Engineering Research Center of Sichuan-Tibet Traditional Medicinal Plant, Chengdu University, Chengdu, 610106, China.
Institute of Cancer Biology and Drug Discovery, Chengdu University, Chengdu, 610106, China.
BMC Complement Med Ther. 2023 Apr 6;23(1):111. doi: 10.1186/s12906-023-03933-w.
Despite rapid developments in immunotherapy and targeted therapy, dacarbazine (DTIC)-based chemotherapy still has been placed at the first-line for advanced melanoma patients who are after failure of immunotherapy or targeted therapy. However, the limited response rate and survival benefit challenge the DTIC-based chemotherapy for advanced melanoma patients.
Two melanoma cell lines, A375 and SK-MEL-28 were cultured with PA and DTIC over a range of concentrations for 72 h and the cell viabilities were detected by CCK8 assay. The Bliss model and ZIP model were used for calculating the synergistic effect of PA and DTIC. DNA double-strand breaks in the two cell lines were examined by the Comet assay, and cell apoptosis was analyzed by flow cytometry. The short hairpin RNA (shRNA)-mediated knockdown, Real-time polymerase chain reaction (RT-PCR) and Western blot were performed for molecular analysis.
In the present study, we report that Protocatechuic aldehyde (PA) synergistically enhances the cytotoxicity of DTIC to two melanoma cell lines, A375 and SK-MEL-28. The combination of PA and DTIC augments DNA double-strand breaks and increases cell apoptosis. Further mechanism study reveals that PA destabilizes MGMT protein (O-6-Methylguanine-DNA Methyltransferase) through the ubiquitin-proteasome process and directly repairs DTIC-induced genetic lesions. Knockdown of MGMT compromises the synergistic effect between PA and DTIC.
Our study demonstrates that the bioactive compound, Protocatechuic aldehyde, synergistically promotes the cytotoxicity of DTIC to melanoma cells through destabilization of MGMT protein. It could be a potential candidate for melanoma chemotherapy.
尽管免疫疗法和靶向治疗有了快速发展，但在免疫治疗或靶向治疗失败后，替莫唑胺（DTIC）为基础的化疗仍被置于晚期黑色素瘤患者的一线治疗。然而，有限的反应率和生存获益对基于 DTIC 的化疗治疗晚期黑色素瘤患者提出了挑战。
用 PA 和 DTIC 在一系列浓度下培养两种黑色素瘤细胞系 A375 和 SK-MEL-28 72 小时，用 CCK8 法检测细胞活力。采用 Bliss 模型和 ZIP 模型计算 PA 和 DTIC 的协同作用。用彗星试验检测两种细胞系的 DNA 双链断裂，用流式细胞术分析细胞凋亡。采用短发夹 RNA（shRNA）介导的敲低、实时聚合酶链反应（RT-PCR）和 Western blot 进行分子分析。
本研究报告了原儿茶醛（PA）协同增强 DTIC 对两种黑色素瘤细胞系 A375 和 SK-MEL-28 的细胞毒性。PA 和 DTIC 的联合作用增强了 DNA 双链断裂并增加了细胞凋亡。进一步的机制研究表明，PA 通过泛素-蛋白酶体途径使 MGMT 蛋白（O-6-甲基鸟嘌呤-DNA 甲基转移酶）不稳定，并直接修复 DTIC 诱导的遗传损伤。MGMT 的敲低削弱了 PA 和 DTIC 之间的协同作用。
本研究表明，生物活性化合物原儿茶醛通过使 MGMT 蛋白不稳定，协同增强 DTIC 对黑色素瘤细胞的细胞毒性。它可能是一种潜在的黑色素瘤化疗候选药物。
