Antisense oligonucleotides targeting hepatic angiotensinogen reduce atherosclerosis and liver steatosis in hypercholesterolemic mice

Abstract

Hepatocyte-derived angiotensinogen (AGT) is the precursor of angiotensin II (AngII). We determined the effects of hepatocyte-specific (N-acetylgalactosamine-conjugated) antisense oligonucleotides targeting AGT (GalNAc AGT ASO) on AngII-mediated blood pressure (BP) regulation and atherosclerosis and compared its effects with losartan, an AngII type 1 (AT1) receptor blocker, in hypercholesterolemic mice. Eight-week-old male low-density lipoprotein (LDL) receptor deficient mice were administered vehicle or GalNAc AGT ASO (1, 2.5, or 5 mg/kg) subcutaneously beginning 2 weeks before the initiation of Western diet feeding. All mice were fed Western diet for 12 weeks. Their systolic BP was monitored by the tail-cuff technique, and the atherosclerotic lesion area was measured by an en face method. Although the effects of all 3 doses of GalNAc AGT ASO on plasma AGT concentrations were similar, GalNAc AGT ASO reduced BP and atherosclerotic lesion size in a dose-dependent manner. Subsequently, we compared the effects of GalNAc AGT ASO (5 mg/kg) with losartan (15 mg/kg/day). Compared to losartan, GalNAc AGT ASO led to more profound increases in plasma renin and reduction in BP but had similar effects on atherosclerosis. Remarkably, GalNAc AGT ASO also reduced liver steatosis, which was not observed in losartan-treated mice. In conclusion, the BP increase and atherosclerosis development in hypercholesterolemic mice are dependent on AngII generated from hepatic AGT. Deleting hepatic AGT improves diet-induced liver steatosis, and this occurs in an AT1 receptor-independent manner.

Angiotensinogen as a Therapeutic Target for Cardiovascular and Metabolic Diseases

“…Inhibition of renin would have an equivalent effect of AGT deletion on diminishing synthesis of all angiotensin peptides while retaining AGT in plasma, while inhibition of AT1 receptors blocks AngII-AT1 receptor interaction and their downstream signaling. 46,65 A second line of evidence for AngII-independent effects was demonstrated in hepatocyte-specific AGT-deleted mice in which plasma concentrations of AGT were restored by infection with adenoassociated viruses expressing either Agt or des(AngI)Agt . Expression of both forms of AGT resulted in equivalent restoration of body weight gain and liver steatosis.…”

“…3,4 Unexpectedly, these studies demonstrated profound effects of liver-derived AGT deletion on the development of Western diet-induced body weight gain and liver steatosis in normocholesterolemic or hypercholesterolemic mice. 3,46,65 These effects are not mimicked by pharmacological inhibition of AT1 receptors, ACE, or renin, and hence suggest that liver-derived AGT exerts effects that are independent of reduction in AngII production. 3,46,65 Demonstration of liver-derived AGT's ability to contribute to disease has facilitated interest in inhibiting AGT synthesis using ASO or small-interfering RNA (siRNA).…”

“…3,46,65 These effects are not mimicked by pharmacological inhibition of AT1 receptors, ACE, or renin, and hence suggest that liver-derived AGT exerts effects that are independent of reduction in AngII production. 3,46,65 Demonstration of liver-derived AGT's ability to contribute to disease has facilitated interest in inhibiting AGT synthesis using ASO or small-interfering RNA (siRNA). While ASO and siRNA therapies predominately target the liver as an inherent characteristic of the technology, liver specificity can be enhanced by the addition of a triantennary N-acetylgalactosamine (GalNAc) moiety that enables hepatocytes to exclusively endocytose the drug.…”

“…There were 2 indications that reductions in obesity and liver steatosis occurred via an AngII-independent mechanism. One is that administration of either aliskiren (a renin inhibitor) 3 or losartan (an AT1 receptor blocker), 46,65 at doses that cause equivalent reductions of blood pressure in hepatocyte-specific Agt -deleted mice, had no discernable reductions in body weight gain, liver weight, or liver triglyceride concentrations. Inhibition of renin would have an equivalent effect of AGT deletion on diminishing synthesis of all angiotensin peptides while retaining AGT in plasma, while inhibition of AT1 receptors blocks AngII-AT1 receptor interaction and their downstream signaling.…”

“…3,46,65 These effects are not mimicked by pharmacological inhibition of AT1 receptors, ACE, or renin, and hence suggest that liver-derived AGT exerts effects that are independent of reduction in AngII production. 3,46,65…”