DataSheet_1_UBE2S and UBE2C confer a poor prognosis to breast cancer via downregulation of Numb.docx

PurposeUbiquitin-conjugating enzymes E2S (UBE2S) and E2C (UBE2C), which mediate the biological process of ubiquitination, have been widely reported in various cancers. Numb, the cell fate determinant and tumor suppressor, was also involved in ubiquitination and proteasomal degradation. However, the interaction between UBE2S/UBE2C and Numb and their roles in the clinical outcome of breast cancer (BC) are not widely elucidated.
MethodsOncomine, Cancer Cell Line Encyclopedia (CCLE), the Human Protein Atlas (HPA) database, qRT-PCR, and Western blot analyses were utilized to analyze UBE2S/UBE2C and Numb expression in various cancer types and their respective normal controls, breast cancer tissues, and breast cancer cell lines. The expression of UBE2S, UBE2C, and Numb in BC patients with different ER, PR, and HER2 status, grades, stages, and survival status was compared. By Kaplan–Meier plotter, we further evaluated the prognostic value of UBE2S, UBE2C, and Numb in BC patients. We also explored the potential regulatory mechanisms underlying UBE2S/UBE2C and Numb through overexpression and knockdown experiments in BC cell lines and performed growth and colony formation assays to assess cell malignancy.
ResultsIn this study, we showed that UBE2S and UBE2C were overexpressed while Numb was downregulated in BC, and in BC of higher grade, stage, and poor survival. Compared to hormone receptor negative (HR−) BC cell lines or tissues, HR+ BC demonstrated lower UBE2S/UBE2C and higher Numb, corresponding to better survival. We also showed that increased UBE2S/UBE2C and reduced Numb predicted poor prognosis in BC patients, as well as in ER+ BC patients. In BC cell lines, UBE2S/UBE2C overexpression decreased the level of Numb and enhanced cell malignancy, while knocking down UBE2S/UBE2C demonstrated the opposite effects.
ConclusionUBE2S and UBE2C downregulated Numb and enhanced BC malignancy. The combination of UBE2S/UBE2C and Numb could potentially serve as novel biomarkers for BC.

Purpose：泛素结合酶E2S（UBE2S）与泛素结合酶E2C（UBE2C）可介导泛素化生物学过程，二者已在多种癌症中被广泛报道。细胞命运决定因子及抑癌蛋白Numb同样参与泛素化与蛋白酶体降解过程，但目前UBE2S/UBE2C与Numb的相互作用，以及二者在乳腺癌（breast cancer, BC）临床结局中的作用尚未得到充分阐明。
Methods：本研究采用Oncomine数据库、癌细胞系百科全书（Cancer Cell Line Encyclopedia, CCLE）、人类蛋白质图谱（Human Protein Atlas, HPA）数据库、实时定量聚合酶链反应（quantitative real-time polymerase chain reaction, qRT-PCR）及蛋白质印迹（Western blot）分析，对多种癌症类型及其对应正常对照、乳腺癌组织与乳腺癌细胞系中UBE2S/UBE2C和Numb的表达水平进行检测。本研究比较了不同雌激素受体（ER）、孕激素受体（PR）、人表皮生长因子受体2（HER2）状态、不同分级、分期及生存状态的乳腺癌患者中UBE2S、UBE2C及Numb的表达差异。通过Kaplan–Meier绘图仪（Kaplan–Meier plotter），本研究进一步评估了UBE2S、UBE2C及Numb在乳腺癌患者中的预后价值。此外，本研究通过在乳腺癌细胞系中开展过表达与敲低实验，探讨了UBE2S/UBE2C与Numb的潜在调控机制，并通过生长实验与集落形成实验评估细胞恶性程度。
Results：本研究结果显示，在乳腺癌组织中，UBE2S与UBE2C呈高表达，而Numb呈低表达；且在高级别、高分期及预后不良的乳腺癌患者中，该表达模式更为显著。与激素受体阴性（HR⁻）乳腺癌细胞系或组织相比，激素受体阳性（HR⁺）乳腺癌中UBE2S/UBE2C的表达水平更低，Numb的表达水平更高，对应更优的生存结局。此外，UBE2S/UBE2C表达升高与Numb表达降低可预测乳腺癌患者及雌激素受体阳性（ER⁺）乳腺癌患者的不良预后。在乳腺癌细胞系中，过表达UBE2S/UBE2C可降低Numb的蛋白水平并增强细胞恶性程度，而敲低UBE2S/UBE2C则可产生相反的效应。
Conclusion：UBE2S与UBE2C可通过下调Numb的表达增强乳腺癌的恶性程度。UBE2S/UBE2C与Numb的联合检测有望成为乳腺癌新型的生物标志物。