| 摘要: | 乳癌在20年來的發生比例逐年上升，而發病年齡逐年上升下降，也因此在臨床上不斷有許多專家學者蒐集分析乳癌治療，以期降低復發率及死亡率。然而在多年來研究顯示發現三陰性乳癌預後差，且大部分患者容易在治療完成後5年內復發，並且比起管腔型乳癌更容易發生腦部轉移、肺部轉移、肝臟轉移。也因此，部分專家經年累月的研究成果發現，部分的三陰性乳癌跟免疫失能、BRCA基因突變有關聯性，也因此，PDL1抑制劑及PARP抑制劑問世並且運用在三陰性乳癌患者身上。但是仍然有部分三陰性乳癌與上述情況無關，化學治療仍為首選藥物，但是治療效果不佳及復發，讓我們去思考藥物抗性的可能，也因此，我們團隊使用大數據系統去分析現有的臨床數據，來推論出在藥物抗性的情況下某些基因蛋白質可能會高度表現，再使用Gene set enrichment analysis (GSEA)數據來分析相對應基因群其所屬的細胞傳遞路徑，比對出可能的傳遞路徑，再取傳遞路徑上的相關蛋白質再反覆驗證，來證明這些乳癌在化療抗藥性時所反應出高表現的基因蛋白的傳遞路徑，試著去逆調控這些基因蛋白來降低抗藥性增加化療藥物療效，另外也可以藉此基因蛋白來預估患者的藥物效果來評估預後。
由於化學療法是當前治療三陰性乳癌（TNBC）患者的主要策略，因此迫切需要發展鑑定生物標誌物以預測化學療法的反應，以使患者避免通過不必要的化學療法治療而遭受痛苦。在這裡，我們發現抑制敲低TNFSF13可以增強Paclitaxel (PTX) 不敏感的MDA-MB231細胞中的PTX效力。此外，Kaplan-Meier分析顯示，在接受蒽環類藥物(Anthracycline)治療的雌激素受體（ER）陰性的乳腺癌患者中，TNFSF13mRNA高表現顯著預測了癌症復發的風險增加。值得注意的是，使用抑制自噬活性的藥物可恢復PTX在TNFSF13處理的HCC1806細胞中的治療效果。這些發現表明，TNFSF13可以作為TNBC患者的預測生物標誌物，他們可以用它來決定是否接受化療。
當肺轉移在三陰性乳癌（TNBC）中發生，將導致TNBC患者的生存率降低。然而，TNBC細胞肺轉移的分子機制仍是未知之數。通過使用電腦計算基因組富集分析（GSEA），我們發現mTORC1相關途徑在Gh高表現上調的雌激素受體（ER）陰性乳腺癌伴肺轉移的原發腫瘤中被高度激活。自噬活性與內源性Gh表現量水平和細胞侵襲能力呈負相關。在高侵襲性的MDA-MB231細胞中，敲弱低Gh並通過合成肽抑製劑阻斷其與PLC-1的結合，可顯著抑制Akt / mTORC1磷酸化並促進自噬啟動，另外即增強Gh的強製表達在侵襲性差的HCC1806細胞中，Akt / mTORC1磷酸化亦增強，但自噬啟動受到抑制。發現使用3-甲基腺嘌呤(3-MA)對藥物用於抑制自噬作用起始的抑製作用可以回復挽救沉默Gh被沉默化的MDA-MB231細胞的侵襲能力和肺轉移潛能。相反，雷帕黴素(Rapamycin)對mTORC1活性的抑制製作用增強了自噬的啟動， 減輕了但減輕在了Gh過度表達之的HCC1806細胞的侵襲能力和肺轉移潛能。 重要的是，高水平Gh表達和低水平自噬活性結合的特徵與ER（-）乳腺癌患者發生肺轉移的風險增加相關。這些發現代表在Gh/PLC-1軸驅動Akt-mTORC1軸的活化後抑制表明自噬作用誘導TNBC轉移及惡化。活性的抑制是由激活的Akt-mTORC1軸在Gh/PLC-1驅動的TNBC轉移中誘導的結合，可顯著抑制Akt / mTORC1磷酸化並促進自噬啟動，即Gh的強製制表達在侵襲性差的HCC1806細胞中，Akt / mTORC1磷酸化增強，但自噬啟動受到抑制。發現3-甲基腺嘌呤對藥物自噬起始的抑製制作用可以挽救沉默Gh的MDA-MB231細胞的侵襲能力和肺轉移潛能。相反，雷帕黴素對mTORC1活性的抑製作用增強了自噬的啟動，但減輕了Gh過表達的HCC1806細胞的侵襲能力和肺轉移潛能。重要的是，高水平Gh表達和低水平自噬活性結合的特徵與ER（-）乳腺癌患者發生肺轉移的風險增加相關。這些發現表明自噬活性的抑制是由激活的Akt-mTORC1軸在Gh/PLC-1驅動的TNBC轉移中誘導的。
The incidences of breast cancer increase yearly in the past 20 years, and younger age of onset has been noticed year by year. However, studies over the years have shown that triple-negative breast cancer has a poor prognosis, and most patients are prone to relapse within 5 years after the completion of treatment, and are more prone to brain metastasis, lung metastasis, and liver metastasis than luminal-type breast cancer. Therefore, experts have found that some triple-negative breast cancers (TNBCs) are associated with immune disability and BRCA gene mutations. Therefore, PDL1 inhibitors and PARP inhibitors have been used in triple-negative breast cancer patients. Although chemotherapy is still preferred, the effect in some patients’ treatment was poor and the disease recurred in a short time, due to drug resistance. Therefore, we analyze data in silico, to identify certain genes that may be highly expressed in the case of drug resistance, and then use Gene Set Enrichment Analysis (GSEA) database to analyze the cell signal transduction pathways of the corresponding gene sets in chemo-resistance events. Those relevant proteins in the pathways may be applied as biomarkers to estimate the patients’ drug effect and to evaluate the prognosis of the TNBCs.
