Mutation Update for Kabuki Syndrome Genes KMT2D and KDM6A and Further Delineation of X‐Linked Kabuki Syndrome Subtype 2

Kabuki syndrome (KS) is a rare but recognizable condition that consists of a characteristic face, short stature, various organ malformations, and a variable degree of intellectual disability. Mutations in KMT2D have been identified as the main cause for KS, whereas mutations in KDM6A are a much less frequent cause. Here, we report a mutation screening in a case series of 347 unpublished patients, in which we identified 12 novel KDM6A mutations (KS type 2) and 208 mutations in KMT2D (KS type 1), 132 of them novel. Two of the KDM6A mutations were maternally inherited and nine were shown to be de novo. We give an up‐to‐date overview of all published mutations for the two KS genes and point out possible mutation hot spots and strategies for molecular genetic testing. We also report the clinical details for 11 patients with KS type 2, summarize the published clinical information, specifically with a focus on the less well‐defined X‐linked KS type 2, and comment on phenotype–genotype correlations as well as sex‐specific phenotypic differences. Finally, we also discuss a possible role of KDM6A in Kabuki‐like Turner syndrome and report a mutation screening of KDM6C (UTY) in male KS patients.     

Molecular,clinical and neuropsychological study in 31 patients with Kabuki syndrome and KMT2D mutations

Kabuki syndrome (KS—OMIM 147920) is a rare developmental disease characterized by the association of multiple congenital anomalies and intellectual disability. This study aimed to investigate intellectual performance in children with KS and link the performance to several clinical features and molecular data. We recruited 31 children with KMT2D mutations who were 6 to 16 years old. They all completed the Weschler Intelligence Scale for Children, fourth edition. We calculated all indexes: the Full Scale Intellectual Quotient (FSIQ), Verbal Comprehension Index (VCI), Perceptive Reasoning Index (PRI), Processing Speed Index (PSI), and Working Memory Index (WMI). In addition, molecular data and several clinical symptoms were studied. FSIQ and VCI scores were 10 points lower for patients with a truncating mutation than other types of mutations. In addition, scores for FSIQ, VCI and PRI were lower for children with visual impairment than normal vision. We also identified a discrepancy in indexes characterized by high WMI and VCI and low PRI and PSI. We emphasize the importance of early identification and intensive care of visual disorders in patients with KS and recommend individual assessment of intellectual profile.     

Insights into the perinatal phenotype of Kabuki syndrome in infants identified by genome-wide sequencing

Increasing use of unbiased genomic sequencing in critically ill infants can expand understanding of rare diseases such as Kabuki syndrome (KS). Infants diagnosed with KS through genome-wide sequencing performed during the initial hospitalization underwent retrospective review of medical records. Human phenotype ontology terms used in genomic analysis were aggregated and analyzed. Clinicians were surveyed regarding changes in management and other care changes. Fifteen infants met inclusion criteria. KS was not suspected prior to genomic sequencing. Variants were classified as Pathogenic (n = 10) or Likely Pathogenic (n = 5) by American College of Medical Genetics and Genomics Guidelines. Fourteen variants were de novo (KMT2D, n = 12, KDM6A, n = 2). One infant inherited a likely pathogenic variant in KMT2D from an affected father. Frequent findings involved cardiovascular (14/15) and renal (7/15) systems, with palatal defects also identified (6/15). Three infants had non-immune hydrops. No minor anomalies were universally documented; ear anomalies, micrognathia, redundant nuchal skin, and hypoplastic nails were common. Changes in management were reported in 14 infants. Early use of unbiased genome-wide sequencing enabled a molecular diagnosis prior to clinical recognition including infants with atypical or rarely reported features of KS while also expanding the phenotypic spectrum of this rare disorder.     

