Novel efficacious microRNA-30c analogs reduce apolipoprotein B secretion in human hepatoma and primary hepatocyte cells

Pradeep Kumar Yadav,Phensinee Haruehanroengra,Sara Irani,Ting Wang,Abulaish Ansari,Jia Sheng,M Mahmood Hussain

doi:10.1016/j.jbc.2022.101813

Abstract

High plasma lipid levels have been demonstrated to increase cardiovascular disease risk. Despite advances in treatments to decrease plasma lipids, additional therapeutics are still needed because many people are intolerant or nonresponsive to these therapies. We previously showed that increasing cellular levels of microRNA-30c (miR-30c) using viral vectors or liposomes reduces plasma lipids and atherosclerosis. In this study, we aimed to synthesize potent miR-30c analogs that can be delivered to hepatoma cells without the aid of viral vectors and lipid emulsions. We hypothesized that modification of the passenger strand of miR-30c would increase the stability of miR-30c and augment its delivery to liver cells. Here, we report the successful synthesis of a series of miR-30c analogs by using different chemically modified nucleosides. In these analogs, we left the active sense strand untouched so that its biological activity remained unaltered, and we modified the passenger strand of miR-30c to enhance the stability and uptake of miR-30c by hepatoma cells through phosphorothiorate linkages and the addition of GalNAc. We show that these analogs significantly reduced apolipoprotein B secretion in Huh-7 human hepatoma cells and human primary hepatocytes without affecting apolipoprotein A1 secretion and cellular lipid levels. Our results provide a proof of concept that the passenger strand of miR-30c can be modified to increase its stability and delivery to cells while retaining the potency of the sense strand. We anticipate these miR-30c analogs will be useful in the development of more efficacious analogs for the treatment of hyperlipidemias and cardiovascular diseases.

Highlights

Atherosclerosis, hardening of the arteries after lipid deposition, is the leading cause of morbidity and mortality in the United States and worldwide
We have reported that overexpression of miR-30c significantly reduces microsomal triglyceride transfer protein (MTP) activity, whereas overexpression of its corresponding anti-miR elevates MTP activity in Huh-7 human hepatoma cells and human primary hepatocytes [25–27]
We show that the miR-30c-3p passenger strand can be modified and duplexed with native miR-30c-5p to augment delivery to hepatoma cells and to reduce apolipoprotein B (apoB) secretion without affecting apolipoprotein A1 (apoA1) secretion

Summary

Introduction

Atherosclerosis, hardening of the arteries after lipid deposition, is the leading cause of morbidity and mortality in the United States and worldwide. Inhibitors of hydroxyl-methyl-glutaryl-coenzyme A reductase, lower plasma lipids by increasing the hepatic expression of low-density lipoprotein receptors and decreasing cholesterol synthesis. Several pharmaceutical companies have developed drugs that potently inhibit MTP activity and lower plasma lipids [12, 13]. These drugs increase hepatic lipids and plasma transaminases [14–16]. MicroRNAs (miRs) are endogenous gene products 22 nucleotides in length that regulate gene expression at the posttranscriptional level They interact with the 30-untranslated regions of target mRNAs and decrease protein synthesis by enhancing mRNA degradation and/or interfering with translation [19, 20]. The 30-strands (miR-30c-3p) derived from the two genes are slightly different but are conserved in humans and mice

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