An atypical Aymé‐Gripp phenotype detected by exome sequencing

Aymé‐Gripp Syndrome (AGS) is an ultra‐rare syndrome characterized by peculiar facial traits combined with early bilateral cataracts, sensorineural hearing loss, and variable neurodevelopmental abnormalities. Only a few cases carrying a pathogenic variant in MAF have been described to date. A significant effort is then required to expand the genotypic and phenotypic spectrum of this condition. In this paper, we report the peculiar case of a 6‐year‐old girl carrying a de novo missense pathogenic variant in MAF, being the first case reported to show a milder phenotype with no cataracts and deafness displayed. Furthermore, we performed a systematic review of previously published cases, focusing on clinical manifestation and genotype.


Patients sharing similar characteristics have been collected by
Fine and Lubinsky since 1983 (Amudhavalli et al., 2020;Aymé & Philip, 1996;Niceta et al., 2020).However, few cases have been reported in literature associated with a causative mutation in MAF, and a significant effort is required to understand the genotypic and phenotypic spectrum (Wang et al., 2022).So far, dominant pathogenic variants in the N-terminus region of MAF have been associated with AGS, and most of these mutations were de novo (Wang et al., 2022).
The MAF gene (OMIM 177075) is mapped on chromosome 16q23 and encodes for a transcription factor involved in T-helper-2 cell differentiation.This protein plays a role in regulating several cellular processes, including embryonic lens fiber cell development, Martina Caiazza, Alberto Budillon, Gioacchino Scarano, and Giuseppe Limongelli have equally contributed.
increased T-cell susceptibility to apoptosis, and chondrocyte terminal differentiation (Amudhavalli et al., 2018).Aminoacidic substitutions in MAF may cause AGS (OMIM 601088) or its allelic condition Cataract 21, multiple types (OMIM 610202), localized within two significant missense PLP cluster regions previously explored by Niceta et al. (2015).Furthermore, heterozygous pathogenic variants in MAF are also known to be associated with isolated ocular findings (i.e., cataract, iris coloboma, microphthalmia, and anterior chamber dysgenesis) in the absence of other findings of AGS, and they are localized at the C-terminal DNA binding domain, containing an "extended homology," a "basic motif," and leucine-zipper regions (Amudhavalli et al., 2018;Niceta et al., 2015).The aim of this case report is to underline the importance of exome sequencing in the diagnosis of rare genetic diseases presenting with mild or atypical phenotypes.

| CASE REPORT
Here we report the case of a 4-year-old girl.Her medical history revealed she was born at term by normal delivery with birth weight and height of 2810 g and 47.0 cm.The head circumference was 30 cm.In addition, she exhibited fetal pericardial effusion that was early treated with pericardiocentesis.Hearing and ophthalmic screening test were normal.On day 4, she was admitted to the pediatric department for further investigation because of dysmorphic features and poor feeding.Karyotype and CGH array were performed but did not reveal any chromosomal abnormalities.Pericardial effusion was treated with ibuprofen and furosemide for 3 months.
During childhood, the patient was tested for QI e WISC-IV and neuropsychiatric evaluation suggested an intellectual disability and a mild delay in motor development.Therefore, physical education and logopedic therapy were recommended.EEG was normal.Neuroimaging (MRI) revealed a mild Chiari I malformation for which she was asymptomatic.Due to moderate scoliosis, spine radiography was performed and showed cervical and dorsal vertebral fusion of the posterior arches (C6-D7), while cerebral and cardiac echography were unremarkable.
At 6 years old, the patient was referred to the Inherited and Rare Cardiovascular Diseases Unit -University of Campania "Luigi Vanvitelli," Naples, Italy -for a cardiological and clinical genetic evaluation.Her pedigree showed no family history of any genetic or malformative condition.During the last evaluation, the patient's weight and height were 15.850 g and 111 cm corresponding to the 5th and 32nd percentile, respectively.The head circumference was 50.5 cm (10th percentile).On physical examination, the patient represents a flat facial appearance, low-set and posteriorly rotated ears, short nose, long philtrum (1.5 cm), narrow mouth, dental abnormalities, and dystrophic nails (Figure 1).Cardiologic examination showed no pericardial effusion.

| SUBJECT AND METHODS
Informed consent was appropriately obtained for genetic investigations through next-generation sequencing (NGS) with a clinical exome platform in trio encompassing 5228 disease-causing genes.A blood sample in EDTA has been collected from the subject and parents.
Genomic DNA was extracted using the Maxwell 16 instrument (Promega, Madison, WI, USA), and DNA quality was assessed using a Nanodrop machine.
Molecular testing has been carried out by analyzing a panel of target genes through an NGS-based procedure.The Alyssa software (Agilent, Santa Clara, CA, USA) has been used to perform sequence data analysis.This tool allows the alignment of the sequences to the reference genome to obtain a list of genomic variants that can be prioritized using a bioinformatic pipeline to highlight likely pathogenic or pathogenic variants.Genome data processing has been performed using a homemade bioinformatic pipeline.The MAF threshold has been set to 5% using the Illumina Variant Interpreter Software.
F I G U R E 1 (a-c) Features include facies from frontal and lateral views and the peculiar characteristic of dystrophic nails.
The systematic review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement (Page et al., 2021).We performed a systematic review of published studies (from 1993 to 2023) by searching in the Pubmed database on April 1, using keywords referring to AGS or MAF gene.
Criteria for inclusion were observational full-length original articles, research letters, case series, and case reports describing individuals of AGS with genetic confirmation.Studies published as abstract, review, preclinical, or non-English studies were excluded.Reference lists of the articles included in the review were manually screened to identify additional studies.
For each reported study, the following information was collected: first author; date of publication; number of patients; sex; age at presentation; clinical manifestations; and genetic variant.

