D'Souza Amber M, Gnanamony Manu, Thomas Maria, Hanley Peter, Kanabar Dipti, de Alarcon Pedro, Muth Aaron, Timchenko Nikolai
Department of Pediatrics, University of Illinois College of Medicine Peoria, 1 Illini Drive, Peoria, IL 61605, USA.
Department of Pharmaceutical Sciences, St. John's University, 8000 Utopia Pkwy, Jamaica, NY 11439, USA.
Cancers (Basel). 2022 Jun 22;14(13):3068. doi: 10.3390/cancers14133068.
Background: Gankyrin, a member of the 26S proteasome, is an overexpressed oncoprotein in hepatoblastoma (HBL) and hepatocellular carcinoma (HCC). Cjoc42 was the first small molecule inhibitor of Gankyrin developed; however, the IC50 values of >50 μM made them unattractive for clinical use. Second-generation inhibitors demonstrate a stronger affinity toward Gankyrin and increased cytotoxicity. The aim of this study was to characterize the in vitro effects of three cjoc42 derivatives. Methods: Experiments were performed on the HepG2 (HBL) and Hep3B (pediatric HCC) cell lines. We evaluated the expression of TSPs, cell cycle markers, and stem cell markers by Western blotting and/or real-time quantitative reverse transcription PCR. We also performed apoptotic, synergy, and methylation assays. Results: The treatment with cjoc42 derivatives led to an increase in TSPs and a dose-dependent decrease in the stem cell phenotype in both cell lines. An increase in apoptosis was only seen with AFM-1 and -2 in Hep3B cells. Drug synergy was seen with doxorubicin, and antagonism was seen with cisplatin. In the presence of cjoc42 derivatives, the 20S subunit of the 26S proteasome was more available to transport doxorubicin to the nucleus, leading to synergy. Conclusion: Small-molecule inhibitors for Gankyrin are a promising therapeutic strategy, especially in combination with doxorubicin.
26S蛋白酶体成员Gankyrin是一种在肝母细胞瘤（HBL）和肝细胞癌（HCC）中过表达的癌蛋白。Cjoc42是首个研发出的Gankyrin小分子抑制剂；然而，其>50μM的半数抑制浓度（IC50）值使其不适合临床应用。第二代抑制剂对Gankyrin表现出更强的亲和力和更高的细胞毒性。本研究的目的是表征三种Cjoc42衍生物的体外作用。方法：在HepG2（HBL）和Hep3B（儿童HCC）细胞系上进行实验。我们通过蛋白质免疫印迹法和/或实时定量逆转录PCR评估了血小板反应蛋白（TSPs）、细胞周期标志物和干细胞标志物的表达。我们还进行了凋亡、协同和甲基化测定。结果：用Cjoc42衍生物处理导致两种细胞系中TSPs增加，干细胞表型呈剂量依赖性降低。仅在Hep3B细胞中用AFM - 1和 - 2处理时观察到凋亡增加。与阿霉素联用可见药物协同作用，与顺铂联用则可见拮抗作用。在存在Cjoc42衍生物的情况下，26S蛋白酶体的20S亚基更易于将阿霉素转运至细胞核，从而导致协同作用。结论：Gankyrin小分子抑制剂是一种有前景的治疗策略，尤其是与阿霉素联合使用时。
