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Complex effects of inhibiting hepatic apolipoprotein B100 synthesis in humans.

ABSTRACT: Mipomersen is a 20mer antisense oligonucleotide (ASO) that inhibits apolipoprotein B (apoB) synthesis; its low-density lipoprotein (LDL)-lowering effects should therefore result from reduced secretion of very-low-density lipoprotein (VLDL). We enrolled 17 healthy volunteers who received placebo injections weekly for 3 weeks followed by mipomersen weekly for 7 to 9 weeks. Stable isotopes were used after each treatment to determine fractional catabolic rates and production rates of apoB in VLDL, IDL (intermediate-density lipoprotein), and LDL, and of triglycerides in VLDL. Mipomersen significantly reduced apoB in VLDL, IDL, and LDL, which was associated with increases in fractional catabolic rates of VLDL and LDL apoB and reductions in production rates of IDL and LDL apoB. Unexpectedly, the production rates of VLDL apoB and VLDL triglycerides were unaffected. Small interfering RNA-mediated knockdown of apoB expression in human liver cells demonstrated preservation of apoB secretion across a range of apoB synthesis. Titrated ASO knockdown of apoB mRNA in chow-fed mice preserved both apoB and triglyceride secretion. In contrast, titrated ASO knockdown of apoB mRNA in high-fat-fed mice resulted in stepwise reductions in both apoB and triglyceride secretion. Mipomersen lowered all apoB lipoproteins without reducing the production rate of either VLDL apoB or triglyceride. Our human data are consistent with long-standing models of posttranscriptional and posttranslational regulation of apoB secretion and are supported by in vitro and in vivo experiments. Targeting apoB synthesis may lower levels of apoB lipoproteins without necessarily reducing VLDL secretion, thereby lowering the risk of steatosis associated with this therapeutic strategy.

SUBMITTER: Reyes-Soffer G

PROVIDER: S-EPMC4944115 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Complex effects of inhibiting hepatic apolipoprotein B100 synthesis in humans.

Reyes-Soffer Gissette G Moon Byoung B Hernandez-Ono Antonio A Dionizovik-Dimanovski Marija M Jimenez Jhonsua J Obunike Joseph J Thomas Tiffany T Ngai Colleen C Fontanez Nelson N Donovan Daniel S DS Karmally Wahida W Holleran Stephen S Ramakrishnan Rajasekhar R Mittleman Robert S RS Ginsberg Henry N HN

Science translational medicine 20160101 323

Mipomersen is a 20mer antisense oligonucleotide (ASO) that inhibits apolipoprotein B (apoB) synthesis; its low-density lipoprotein (LDL)-lowering effects should therefore result from reduced secretion of very-low-density lipoprotein (VLDL). We enrolled 17 healthy volunteers who received placebo injections weekly for 3 weeks followed by mipomersen weekly for 7 to 9 weeks. Stable isotopes were used after each treatment to determine fractional catabolic rates and production rates of apoB in VLDL, I ...[more]

PMID: 26819195

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Project description:Glucosamine impairs hepatic apolipoprotein B100 (apoB100) production by inducing endoplasmic reticulum (ER) stress and enhancing cotranslational and posttranslational apoB100 degradation (Qiu, W., R. K. Avramoglu, A. C. Rutledge, J. Tsai, and K. Adeli. Mechanisms of glucosamine-induced suppression of the hepatic assembly and secretion of apolipoprotein B-100-containing lipoproteins. J. Lipid Res. 2006. 47: 1749-1761). Here, we report that glucosamine also regulates apoB100 protein synthesis via ER-stress-induced PERK activation. Short-term (4 h) glucosamine treatment of HepG2 cells reduced both cellular (by 62%) and secreted apoB100 (by 43%) without altering apoB100 mRNA. Treatment with proteasomal inhibitors only partially prevented the suppressive effects of glucosamine, suggesting that mechanisms other than proteasomal degradation may also be involved. Glucosamine-induced ER stress was associated with a significantly reduced apoB100 synthesis with no significant change in posttranslational decay rates, suggesting that glucosamine exerted its effect early during apoB biosynthesis. The role of PERK and its substrate, alpha-subunit of eukaryotic initiation factor 2 (eIF2alpha), in the suppressive effects of glucosamine on apoB synthesis was then investigated. Coexpression of apoB15 (normally resistant to intracellular degradation) with wild-type double stranded (ds) RNA activated protein kinase (PKR)-like endoplasmic reticulum kinase (PERK) in COS-7 cells resulted in a dramatic reduction in the levels of newly synthesized apoB15. Interestingly, cotransfection with apoB15 and a kinase inactive PERK mutant (K618A) increased apoB15 expression. In addition, short-term glucosamine treatment stimulated an increase in phosphorylation of PERK and eIF2alpha. Taken together, these data suggest that in addition to the induction of ER-associated degradation and other degradative pathways, ER stress is associated with suppression of apoB synthesis via a PERK-dependent mechanism.

Project description:RATIONALE:Genome-wide association studies identified single-nucleotide polymorphisms near the SORT1 locus strongly associated with decreased plasma LDL-C (low-density lipoprotein cholesterol) levels and protection from atherosclerotic cardiovascular disease and myocardial infarction. The minor allele of the causal SORT1 single-nucleotide polymorphism locus creates a putative C/EBP? (CCAAT/enhancer-binding protein ?)-binding site in the SORT1 promoter, thereby increasing in homozygotes sortilin expression by 12-fold in liver, which is rich in this transcription factor. Our previous studies in mice have showed reductions in plasma LDL-C and its principal protein component, apoB (apolipoprotein B) with increased SORT1 expression, and in vitro studies suggested that sortilin promoted the presecretory lysosomal degradation of apoB associated with the LDL precursor, VLDL (very-low-density lipoprotein). OBJECTIVE:To determine directly that SORT1 overexpression results in apoB degradation and to identify the mechanisms by which this reduces apoB and VLDL secretion by the liver, thereby contributing to understanding the clinical phenotype of lower LDL-C levels. METHODS AND RESULTS:Pulse-chase studies directly established that SORT1 overexpression results in apoB degradation. As noted above, previous work implicated a role for lysosomes in this degradation. Through in vitro and in vivo studies, we now demonstrate that the sortilin-mediated route of apoB to lysosomes is unconventional and intersects with autophagy. Increased expression of sortilin diverts more apoB away from secretion, with both proteins trafficking to the endosomal compartment in vesicles that fuse with autophagosomes to form amphisomes. The amphisomes then merge with lysosomes. Furthermore, we show that sortilin itself is a regulator of autophagy and that its activity is scaled to the level of apoB synthesis. CONCLUSIONS:These results strongly suggest that an unconventional lysosomal targeting process dependent on autophagy degrades apoB that was diverted from the secretory pathway by sortilin and provides a mechanism contributing to the reduced LDL-C found in individuals with SORT1 overexpression.

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Complex effects of inhibiting hepatic apolipoprotein B100 synthesis in humans.

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Complex effects of inhibiting hepatic apolipoprotein B100 synthesis in humans.

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