Metastatic breast cancer is responsible for the death of the majority of breast cancer patients. In fact, metastatic BC is the 2nd leading cause of cancer-related deaths in women in the USA and worldwide. Triple negative breast cancer (TNBC), which lacks expression of hormone receptors (ER-α and PR) and ErbB2/HER2, is especially lethal due to its highly metastatic behavior, propensity to recur rapidly, and for its resistance to standard of care therapies, through mechanisms that remain incompletely understood. WAVE3 has been established as a promoter of TNBC development and metastatic progression. In this study, we investigated the molecular mechanisms whereby WAVE3 promotes therapy-resistance and cancer stemness in TNBC, through the regulation of β-catenin stabilization. The Cancer Genome Atlas dataset was used to assess the expression of WAVE3 and β-catenin in breast cancer tumors. Kaplan–Meier Plotter analysis was used to correlate expression of WAVE3 and β-catenin with breast cancer patients’ survival probability. MTT assay was used to quantify cell survival. CRISPR/Cas9-mediated gene editing, 2D and 3D tumorsphere growth and invasion assays, Immunofluorescence, Western blotting, Semi-quantitative and real-time quantitative PCR analyses were applied to study the WAVE3/β-catenin oncogenic signaling in TNBC. Tumor xenograft assays were used to study the role of WAVE3 in mediating chemotherapy resistance of TNBC tumors. Genetic inactivation of WAVE3 in combination of chemotherapy resulted in inhibition of 2D growth and 3D tumorsphere formation and invasion of TNBC cells in vitro, as well as tumor growth and metastasis in vivo. In addition, while re-expression of phospho-active WAVE3 in the WAVE3-deficient TNBC cells restored the oncogenic activity of WAVE3, re-expression of phospho-mutant WAVE3 did not. Further studies revealed that dual blocking of WAVE3 expression or phosphorylation in combination with chemotherapy treatment inhibited the activity and expression and stabilization of β-catenin. Most importantly, the combination of WAVE3-deficiency or WAVE3-phospho-deficiency and chemotherapy suppressed the oncogenic behavior of chemoresistant TNBC cells, both in vitro and in vivo. We identified a novel WAVE3/β-catenin oncogenic signaling axis that modulates chemoresistance of TNBC. This study suggests that a targeted therapeutic strategy against WAVE3 could be effective for the treatment of chemoresistant TNBC tumors.

中文翻译：

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WAVE3/β-连环蛋白致癌信号调节三阴性乳腺癌的化疗耐药性

转移性乳腺癌是导致大多数乳腺癌患者死亡的原因。事实上，转移性 BC 是美国和全世界女性癌症相关死亡的第二大主要原因。三阴性乳腺癌 (TNBC) 缺乏激素受体（ER-α 和 PR）和 ErbB2/HER2 的表达，由于其高度转移行为、快速复发倾向以及对标准护理疗法的抵抗力，因此特别致命，通过仍未完全理解的机制。WAVE3 已被确定为 TNBC 发展和转移进展的促进者。在这项研究中，我们研究了 WAVE3 通过调节 β-连环蛋白稳定性来促进 TNBC 治疗耐药性和癌症干性的分子机制。癌症基因组图谱数据集用于评估 WAVE3 和 β-连环蛋白在乳腺癌肿瘤中的表达。Kaplan-Meier Plotter 分析用于将 WAVE3 和 β- 连环蛋白的表达与乳腺癌患者的生存概率相关联。MTT测定用于量化细胞存活。CRISPR/Cas9 介导的基因编辑、2D 和 3D 肿瘤球生长和侵袭测定、免疫荧光、蛋白质印迹、半定量和实时定量 PCR 分析被应用于研究 TNBC 中的 WAVE3/β-catenin 致癌信号传导。肿瘤异种移植试验用于研究 WAVE3 在介导 TNBC 肿瘤化疗耐药中的作用。WAVE3 基因失活联合化疗导致 TNBC 细胞体外 2D 生长和 3D 肿瘤球形成和侵袭受到抑制，以及体内肿瘤的生长和转移。此外，虽然在 WAVE3 缺陷型 TNBC 细胞中重新表达磷酸化活性 WAVE3 可恢复 WAVE3 的致癌活性，但重新表达磷酸化突变体 WAVE3 却没有。进一步的研究表明，WAVE3 表达或磷酸化的双重阻断与化疗治疗相结合，可抑制 β-连环蛋白的活性、表达和稳定。最重要的是，WAVE3 缺乏症或 WAVE3 磷酸化缺乏症与化疗相结合，在体外和体内均抑制了化疗耐药 TNBC 细胞的致癌行为。我们发现了一种新的 WAVE3/β-catenin 致癌信号轴，可调节 TNBC 的化疗耐药性。这项研究表明针对 WAVE3 的靶向治疗策略可有效治疗化疗耐药的 TNBC 肿瘤。