Thymoquinone Inhibits Growth of Acute Myeloid Leukemia Cells through Reversal SHP-1 and SOCS-3 Hypermethylation: In Vitro and In Silico Evaluation

Abstract

Epigenetic silencing of tumor suppressor genes (TSGs) plays an essential role in cancer pathogenesis, including acute myeloid leukemia (AML). All of <i>SHP-1</i>, <i>SOCS-1</i>, and <i>SOCS-3</i> are TSGs that negatively regulate JAK/STAT signaling. Enhanced re-expression of TSGs through de-methylation represents a therapeutic target in several cancers. Thymoquinone (TQ) is a major component of <i>Nigella sativa</i> seeds with anticancer effects against several cancers. However, the effects of TQ on DNA methylation are not entirely understood. This study aimed to evaluate the ability of TQ to re-express <i>SHP-1</i>, <i>SOCS-1</i>, and <i>SOCS-3</i> in MV4-11 AML cells through de-methylation. Cytotoxicity, apoptosis, and cell cycle assays were performed using WSTs-8 kit, Annexin V-FITC/PI apoptosis detection kit, and fluorometric-red cell cycle assay kit, respectively. The methylation of <i>SHP-1</i>, <i>SOCS-1</i>, and <i>SOCS-3</i> was evaluated by pyrosequencing analysis. The expression of <i>SHP-1</i>, <i>SOCS-1</i>, <i>SOCS-3</i>, <i>JAK2</i>, <i>STAT3</i>, <i>STAT5A</i>, <i>STAT5B</i>, <i>FLT3</i>-ITD, <i>DNMT1</i>, <i>DNMT3A</i>, <i>DNMT3B</i>, <i>TET2</i>, and <i>WT1</i> was assessed by RT-qPCR. The molecular docking of TQ to <i>JAK2</i>, <i>STAT3</i>, and <i>STAT5</i> was evaluated. The results revealed that TQ significantly inhibited the growth of MV4-11 cells and induced apoptosis in a dose- and time-dependent manner. Interestingly, the results showed that TQ binds the active pocket of <i>JAK2</i>, <i>STAT3</i>, and <i>STAT5</i> to inhibit their enzymatic activity and significantly enhances the re-expression of <i>SHP-1</i> and <i>SOCS-3</i> through de-methylation. In conclusion, TQ curbs MV4-11 cells by inhibiting the enzymatic activity of JAK/STAT signaling through hypomethylation and re-expression of JAK/STAT negative regulators and could be a promising therapeutic candidate for AML patients.