Functional characterization of AIP gene variants causing gigantism

Ekaterina Chaban; Sayka Barry; Atrin Hamidzadeh; Vishnou Mourougavelou; Kesson Magid; Tom Rice; Oniz Suleyman; Afiya Andrews; Dias Renuka P.; Goldstone Anthony P.; Marta Korbonits

doi:10.1530/endoabs.109.P210

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Endocrine Abstracts (2025) 109 P210 | DOI: 10.1530/endoabs.109.P210

SFEBES2025 Poster Presentations Neuroendocrinology and Pituitary (48 abstracts)

Functional characterization of AIP gene variants causing gigantism

Ekaterina Chaban ¹ , Sayka Barry ¹ , Atrin Hamidzadeh ¹ , Vishnou Mourougavelou ¹ , Kesson Magid ¹ , Tom Rice ¹ , Oniz Suleyman ¹ , Afiya Andrews ¹ , Renuka P. Dias ² , Anthony P. Goldstone ³ & Márta Korbonits ¹

Author affiliations

¹Centre for Endocrinology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, United Kingdom; ²Department of Diabetes and Endocrinology, Birmingham Children’s Hospital, Birmingham, United Kingdom. Institute of Applied Health Research, University of Birmingham, Birmingham, United Kingdom; ³PsychoNeuroEndocrinology Research Group, Department of Brain Sciences, Faculty of Medicine, Imperial College London, and Dept. of Endocrinology, Hammersmith Hospital, Imperial College Healthcare NHS Trust, London, United Kingdom

Introduction: AIP (Aryl Hydrocarbon Receptor Interacting Protein) predisposes to young-onset growth hormone secreting adenomas. Mutations in the AIP gene are known to be associated with young-onset pituitary tumours. The pathogenicity of some AIP missense variants is unclear, and this is relevant for clinical genetic counselling.

Aim: This study evaluated the protein stability of AIP missense variants identified in patients to determine their pathogenic role.

Method: Site-directed mutagenesis performed using Agilent’s QuikChange Kit. Functional evaluation of the AIP variants was done by cycloheximide chase assays in HEK293T cells transfected with wild type (WT) and the five AIP variants and subsequent immunoblotting. The results were analysed using a one-phase decay equation and the degradation constants of the AIP variants were compared to the WT AIP. We applied ACMG scores (pathogenic (P), likely pathogenic (LP), variant of uncertain significance (VUS), likely benign (LB), benign (B)) and Revel scores (suggesting pathogenic above 0.7). AIP variants were also analysed via computational protein stability prediction tools such as DDMut, mCSM and DUET.

Results: The results of the half-life experiments for five AIP variants showed that variant c.38T>A (p.I13N) and c.47G>C (p.R16P) have a shorter half-life, while variant c.68G>A (p.G23E) have intermediate and both c.325G>A (p.A109T) and c.491A>G (p.Q164R) variants resembled wild type with long half-lives. In silico predictions for all five variants (p.I13N ACMG:VUS, Revel 0.936), p.G23E (ACMG:B, Revel:0.816), p.A109T (ACMG:LB, Revel:0.457), p.Q164R (ACMG:LB, Revel:0.265) and p.R16P (ACMG:VUS, Revel:0.744)) were consistent with the results of the half-life experiment. The variants p.I13N and p.R16P, with shorter half-lives, have significantly higher degradation rates than WT and are predicted to affect protein stability by in silico prediction tools.

Conclusion: The phenotype of the patients, half-life data and in silico predictions confirm that the variants p.I13N and p.R16P are most likely to be pathogenic.