探討X814在犬乳腺腫瘤上的抗癌效果

Abstract

X814是常見的中草藥，在近年來許多研究發現其具有多重作用，包含免疫調節、抗腫瘤、抗微生物、抗血管新生與刺激免疫反應等。而目前並沒有任何文獻探討X814在犬隻腫瘤上的影響。犬乳腺腫瘤為母犬中最為好發的腫瘤。七歲以上患有腫瘤的母狗中，有52%以上的機率為乳腺腫瘤，而其中又有50%的腫瘤為惡性。手術為犬乳腺腫瘤的主要治療方法，但是術後依然有50%的機率會發生復發或轉移。因此，發展新的、具抗犬乳腺腫瘤的藥物相當的重要。本研究旨在探討X814的粗萃物，以酒精與正己烷溶劑製備後，評估其在犬乳腺腫瘤上的抗腫瘤效果。結果顯示該藥的酒精萃取物(X.E.E.)與正己烷萃取物(X.H.E.)能引起犬乳腺腫瘤細胞發生壞死性死亡、自體吞噬、與細胞周期停滯；也會抑制細胞聚落形成、移行與增生的能力。在蛋白質表現方面，X.E.E.與X.H.E.會增加犬乳腺腫瘤細胞的LC3Ⅱ蛋白的表現，而在許多文獻皆指出LC3Ⅱ的表現量與細胞自體吞噬作用具有正相關。另外，細胞質中的p53與p-AKT的表現皆有下降，可能與細胞壞死性死亡、自體吞噬、細胞周期停滯、細胞移行能力下降有關。由於細胞自體吞噬的作用在腫瘤上扮演著一體兩面的腳色，我們進一步探討X.E.E.與X.H.E.所引起細胞自體吞噬的影響。研究結果顯示以自體吞噬抑制劑bafilomycin A1或chloroquine與X.E.E.或X.H.E.共同作用後，能更加強腫瘤細胞的死亡效果。前述結果顯示X814萃取物X.E.E.與X.H.E.具有治療犬乳腺腫瘤的潛力。

X814 is an herbal plant shown to possess a number of beneficial anti-cancer and immunomodulatory properties in recent reports. Though potential anti-microbial, anti-angiogenesis, and immune-stimulation activities have been demonstrated, the effects of X814 on canine tumors have not been well elucidated. Canine mammary gland tumors (cMGT) are the most frequently diagnosed neoplasms in female dogs. Roughly 52% of canine mammary tumors are diagnosed in female dogs older than seven years. About 50% of the cases are diagnosed as malignant mammary carcinoma. Although surgical excision is now viewed as the most effective modality for treatment of cMGT, 50% of them relapsed after surgery, and the two-year survival rate of malignant cases were less than 50%. Therefore, developing potential anti-cMGT therapies becomes much more important. The crude extracts of X814 were firstly prepared in ethanol and hexane solvents to evaluate their anti-proliferation effects on cMGT cell lines. The results showed both the EtOH extract (X.E.E.) and hexane extract (X.H.E.) of X814 could inhibit cell proliferation, colony formation, and cell migration in cMGT cells. The reduction of tumor cell growth was resulted from necrosis, autophagy, and cell cycle arrest. Moreover, cMGT cells treated with X.E.E. and X.H.E. showed increased expression of autophagy-related proteins, LC3Ⅱ, and decreased cytoplasmic p53 and p-AKT in a dose-dependent manner. Autophagy seemed to play a protective role in X.E.E and X.H.E.-treated cMGT cells since using autophagy inhibitor, bafilomycin A1 or chloroquine, enhanced the cell death. Collectively, our results indicated that X814 has potential anticancer effects on cMGT.