囊胺酸双氧酶1（CDO1）在许多人类癌症中，包括乳腺癌（BC），经常发生甲基化并且其表达量降低。然而，CDO1在BC中失活的功能和机制方面还知之甚少，而且血清CDO1甲基化的诊断意义尚不清楚。我们对公开可获得的数据库进行了生物信息学分析，采用MassARRAY EpiTYPER甲基化测序技术在BC组织与正常相邻组织（NATs）中鉴定了CDO1启动子中甲基化差异较大的位点。随后，我们设计了针对这些CpG位点的特异引物和探针，利用MethyLight检测方法测定CDO1启动子的甲基化参考百分比（PMR）。此外，我们还利用LentiCRISPR/dCas9-Tet1CD基因编辑系统和CDO1过表达策略来研究CDO1在BC中的功能和机制。最后，我们评估了CDO1作为BC血清甲基化生物标志物的早期诊断价值。BC组织中CDO1启动子呈高甲基化状态，与恶性预后相关（p < .05）。基于CRISPR/dCas9的靶向去甲基化系统显著降低了CDO1启动子的PMR并增加了BC细胞中CDO1的表达。结果表明，这导致了细胞增殖、迁移和侵袭的抑制。此外，我们发现CDO1通过抑制细胞周期、促进细胞凋亡和铁死亡发挥了抑制肿瘤的作用。此外，我们采用MethyLight检测BC血清中的CDO1 PMR，并发现血清CDO1甲基化是BC早期诊断的一种有效无创生物标志物。CDO1在BC中呈高甲基化状态，并具有抑制肿瘤的作用。调控异常CDO1甲基化的表观遗传编辑可能在BC的临床治疗和预后中发挥关键作用。此外，血清CDO1甲基化有望成为BC早期诊断和管理的有价值的生物标志物。(c) 2023 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.

Cysteine dioxygenase 1 (CDO1) is frequently methylated, and its expression is decreased in many human cancers including breast cancer (BC). However, the functional and mechanistic aspects of CDO1 inactivation in BC are poorly understood, and the diagnostic significance of serum CDO1 methylation remains unclear.We performed bioinformatics analysis of publicly available databases and employed MassARRAY EpiTYPER methylation sequencing technology to identify differentially methylated sites in the CDO1 promoter of BC tissues compared to normal adjacent tissues (NATs). Subsequently, we developed a MethyLight assay using specific primers and probes for these CpG sites to detect the percentage of methylated reference (PMR) of the CDO1 promoter. Furthermore, both LentiCRISPR/dCas9-Tet1CD-based CDO1-targeted demethylation system and CDO1 overexpression strategy were utilized to detect the function and underlying mechanism of CDO1 in BC. Finally, the early diagnostic value of CDO1 as a methylation biomarker in BC serum was evaluated.CDO1 promoter was hypermethylated in BC tissues, which was related to poor prognosis (p < .05). The CRISPR/dCas9-based targeted demethylation system significantly reduced the PMR of CDO1 promotor and increased CDO1 expression in BC cells. Consequently, this leads to suppression of cell proliferation, migration and invasion. Additionally, we found that CDO1 exerted a tumour suppressor effect by inhibiting the cell cycle, promoting cell apoptosis and ferroptosis. Furthermore, we employed the MethyLight to detect CDO1 PMR in BC serum, and we discovered that serum CDO1 methylation was an effective non-invasive biomarker for early diagnosis of BC.CDO1 is hypermethylated and acts as a tumour suppressor gene in BC. Epigenetic editing of abnormal CDO1 methylation could have a crucial role in the clinical treatment and prognosis of BC. Additionally, serum CDO1 methylation holds promise as a valuable biomarker for the early diagnosis and management of BC.© 2023 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.