLM transcription. The concurrent targeting of BLM and PARP1 emerges as a promising therapeutic approach for PCa treatment, holding potential clinical significance.
前列腺癌（PCa）是一种常见的恶性疾病，在全球范围内影响着大量男性。布卢姆综合征蛋白（BLM）解旋酶的高表达已成为一种有前途的癌症生物标志物，与 PCa 的发生和发展有关。然而，BLM 在 PCa 中调控的确切分子机制仍不清楚。
采用免疫组织化学（IHC）分析人标本中 BLM 的表达。合成了含有 BLM 启动子区域的 5′-生物素标记 DNA 探针，用于下拉 BLM 启动子结合蛋白。使用一系列测定法进行功能研究，包括 CCK-8、EdU 掺入、克隆形成、划痕实验、transwell 迁移、碱性彗星实验、异种移植小鼠模型和 H&E 染色。使用各种技术进行机制研究，包括链亲和素-琼脂糖介导的 DNA 下拉、质谱（MS）、免疫荧光（IF）、双荧光素酶报告基因检测系统、RT-qPCR、ChIP-qPCR、共免疫沉淀（co-IP）和 Western blot。
结果显示，BLM 在人前列腺癌组织中显著上调，其过表达与 PCa 患者的不良预后相关。BLM 表达增加与晚期临床分期（P=0.022）和 Gleason 分级（P=0.006）显著相关。体外实验表明，BLM 敲低对细胞增殖、克隆形成、侵袭和迁移具有抑制作用。此外，PARP1（多聚（ADP-核糖）聚合酶 1）被鉴定为 BLM 启动子结合蛋白。进一步的研究表明，下调 PARP1 导致 BLM 启动子活性和表达增加，而过表达 PARP1 则产生相反的效果。通过机制研究，我们阐明了 PARP1 与 HSP90AB1（热休克蛋白 alpha 家族 B 类）之间的相互作用通过抵消 PARP1 对 BLM 的抑制影响，增强了 BLM 的转录调控。此外，奥拉帕利联合 ML216 的联合治疗在体外对细胞增殖、克隆形成、侵袭和迁移具有更强的抑制作用，并在体内对 PC3 异种移植肿瘤的增殖具有更好的抑制作用。
本研究结果强调了 BLM 过表达作为 PCa 预后生物标志物的重要性，同时也表明 PARP1 对 BLM 转录具有负向调节作用。BLM 和 PARP1 的联合靶向治疗可能成为治疗 PCa 的一种有前途的治疗方法，具有潜在的临床意义。