Burkitt lymphoma in a patient with Kabuki syndrome carrying a novel KMT2D mutation

Kabuki syndrome (KS) is an extremely rare genetic disorder, mainly caused by germline mutations at specific epigenetic modifier genes, including KMT2D. Because the tumor suppressor gene KMT2D is also frequently altered in many cancer types, it has been suggested that KS may predispose to the development of cancer. However, KS being a rare disorder, few data are available on the incidence of cancer in KS patients. Here, we report the case of a 5‐year‐old boy affected by KS who developed Burkitt lymphoma (BL). Genetic analysis revealed the presence of a novel heterozygous mutation in the splice site of the intron 4 of KMT2D gene in both peripheral blood‐extracted DNA and tumour cells. In addition, the tumour sample of the patient was positive for the classical somatic chromosomal translocation t(8;14) involving the c‐MYC gene frequently identified in BL. We propose that the mutated KMT2D gene contributes to the development of both KS and BL observed in our patient and we suggest that strict surveillance must be performed in KS patients.     

Identification of a KDM6A somatic mutation responsible for Kabuki syndrome by excluding a conflicting KMT2D germline variant through episignature analysis

Kabuki syndrome (KS) is a congenital disorder caused by mutations in either KMT2D on chromosome 12 or KDM6A on chromosome X, encoding a lysine methyltransferase and a lysine demethylase, respectively. A 9-year-4-month-old male patient with a normal karyotype presented with KS and autism spectrum disorder. Genetic testing for KS was conducted by Sanger sequencing and episignature analysis using DNA methylation array data. The patient had a mosaic stop-gain variant in KDM6A and a heterozygous missense variant (rs201078160) in KMT2D. The KDM6A variant is expected to be deleterious. The KMT2D variant pathogenicity has been inconsistently reported in the ClinVar database. Using biobanking resources, we identified two heterozygous individuals possessing the rs201078160 variant. In a subsequent episignature analysis, the KS patient showed the KS episignature, but two control individuals with the rs201078160 variant did not. Our results indicate that the mosaic stop-gained variant in KDM6A, but not the rs201078160 variant in KMT2D, is responsible for the KS phenotype in the patient. This study further demonstrated the utility of DNA methylation information in diagnosing rare genetic diseases and emphasized the importance of a reference dataset containing both genotype and DNA methylation information.     

Immunologic assessment and KMT2D mutation detection in Kabuki syndrome

Kabuki or Niikawa–Kuroki syndrome (KS) is a rare disorder with multiple malformations and recurrent infections, especially otitis media. This study aimed to investigate the genetic defects in Kabuki syndrome and determine if immune status is related to recurrent otitis media. Fourteen patients from 12 unrelated families were enrolled in the 9‐year study period (2005–2013). All had Kabuki faces, cleft palate, developmental delay, mental retardation, and the short fifth finger. Recurrent otitis media (12/14) and hearing impairment (8/14) were also more common features. Immunologic analysis revealed lower memory CD19+ cells (11/13), lower memory CD4+ cells (8/13), undetectable anti‐HBs antibodies (7/13), and antibody deficiency (7/13), including lower IgA (4), IgG (2), and IgG2 (1). Naïve emigrant lymphocytes, lymphocyte proliferation function, complement activity, and superoxide production in polymorphonuclear cells were all normal. All the patients had KMT2D mutations and 10 novel mutations of R1252X, R1757X,Y1998C, P2550R fs2604X, Q4013X, G5379X, E5425K, R5432X, R5432W, and R5500W. Resembling the phenotype of common variable immunodeficiency, KS patients with antibody deficiency, decreased memory cells, and poor vaccine response increased susceptibility to recurrent otitis media. Large‐scale prospective studies are warranted to determine if regular immunoglobulin supplementation decreases the frequency of otitis media and severity of hearing impairment.     

