老藥新用開發潛在有效治療膠質母細胞瘤藥物

Drug Repositioning for Potential Effective Glioblastoma Treatment

朱晟瑋(Cheng-Wei Chu)

指導教授 ： 洪義人

高雄醫學大學/醫學院/醫學研究所博士班/博士(2019年)

摘要

多型性神經膠質母細胞瘤 (Glioblastoma multiforme, GBM)是一種最常見也 是最致命的原發性腦癌，患者預後極差，其中位生存期僅 14-15 個月。儘管用分
子做為標靶的治療策略時有進展，仍需不斷尋求及探索新的方法。利用酵母菌雙 雜交系統，以 GSK3β 做為釣餌，並於 cDNA 基因庫找到 Bcl2L12。BCL2L12 與 GSK3β 的結合區域是藉由 C -端 BH2 模體外的中間片段 (153-191 a.a.) 負責， GSK3β 主要是對於 BCL2L12 的 BH3 相似區段在 S156 的位置上產生磷酸化。我 們研究顯示Bcl2L12參與抗細胞凋亡與自噬作用主要透過BH3相似區段、GSK3β 調控 S156 磷酸化以及 ABT-737 合併 TMZ 處理 GBM。除了特定癌症標靶藥物 的蛋白質，我們從資料庫裡蒐集了 356 個 GBM 病人的基因表現（腫瘤與非腫 瘤組織），並且利用 Connectivity map (C map) 分析，評估 79 種藥物在 GBM 細 胞抗腫瘤作用，經由這些篩選方式，找到精神疾病用藥 Thioridazine (THD)。THD 不僅上調 AMPK 活性亦藉由 Wnt/β-catenin 訊息傳遞增強 P62 調控 GBM 誘導自 噬作用所引起細胞凋亡。總而言之，我們研究強調 GSK3β 調節 Bcl2L12 或 Fzd/GSK3β/β-catenin/P62 之間的交互作用(crosstalk)，其可利用精神藥物與 TMZ 合併治療有利於 GBM 患者的的最佳治療策略。

關鍵字

多型性神經膠質母細胞瘤 ； 酵母菌雙雜交系統

並列摘要

Glioblastoma multiforme(GBM) is the most common and deadliest type of primary brain tumor, and is characterized by very poor prognosis with an overall survival of 14-15 month. Despite some development of the molecularly targeting strategies, exploration of a new approach is timely needed. From large-scale yeast twohybrid screening, we had showed that BCL2L12 was a GSK3β binding partner in a testis cDNA library. Bcl2L12 middle fragment (153-191 residues) which locates outside of Bcl2L12 C-terminal BH2 motif is responsible for the binding to GSK3β, and GSK3β‑mediated Ser156 phosphorylation modulated a BH3‑like domain in BCL2L12. Our findings indicated that BCL2L12 may participate in anti‑apoptosis and autophagy via a BH3‑like domain and GSK3β‑mediated phosphorylation at Ser156, and the implications for the possible combination use of ABT-737 and TMZ in GBM. In addition to identify specific protein for drug targeting, we collected 356 GBM gene signatures (tumor vs. non-tumor tissues) from public databases and queried the Connectivity Map, evaluated the in vitro anti-tumor effects of 79 drugs in GBM cell lines, and narrowed down to a metabolic drug repurposing, Thioridazine (THD). THD induced autophagy in GBM cells by not only upregulating AMPK activity, but also enhancing P62-mediated autophagy and apoptosis through Wnt/β-catenin signaling. Collectively, our studies emphasize either GSK3β regulates Bcl2L12 or in a crosstalk in a Fzd/GSK3β/β-catenin/P62 regulatory loop which may exploit psychotropic agents with TMZ combination treatment can benefit for the design of novel therapeutic strategies for GBM patients.

並列關鍵字

Glioblastoma multiforme ； yeast two-hybrid ； BCL2L12 ； Thioridazine ； Wnt/β-catenin

參考文獻

1. Wen, P. Y.; Kesari, S., Malignant gliomas in adults. The New England journal of medicine 2008, 359, (5), 492-507.

2. Van Meir, E. G.; Hadjipanayis, C. G.; Norden, A. D.; Shu, H. K.; Wen, P. Y.; Olson, J. J., Exciting new advances in neuro-oncology: the avenue to a cure for malignant glioma. CA: a cancer journal for clinicians 2010, 60, (3), 166-93.

3. Liu, C.; Li, Y.; Semenov, M.; Han, C.; Baeg, G. H.; Tan, Y.; Zhang, Z.; Lin, X.; He, X., Control of beta-catenin phosphorylation/degradation by a dual-kinase mechanism. Cell 2002, 108, (6), 837-47.

4. Anastas, J. N.; Moon, R. T., WNT signalling pathways as therapeutic targets in cancer. Nature reviews. Cancer 2013, 13, (1), 11-26.

5. McCord, M.; Mukouyama, Y. S.; Gilbert, M. R.; Jackson, S., Targeting WNT Signaling for Multifaceted Glioblastoma Therapy. Frontiers in cellular neuroscience 2017, 11, 318.