Key Laboratory of Marine Drugs, The Ministry of Education of China, School of Medicine and Pharmacy, Ocean University of China, Qingdao 266003, People's Republic of China.
Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266200, People's Republic of China.
Anticancer Agents Med Chem. 2022;22(6):1091-1101. doi: 10.2174/1871520621666210608110435.
Chronic Myeloid Leukemia (CML) is a myeloproliferative disease caused by BCR-ABL oncoprotein. Tyrosine kinase inhibitors have been developed to inhibit the activity of BCR-ABL; however, drug resistance and side effect occur in clinic application. Therefore, it is urgent to find novel drugs for CML treatment. Under the guidance of cytotoxic activity, crude extracts of 55 fungal strains from the medicinal mangrove Acanthus ilicifolius were evaluated, and one potent cytotoxic natural compound, brefeldin A (BFA), was discovered from Penicillium sp. (HS-N-29).
This study was aimed to determine the cytotoxic activity of BFA and the effect on the activation and expression of BCR-ABL in K562 cells.
We evaluated cytotoxic activity by MTT assay and soft agar clone assay; apoptosis and cell cycle distribution by Muse cell analyzer. The protein level of BCR-ABL and signaling molecules was detected by western blotting, and the mRNA level of BCR-ABL was determined by RT-PCR.
BFA inhibited cell proliferation, induced G2/M cell cycle arrest, and stimulated cell apoptosis in K562 cells. Importantly, for the first time, we revealed that BFA inhibited the activation of BCR-ABL and consequently inhibited the activation of its downstream signaling molecules in K562 cells. Moreover, we found BFA degraded BCR-ABL without affecting its transcription in K562 cells, and BFA-induced BCR-ABL degradation was related to caspase activation, while not to autophagy or ubiquitinated proteasome degradation pathway.
Our present results indicate that BFA acts as a dual functional inhibitor and degrader of BCR-ABL, and BFA is a potential compound for chemotherapeutics to overcome CML.
慢性髓性白血病（CML）是一种由 BCR-ABL 癌蛋白引起的骨髓增殖性疾病。已开发出酪氨酸激酶抑制剂来抑制 BCR-ABL 的活性；然而，在临床应用中会出现耐药性和副作用。因此，迫切需要寻找治疗 CML 的新型药物。在细胞毒性活性的指导下，评估了来自药用红树林老鼠簕的 55 株真菌菌株的粗提物，从青霉属（HS-N-29）中发现了一种有效的细胞毒性天然化合物布雷菲德菌素 A（BFA）。
本研究旨在确定 BFA 的细胞毒性活性及其对 K562 细胞中 BCR-ABL 的激活和表达的影响。
我们通过 MTT 测定法和软琼脂克隆测定法评估细胞毒性活性；通过 Muse 细胞分析仪评估细胞凋亡和细胞周期分布。通过 Western blot 检测 BCR-ABL 和信号分子的蛋白水平，通过 RT-PCR 检测 BCR-ABL 的 mRNA 水平。
BFA 抑制 K562 细胞的增殖，诱导 G2/M 细胞周期停滞，并刺激细胞凋亡。重要的是，我们首次揭示 BFA 抑制了 K562 细胞中 BCR-ABL 的激活，从而抑制了其下游信号分子的激活。此外，我们发现 BFA 抑制 BCR-ABL 的转录，而不影响其转录，BFA 诱导的 BCR-ABL 降解与 caspase 激活有关，而与自噬或泛素蛋白酶体降解途径无关。
我们的研究结果表明，BFA 作为 BCR-ABL 的双重功能抑制剂和降解剂，是克服 CML 的化疗药物的潜在化合物。
