Rare variants in a PCSK9 distal-enhancer as drivers of LDL-cholesterol levels

To validate our findings, we used an advanced cellular model of human induced pluripotent stem cells (hiPSCs) stably expressing a CRISPR-interference system. This system enables, when combined with guide-RNAs, to target regions of interest and silence regulated genes. RESULTS.

We found that the cumulative burden of rare genetic variants in 18 among 53,039 ATAC-seq peaks in human liver, is significantly associated with LDL-cholesterol. We focus our proof-of-concept study on a peak located ~17 kb upstream PCSK9, strongly associated with LDL-cholesterol (p=5.85e-52), apolipoprotein-B (p=3.82e-49) and PCSK9 plasma levels (p=1.15e-67). We showed that this regulatory region is also active in hiPSCs and hiPSC-derived liver organoids.

When transfected with guide-RNAs targeting this region, we showed that CRISPRi-hiPSCs present a hypermethylation of the targeted region which in terms leads to a substantial decrease in PCSK9 gene expression (-48%), intracellular protein (-65%) and secretion (-36%) (p<0.001). CONCLUSIONS. This study identifies a non-coding genetic region upstream PCSK9 where rare variants lead to decreased PCSK9 expression and lowered LDL-cholesterol.

Such findings advocate for the genetic screening of non-coding regulatory regions in patients with unidentified genetic causes of extreme dyslipidemia and open novel perspectives for genetic regulatory therapeutic intervention.