A novel Gly436Glu variant in the LPL gene identified in a Saudi Arabian patient with severe hypertriglyceridemia and recurrent pancreatitis
Article information
Abstract
Familial chylomicronemia syndrome (FCS) is a rare autosomal recessive disorder characterized by severe hypertriglyceridemia and recurrent pancreatitis, often manifesting in childhood. The condition results from variants in the lipoprotein lipase (LPL) gene, which lead to impaired fat metabolism. We report the case of an 18-year-old Saudi male with a lifelong history of hypertriglyceridemia and recurrent episodes of pancreatitis. Laboratory investigations revealed severe hypertriglyceridemia and low high-density lipoprotein cholesterol, consistent with FCS. A comprehensive evaluation excluded secondary causes of hyperlipidemia, suggesting a potential genetic basis for the condition. Whole-exome sequencing identified a novel homozygous missense variant (c.1307G> A; p.Gly436Glu) in the LPL gene. Bioinformatics analysis predicted this variant to be deleterious, potentially disrupting the structure and stability of the LPL enzyme and impairing its ability to hydrolyze dietary fats. This finding suggested a causal link between the variant and the patient’s FCS phenotype. This case highlights the importance of molecular diagnosis in FCS, enabling the identification of causative genetic alterations and improving our understanding of the link between LPL dysfunction and severe metabolic disorders.
INTRODUCTION
The human lipoprotein lipase (LPL) gene, located on chromosome 8p22, spans approximately 30 kB and comprises 10 exons. It encodes the LPL protein, a 475-amino acid enzyme expressed in adipose tissue and muscles. This enzyme hydrolyzes triglyceride (TG)-rich lipoproteins, including chylomicrons and very low-density lipoprotein cholesterol (LDL-C), playing a vital role in TG metabolism [1]. Loss-of-function variants in LPL lead to chylomicron accumulation and subsequent hypertriglyceridemia (HTG) due to LPL deficiency (LPLD). Severe HTG, with TG levels > 11.4 mmol/L, often indicates an underlying genetic cause and can lead to acute pancreatitis. Familial chylomicronemia syndrome (FCS), or type I hyperlipoproteinemia, is a rare autosomal recessive disorder affecting 1–10 per million individuals [2]. Approximately 80% of FCS cases result from biallelic loss-of-function variants in the LPL gene, whereas others arise from variants that affect LPL function [2]. FCS typically presents during infancy or adolescence with symptoms such as severe HTG, recurrent acute pancreatitis, failure to thrive, eruptive xanthomas, lipemia retinalis, and hepatosplenomegaly [2,3]. Long-term management includes a fat-restricted diet to maintain TG levels < 11.4 mmol/L, which is the threshold for pancreatitis [2]. Saudi guidelines recommend fibrates and eicosapentaenoic acid ethyl ester for severe HTG ( > 5.65 mmol/L) [4]. However, fibrates and niacin show limited efficacy in FCS as they do not reduce chylomicrons [2]. Gene therapy with alipogene tiparvovec is available for LPLD, but it is only suitable for patients with a confirmed genetic diagnosis. Consequently, molecular diagnosis of FCS is crucial, emphasizing the importance of genetic testing in clinical practice. In this report, we described a novel LPL gene missense variant in a Saudi Arabian patient with severe HTG and recurrent pancreatitis.
CASE REPORT
Case presentation and pedigree
We followed the Case Report (CARE) checklist while writing this report [5]. The patient was an 18-year-old Saudi male with a known history of HTG since childhood. He had a long history of recurrent non-biliary pancreatitis secondary to HTG, resulting in multiple hospital admissions. These episodes occurred every 2 to 3 years since birth and were managed conservatively with pancreatic rest through nil per oral and the administration of anti-lipid medications. The pedigree of the probands is shown in Fig. 1.
A pedigree of the patient’s family. The double line represents parental consanguinity.
This study was approved (IRB No. 06/21) by the ethics committee of Alborg Diagnostics, Saudi Arabia. Informed consent was obtained from the patient and his family members prior to genetic testing.
Lab investigations and course of admissions
The patient was initially diagnosed with hyperlipidemia 20 days after birth during hospitalization for fever, reduced activity, and decreased oral intake, with an unremarkable physical examination. The chemistry and hormonal analysis was conducted using the Atellica system. The reagents employed included both colourimetric and enzymatic solutions. A homogeneous enzymatic reagents were utilized for the analysis of cholesterol, high-density lipoprotein cholesterol (HDL-C), LDL-C, and TG. For liver enzymes, ultraviolet (UV)-based kinetic reagents were applied. Hemoglobin A1c (HbA1c) measurements were performed using a Bio-Rad assay, for both control and calibration. Complete blood count analysis was carried out using the Sysmex system. Laboratory investigations performed at that time revealed a lipemic sample. Results showed high total cholesterol (TC) (34.5 mmol/L) and TG (> 35 mmol/L), low HDL-C (0.1 mmol/L), and normal LDL-C ( < 0.1 mmol/L). His presentation was accompanied by severe anemia (hemoglobin, 6 g/dL) and thrombocytosis (platelet count, 706,000/μL), while lipase (10 IU/L) and alanine transaminase (25 U/L) levels were within normal ranges. The patient was treated with a blood transfusion and gemfibrozil.
