Sheila and David Fuente Graduate Program in Cancer Biology, Sylvester Comprehensive Cancer Center, University of Miami, Miller School of Medicine, Miami, FL, USA.
Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, 3970 Reservoir Rd NW, Washington, DC, NRB W412, USA.
Breast Cancer Res Treat. 2021 Jan;185(2):331-341. doi: 10.1007/s10549-020-05962-2. Epub 2020 Oct 31.
The study aimed to investigate the role of spindle assembly checkpoint (SAC) in cancer cells with compromised genomic integrity. Chromosomal instability (CIN) gives cancer cells an adaptive advantage. However, maintaining the balance of this instability is crucial for the survival of cancer cells as it could lead them to the mitotic catastrophe. Therefore, cancer cells adapt to the detrimental effects of CIN. We hypothesized that changes in SAC might be one such adaptation mechanism. The focus of the study was BUB1B, an integral part of the checkpoint.
Clinical datasets were analyzed to compare expression levels of SAC genes in normal tissue vs. breast carcinoma. The effects of the reduction of BUB1B expression was examined utilizing RNA interference method with siRNAs. In vitro viability, clonogenicity, apoptosis, and SAC activity levels of a variety of breast cancer (BrCa) cell lines, as well as in vivo tumorigenicity of the triple-negative breast cancer (TNBC) cell line MDA-MB-468, were tested. Additionally, the chromosomal stability of these cells was tested by immunofluorescence staining and flow cytometry.
In clinical breast cancer datasets, SAC genes were elevated in BrCa with BUB1B having the highest fold change. BUB1B overexpression was associated with a decreased probability of overall survival. The knockdown of BUB1B resulted in reduced viability and clonogenicity in BrCa cell lines and a significant increase in apoptosis and cell death. However, the viability and apoptosis levels of the normal breast epithelial cell line, MCF12A, were not affected. BUB1B knockdown also impaired chromosome alignment and resulted in acute chromosomal abnormalities. We also showed that BUB1B knockdown on the MDA-MB-468 cell line decreases tumor growth in mice.
A functional spindle assembly checkpoint is essential for the survival of BrCa cells. BUB1B is a critical factor in SAC, and therefore breast cancer cell survival. Impairment of BUB1B has damaging effects on cancer cell viability and tumorigenicity, especially on the more aggressive variants of BrCa.
本研究旨在探讨纺锤体组装检查点（SAC）在基因组完整性受损的癌细胞中的作用。染色体不稳定性（CIN）赋予癌细胞适应优势。然而，维持这种不稳定性的平衡对于癌细胞的存活至关重要，因为它可能导致它们发生有丝分裂灾难。因此，癌细胞会适应 CIN 的有害影响。我们假设 SAC 的变化可能是一种适应机制。本研究的重点是 BUB1B，它是检查点的一个组成部分。
分析临床数据集，比较正常组织与乳腺癌中 SAC 基因的表达水平。利用 siRNA 的 RNA 干扰方法，检测 BUB1B 表达减少的影响。检测多种乳腺癌（BrCa）细胞系的体外存活率、集落形成能力、细胞凋亡和 SAC 活性水平，以及三阴性乳腺癌（TNBC）细胞系 MDA-MB-468 的体内致瘤性。此外，通过免疫荧光染色和流式细胞术检测这些细胞的染色体稳定性。
在临床乳腺癌数据集，SAC 基因在 BrCa 中升高，其中 BUB1B 的倍数变化最高。BUB1B 过表达与总生存率降低相关。BUB1B 敲低导致 BrCa 细胞系的存活率和集落形成能力降低，细胞凋亡和细胞死亡显著增加。然而，正常乳腺上皮细胞系 MCF12A 的存活率和细胞凋亡水平不受影响。BUB1B 敲低也会损害染色体排列，导致急性染色体异常。我们还表明，MDA-MB-468 细胞系的 BUB1B 敲低会降低小鼠肿瘤的生长。
功能性纺锤体组装检查点对于 BrCa 细胞的存活至关重要。BUB1B 是 SAC 的关键因素，因此也是乳腺癌细胞存活的关键因素。BUB1B 受损对癌细胞的存活率和致瘤性有破坏性影响，特别是对更具侵袭性的 BrCa 变体。
