A case series of a mother and two daughters with a GLI2 gene deletion demonstrating variable expressivity and incomplete penetrance

Abstract This case series and review of the literature support that patients with pathogenic variants of the GLI2 gene demonstrate an autosomal dominant inheritance pattern, variable expressivity, and incomplete penetrance.


| INTRODUCTION
In humans, mutations in GLI2 gene, a mediator of the Sonic Hedgehog (SHH) pathway, can result in holoprosencephaly (HPE), which is the failure of midline cleavage of the forebrain during embryonal development. Clinical manifestations can range from malformed or absent cerebral tissue to hypotelorism, or narrowly spaced eyes, to cyclopia. In murine models, SHH has been shown to be expressed in the ventral diencephalon as well as the midline structures such as the notochord and floor plate of the notochord, identifying its role in pituitary and limb development, respectively. 1 Thus, the GLI2 gene has been implicated to be involved in forebrain, pituitary, and limb development. 2,3,8,9,11 In this series, we present a family, two daughters and a mother, with an identical GLI2 gene mutation. Sisters 1 and 2 present with the classic dyad of findings in a patient with Culler Jones syndrome of growth hormone (GH) deficiency and postaxial polydactyly, and their mother also demonstrated postaxial polydactyly at birth but otherwise has an unremarkable phenotype.
We also reviewed the literature for previously described families of two members or more that have a pathogenic GLI2 gene mutation. To our knowledge, there are only seven previously described families of two or more members with the GLI2 gene mutation or deletion, collectively they total thirty-one patients. The content of this manuscript is not considered research at our institution and instead falls in the realm of routine clinical care. Informed consent for publication was obtained from the parents.
also Ashkenazi Jewish ancestry in the family. She was referred to genetics for evaluation of the GLI2 gene deletion, which was also later confirmed on testing in her younger sister and mother, in the setting of an isolated GH deficiency and ectopic posterior pituitary. The patient was born by induced vaginal delivery at 40 + 3 weeks gestation, birth weight (BW) of 3.4 kg (41st centile). Mother was positive for Group B Streptococcus (GBS), which was adequately treated with two doses of antibiotics. Her newborn screen was normal. She was transferred to the neonatal intensive care unit (NICU) for observation for jaundice and hypoglycemia and removal of right hand postaxial polydactyly on the first day of life, and she was discharged at 7 days. She was identified to have failure to thrive at around 6 months of life despite adequate caloric intake; laboratory work-up showed low IGF-1, and follow-up laboratory studies confirmed a diagnosis of GH deficiency and was subsequently placed on daily subcutaneous somatotropin injections. At around 1 year of life, the patient had gastroparesis demonstrated by gastric emptying study and reflux demonstrated by esophagoduodenoscopy so she was started on lansoprazole, metoclopramide, and erythromycin. Due to persistent difficulty in obtaining adequate oral calories, a gastric tube was placed at 19 months and then removed at 4 years when oral intake became adequate. Array-based comparative genomic hybridization (aCGH) was performed and showed a small 90kb interstitial deletion of chromosome 2 including a portion of the GLI2 gene. Brain magnetic resonance imaging (MRI) without contrast showed an ectopic posterior pituitary gland and Chiari I malformation. The patient's examination was significant for hypotelorism, bilateral epicanthal folds, a flat nasal bridge, a bulbous nasal tip, and bilateral clinodactyly of digits 2, 3, 4, and 5. The patient at 11 years old had a weight of 41.6 kg (65th centile) and length of 141.7 cm (31st centile) and was tracking appropriately on both curves. There were no concerns with development or cognition.
Sister 2 is an 8-year-old girl born to the same parents. She was referred to genetics for a more thorough work-up for her short stature, history of failure to thrive, and a sister known to have a previous genetic work-up which revealed a GLI2 gene deletion. She was confirmed to have the same deletion on chromosome 2 as her sister. Of note, her father has a stature of 69 inches, and her mother a stature of 61.5 inches. The patient was born by spontaneous vaginal delivery at 40 + 0 weeks gestation, BW of 3.2 kg (21st centile). Her natal stay was significant for transfer to the NICU for observation for jaundice and hypoglycemia and removal of left hand postaxial polydactyly on the first day of life. She was discharged home after 48 hours. Around 1 year of life, chromosomal microarray identified a heterozygous deletion for the GLI2 gene so an arginine/glucagon stimulation test for GH deficiency was performed and showed a GH peak >10 ng/mL (peak: 11.8 ng/mL) and a cortisol peak >18 mcg/ dL (peak: 33.1 mcg/dL), which ruled out GH deficiency and adrenal insufficiency, respectively, at that time. Given persistent short stature, patient was referred to genetics at 5 years of life, and repeat whole-genome aCGH and single nucleotide polymorphism analysis was performed, and the patient was found to have an interstitial deletion of at least 90 kb within cytogenetic band 2q14.2 involving the GLI2 gene. The patient's examination was significant for height and weight less than the 3rd centile and bilateral clinodactyly of digits 4 and 5. Around 7 years of life, a brain MRI with and without contrast was performed, which showed a hypoplastic anterior pituitary and ectopic posterior pituitary Figure 1. The patient's growth velocity had averaged about 5.2 cm/year in the last year, and she remained below the 3rd centile for height and weight. Given the less than expected growth in the setting of GLI2 deletion and imaging findings, the patient was started on a trial of recombinant human GH dosing given the high likelihood of relative GH deficiency. The mother has a history of postaxial polydactyly but is otherwise phenotypically normal. Whole-genome aCGH revealed identical, heterozygous, interstitial 90 kb deletion within cytogenetic band 2q14.2 involving exons 3-13 of the GLI2 gene (ISCN: arr[GRCh37] 2q14.2 [121696090_121786360] x1) identical to that found in her daughters, which is classified as a pathogenic variant.

