Tamoxifen is a successful endocrine therapy drug for estrogen receptor–positive (ER+) breast cancer. However, resistance to tamoxifen compromises the efficacy of endocrine treatment. In the present study, we identified potential tamoxifen resistance–related gene markers and investigated their mechanistic details. First, we established two ER + breast cancer cell lines resistant to tamoxifen, named MCF-7/TMR and BT474/TMR. Gene expression profiling showed that CXXC finger protein 4 (CXXC4) expression is lower in MCF-7/TMR cells than in MCF-7 cells. Furthermore, CXXC4 mRNA and protein expression are lower in the resistant cell lines than in the corresponding parental cell lines. We also investigated the correlation between CXXC4 and endocrine resistance in ER + breast cancer cells. CXXC4 knockdown accelerates cell proliferation in vitro and in vivo and renders breast cancer cells insensitive to tamoxifen, whereas CXXC4 overexpression inhibits cancer cell growth and increases tamoxifen sensitivity of resistant cells. In addition, we demonstrated that CXXC4 inhibits Wnt/β-catenin signaling in cancer cells by modulating the phosphorylation of GSK-3β, influencing the integrity of the β-catenin degradation complex. Silencing the CXXC4 gene upregulates expression of cyclinD1 and c-myc (the downstream targets of Wnt signaling) and promotes cell cycle progression. Conversely, ectopic expression of CXXC4 downregulates the expression of these proteins and arrests the cell cycle in the G0/G1 phase. Finally, the small-molecule inhibitor XAV939 suppresses Wnt signaling and sensitizes resistant cells to tamoxifen. These results indicate that components of Wnt pathway that are early in response to tamoxifen could be involved as an intrinsic factor of the transition to endocrine resistance, and inhibition of Wnt signaling may be an effective therapeutic strategy to overcome tamoxifen resistance.

他莫昔芬是一种成功的内分泌治疗药物，可治疗雌激素受体阳性（ER +）乳腺癌。然而，对他莫昔芬的抗性损害了内分泌治疗的功效。在本研究中，我们鉴定了潜在的他莫昔芬抗药性相关基因标记，并研究了它们的机理细节。首先，我们建立了两种对他莫昔芬具有抗性的ER +乳腺癌细胞系，分别为MCF-7 / TMR和BT474 / TMR。基因表达谱分析表明，MCF-7 / TMR细胞中的CXXC手指蛋白4（CXXC4）表达低于MCF-7细胞。此外，抗性细胞系中的CXXC4 mRNA和蛋白质表达低于相应的亲本细胞系。我们还研究了ER +乳腺癌细胞中CXXC4与内分泌抗性之间的相关性。CXXC4敲低可加速细胞增殖体外和体内导致乳腺癌细胞对他莫昔芬不敏感，而CXXC4的过表达抑制癌细胞的生长并增加他莫昔芬对耐药细胞的敏感性。此外，我们证明了CXXC4通过调节GSK-3β的磷酸化来抑制癌细胞中Wnt /β-catenin信号传导，从而影响β-catenin降解复合物的完整性。沉默CXXC4基因上调cyclinD1和c-myc（Wnt信号下游靶标）的表达并促进细胞周期进程。相反，CXXC4的异位表达下调了这些蛋白质的表达，并使细胞周期停滞在G0 / G1期。最后，小分子抑制剂XAV939抑制Wnt信号转导并使耐药细胞对他莫昔芬敏感。