Since chemotherapy is a main strategy to treat TNBC patients currently, identifying a biomarker to predict chemotherapeutic responses is urgently needed for patients to avoid suffering through unnecessary chemotherapeutic treatments. Here, we found that the endogenous expression of TNFSF13 in a panel of TNBC cell lines highly correlated with paclitaxel (PTX) and doxorubicin IC50, whereas knocking down TNFSF13 enhanced PTX effectiveness in PTX-insensitive MDA-MB231 cells. Moreover, Kaplan-Meier analysis revealed that higher TNFSF13mRNA expression significantly predicted an increased risk for cancer recurrence in estrogen receptor (ER)-negative breast cancer patients receiving an anthracycline-based treatment. The in silico analysis and Western blotting demonstrated that TNFSF13 expression inversely associated with the activity of the Akt-mTOR pathway, which acts as a negative regulator of autophagy activity. Significantly, the pharmaceutical inhibition of autophagy activity restored the therapeutic effectiveness of PTX in TNFSF13-treated HCC1806 cells. These findings suggest that TNFSF13 can serve as a predictive biomarker for TNBC patients to decide whether to receive chemotherapy.
Lung metastasis is commonly found in TNBC patients and leads to a reduced survival rate. However, the molecular mechanism underlying the lung metastasis of TNBC cells remains largely unknown. By using in silico GSEA, we found that the mTORC1-related pathway was predicted to be highly activated in Gh-upregulated primary tumors from estrogen receptor (ER)-negative breast cancer patients with lung metastasis. Knocking down Gh, as well as blocking its binding with PLC-1 by a synthetic peptide inhibitor, in the highly invasive MDA-MB231 cells dramatically suppressed Akt/mTORC1 phosphorylation and promoted autophagy initiation. On the other hand, the forced expression of Gh in the poorly invasive HCC1806 cells enhanced Akt/mTORC1 phosphorylation but suppressed autophagy initiation. The pharmaceutical inhibition of autophagy initiation by 3-methyladenine was found to rescue the cell invasion ability and lung metastatic potential of Gh-silenced MDA-MB231 cells. In contrast, the inhibition of mTORC1 activity by rapamycin enhanced autophagy initiation but mitigated the cell invasion ability and lung metastatic potential of Gh-overexpressing HCC1806 cells. Significantly, the signature of a combination of high-level Gh expression and low-level autophagy activity correlated with an increased risk of lung metastasis for ER(-) breast cancer patients. These findings demonstrate that the inhibition of autophagy activity is induced by the activated Akt-mTORC1 axis in Gh/PLC-1-driven TNBC metastasis. |
