乳腺癌细胞的长期药物选择和癌症干细胞特征的富集。
Prolonged drug selection of breast cancer cells and enrichment of cancer stem cell characteristics.
机构信息
Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
出版信息
J Natl Cancer Inst. 2010 Nov 3;102(21):1637-52. doi: 10.1093/jnci/djq361. Epub 2010 Oct 8.
BACKGROUND
Cancer stem cells are presumed to have virtually unlimited proliferative and self-renewal abilities and to be highly resistant to chemotherapy, a feature that is associated with overexpression of ATP-binding cassette transporters. We investigated whether prolonged continuous selection of cells for drug resistance enriches cultures for cancer stem-like cells.
METHODS
Cancer stem cells were defined as CD44+/CD24⁻ cells that could self-renew (ie, generate cells with the tumorigenic CD44+/CD24⁻ phenotype), differentiate, invade, and form tumors in vivo. We used doxorubicin-selected MCF-7/ADR cells, weakly tumorigenic parental MCF-7 cells, and MCF-7/MDR, an MCF-7 subline with forced expression of ABCB1 protein. Cells were examined for cell surface markers and side-population fractions by microarray and flow cytometry, with in vitro invasion assays, and for ability to form mammospheres. Xenograft tumors were generated in mice to examine tumorigenicity (n = 52). The mRNA expression of multidrug resistance genes was examined in putative cancer stem cells and pathway analysis of statistically significantly differentially expressed genes was performed. All statistical tests were two-sided.
RESULTS
Pathway analysis showed that MCF-7/ADR cells express mRNAs from ABCB1 and other genes also found in breast cancer stem cells (eg, CD44, TGFB1, and SNAI1). MCF-7/ADR cells were highly invasive, formed mammospheres, and were tumorigenic in mice. In contrast to parental MCF-7 cells, more than 30% of MCF-7/ADR cells had a CD44+/CD24⁻ phenotype, could self-renew, and differentiate (ie, produce CD44+/CD24⁻ and CD44+/CD24+ cells) and overexpressed various multidrug resistance-linked genes (including ABCB1, CCNE1, and MMP9). MCF-7/ADR cells were statistically significantly more invasive in Matrigel than parental MCF-7 cells (MCF-7 cells = 0.82 cell per field and MCF-7/ADR = 7.51 cells per field, difference = 6.69 cells per field, 95% confidence interval = 4.82 to 8.55 cells per field, P < .001). No enrichment in the CD44+/CD24⁻ or CD133+ population was detected in MCF-7/MDR.
CONCLUSION
The cell population with cancer stem cell characteristics increased after prolonged continuous selection for doxorubicin resistance.
背景
癌症干细胞被认为具有几乎无限的增殖和自我更新能力，并且对化疗高度耐药，这一特征与三磷酸腺苷结合盒转运蛋白的过度表达有关。我们研究了长期连续选择耐药细胞是否会富集具有癌症干细胞样特性的细胞。
方法
癌症干细胞被定义为能够自我更新（即产生具有肿瘤形成性 CD44+/CD24⁻表型的细胞）、分化、侵袭和体内形成肿瘤的 CD44+/CD24⁻细胞。我们使用多柔比星选择的 MCF-7/ADR 细胞、弱致瘤性亲本 MCF-7 细胞和 MCF-7/MDR（一种强制表达 ABCB1 蛋白的 MCF-7 亚系）进行研究。通过微阵列和流式细胞术检查细胞表面标志物和侧群分数，进行体外侵袭试验，并检查形成乳腺球体的能力。在小鼠中生成异种移植肿瘤以检查致瘤性（n=52）。在假定的癌症干细胞中检查多药耐药基因的 mRNA 表达，并对统计学上差异表达基因进行途径分析。所有统计检验均为双侧检验。
结果
途径分析表明，MCF-7/ADR 细胞表达 ABCB1 和其他在乳腺癌干细胞中也发现的基因的 mRNA（例如 CD44、TGFB1 和 SNAI1）。MCF-7/ADR 细胞具有高侵袭性，形成乳腺球体，并在小鼠中具有致瘤性。与亲本 MCF-7 细胞相比，超过 30%的 MCF-7/ADR 细胞具有 CD44+/CD24⁻表型，能够自我更新并分化（即产生 CD44+/CD24⁻和 CD44+/CD24+细胞），并过度表达各种多药耐药相关基因（包括 ABCB1、CCNE1 和 MMP9）。MCF-7/ADR 细胞在 Matrigel 中的侵袭性明显高于亲本 MCF-7 细胞（MCF-7 细胞=0.82 个细胞/视野，MCF-7/ADR=7.51 个细胞/视野，差异=6.69 个细胞/视野，95%置信区间=4.82 至 8.55 个细胞/视野，P<0.001）。在 MCF-7/MDR 中未检测到 CD44+/CD24⁻或 CD133+群体的富集。
结论
在长期连续选择多柔比星耐药后，具有癌症干细胞特征的细胞群体增加。
Zotero 插件