Gene therapy for rhesus monkeys heterozygous for LDL receptor deficiency by balloon catheter hepatic delivery of helper-dependent adenoviral vector

Abstract

Autosomal dominant familial hypercholesterolemia (FH) is a monogenic life-threatening disease. We tested the efficacy of low-density lipoprotein receptor (LDLR) gene therapy using helper-dependent adenoviral vector (HDAd) in a nonhuman primate model of FH, comparing intravenous injection versus intrahepatic arterial injection in the presence of balloon catheter-based hepatic venous occlusion. Rhesus monkeys heterozygous for mutant LDLR gene (LDLR+/−) developed hypercholesterolemia while on a high cholesterol diet. We treated them with HDAd-LDLR either by intravenous delivery, or by catheter-based intra-hepatic artery injection. Intravenous injection of ≤1.1×1012 viral particles (vp)/kg failed to have any effect on plasma cholesterol. Increasing the dose to 5×1012 vp/kg led to a 59% lowering of the plasma cholesterol that lasted for 30 days before it returned to pretreatment levels by day 40. A further increase in dose to 8.4×1012 vp/kg resulted in severe lethal toxicity. In contrast, direct hepatic artery injection following catheter-based hepatic venous occlusion enabled the use of a reduced HDAd-LDLR dose of 1×1012 vp/kg that lowered plasma cholesterol within a week, and reached a nadir of 59% pretreatment level on days 20 to 48 after injection. Serum alanine aminotransaminase (ALT) remained normal until day 48 when it went up slightly and stayed mildly elevated on day 72 before it returned to normal on day 90. In this monkey, the HDAd-LDLR-induced trough of hypocholesterolemia started trending upwards on day 72 and returned to pretreatment levels on day 120. We measured the LDL apolipoprotein B turnover rate at 10 days before, and again 79 days after, HDAd-LDLR treatment in two monkeys that exhibited a cholesterol lowering response. HDAd-LDLR therapy increased the LDL fractional catabolic rate by 78% and 50%, respectively, in the two monkeys, coincident with an increase in hepatic LDLR mRNA expression. In conclusion, HDAd-mediated LDLR gene delivery to the liver using a balloon catheter occlusion procedure is effective in reversing hypercholesterolemia in a nonhuman primate FH model; however, the unsustainability of the hypocholesterolemic response during 3–4 months of follow up and heterogeneous response to the treatment remains a challenge.

Management of patients with familial hypercholesterolaemia

“…This approach was successful in decreasing LDL-C levels, but the cholesterol-lowering response during 3-4 months of follow-up was not sustained, and the heterogeneous response to treatment observed showed little promise. 105 …”

Review of the scientific evolution of gene therapy for the treatment of homozygous familial hypercholesterolaemia: past, present and future perspectives

“…Nevertheless, not all forms of gene therapy tried in the past were efficacious. Next, we will review the scientific evolution of gene therapy for HoFH from the initial approaches using ex vivo transduction with retroviruses in the 1990s 44 and the first clinical trial based in this technology in 1995 45 to the in vivo adenoviral [46][47][48][49][50] and helper-dependent adenoviral (HD-Ad) transduction [51][52][53] and the development of the adeno-associated virus (AAV) vectors [54][55][56][57][58][59][60] that have allowed the current clinical trial in development since 2016 ( figure 3). The limitations of these strategies and their replacement by advanced approaches will also be discussed.…”

“…52 However, although LDLR overexpression by HD-Ad in rhesus macaques heterozygous for a nonsense mutation in the LDLR gene (LDLR +/-) effectively reversed hypercholesterolaemia, the heterogeneous and unsustained long-term response precludes its future application. 53…”

Helper-dependent adenovirus-mediated gene transfer of a secreted LDL receptor/transferrin chimeric protein reduces aortic atherosclerosis in LDL receptor-deficient mice

“…The harvested cells were lysed with three freezethaw cycles and incubated in 2 M MgCl 2 and DNase I for 1 h at 37°C. Subsequently, cellular debris were spun down and lysate was subjected to ultracentrifugation, as described elsewhere [14]. Vector concentration was measured as virus particle number determining absorbance at 260 nm.…”

“…Significant progresses in gene transfer technology have encouraged to develop LDLR gene therapy approaches for FH treatment [9][10][11][12]. In experimental animal models of FH, VLDL receptor overexpression following viral vector-based gene transfer is associated with long-term stable correction of hyperlipidemia, with consequent attenuation of atherosclerosis progression and in certain cases even with lesions regression [13,14]. We also observed that helper-dependent adenoviral (HD-Ad)-mediated human apolipoprotein A-I (hApoA-I) overexpression reduces aortic atherosclerosis development in ApoE-and LDLR-deficient mice [15,16].…”