Subsequently, the patient was followed-up every 1 to 2 years. Severe HTG was evident throughout his life, fluctuating between 9.4 and 48.5 mmol/L (Table 1). The latest episode of acute pancreatitis led to hospitalization at the age of 18 years. Laboratory results included high TC (5 mmol/L) and TG (19.9 mmol/L), low HDL-C (0.2 mmol/L), and normal LDL-C (0.2 mmol/L). He was treated with atorvastatin (40 mg), ezetimibe (10 mg), omega-3, fenofibrate, insulin-dextrose infusion, and intravenous fluid therapy, after which his symptoms resolved, and he was discharged in good condition. Secondary causes of hyperlipidemia were ruled out based on normal laboratory investigations (HbA1c, thyroid-stimulating hormone, urea and electrolytes, liver function test).
Blood lipid profile of the familial chylomicronemia syndrome patient since birth
Physical examination revealed soft and lax epigastric and umbilical areas, with no evidence of tendon xanthomas, corneal ulcers, or xanthelasmas. Respiratory and neurological examinations were unremarkable. Fig. 2 illustrates a computed tomography scan of the abdomen and pelvis showing the characteristic features of acute pancreatitis. It revealed pan-creatic and peripancreatic edema and a non-enhancing area in the neck and body of the pancreas, accompanied by peripancreatic and epigastric fluid accumulation and mesenteric fat stranding. No evidence of biliary or pancreatic ductal dilatation, cyst formation, necrosis, or vascular complications was observed. The gallbladder was neither calculable nor inflamed. The liver, spleen, bowel loops, kidneys, and adrenal glands were unremarkable, in addition to borderline splenomegaly. Considering the severity of HTG, the patient was referred to the Lipid Clinic at King Abdulaziz University Hospital for further investigation and a diagnosis of familial chylomicronemia (LPLD) was considered.
Key radiological features of acute pancreatitis. (A) 1. Pancreatic oedema; 2. Peripancreatic fluid and fat stranding. (B) 3. A hypoenhancing area in the pancreatic neck indicating reduced perfusion or necrosis; 4. Perigastric fluid accumulation.
Molecular investigation
After obtaining written informed consent, blood samples were collected in ethylenediaminetetraacetic acid (EDTA) tubes, and genomic DNA was extracted. The exonic regions of over 20,000 genes and exon-intron boundaries (±15 nucleotides) were enriched using the Agilent SureSelect V6 kit (Agilent Technologies). Whole-exome sequencing (WES) was performed using an Illumina NextSeq sequencer, achieving an average coverage depth of 100–130X. Raw sequencing data were aligned to the hg19 genome assembly, and variant calling and annotation were performed using commercial bioinformatics tools. Variants with poor quality or high population frequency (> 1.5%, unless pathogenic) were excluded. The remaining variants were assessed following American College of Medical Genetics and Genomics (ACMG) guidelines.
WES identified homozygous LPL gene variants: LPL (NM_000 237.3): c.1307G>A (p.Gly436Glu), Chr8(GRCh37): g.19818579G>A, database of single nucleotide polymorphisms (dbSNP): rs207 0034186. This substitution resulted in the replacement of glycine with glutamic acid at position 436 (Fig. 3A). This variant has not been reported in published literature and also in public genetic databases like Genome Aggregation Database (gnomAD), Exome Aggregation Consortium (ExAC), 1,000 genomes, and dbSNP. However, ClinVar database contains an entry for this variant with the “variant of uncertain significance (VUS)” classification (Variation ID: 986276), but no details about phenotype were mentioned. The computational biology prediction methods, like Combined Annotation Dependent Depletion (CADD) predicted the score of 29.4, additionally 16 other pathogenicity prediction algorithms developed to predict the effect of missense changes (BayesDel, MetaLR, Met-aSVM, PolyPhen-2, EIGEN, MutationTaster, FATHMM-MKL, MetaRNN, REVEL, M-CAP, LRT, Mutation Assessor, MVP, MutPred, PROVEAN, and SIFT) have also suggested that this variant is deleterious; likely to disrupt the function of the LPL protein. As per ACMG guidelines, this variant is classified as ‘likely pathogenic’ based on ACMG criteria, PM2 (moderate) for extremely low population frequency, PM5 (moderate) for a different pathogenic missense change at the same position, PP3 (supporting) for unanimous deleterious predictions by in silico tools, and PP2 (supporting) for occurring in a gene where missense variants are a common disease mechanism.
