γ-生育三烯酚与厄洛替尼或吉非替尼联合治疗对乳腺肿瘤细胞的增殖抑制和凋亡诱导作用增强。

Enhanced antiproliferative and apoptotic response to combined treatment of gamma-tocotrienol with erlotinib or gefitinib in mammary tumor cells.

机构信息

College of Pharmacy, University of Louisiana at Monroe, 700 University Ave, Monroe, Louisiana 71209, USA.

出版信息

BMC Cancer. 2010 Mar 8;10:84. doi: 10.1186/1471-2407-10-84.

DOI:10.1186/1471-2407-10-84

PMID:20211018

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC2841143/

Abstract

BACKGROUND

Aberrant ErbB receptor signaling is associated with various types of malignancies. gamma-Tocotrienol is a member of the vitamin E family of compounds that displays potent anticancer activity that is associated with suppression in ErbB receptor phosphorylation and mitogenic signaling. Erlotinib and gefitinib are tyrosine kinase inhibitors that block ErbB1 receptor activation, whereas trastuzumab is a monoclonal antibody that has been designed to specifically inhibit ErbB2 receptor activation. However, the clinical effectiveness of these agents have been disappointing because of cooperation between different ErbB family members that can rescue cancer cells from agents directed against a single ErbB receptor subtype. It was hypothesized that targeting multiple ErbB receptor subtypes with combined treatment of gamma-tocotrienol and ErbB receptor inhibitors would provide greater anticancer effects than monotherapy targeting only a single ErbB receptor subtype.

METHODS

Highly malignant mouse +SA mammary epithelial cells were maintained in culture on serum-free defined media containing 10 ng/ml EGF as a mitogen. Cell viability wase determined by MTT assay, whereas Western blot and immunofluorescent staining was used to determine treatment effects on ErbB receptor subtype level and activation. Treatment-induced apoptosis was determined using annexin V staining and Western blot analysis of cleaved caspase-3 and PARP levels.

RESULTS

Treatment with 3.5 microM gamma-tocotrienol, 0.5 microM erlotinib or 1.0 microM gefitinib alone, significantly inhibited +SA tumor cell growth. Combined treatment with subeffective doses of erlotinib (0.25 microM) or gefitinib (0.5 microM) with subeffective doses of gamma-tocotrienol (0.5-3.0 microM) significantly inhibited the growth and induced apoptosis in a dose-responsive manner. Trastuzumab treatment alone or in combination had no effect on +SA cell growth and viability. Combined treatment of gamma-tocotrienol with erlotinib or gefitinib also cause a large decrease in ErbB3, ErbB4, and to a lesser extent ErbB2 receptor levels, and EGF-dependent ErbB2-4 tyrosine phosphorylation (activation), but had no effect on ErbB1 receptor levels or activation.

CONCLUSION

Combination treatment of gamma-tocotrienol with specific ErbB receptor inhibitors is more effective in reducing mammary tumor cell growth and viability than high dose monotherapy, suggesting that targeting multiple ErbB receptors with combination therapy may significantly improve the therapeutic response in breast cancer patients.

摘要

背景

异常的 ErbB 受体信号与各种类型的恶性肿瘤有关。γ-生育三烯酚是维生素 E 家族的一种化合物，具有强大的抗癌活性，与 ErbB 受体磷酸化和有丝分裂信号的抑制有关。厄洛替尼和吉非替尼是酪氨酸激酶抑制剂，可阻断 ErbB1 受体的激活，而曲妥珠单抗是一种设计用于特异性抑制 ErbB2 受体激活的单克隆抗体。然而，这些药物的临床效果令人失望，因为不同的 ErbB 家族成员之间存在合作，这可以使癌细胞免受针对单一 ErbB 受体亚型的药物的影响。研究假设，用 γ-生育三烯酚和 ErbB 受体抑制剂联合治疗靶向多种 ErbB 受体亚型，将比仅针对单一 ErbB 受体亚型的单一疗法提供更大的抗癌效果。

方法

在含有 10ng/ml EGF 的无血清定义培养基中，将高度恶性的小鼠+SA 乳腺上皮细胞维持在培养物中作为有丝分裂原。通过 MTT 测定法测定细胞活力，通过 Western blot 和免疫荧光染色测定 ErbB 受体亚型水平和激活的治疗效果。通过 Annexin V 染色和 Western blot 分析 cleaved caspase-3 和 PARP 水平来测定治疗诱导的细胞凋亡。

结果

单独用 3.5μMγ-生育三烯酚、0.5μM 厄洛替尼或 1.0μM 吉非替尼治疗，显著抑制+SA 肿瘤细胞生长。用亚效剂量的厄洛替尼（0.25μM）或吉非替尼（0.5μM）联合亚效剂量的γ-生育三烯酚（0.5-3.0μM）联合治疗，以剂量反应方式显著抑制生长并诱导细胞凋亡。曲妥珠单抗单独或联合治疗对+SA 细胞生长和活力没有影响。γ-生育三烯酚与厄洛替尼或吉非替尼联合治疗也导致 ErbB3、ErbB4 的水平大幅下降，而 ErbB2 的水平略有下降，并且 EGF 依赖性 ErbB2-4 酪氨酸磷酸化（激活），但对 ErbB1 受体水平或激活没有影响。