Ding Rui, He Ming, Huang Huoying, Chen Jing, Huang Mingxing, Su Yonghui
Department of General Surgery, The Fifth Affiliated Hospital, Sun Yat-Sen University, Zhuhai, Guangdong 519000, China.
Department of Biological Engineering, School of Biomedical and Pharmaceutical Science, Guangdong University of Technology, Guangzhou, Guangdong 510006, China.
Asian Biomed (Res Rev News). 2022 Aug 31;16(4):201-211. doi: 10.2478/abm-2022-0024. eCollection 2022 Aug.
Venomous arthropods have substances in their venom with antiproliferative potential for neoplastic cells.
To identify a polypeptide from (antlion) with antiproliferative activity against neoplastic cells, and to elucidate the molecular mechanism of the activity.
We used gel filtration and ion exchange chromatography to purify a polypeptide with antiproliferative activity against MG-63 human osteosarcoma cells from a proteinaceous extract of antlion. The polypeptide was sequenced and the stability of its antiproliferative activity was tested under a range of conditions in vitro. An 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine the antiproliferative activity of the polypeptide against the MG-63 osteosarcoma cells and MC3T3-E1 mouse calvarial osteoblasts, which were used as a non-neoplastic control. We used western blotting to compare the levels of expression of heat shock transcription factor 1 (HSF1), heat shock protein 90 (HSP90), cyclin-dependent kinase 4 (CDK4), and protein kinase B alpha (ATK1) in MG-63 osteosarcoma cells and their mouse homologs in MC3T3-E1 osteoblasts after their treatment with the antlion antiproliferative polypeptide (ALAPP).
The 85-amino-acid ALAPP has a 56% sequence identity with the human heat shock factor binding protein 1 (HSBP1). The antiproliferative activity of the polypeptide is relatively insensitive to temperature, pH, and metal ions. ALAPP has a strong concentration-dependent antiproliferative activity against MG-63 osteosarcoma cells compared with its effect on MC3T3-E1 osteoblasts. ALAPP significantly upregulates the expression of HSF1 in MC3T3-EL osteoblasts, but not in MG-63 osteosarcoma. ALAPP significantly downregulated the expression of HSP90, CDK4, and AKT1 expression in MG-63 osteosarcoma, but not in the osteoblasts.
ALAPP has significant antiproliferative activity against MG-63 osteosarcoma cells, but not nonneoplastic MC3T3-E1 osteoblasts. We speculate that non-neoplastic cells may evade the antiproliferative effect of ALAPP by upregulating HSF1 to maintain their HSP90, CDK4, and AKT1 expression at a relatively constant level.
有毒节肢动物的毒液中含有对肿瘤细胞具有抗增殖潜力的物质。
从蚁狮中鉴定出一种对肿瘤细胞具有抗增殖活性的多肽，并阐明其活性的分子机制。
我们使用凝胶过滤和离子交换色谱法从蚁狮的蛋白质提取物中纯化出一种对MG-63人骨肉瘤细胞具有抗增殖活性的多肽。对该多肽进行测序，并在一系列体外条件下测试其抗增殖活性的稳定性。使用3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐(MTT) 法测定该多肽对MG-63骨肉瘤细胞和MC3T3-E1小鼠颅骨成骨细胞的抗增殖活性，后者用作非肿瘤对照。我们使用蛋白质免疫印迹法比较了MG-63骨肉瘤细胞及其在MC3T3-E1成骨细胞中的小鼠同源物在用蚁狮抗增殖多肽 (ALAPP) 处理后热休克转录因子1 (HSF1)、热休克蛋白90 (HSP90)、细胞周期蛋白依赖性激酶4 (CDK4) 和蛋白激酶Bα (ATK1) 的表达水平。
含85个氨基酸的ALAPP与人类热休克因子结合蛋白1 (HSBP1) 的序列同一性为56%。该多肽的抗增殖活性对温度、pH和金属离子相对不敏感。与对MC3T3-E1成骨细胞的作用相比，ALAPP对MG-63骨肉瘤细胞具有强烈的浓度依赖性抗增殖活性。ALAPP显著上调MC3T3-EL成骨细胞中HSF1的表达，但在MG-63骨肉瘤细胞中则不然。ALAPP显著下调MG-63骨肉瘤细胞中HSP90、CDK4和AKT1的表达，但在成骨细胞中则不然。
ALAPP对MG-63骨肉瘤细胞具有显著的抗增殖活性，但对非肿瘤性MC3T3-E1成骨细胞则无此活性。我们推测非肿瘤细胞可能通过上调HSF1来逃避ALAPP的抗增殖作用，从而将其HSP90、CDK4和AKT1的表达维持在相对恒定的水平。