Analysis of a novel Gly436Glu mutation in the lipoprotein lipase (LPL) gene. (A) Electropherogram showing the c.1307G>A missense mutation leading to a Gly436Glu amino acid substitution. (B) Secondary structure analysis of the LPL protein, highlighting alterations in loop length and sheet extensions at positions Cys445 and Leu453 (red arrow). (C, D) Three-dimensional structural models of wild-type (C) and mutant (D) LPL proteins generated using AlphaFold (DeepMind Technologies), demonstrating significant structural deviation at the mutation site. Root mean square deviation analysis reveals an alteration of 2.20 Å.
Structural analysis using bioinformatic tools revealed localized changes in the LPL protein. NetSurfP secondary structure analysis revealed alterations in loop lengths and sheet extensions near residues Cys445 and Leu453 (Fig. 3B). AlphaFold modeling (DeepMind Technologies) and root mean square deviation (RMSD) analysis revealed structural deviations at the variant site (RMSD= 2.20 Å) (Fig. 3C, D). Thermodynamic stability assessment with Design and Understanding of Engineered Thermostability (DUET) predicted destabilization (ΔΔ G= ‒2.295 kcal/mol), suggesting impaired protein stability and function.
DISCUSSION
LPLD is a metabolic disorder characterized by the defective hydrolysis of dietary fats and impaired plasma clearance of chylomicrons, leading to severe HTG. Most patients with LPLD present clinical symptoms within the first decade of life, with only 25% manifesting in the neonatal period [6]. Currently, 490 variants in the LPL gene have been reported in the Human Gene Mutation Database (HGMD) (www.hgmd.cf.ac.uk). In this report, we describe a novel variant (Gly436Glu; rs20700 34186) in a Saudi Arabian patient with severe HTG and recurrent pancreatitis caused by LPLD. This variant occurs in the eighth exon of the LPL gene, resulting in a protein with different amino acid sequence but with preserved length.
The Gly436Glu variant in LPL highlights an intricate connection between genetic variants and metabolic dysfunctions. Structural analyses including RMSD deviations, secondary structure changes, and thermodynamic stability provided comprehensive insights into the functional impact of this variant. The RMSD value of 2.20 Å at the variant site indicated significant local deviations caused by the substitution of a small glycine with a bulkier glutamic acid. This substitution alters the spatial arrangement, potentially impairing the interaction of the enzyme with lipid substrates and its catalytic activity, which are essential for TG hydrolysis. Similar structural modifications have been linked to changes in enzyme kinetics and substrate affinity [7].
Although the overall secondary structure remained unchanged, localized alterations at the key residues Cys445 and Leu453, which affect secondary elements, could disrupt the regulatory and substrate-binding functions of the enzyme. The predicted destabilizing effect (ΔΔG= –2.295 kcal/mol) underscores decreased mutant protein stability, potentially causing misfolding or degradation. Previous studies have demonstrated how disruption of protein stability influences functional efficacy, disease progression, and evolutionary mechanisms [8].
This destabilization directly affects the three-dimensional structure of the LPL enzyme, especially around its active site, thereby reducing its affinity for TG-rich lipoproteins. Consequently, LPL’s enzymatic activity—which hydrolyze TGs to free fatty acids and glycerol—is compromised. TG accumulation in the bloodstream leads to severe HTG, which increases the risk of acute pancreatitis. Pancreatic lipases hydrolyze elevated TGs, release cytotoxic free fatty acids, triggering inflammation [9,10].
These structural and functional disruptions align with the clinical manifestations, including recurrent pancreatitis, emphasizing the critical role of LPL in lipid metabolism. The bioinformatic findings further validated genetic studies linking similar variants to lipid metabolism disorders, thus reinforcing the disease mechanism associated with the LPL Gly436Glu variant.
This novel finding permits future research to explore targeted therapies for this variant that could potentially prevent HTG. Additionally, this could offer a personalized approach for managing lipid disorders tailored to the genetic makeup of individuals. However, rigorous randomized controlled trials are necessary to translate these findings into effective therapeutic strategies.
In conclusion, we identified a novel LPL variant (Gly436Glu) in a patient with severe HTG and pancreatitis. Our analysis suggests that this variant disrupts the structure of the enzyme, impairing its ability to hydrolyze fats. This impairment leads to HTG and increased risk of pancreatitis. This study highlights the link between LPL variants and metabolic disorders and provides insights into disease mechanisms. These findings support the established role of protein structure in enzyme function and strengthen our understanding of LPLD. This information will inform future research on therapeutic strategies for patients with similar LPL variants.
Notes
No potential conflict of interest relevant to this article was reported.
AUTHOR CONTRIBUTIONS
Conception or design: ZAA.
Acquisition, analysis, or interpretation of data: DAN, HD, NAS.
Drafting the work or revising: DAN, NAS, ZAA.
Final approval of the manuscript: DAN, HD, NAS, ZAA.