Novel KDM6A (UTX) mutations and a clinical and molecular review of the X‐linked Kabuki syndrome (KS2)

We describe seven patients with KDM6A (located on Xp11.3 and encodes UTX) mutations, a rare cause of Kabuki syndrome (KS2, MIM 300867) and report, for the first time, germ‐line missense and splice‐site mutations in the gene. We demonstrate that less than 5% cases of Kabuki syndrome are due to KDM6A mutations. Our work shows that similar to the commoner Type 1 Kabuki syndrome (KS1, MIM 147920) caused by KMT2D (previously called MLL2) mutations, KS2 patients are characterized by hypotonia and feeding difficulties during infancy and poor postnatal growth and short stature. Unlike KS1, developmental delay and learning disability are generally moderate–severe in boys but mild–moderate in girls with KS2. Some girls may have a normal developmental profile. Speech and cognition tend to be more severely affected than motor development. Increased susceptibility to infections, join laxity, heart, dental and ophthalmological anomalies are common. Hypoglycaemia is more common in KS2 than in KS1. Facial dysmorphism with KDM6A mutations is variable and diagnosis on facial gestalt alone may be difficult in some patients. Hypertrichosis, long halluces and large central incisors may be useful clues to an underlying KDM6A mutation in some patients.     

Molecular genetic analysis in 14 Czech Kabuki syndrome patients is confirming the utility of phenotypic scoring

Kabuki syndrome (KS) is a dominantly inherited disorder mainly due to de novo pathogenic variation in KMT2D or KDM6A genes. Initially, a representative cohort of 14 Czech cases with clinical features suggestive of KS was analyzed by experienced clinical geneticists in collaboration with other specialties, and observed disease features were evaluated according to the ‘MLL2‐Kabuki score’ defined by Makrythanasis et al. Subsequently, the aforementioned genes were Sanger sequenced and copy number variation analysis was performed by MLPA, followed by genome‐wide array CGH testing. Pathogenic variants in KMT2D resulting in protein truncation in 43% (6/14; of which 3 are novel) of all cases were detected, while analysis of KDM6A was negative. MLPA analysis was negative in all instances. One female patient bears a 6.6 Mb duplication of the Xp21.2–Xp21.3 region that is probably disease causing. Subjective KS phenotyping identified predictive clinical features associated with the presence of a pathogenic variant in KMT2D. We provide additional evidence that this scoring approach fosters prioritization of patients prior to KMT2D sequencing. We conclude that KMT2D sequencing followed by array CGH is a diagnostic strategy with the highest diagnostic yield.     

MLL2 mosaic mutations and intragenic deletion–duplications in patients with Kabuki syndrome

Kabuki syndrome (KS) is a rare multi‐system disorder that can result in a variety of congenital malformations, typical dysmorphism and variable learning disability. It is caused by MLL2 point mutations in the majority of the cases and, rarely by deletions involving KDM6A. Nearly one third of cases remain unsolved. Here, we expand the known genetic basis of KS by presenting five typical patients with the condition, all of whom have novel MLL2 mutation types– two patients with mosaic small deletions, one with a mosaic whole‐gene deletion, one with a multi‐exon deletion and one with an intragenic multi‐exon duplication. We recommend MLL2 dosage studies for all patients with typical KS, where traditional Sanger sequencing fails to identify mutations. The prevalence of such MLL2 mutations in KS may be comparable with deletions involving KDM6A. These findings may be helpful in understanding the mutational mechanism of MLL2 and the disease mechanism of KS.     

Under the mask of Kabuki syndrome: Elucidation of genetic-and phenotypic heterogeneity in patients with Kabuki-like phenotype

Kabuki syndrome is mainly caused by dominant de-novo pathogenic variants in the KMT2D and KDM6A genes. The clinical features of this syndrome are highly variable, making the diagnosis of Kabuki-like phenotypes difficult, even for experienced clinical geneticists. Herein we present molecular genetic findings of causal genetic variation using array comparative genome hybridization and a Mendeliome analysis, utilizing targeted exome analysis focusing on regions harboring rare disease-causing variants in Kabuki-like patients which remained KMT2D/KDM6A-negative. The aCGH analysis revealed a pathogenic CNV in the 14q11.2 region, while targeted exome sequencing revealed pathogenic variants in genes associated with intellectual disability (HUWE1, GRIN1), including a gene coding for mandibulofacial dysostosis with microcephaly (EFTUD2). Lower values of the MLL2-Kabuki phenotypic score are indicative of Kabuki-like phenotype (rather than true Kabuki syndrome), where aCGH and Mendeliome analyses have high diagnostic yield. Based on our findings we conclude that for new patients with Kabuki-like phenotypes it is possible to choose a specific molecular testing approach that has the highest detection rate for a given MLL2-Kabuki score, thus fostering more precise patient diagnosis and improved management in these genetically- and phenotypically heterogeneous clinical entities.     

