Serban Radian

FTO gene associates to metabolic syndrome in women with polycystic ovary syndrome

The FTO (Fat mass and obesity associated) locus has recently been associated with obesity and type 2 diabetes (T2D) in humans. To understand the role of the FTO gene in polycystic ovary syndrome (PCOS) we genotyped single nucleotide polymorphism (SNP) rs1421085 (C/T) in women with PCOS (n=207) and controls (n=100) from a Central European population. The homozygous C/C genotype showed increased prevalence in PCOS patients either obese or with metabolic syndrome (MetS) compared to lean PCOS patients or controls (27.6%, 38.9%, 22.3%, and 16.3%, respectively). In logistic regression, this genotype strongly associated with MetS (P<0.0001, OR 3.2, 95% CI 1.8-5.7) and impaired fasting glucose (IFG) with P<0.0007, OR 7.7, 95% CI 2.1-28.6, independently of BMI or age, and to AUC(gluc) during OGTT (P<0.0001, alpha=0.99), indicating an influential role of the FTO gene in the glucose intolerance component of MetS.

Increased Population Risk of AIP-related Acromegaly and Gigantism in Ireland.

The aryl hydrocarbon receptor interacting protein (AIP) founder mutation R304* (or p.R304*; NM_003977.3:c.910C>T, p.Arg304Ter) identified in Northern Ireland (NI) predisposes to acromegaly/gigantism; its population health impact remains unexplored. We measured R304* carrier frequency in 936 Mid Ulster, 1,000 Greater Belfast (both in NI) and 2,094 Republic of Ireland (ROI) volunteers and in 116 NI or ROI acromegaly/gigantism patients. Carrier frequencies were 0.0064 in Mid Ulster (95%CI = 0.0027–0.013; P = 0.0005 vs. ROI), 0.001 in Greater Belfast (0.00011–0.0047) and zero in ROI (0–0.0014). R304* prevalence was elevated in acromegaly/gigantism patients in NI (11/87, 12.6%, P < 0.05), but not in ROI (2/29, 6.8%) versus non‐Irish patients (0–2.41%). Haploblock conservation supported a common ancestor for all the 18 identified Irish pedigrees (81 carriers, 30 affected). Time to most recent common ancestor (tMRCA) was 2550 (1,275–5,000) years. tMRCA‐based simulations predicted 432 (90–5,175) current carriers, including 86 affected (18–1,035) for 20% penetrance. In conclusion, R304* is frequent in Mid Ulster, resulting in numerous acromegaly/gigantism cases. tMRCA is consistent with historical/folklore accounts of Irish giants. Forward simulations predict many undetected carriers; geographically targeted population screening improves asymptomatic carrier identification, complementing clinical testing of patients/relatives. We generated disease awareness locally, necessary for early diagnosis and improved outcomes of AIP‐related disease.

Association of insulin receptor genetic variants with polycystic ovary syndrome in a population of women from Central Europe

To assess the role of the insulin receptor gene in polycystic ovary syndrome (PCOS) we performed a case-control study in a female population (n=226) from Central Europe by examining the genetic associations of single nucleotide polymorphisms (rs8107575, rs2245648, rs2245649, rs2963, rs2245655, and rs2962) and inferred haplotypes around exon 9 of this gene. The ancestral T allele of single nucleotide polymorphism rs2963 or the corresponding haplotype (GGTC-C) showed association with PCOS with odds ratio 2.99, 95% confidence interval 1.4-6.3, independent of obesity but related to the presence of Acanthosis nigricans and insulin resistance, metabolic syndrome, or hyperandrogeny, thus providing a frame for future fine mapping of the susceptibility loci in PCOS.

In-frame seven amino-acid duplication in AIP arose over the last 3000 years, disrupts protein interaction and stability and is associated with gigantism.

Objective Mutations in the aryl hydrocarbon receptor-interacting protein (AIP) gene are associated with pituitary adenoma, acromegaly and gigantism. Identical alleles in unrelated pedigrees could be inherited from a common ancestor or result from recurrent mutation events. Design and methods Observational, inferential and experimental study, including: AIP mutation testing; reconstruction of 14 AIP-region (8.3 Mbp) haplotypes; coalescent-based approximate Bayesian estimation of the time to most recent common ancestor (tMRCA) of the derived allele; forward population simulations to estimate current number of allele carriers; proposal of mutation mechanism; protein structure predictions; co-immunoprecipitation and cycloheximide chase experiments. Results Nine European-origin, unrelated c.805_825dup-positive pedigrees (four familial, five sporadic from the UK, USA and France) included 16 affected (nine gigantism/four acromegaly/two non-functioning pituitary adenoma patients and one prospectively diagnosed acromegaly patient) and nine unaffected carriers. All pedigrees shared a 2.79 Mbp haploblock around AIP with additional haploblocks privately shared between subsets of the pedigrees, indicating the existence of an evolutionarily recent common ancestor, the ‘English founder’, with an estimated median tMRCA of 47 generations (corresponding to 1175 years) with a confidence interval (9–113 generations, equivalent to 225–2825 years). The mutation occurred in a small tandem repeat region predisposed to slipped strand mispairing. The resulting seven amino-acid duplication disrupts interaction with HSP90 and leads to a marked reduction in protein stability. Conclusions The c.805_825dup allele, originating from a common ancestor, associates with a severe clinical phenotype and a high frequency of gigantism. The mutation is likely to be the result of slipped strand mispairing and affects protein–protein interactions and AIP protein stability.

