Department of Breast Medical Oncology, The University of Texas M, D, Anderson Cancer Center, Houston, TX 77030, USA.
Breast Cancer Res. 2010;12(6):R96. doi: 10.1186/bcr2777. Epub 2010 Nov 16.
The human epidermal growth factor receptor 2 (HER2) is a validated therapeutic target in breast cancer. Heterodimerization of HER2 with other HER family members results in enhanced tyrosine phosphorylation and activation of signal transduction pathways. HER2 overexpression increases the translation of fatty acid synthase (FASN), and FASN overexpression markedly increases HER2 signaling, which results in enhanced cell growth. However, the molecular mechanism and regulation of HER2 and FASN interaction are not well defined. Lapatinib is a small-molecule tyrosine kinase inhibitor that blocks phosphorylation of the epidermal growth factor receptor and HER2 in breast cancer cells, resulting in apoptosis. We hypothesized that FASN is directly phosphorylated by HER2, resulting in enhanced signaling and tumor progression in breast cancer cells.
Using mass spectrometry, we identified FASN as one of the proteins that is dephosphorylated by lapatinib in SKBR3 breast cancer cells. Immunofluorescence, immunoprecipitation, Western blotting, a kinase assay, a FASN enzymatic activity assay, an invasion assay, a cell viability assay and zymography were used to determine the role of FASN phosphorylation in invasion of SKBR3 and BT474 cells. The FASN inhibitor C75 and small interfering RNA were used to downregulate FASN expression and/or activity.
Our data demonstrated that FASN is phosphorylated when it is in complex with HER2. FASN phosphorylation was induced by heregulin in HER2-overexpressing SKBR3 and BT474 breast cancer cells. Heregulin-induced FASN phosphorylation resulted in increased FASN enzymatic activity, which was inhibited by lapatinib. The FASN inhibitor C75 suppressed FASN activity by directly inhibiting HER2 and FASN phosphorylation. Blocking FASN phosphorylation and activity by lapatinib or C75 suppressed the activity of matrix metallopeptidase 9 and inhibited invasion of SKBR3 and BT474 cells.
FASN phosphorylation by HER2 plays an important role in breast cancer progression and may be a novel therapeutic target in HER2-overexpressing breast cancer cells.
人表皮生长因子受体 2（HER2）是乳腺癌的一个经过验证的治疗靶点。HER2 与其他 HER 家族成员异二聚化导致酪氨酸磷酸化增强和信号转导途径激活。HER2 过表达增加脂肪酸合酶（FASN）的翻译，而 FASN 过表达显著增加 HER2 信号，导致细胞生长增强。然而，HER2 和 FASN 相互作用的分子机制和调节尚不清楚。拉帕替尼是一种小分子酪氨酸激酶抑制剂，可阻断乳腺癌细胞中表皮生长因子受体和 HER2 的磷酸化，导致细胞凋亡。我们假设 FASN 被 HER2 直接磷酸化，导致乳腺癌细胞中信号增强和肿瘤进展。
使用质谱法，我们鉴定出 FASN 是拉帕替尼使 SKBR3 乳腺癌细胞中去磷酸化的蛋白质之一。免疫荧光、免疫沉淀、Western blot、激酶测定、FASN 酶活性测定、侵袭测定、细胞活力测定和胶内酶谱分析用于确定 FASN 磷酸化在 SKBR3 和 BT474 细胞侵袭中的作用。使用 FASN 抑制剂 C75 和小干扰 RNA 下调 FASN 表达和/或活性。
我们的数据表明，当 FASN 与 HER2 复合物时，FASN 被磷酸化。在 HER2 过表达的 SKBR3 和 BT474 乳腺癌细胞中，由人表皮生长因子样生长因子诱导 FASN 磷酸化。人表皮生长因子样生长因子诱导的 FASN 磷酸化导致 FASN 酶活性增加，而拉帕替尼可抑制该活性。FASN 抑制剂 C75 通过直接抑制 HER2 和 FASN 磷酸化来抑制 FASN 活性。拉帕替尼或 C75 通过抑制基质金属蛋白酶 9 的活性并抑制 SKBR3 和 BT474 细胞的侵袭来抑制 FASN 磷酸化和活性。
HER2 对 FASN 的磷酸化在乳腺癌进展中起重要作用，可能是 HER2 过表达乳腺癌细胞的一个新的治疗靶点。
