Novel functional APOB mutations outside LDL-binding region causing familial hypercholesterolaemia

Area of Science:

• Genetics
• Cardiovascular Medicine
• Biochemistry

Background:

• Familial hypercholesterolaemia (FH) is a genetic disorder causing high LDL cholesterol and premature cardiovascular disease.
• While LDLR and PCSK9 mutations are known causes, many FH patients lack identifiable mutations in these genes.
• APOB mutations are less common, with p.R3527Q being the most frequent.

Purpose of the Study:

• To identify the genetic cause of hypercholesterolaemia in 65 patients with suspected FH but no identified mutations in LDLR, PCSK9, or specific APOB exons.
• To investigate the functional impact of newly identified APOB mutations on LDL internalization and cellular growth.

Main Methods:

• Whole sequencing of the APOB gene using pyrosequencing.
• Flow cytometry using fluorescently labeled LDL to assess LDL internalization in lymphocytes and HepG2 cells.
• Proliferation assays with U937 cells.

Main Results:

• Ten putative mutations in the APOB gene were identified.
• APOB variants p.Arg1164Thr and p.Gln4494del showed a 40% decrease in LDL internalization, similar to the known APOB3527 variant.
• These variants also led to reduced U937 cell growth.
• APOB p.Tyr1247Cys was found to be neutral, and three other alterations were classified as polymorphisms.

Conclusions:

• Whole APOB sequencing should be incorporated into routine diagnostic protocols for FH.
• Identifying novel APOB mutations can improve the diagnosis of FH and enhance cardiovascular risk assessment in affected individuals.