In vivo bioassay to test the pathogenicity of missense human AIP variants

Background Heterozygous germline loss-of-function mutations in the aryl hydrocarbon receptor-interacting protein gene (AIP) predispose to childhood-onset pituitary tumours. The pathogenicity of missense variants may pose difficulties for genetic counselling and family follow-up. Objective To develop an in vivo system to test the pathogenicity of human AIP mutations using the fruit fly Drosophila melanogaster. Methods We generated a null mutant of the Drosophila AIP orthologue, CG1847, a gene located on the Xchromosome, which displayed lethality at larval stage in hemizygous knockout male mutants (CG1847exon1_3 ). We tested human missense variants of ‘unknown significance’, with ‘pathogenic’ variants as positive control. Results We found that human AIP can functionally substitute for CG1847, as heterologous overexpression of human AIP rescued male CG1847exon1_3 lethality, while a truncated version of AIP did not restore viability. Flies harbouring patient-specific missense AIP variants (p.C238Y, p.I13N, p.W73R and p.G272D) failed to rescue CG1847exon1_3 mutants, while seven variants (p.R16H, p.Q164R, p.E293V, p.A299V, p.R304Q, p.R314W and p.R325Q) showed rescue, supporting a non-pathogenic role for these latter variants corresponding to prevalence and clinical data. Conclusion Our in vivo model represents a valuable tool to characterise putative disease-causing human AIP variants and assist the genetic counselling and management of families carrying AIP variants.

Dense mapping of the region of insulin gene VNTR in polycystic ovary syndrome in a population of women from Central Europe

INTRODUCTION Insulin gene VNTR was associated with polycystic ovary syndrome (PCOS) in some studies but not in others. This couldb be due to the heterogeneity of the definition of PCOS and/or the use of inappropriate gene mapping strategies. MATERIAL AND METHODS In this investigation, the association of VNTR with PCOS was explored in a population of women from Central Europe (377 cases and 105 controls) in whom PCOS was diagnosed according to Rotterdam criteria. Seven SNPs: rs3842756 (G/A), rs3842755 (G/T), rs3842754 (C/T), rs3842753 (A/C), rs3842752 (C/T), rs3842748 (G/C), and rs689 (T/A) were genotyped in a portion of the population (160 cases and 95 controls) by sequencing or by SSO-PCR. Analysis of linkage disequilibrium (LD) pattern allowed selecting three tagSNPs (rs3842754, rs3842748, and rs689), which were genotyped in the rest of the population by KASPar. RESULTS Six haplotypes were reconstructed, among which three (h1, h2 and h6) were more frequent. Statistical analysis allowed observation of the association of the SNP rs3842748, through its GC genotype, with obesity in PCOS (P = 0.049; OR CI95% 1,59 [1.00-2.51]) and in classical PCOS (YPCOS) (P = 0.010), as well as the correlation of the SNP rs689 and the pair of haplotypes h1/h1 with higher levels of testosteronaemia in the PCOS group, although this was at the limit of significance (P = 0.054) CONCLUSION: These results are in accordance with some studies in literature and highlight the role of insulin gene VNTR in complex metabolic disorders.

Acromegaly treatment in Romania. How close are we to disease control

INTRODUCTION In Romania, no nationwide data for acromegaly treatment and control rate are available. Our objective was to assess the acromegaly control rate in a tertiary referral centre, which covers an important part of Romanian territory and population of patients with acromegaly. MATERIALS AND METHODS We reviewed the records of all 164 patients (49 males and 115 females; median age 55 [47, 63.5] years) with newly or previously diagnosed acromegaly, who have been assessed at least once in our tertiary referral centre between January 1, 2012 and March 31, 2016. This sample represents 13.6% of the total expected 1200 Romanian patients with acromegaly and covers 82.9% of the counties in Romania. Control of acromegaly was defined as a random serum growth hormone (GH) < 1 ng/mL and an age-normalised serum insulin-like growth factor-I (IGF-I) value. The GH and IGF-I values used for calculation of the control rate were those at the last evaluation. The same assays for GH and IGF-I measurement were used in all patients. RESULTS There were 147 treated and 17 untreated patients. Of the 147 patients assessed after therapy, 137 (93.2%) had pituitary surgery, 116 (78.9%) were on medical treatment at the last evaluation, and 67 (45.5%) had radiotherapy. Seventy-one (48.3%) had a random GH < 1 ng/mL, 54 (36.7%) had a normalised, age-adjusted IGF-I, and 42 (28.6%) had both normal random serum GH and IGF-I. CONCLUSIONS In Romania, acromegaly benefits from the whole spectrum of therapeutic interventions. However, the control rate remains disappointing.

Creation of a locus-specific database for AIP mutations

response to the increasing number of sequence variations reported in the human genome and the consequent necessity to establish their clinical significance. Several genes causing endocrine syndromes have LSDBs available (e.g. MEN1, VHL, RET, GNAS, PRKAR1A, the SDH subunits). Germline mutations in the AIP gene are found in about 20% of familial isolated pituitary adenoma (FIPA) patients. The majority of these variants are of unknown pathogenicity.