Angiopoietin-like protein 4 (ANGPTL4) is an inhibitor of endothelial lipase (EL) while the ANGPTL4/8 complex has reduced EL-inhibitory activity

Abstract

We previously demonstrated that angiopoietin-like protein 8 (ANGPTL8) forms ANGPTL3/8 and ANGPTL4/8 complexes that increase with feeding to direct fatty acids (FA) toward adipose tissue through differential modulation of lipoprotein lipase (LPL) activity. Each complex correlated inversely with high density lipoprotein cholesterol (HDL) in control subjects. We thus investigated ANGPTL3/8 and ANGPTL4/8 levels in type 2 diabetes patients, who can present with decreased HDL. While ANGPTL3/8 levels in type 2 diabetes patients were similar to those previously observed in normal controls, ANGPTL4/8 levels were roughly twice as high as those in control subjects. Concentrations of ANGPTL3/8 and ANGPTL4/8 in type 2 diabetes patients were inversely correlated with HDL, with the correlation being significant for ANGPTL4/8. We therefore measured the ability of the various ANGPTL proteins and complexes to inhibit endothelial lipase (EL), the enzyme which hydrolyzes phospholipids (PL) in HDL. While confirming ANGPTL3 as an EL inhibitor, we found that ANGPTL4 was a more potent EL inhibitor than ANGPTL3. Interestingly, we observed that while ANGPTL3/8 had increased EL-inhibitory activity compared to ANGPTL3 alone, ANGPTL4/8 exhibited decreased potency in inhibiting EL compared to ANGPTL4 alone. Together, these results show for the first time that ANGPTL4 is a more potent EL inhibitor than ANGPTL3 and suggest a possible reason for why ANGPTL4/8 levels are correlated inversely with HDL.

An anti-ANGPTL3/8 antibody decreases circulating triglycerides by binding to a LPL-inhibitory leucine zipper-like motif

“…In light of our recent observations that ANGPTL3/8 can also inhibit EL activity and do so more potently than ANGPTL3 alone ( 62 ), we performed additional experiments to determine the effect of the anti-ANGPTL3/8 antibody and evinacumab on ANGPTL3/8-mediated inhibition of EL. Interestingly, we observed that the anti-ANGPTL3/8 antibody and evinacumab both comparably blocked ANGPTL3/8-mediated EL inhibition and did so much less completely and potently (IC 50 values of 53 nM and 78 nM, respectively) than they blocked ANGPTL3/8-mediated LPL inhibition ( Fig.…”

“…A human endothelial lipase (EL) activity assay was optimized to allow measurement of the phospholipase A1 activity of EL, which preferentially hydrolyzes phospholipids (PLs) at the sn -1 position and was performed as previously described ( 62 ), with some minor modifications. After overnight incubation of EL-stable expression cells at 37°C, the medium was replaced with 80 μl of medium (OptiMEM, Invitrogen) containing human ANGPTL3/8 complex or human ANGPTL3 at their respective IC 80 concentrations (38 nM or 328 nM) that were each preincubated with increasing concentrations of either the anti-ANGPTL3/8 antibody or evinacumab.…”

The Potential of ANGPTL8 Antagonism to Simultaneously Reduce Triglyceride and Increase HDL-Cholesterol Plasma Levels

“…Future studies to identify human SNPs that result in a complete ANGPTL8 deficiency will further confirm the hypothesis of ANGPTL8-antagonism based therapeutics. Future drug development requires a clear mechanistic understanding of how the ANGPTL3-4-8 system works in regulating EL ( 9 , 15 ), as what it does to LPL ( 2 – 5 ).…”

“…Thus, the hypothesized model ( Figure 1 ) appears to be oversimplified, because it lacks ANGPTL4. The hypothesis holds true, however, whether or not ANGPTL8 increases ANGPTL3's EL-inhibiting activity ( 9 , 15 ), because when ANGPTL3-8 complexes are disrupted, fewer ANGPTL3-8 complexes and more free ANGPTL3 still lead to higher LPL and lower EL activities, respectively.…”

“…Konrad's group recently showed that ANGPTL4 is also a potent EL inhibitor, and that ANGPTL8 reduces ANGPTL4's but increases ANGPTL3's EL-inhibiting activity ( 15 ). Thus, the hypothesized model ( Figure 1 ) appears to be oversimplified, because it lacks ANGPTL4.…”