| RESULTS
Based on the clinical and physical presentation, genetic counseling has been performed and molecular testing was suggested.Genetic testing identified a heterozygous de novo missense variant in MAF (NM_005360.5:c.161C> G; p.Ser54Trp).According to the American College of Medical Genetics classification, this variant was classified as likely pathogenic.The following criteria were met: PM1; PM5; PM2; and PP3.Thus, the genetic result confirmed the clinical diagnosis of AGS.AGS patients with the same variant have been already reported in the literature.No other clinically relevant variants were identified.Figure 2 summarizes the family pedigree.Moreover, this variant was predicted to be probably damaging by Polyphen.
However, other substitutions have been identified for the amino acid residues 54 of the MafB protein.All identified mutations (Zankl et al., 2012) cluster within the amino-terminal transcriptional activation domain of MafB, which is a Pro-Ser-Thr-rich region, and affect one of these three residues.

| DISCUSSION
To our knowledge, this is the first case of a mild AGS in which two of the main characteristics of the disease were absent (i.e., sensorineural hearing loss and cataract).Nevertheless, facial traits, combined with other clinical features (i.e., intellectual disability, pericarditis, and spine anomalies) were strongly suggestive of AGS (König et al., 2022;Niceta et al., 2020).In literature, hearing loss is always bilateral and is mostly diagnosed early in childhood.On the other hand, cataract is often present in childhood, but may even occur later (Amudhavalli et al., 2018(Amudhavalli et al., , 2020;;Chaudhry et al., 2021;Niceta et al., 2020).Thus, as an age-F I G U R E 2 Pedigree of the family of propositus.Genotype and phenotype are defined according to the legend inset.Open symbols represent subjects with negative genotypes and phenotypes.II-3 and II-4 represent subjects with negative genotypes.
T A B L E 1 Clinical features with HPO terms of the current case and the previous 27 patients reported in chronological order of identification (Alkhunaizi et al., 2019;Anand et al., 2018;Chaudhry et al., 2021;Gripp et al., 1996;Javadiyan et al., 2017;Jung et al., 2021;Keppler-Noreuil et al., 2007;Nakane et al., 2002;Xu et al., 2021).Table 1, our review of the current literature provides a clear overview of the possible scenarios that could be manifested in AGS patient together with the relative accountable genotypic variations.Among them, a male patient carrying p.Ser54Trp aminoacidic change was previously described (Amudhavalli et al., 2018).At the time of the report, this patient was 12.5 years old and showed all the characteristics of the tetrad (intellectual disability, deafness, cataracts, and the peculiar face).PE tube placement was necessitated by hearing loss.Afterward, the proband experienced mild sensorineural hearing loss in the left ear and moderate-severe hearing loss in the right ear.During an ophthalmology evaluation, a "white spot on his left retina" was detected and kept under clinical observation and, soon after, at the age of 11 years and 11 months, bilateral early cataracts were observed.
Moreover, he did not show pericardial effusion but a Chiari I malformation and seizures were reported.Since then, no strong similarity between this patient and our case could be assessed.
the patient will require long-term surveillance for the possible development of this condition during life-term.Through AGS patients resulted characterized by well-known and homogeneous features, such as Down-like facies, deafness, and cataract, as well as a wide intrafamilial and interfamilial variability that suggests the absence of genotype-phenotype correlations.This is confirmed by our patient, the only one with a such mild phenotype.Our results furtherly support a clear trend in increasing the number of molecular diagnoses due to the worldwide adoption of NGS techniques, helping us to better understand the progression of this disease in the near future.6| CONCLUSIONWe described a rare case of a young girl with AGS, carrying a disease-causing mutation in MAF.This study highlights the obstacles to a definite diagnosis when crucial clinical features in rare diseases are missing, and it confirms how similar genotypes may underlie diverse potential clinical profiles of the same disorder.In conclusion, we underlie the importance of suspecting and properly investigating any possible phenotype suggestive of a genetic and/or rare disease, even in the presence of a mild phenotype, through the collaboration of a multidisciplinary team, including different specialists and geneticists with specific expertise in inherited and rare diseases.AUTHOR CONTRIBUTIONSMartina Caiazza, Alberto Budillon, and Giuseppe Limongelli contributed to the conception and design of the work.Giulio Piluso, Vincenzo Nigro, and Francesca Del Vecchio Blanco contributed to clinical or genetic studies.Martina Caiazza, Gioacchino Scarano, Giustina Aruta, Emanuele Monda, and Giuseppe Limongelli drafted the manuscript.All the authors critically revised the manuscript.All gave final approval and agreed to be accountable for all aspects of work ensuring integrity and accuracy.All authors have read and agreed to the published version of the manuscript.