Combination of Tigecycline and Homoharringtonine Synergistically Enhances Anti-Leukemia and Anti-MDS Effect Followed By Inhibiting Mitochondrial Translation through AKT/Mtorpathway and Downregulation of Anti-Apoptotic Proteins

Title:Combination of Tigecycline and HomoharringtonineSynergistically Enhances anti-Leukemia and anti-MDS Effect Followed by Inhibiting Mitochondrial Translation through AKT/mTORPathway and Downregulation of anti-Apoptotic Proteins.

Abstract

Antimicrobial tigecycline was identified toxic to leukemia stem and progenitor cells compared to their normal counterparts and also showed antileukemic activity in mouse models of human leukemia through inhibiting mitochondrial translation. We aimed to investigate the effect of several antineoplastic drugs (Idarubicin, Homoharringtonine, Chidamide)in combination with tigecycline on the proliferation of myeloid leukemia cells and MDS cells, to select the most efficient combination group and explore the associated mechanisms of these combination therapies.

Cell proliferation was tested by MTT assay and CFU-GM assay. Cell apoptosis was evaluated by Annexin V and PI staining in cell culture, TUNEL assay and transmission electron microscopy in animal study.The tumor volume in mouse model was measured with a vernier caliper. Molecular studies were conducted using microarray expression analysis, which was used to explore associated pathways. Real-time quantitative reverse transcription-PCR, western blot and immunohistochemistry were used to assess regulation of AKT/mTORpathway.

Among three antineoplastic drugs agents in combining with tigecycline, the combination of tigecycline and homoharringtonineinduced synergistic cell death in SKM-1 cells, and this effect was verified in MDS-L, molml13 cells and primary cells isolated from patients diagnosed with MDS. Importantly, tumor growth inhibition in this combination was found to be higher than in single agent or controls in vivo. Moreover, combination of the two agents induced apoptosis and depression of the AKT/mTORpathway in both MDS and AML cell culture and animal studies.

The findings demonstrated that combination of tigecycline and homoharringtoninehad synergistic anti-MDS and anti-AML effects.

1. AKT phosphorylation down-regulates phosphorylation of PRAS40, further downregulating protein levels of p-raptor and mTORC1.Down-regulation of AKT phosphorylation down-regulates the mTOR phosphorylation, followed by phosphorylation of 4E-BP1.The unactivated 4E-BP1 binds to eIF4E, enhancing the inhibition of translation.

2. Down-regulation of p-rictor and down-regulation of mTORC2 resulted in down-regulation of phosphorylation of AKT ser473.Down-regulation of AKT phosphorylation down-regulates NF-kB phosphorylation, followed by BCL-2 expression.Tune to promote cell death.

3. Mitochondrial translation was inhibited.The subunit Cox-1 expression of the respiratory complex IV in the electron transport chain encoded by the gene in the mitochondria is down-regulated. The nuclear gene-encoded subunit Cox-4 expression was not affected.

These effects were mainly attributed to the inhibition of mitochondrial translation of AKT/mTORpathway inhibitors and downregulation of anti-apoptotic proteins.

Keywords: Tigecycline, Homoharringtonine, AKT,mTOR,Myelodysplastic syndromes, Acute myeloid leukemia