結締組織生長因子抑制癌細胞代謝與自噬作用之研究

Connective Tissue Growth Factor inhibits Cancer Metabolism and Autophagy

賴瑋婷(Wei-Ting Lai)

指導教授 ： 郭彥彬 ； 江俊斌

國立臺灣大學/醫學院/臨床牙醫學研究所/博士(2017年)

https://doi.org/10.6342/NTU201701485

摘要

口腔癌居臺灣男性十大癌症中標準化死亡率第四位。也是癌症死亡年增率最高的癌症。瞭解口腔癌的致癌機轉及發展具潛力的癌症治療藥物對於口腔癌未來的治療有所助益。
首先，我們分析了82例口腔癌病人檢體中「運動失調毛細血管擴張症突變相互作用因子 (ataxia telangiectasia mutated interactor, ATMIN)」的表現量。發現檢體中有較高之ATMIN 表現的病人，其淋巴轉移程度比表現低的患者高。其生存期比表現低患者為短。口腔癌實驗動物模式研究結果顯示，抑制實驗動物口腔癌中ATMIN 表現可減少腫瘤轉移，增加動物生存時間。ATMIN除了能作為口腔癌腫瘤標記外，透過抑制ATMIN或其下游可能達到更好的口腔癌治療效果。
第二，我在先前的研究中發現結締組織生長因子(CTGF)可降低糖解作用及氧化磷酸化功能，減少細胞內腺核苷三磷酸產生，但在正常的上皮細胞中並沒有觀察到此現象。進一步研究顯示CTGF經由泛素-蛋白酶體路徑(ubiquitin proteasome pathway) 減少粒線體轉錄因子A (mtTFA)蛋白表現，而減少癌細胞粒線體DNA之拷貝數，降低糖解作用及氧化磷酸化功能，減少細胞內腺核苷三磷酸產生。最後抑制口腔癌細胞之浸襲和轉移能力。
腫瘤細胞處於營養缺乏、化療或標靶藥物時等壓力時，可啟動自噬作用 (autophagy)促進癌細胞存活或產生抗藥性。本研究以營養缺乏模式研究發現CTGF可干擾後期自噬作用中自噬體(autophagosome)與溶酶體融合步驟而抑制自噬作用發生。進一步研究發現，CTGF可以經由抑制自噬作用提高上皮生長因子接受器的單株抗體阻斷劑Erbitux抑制實驗動物腫瘤的敏感性。
最後，我發現CTGF轉殖株中Rab5A轉錄活性減少，藉由轉殖Rab5A表現質體或持續活化Rab5A可回復CTGF抑制的自噬作用及自噬體與溶酶體融合阻斷。在口腔鱗狀上皮細胞癌病人檢體發現末期(第四期)檢體Rab5A表現比早期和中期(第一到三期)病人的檢體高。檢體中Rab5A表現的量和CTGF表現的量呈負相關。
本研究結果除了提供ATMIN和Rab5A能作為良好的口腔癌腫瘤標記外。CTGF並具有發展成小分子蛋白藥物以作為癌症治療輔助藥物之潛力。

關鍵字

口腔癌 ； ATMIN ； 轉移 ； CTGF ； 癌症代謝 ； 自噬作用 ； Rab5A

並列摘要

In Taiwan, oral cancer (in particular oral squamous cell carcinoma, OSCC) is the fourth leading cause of death in males, and is one of the causes with high death rate worldwide. In this study, we seek to uncover the underlying mechanism of OSCC and help the development of new therapeutic strategies for OSCC.
First, we analyzed 82 OSCC cases and confirmed that OSCC patients with high ataxia telangiectasia mutated interactor (ATMIN) expression showed poor differentiation, higher TNM stages, greater lymph-node metastasis, and shorter accumulated survival time. Evidence from a buccal orthotopic implantation mouse model showed that silencing of ATMIN expression reduced lymph node metastasis and prolongs the survival of mice. ATMIN is now considered as a poor prognosis biomarker, and our study results help to identify possible therapeutic targets of downstream genes for designing effective therapeutic strategies for OSCC.
Second, my previous studies showed that connective tissue growth factor (CTGF/CCN2) significantly blocked glycolysis, mitochondrial oxidative phosphorylation (OXPHOS), and adenosine triphosphate (ATP) production. Moreover, CTGF showed no effects in normal bronchial and oral epithelial cells. We demonstrated that CTGF decreased glycolysis, mitochondrial oxidative phosphorylation, ATP generation and mitochondrial DNA (mtDNA) copy number by increasing the degradation of mitochondrial transcription factor A (mtTFA) through ubiquitin proteasome pathway and in turn reduced migration and invasion of OSCC cells.
Autophagy can promote cancer cell survival or resistance under conditions of poor nutrient supply, chemotherapy or target therapy. In this study, we demonstrated that CTGF inhibited late stage autophagy through blocking autophagosome-lysosome fusion under starvation condition. We further showed that cotreatment of CTGF could markedly enhance the therapeutic efficiency of Erbitux in vivo
Finally, we showed that overexpression of CTGF significantly decreased Rab5A expression, which is essential for completion of autophagy. Furthermore, expression of a wild type or constitutive active form of Rab5A could restore CTGF-inhibited autophagy flux. We investigated the clinical importance of CTGF-Rab5A axis in OSCC patients, the results showed that high Rab5A mRNA expression was significantly associated with an advanced clinical pathological TNM stage. A reverse correlation between CTGF and Rab5A was also demonstrated.
The findings in this study demonstrated that ATMIN and Rab5A can be poor prognostic biomarkers in OSCC. I believe that CTGF has the potential to be developed as an additive therapeutic agent for cancer treatment in the future.

並列關鍵字

OSCC ； ATMIN ； metastasis ； CTGF ； cancer metabolism ； autophagy ； Rab5A

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