Large-scale functional characterization of low-density lipoprotein receptor gene variants improves risk assessment in cardiovascular disease
Authors: Islam Mohammad Majharul , Tamlander Max , Hlushchenko Iryna , Ripatti Samuli , Pfisterer Simon G. 
Published: December 2023

AbstractAimsLack of functional information for low-density lipoprotein (LDL) receptor (LDLR) mutations limits the use of genetics for early diagnosis, risk assessment and clinical decision making in familial hypercholesterolemia (FH). The goal of this study was an in-depth and large-scale functional characterization ofLDLRvariants to overcome this problem.MethodsOpen-source robotic tools were integrated with multiplexed high-content microscopy, image and data analysis into a novel semi-automated analysis pipeline for the characterization ofLDLRvariants to quantify LDL uptake, LDLR localization and expression.Results315LDLRcoding variants were functionally characterized in this study and collapsed into four functional groups based on their residual LDL uptake activity (“Loss-of-function”, 0-10% activity; “defective”, 10-30%; “mildly-defective”, 30-70%; and “non-defective”, > 90%). Integration of the activity groups with whole-exome sequencing and clinical data from UK biobank demonstrated that considering LDLR activity levels improved risk assessment in dyslipidaemia and cardiovascular disease (CVD). Individuals carryingLDLRvariants from the loss-of-function and defective groups displayed increased odds ratios for CVD (OR=6.1, 95% CI = 1.5 - 24.4; OR = 1.83, 95% CI = 1.2 - 2.7) as compared to the non-defective group. Also, plasma LDL-cholesterol, utilization of lipid-lowering drugs and combination therapy were higher in the loss-of-function (OR = 15.4, 95% CI = 3.8 - 61.7; OR = 7.6, 95% CI = 1.8 - 31.8; OR = 96.8, 95% CI = 22.6 - 414.1), defective (OR = 5.9, 95% CI = 4.1 - 8.6; OR = 3.5, 95% CI = 2.5 - 4.9; OR = 15.6, 95% CI = 8.4 - 29.1) and mildly-defective group (OR = 2.0, 95% CI = 1.5 - 2.7; OR = 2.0, 95% CI = 1.6 - 2.4; OR = 1.9, 95% CI = 1.0 - 3.4) as compared to the non-defective group. Especially, the loss-of-function group displayed higher CVD risk, increased LDL-C and combination therapy usage as compared to the ClinVar pathogenic group for the same subjects. Furthermore, the functional data indicates that prediction tools tend to overestimate the fraction of pathogenicLDLRvariants.ConclusionSystematic functional data forLDLRvariants paves the way for improved diagnosis, risk assessment and treatment optimization for FH patients, enabling a better utilization of genetic data in clinical decision making.Translational perspectiveA loss-of-functionLDLRvariant leads to lifelong exposure of elevated LDL-C. Whilst sequencing of theLDLRgene is included in the genetic assessment of FH patients, mostLDLRvariants lack information about functional consequences at the cellular level. This limits the utility of genetic tools in the diagnosis and treatment of FH. This study overcomes this problem, providing functional information for a large set ofLDLRvariants. Integration with genetic and clinical data from UK biobank enables links between functional and clinical effects, making it easier to diagnose FH and estimate a patient’s cardiovascular risk.