Induction of DNMT1-dependent demethylation of SHP-1 by the natural flavonoid compound Baicalein overcame Imatinib-resistance in CML CD34+ cells

Abstract

Abstract Background The most significant cause of treatment failure in chronic myeloid leukemia (CML) is a persistent population of minimal residual cells. Emerging evidences showed that methylation of SHP-1 contributed to Imatinib (IM) resistance. Baicalein was reported to have an effect on reversal of chemotherapeutic agents resistance. However, the molecular mechanism of Baicalein on JAK2/STAT5 signaling inhibition against drug resistance in bone marrow (BM) microenvironment that had not been clearly revealed. Methods We co-cultured hBMSCs and CML CD34+ cells as a model of SFM-DR. Further researches were performed to clarify the reverse mechanisms of Baicalein on SFM-DR model and engraftment model. The apoptosis, cytotoxicity, proliferation, GM-CSF secretion, JAK2/STAT5 activity, the expression of SHP-1 and DNMT1 were analyzed. To validate the role of SHP-1 on the reversal effect of Baicalein, the SHP-1 gene was over-expressed by pCMV6-entry shp-1 and silenced by SHP-1 shRNA, respectively. Meanwhile, the DNMT1 inhibitor decitabine was used. The methylation extent of SHP-1 was evaluated using MSP and BSP. The molecular docking was replenished to further explore the binding possibility of Baicalein and DNMT1. Results BCR/ABL-independent activation of JAK2/STAT5 signaling was involved in IM resistance in CML CD34+ subpopulation. Baicalein significantly reversed BM microenvironment-induced IM resistance not through reducing GM-CSF secretion, but interfering DNMT1 expression and activity. Baicalein induced DNMT1-mediated demethylation of the SHP-1 promoter region, and subsequently activated SHP-1 re-expression, which resulted in an inhibition of JAK2/STAT5 signaling in resistant CML CD34+ cells. Molecular docking model indicated that DNMT1 and Baicalein had binding pockets in 3D structures, which further supported Baicalein might be a small-molecule inhibitor targeting DNMT1. Conclusions The mechanism of Baicalein on improving the sensitivity of CD34+ cells to IM might be correlated with SHP-1 demethylation by inhibition of DNMT1 expression. These findings suggested that Baicalein could be a promising candidate by targeting DNMT1 to eradicate minimal residual disease in CML patients.