Abstract
摘要:表皮生長因子受體抑制劑(EGFR TKIs) 對於活化型突變的表皮生長因子受體(EGFR)有非常好的抑制作用,但是抗藥性會侷限這類藥物的臨床使用。當EGFR發生T790M突變時,第一和第二代的EGFR TKIs即會失效。第三代EGFR TKI(osimertinib) 除了針對T790M突變有顯著的抑制效果外,osimertinib也能當成第一線藥物用於EGFR活化型突變的病患。我們證實STC2會誘導細胞自噬作用,進而造成osimertinib抗藥性。抑制STC2則能觀察到細胞自噬作用受到抑制且osimertinib敏感性增加。我們也發現而STC2的作用和訊息傳遞分子-ERK1/2的活化有關。在此計畫中,我們利用磷酸化蛋白晶片分析STC2是否活化特定受體,而導致ERK1/2磷酸化。結果發現在STC2大量表現的細胞,AXL磷酸化也會顯著增加。抑制AXL能反轉STC2造成的抗藥性,使抗藥性細胞對於osimertinib的感受性增加。這些結果顯示AXL參與STC2造成的抗藥性。先前研究證實AXL與EMT調控因子會相互調控彼此的表現;我們也發現到STC2會使EMT相關蛋白的表現量增加。這些證據加深了STC2與AXL彼此之間的關聯。我們將利用細胞株、動物實驗與病患檢體研究osimeritnib的抗藥性機制。從已抗藥病人的惡性肋膜積液中分離出抗藥性細胞,除了可用來測試AXL抑制劑與osimeritnib併用的效果外,此平台更能做為細胞實驗與臨床應用間的橋梁。因此,我們將探討STC2/AXL/ERK訊息傳遞路徑與EMT相關蛋白在抗藥性中扮演的作用。並且評估STC2、AXL與EMT相關蛋白在使用osimertinib的病人身上是否具有臨床意義與應用價值。透過分析STC2/AXL/ERK的上下游機轉,我們期待找到克服osimertinib抗藥性的機制,這對於肺癌病人會有很重大的意義。目標: (1) 探討AXL是否為STC2造成osimertinib抗藥性的關鍵蛋白。 (2) 探討STC2調控AXL表現的機制。(3) 探討EMT相關蛋白是否參與STC2 調控AXL的表現以及osimertinib抗藥性。(4) 利用分離自對osimertinib 抗藥病人的肺癌細胞 及檢測臨床檢體來證實STC2、AXL與EMT相關的蛋白表現量對osimertinib抗藥性的作用。
Abstract: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are effective against EGFR-mutant lung cancers, but there is issue with acquired resistance. Acquired EGFR T790M mutation leads to treatment failure of first and second generations EGFR TKIs (gefitinib, erlotinib, and afatinib). Osimertinib (AZD9291), a third-generation irreversible EGFR TKI, is developed against T790M mutation, and exhibits potent activity in EGFR mutant advanced NSCLC patients of treatment-naive and acquired EGFR T790M mutation. In our previous study, we showed that stanniocalcin 2 (STC2) conferred osimertinib resistance. Overexpression of STC2 increases cytoprotective autophagy against osimertinib-induced cytotoxicity, whereas knockdown of STC2 decreases autophagy and augments the cytotoxic effects of osimertinib. Moreover, STC2 induces autophagy through the ERK1/2-dependent pathway, which is a crucial modulator of osimertinib resistance. In this proposed study, we launched phospho-receptor tyrosine kinase array between mock- and STC2-overexpressing cells to find out the candidate receptor tyrosine kinase contributing to ERK activation in STC2-overexpressing cells. We showed that AXL was highly activated in STC2-overexpressing cells. Inhibition of AXL restored osimertinib sensitivity in osimertinib-resistant cells. Most importantly, AXL inhibition reverted STC2-mediated osimertinib resistance. These results suggest that AXL is involved in STC2-mediated osimertinib resistance. Previous reports demonstrated that AXL and EMT regulators reciprocally regulate each other. We also demonstrated EMT markers were up-regulated in STC2-overexpressing osimertinib-resistant cells. These observations further indicate that there is a potential correlation between STC2 and AXL. We'll investigate the osimertinib resistant mechanisms with cell lines, animal models, and clinical lung cancer tissues. Importantly, we have isolated lung cancer cells from pleural effusions of patients whose tumors are resistant to osimertinib. These cancer cells are excellent platforms in evaluating combination effects of osimertinib and AXL inhibitors. This patient-derived cell model provides a bridge between cell line study and clinical relevance. We will elucidate the role of STC2/AXL/ERK axis and EMT markers in resistance to third-generation EGFR TKI (osimertinib), and verify the clinical relevance of STC2, AXL and EMT-related proteins in osimertinib resistance of lung cancer patients. We will also validate the expression of possible genes involved in regulation of STC2/AXL/ERK axis in tissue samples of lung cancer patients who received osimertinib. Through this study, we hope to find the ways to overcome the osimertinib resistance and to treat patients of NSCLC with resistance to osimertinib. Our specific Aims:(1) Elucidate the essential role of AXL in STC2-induced osimertinib resistance. (2) Elucidate the mechanism how STC2 up-regulates AXL expression.(3) Investigated whether EMT-related proteins are essential for STC2-upregulated AXL expression and osimertinib resistance.(4) Confirm the role of STC2, AXL, and EMT-related proteins in osimertinib resistance using clinical specimens and osimertinib-resistant cells derived from patients.
Keyword(s)
STC2
AXL
肺癌
osimertinib
抗藥性
STC2
AXL
lung cancer
osimertinib
resistance