Novel ABCG5 and ABCG8 Variants in Sitosterolemia: Insights Into Haemolysis, Calcium Dysregulation and Therapeutic...

Sitosterolemia is a rare autosomal recessive lipid metabolic disorder caused by mutations in ABCG5 or ABCG8, leading to pathological accumulation of dietary plant sterols. The condition is clinically heterogeneous, presenting with xanthomas, premature atherosclerosis and haematological abnormalities such as stomatocytosis, haemolytic anaemia and thrombocytopenia, making diagnosis particularly challenging. We investigated four patients with unexplained inherited haemolytic anaemia and a history of recurrent transfusions to identify molecular and functional mechanisms and explore therapeutic options. Haematological work-up, including CBC, peripheral smear, reticulocyte count, EMA binding assay, red cell enzyme activity and HPLC for haemoglobinopathies, was performed, along with biochemical assays for liver, renal and lipid function. Plasma sterol levels were quantified using GC-MS, whereas functional assays included intracellular calcium flux under osmotic stress, red cell density profiling and ROS measurement. Whole-exome sequencing identified homozygous nonsense variants: a novel ABCG8 (p.Trp584∗) in two siblings, ABCG5 (p.Arg243∗) in one patient and ABCG8 (p.Ser129∗) in another. Two patients showed markedly elevated plasma phytosterols, predominantly stigmasterol and β-sitosterol. Ezetimibe therapy (10 mg/day for 6 months) effectively reduced sterol levels but failed to normalise haemoglobin or significantly reduce overhydrated red cell populations. Functional studies demonstrated elevated intracellular Ca²⁺, enhanced influx under stress, increased ROS and a persistent population of stomatocytes. In silico prediction tools confirmed the pathogenicity of the identified variants. Persistent haemolysis despite sterol-lowering therapy suggests irreversible red cell membrane damage due to sterol incorporation, altered fluidity, oxidative stress and calcium dysregulation. Our findings indicate that combining ezetimibe with Ca²⁺-modulating therapy may represent a novel strategy to reduce haemolysis in sitosterolemia.