背景:增加癌症干细胞(CSC)含量和SOX2过表达是激素依赖性乳腺癌对治疗产生耐药性的共同特征,这仍然是一个重要的临床挑战。SOX2具有作为治疗抵抗的生物标志物和作为治疗靶标的潜力,但是靶向转录因子也具有挑战性。这里,我们研究了不同多金属氧酸盐(POM)衍生物对他莫昔芬耐药乳腺癌细胞中SOX2转录因子的潜在抑制作用.
方法:合成了各种POM衍生物,并通过红外光谱表征,粉末X射线衍射图和核磁共振波谱。雌激素受体(ER)阳性乳腺癌细胞,和他们的同行,对他莫昔芬的激素疗法产生了抗药性,用POM处理,并通过凝胶阻滞和染色质免疫沉淀评估其后果,以确定SOX2与DNA的结合。对增殖的影响,迁移,使用显微镜监测和定量侵袭和致瘤性,克隆形成,transwell,伤口愈合试验,流式细胞术和体内鸡绒毛尿囊膜(CAM)模型。应用使用CRISPR-Cas9基因组编辑的慢病毒稳定基因沉默和基因敲除的产生来验证所选择的POM的抑制作用。癌症干细胞亚群通过乳腺球形成试验进行定量,ALDEFLUOR活性和CD44/CD24染色。流式细胞术和蛋白质印迹用于测量活性氧(ROS)和细胞凋亡。
结果:POM阻断了内源性SOX2的体外结合活性。[P2W18O62]6-(PW)Wells-Dawson型阴离子在抑制他莫昔芬抗性的各种细胞系模型中最有效。10µMPW还减少了癌细胞的迁移和侵袭,以及SNAI2表达水平。通过降低CSC含量治疗具有PW受损肿瘤形成的他莫昔芬抗性细胞,以SOX2依赖的方式,导致体内干细胞消耗。机械上,PW诱导活性氧(ROS)的形成并抑制Bcl-2,导致他莫昔芬耐药细胞死亡。PW处理的他莫昔芬抗性细胞显示恢复对他莫昔芬的敏感性。
结论:一起,这些观察结果强调了PW作为SOX2抑制剂的潜在用途,以及靶向SOX2治疗他莫昔芬耐药乳腺癌的治疗相关性.
BACKGROUND: Increased cancer stem cell (CSC) content and SOX2 overexpression are common features in the development of resistance to therapy in hormone-dependent breast cancer, which remains an important clinical challenge. SOX2 has potential as biomarker of resistance to treatment and as therapeutic target, but targeting transcription factors is also challenging. Here, we examine the potential inhibitory effect of different polyoxometalate (POM) derivatives on SOX2 transcription factor in tamoxifen-resistant breast cancer cells.
METHODS: Various POM derivatives were synthesised and characterised by infrared spectra, powder X-ray diffraction pattern and nuclear magnetic resonance spectroscopy. Estrogen receptor (ER) positive breast cancer cells, and their counterparts, which have developed resistance to the hormone therapy tamoxifen, were treated with POMs and their consequences assessed by gel retardation and chromatin immunoprecipitation to determine SOX2 binding to DNA. Effects on proliferation, migration, invasion and tumorigenicity were monitored and quantified using microscopy, clone formation, transwell, wound healing assays, flow cytometry and in vivo chick chorioallantoic membrane (CAM) models. Generation of lentiviral stable gene silencing and gene knock-out using CRISPR-Cas9 genome editing were applied to validate the inhibitory effects of the selected POM. Cancer stem cell subpopulations were quantified by mammosphere formation assays, ALDEFLUOR activity and CD44/CD24 stainings. Flow cytometry and western blotting were used to measure reactive oxygen species (ROS) and apoptosis.
RESULTS: POMs blocked in vitro binding activity of endogenous SOX2. [P2W18O62]6- (PW) Wells-Dawson-type anion was the most effective at inhibiting proliferation in various cell line models of tamoxifen resistance. 10 µM PW also reduced cancer cell migration and invasion, as well as SNAI2 expression levels. Treatment of tamoxifen-resistant cells with PW impaired tumour formation by reducing CSC content, in a SOX2-dependent manner, which led to stem cell depletion in vivo. Mechanistically, PW induced formation of reactive oxygen species (ROS) and inhibited Bcl-2, leading to the death of tamoxifen-resistant cells. PW-treated tamoxifen-resistant cells showed restored sensitivity to tamoxifen.
CONCLUSIONS: Together, these observations highlight the potential use of PW as a SOX2 inhibitor and the therapeutic relevance of targeting SOX2 to treat tamoxifen-resistant breast cancer.