| GENETIC TESTING
Whole-genome array-based comparative genomic hybridization (aCGH) and genotype analyses were performed on a custom-designed oligonucleotide microarray (GenomeDx v5) by GeneDx. The array design is based on human genome build GRCh37/UCSC hg19, and results are reported according to ISCN guidelines. The array contains approximately 11 8000 probes that provide copy number data and 6 6000 probes that generate genotype information through analysis of SNPs. Reported boundaries correspond to deviating probes, which are dependent on array design and have the inherent limitation of not reflecting exact aberration breakpoints. The array detects copy number changes of >200 kb, on average, across the entire unique sequence of the human genome and between 500 bp and 15 kb in more than 200 targeted regions for testing performed on blood samples, which was the specimen type for the patients in this report.

| DISCUSSION
After literature review identified previously published cases of families with a GLI2 gene mutation, Table 1 was created to compare the characteristics with that of the patients in this case series. Some of the most commonly reported characteristics, including anterior pituitary lobe hypoplasia, ectopic posterior pituitary lobe identified on MRI, the resulting pituitary deficiencies, postaxial polydactyly, and facial dysmorphisms, most commonly cleft lip and palate, were given their own column in order to facilitate comparing and contrasting the different families' characteristics. True holoprosencephaly was only identified in 2/37 patients and was the rarest characteristic identified on this review, although not all patients had imaging since it was obtained when clinically indicated in most cases Table 2. [4][5][6][7]10 Over half (20/37) of the GLI2 gene mutation patients in this study were identified to have polydactyly, making it the most common characteristic. [4][5][6][7]10 The proband and her younger sister both presented with the classic dyad of Culler Jones, the mild presentation of GLI2 gene mutation characterized by GH deficiency and postaxial polydactyly, while their mother had postaxial polydactyly and otherwise asymptomatic for pituitary deficiencies. Our case exemplified the variations in presentation among family members with the same genotype. On examination of previously published families expressing an identical GLI2 mutation in the literature Table 1, the expression of the gene mutation varies widely among family members from normal phenotype to pituitary, facial, and limb abnormalities, confirming the pattern identified with the cases presented in this case series. [4][5][6][7]10 This case series and literature review support that patients with pathogenic variants of the GLI2 gene demonstrate autosomal dominant inheritance, variable expressivity, and incomplete penetrance. However, many of the parents in the literature did not receive brain imaging since it was not clinically indicated so it is possible that the findings noted are under-representative of the severity of the phenotypes. 4-7, 10 We recommend brain MRI T A B L E 2 Ratio of common features observed in GLI2 gene mutation or deletion in study population imaging in such patients in order to better characterize pituitary deficiencies and associated hormonal deficiencies and to better inform possible clinical intervention.