Expanding the phenotype and the genotype of Stromme syndrome: A novel variant of the CENPF gene and literature review

This report describes siblings with Stromme syndrome, a rare genetic condition that primarily presents with a triad of intestinal atresia, cranial and ocular malformations, and other organ systems could be involved. This clinical triad was initially named after the first person to describe it in 1993. Here, we report a family with two siblings who presented with unusual intestinal atresia and ocular and CNS abnormalities. The first patient is a 6-year-old-boy with apple peel duodeno-jejunal atresia, unilateral microphthalmia and microcephaly. The second patient, a younger brother, presented with intestinal atresia, corneal opacity and alobar holoprosencephaly and passed away at the age of 3 months. Exome sequencing showed a novel homozygous variant in the CENPF gene, NM_016343.3: c.1195-2 A > G that was detected in both of the affected siblings. This is a report and literature review of CENPF-related ciliopathy, which may result in Stromme syndrome. As this is the fourth report linking the CENPF gene variant with Stromme syndrome and first reported case presented with holoprosencephaly, it will expand the current knowledge on the genotype and the phenotype of Stromme syndrome.     

Analysis of phenotype and genotype information for the diagnosis of Marfan syndrome

Marfan syndrome is considered a clinical diagnosis. Three diagnostic classifications comprising first, Marfan genotype with a causative FBN1 gene mutation; second, Marfan phenotype with clinical criteria of the original Ghent nosology (Ghent-1); and third, phenotype with clinical criteria of its current revision (Ghent-2) in 300 consecutive persons referred for confirmation or exclusion of Marfan syndrome (150 men, 150 women aged 35 ± 13 years) were used. Sequencing of TGBR1/2 genes was performed in 128 persons without FBN1 mutation. Marfan genotype was present in 140, Ghent-1 phenotype in 139, and Ghent-2 phenotype in 124 of 300 study patients. Marfan syndrome was confirmed in 94 and excluded in 129 persons consistently by all classifications, but classifications were discordant in 77 persons. With combined genotype and phenotype information confirmation of Marfan syndrome was finally achieved in 126 persons by Ghent-1 and in 125 persons by Ghent-2 among 140 persons with Marfan genotype, and exclusion was accomplished in 139 persons by Ghent-1 and in 141 persons by Ghent-2 among 160 persons without Marfan genotype. In total, genotype information changed final diagnoses in 22 persons with Ghent-1, and in 32 persons with Ghent-2. It is concluded that genotype information is essential for diagnosis or exclusion of Marfan syndrome.     

Genome‐wide DNA methylation and RNA analyses enable reclassification of two variants of uncertain significance in a patient with clinical Kabuki syndrome

Nontruncating sequence variants represent a major challenge in variant interpretation and classification. Here, we report a patient with features of Kabuki syndrome who carries two rare heterozygous variants in KMT2D: c.12935C>T, p.(Ser4312Phe) and c.15785‐10T>G. The clinical significance of these variants were discordantly interpreted by different diagnostic laboratories. Parental testing showed that the missense variant was inherited from the father with a mild Kabuki phenotype and the intronic variant from the mother with mosaic status. Through genome‐wide DNA methylation analysis of peripheral blood, we confirmed that the proband exhibited a previously described episignature of Kabuki syndrome. Parental samples had normal DNA methylation profiles, thus ruling out the involvement of the paternally inherited missense variant. RNA analysis revealed that the intronic change resulted in exon 49 skipping and frameshift, thereby providing a molecular diagnosis of Kabuki syndrome. This study demonstrates the utility of epigenomic and RNA analyses in resolving ambiguous clinical cases.