MUC4 Overexpression Augments Cell Migration and Metastasis through EGFR Family Proteins in Triple Negative Breast Cancer Cells

IntroductionCurrent studies indicate that triple negative breast cancer (TNBC), an aggressive breast cancer subtype, is associated with poor prognosis and an early pattern of metastasis. Emerging evidence suggests that MUC4 mucin is associated with metastasis of various cancers, including breast cancer. However, the functional role of MUC4 remains unclear in breast cancers, especially in TNBCs.
MethodIn the present study, we investigated the functional and mechanistic roles of MUC4 in potentiating pathogenic signals including EGFR family proteins to promote TNBC aggressiveness using in vitro and in vivo studies. Further, we studied the expression of MUC4 in invasive TNBC tissue and normal breast tissue by immunostaining.
ResultsMUC4 promotes proliferation, anchorage-dependent and-independent growth of TNBC cells, augments TNBC cell migratory and invasive potential in vitro, and enhances tumorigenicity and metastasis in vivo. In addition, our studies demonstrated that MUC4 up-regulates the EGFR family of proteins, and augments downstream Erk1/2, PKC-γ, and FAK mediated oncogenic signaling. Moreover, our studies also showed that knockdown of MUC4 in TNBC cells induced molecular changes suggestive of mesenchymal to epithelial transition. We also demonstrated in this study, for the first time, that knockdown of MUC4 was associated with reduced expression of EGFR and ErbB3 (EGFR family proteins) in TNBC cells, suggesting that MUC4 uses an alternative to ErbB2 mechanism to promote aggressiveness. We further demonstrate that MUC4 is differentially over-expressed in invasive TNBC tissues compared to normal breast tissue.
ConclusionsMUC4 mucin expression is associated with TNBC pathobiology, and its knockdown reduced aggressiveness in vitro, and tumorigenesis and metastasis in vivo. Overall, our findings suggest that MUC4 mucin promotes invasive activities of TNBC cells by altering the expression of EGFR, ErbB2, and ErbB3 molecules and their downstream signaling.

引言
现有研究表明，三阴性乳腺癌（triple negative breast cancer, TNBC）作为一种侵袭性乳腺癌亚型，与不良预后及早期转移模式密切相关。越来越多的证据显示，黏蛋白MUC4（MUC4 mucin）与包括乳腺癌在内的多种癌症的转移相关。然而，黏蛋白MUC4在乳腺癌尤其是三阴性乳腺癌中的功能作用仍不明确。

方法
本研究通过体外（in vitro）与体内（in vivo）实验，探究了黏蛋白MUC4在激活包括表皮生长因子受体家族蛋白（EGFR family proteins）在内的致病信号以促进三阴性乳腺癌侵袭性中的功能与机制作用。此外，我们通过免疫染色法检测了侵袭性三阴性乳腺癌组织与正常乳腺组织中MUC4的表达水平。

结果
黏蛋白MUC4可促进三阴性乳腺癌细胞的增殖、锚定依赖性与非锚定依赖性生长，在体外增强三阴性乳腺癌细胞的迁移与侵袭能力，并在体内提升其成瘤性与转移能力。此外，本研究证实黏蛋白MUC4可上调表皮生长因子受体家族蛋白的表达，并增强下游Erk1/2、蛋白激酶Cγ（PKC-γ）与黏着斑激酶（FAK）介导的致癌信号通路。进一步研究显示，在三阴性乳腺癌细胞中敲低MUC4可诱导分子层面的变化，提示细胞发生间质上皮转化（mesenchymal to epithelial transition, MET）。本研究还首次证实，在三阴性乳腺癌细胞中敲低MUC4会伴随表皮生长因子受体（EGFR）与ErbB3的表达下调，提示黏蛋白MUC4可通过不同于ErbB2的机制促进三阴性乳腺癌的侵袭性。我们还进一步证实，相较于正常乳腺组织，侵袭性三阴性乳腺癌组织中MUC4呈差异性高表达。

结论
黏蛋白MUC4的表达与三阴性乳腺癌的病理生物学特性相关，敲低MUC4可在体外降低其侵袭能力，并在体内抑制成瘤与转移过程。综上，本研究结果提示，黏蛋白MUC4可通过调控表皮生长因子受体、ErbB2与ErbB3分子的表达及其下游信号通路，促进三阴性乳腺癌细胞的侵袭活性。