Descriptive analysis of mutation-positive ED cases

Descriptive statistics for the 20 positive cases are presented in Table 1. Hypotrichosis was observed in all but one case (ED16), and anhidrosis/hypohidrosis was present in all but two cases (ED16 and ED23). Hypodontia/anodontia was observed in all cases except three (ED6, 9, and 23). For ED23, accurate tooth assessment is challenging because of delayed dentition. Altogether cases carrying mutations in the EDA and EDAR genes were 16, accounting for 80% (16/20) of the total positive cases. Among these, EDA accounted for the majority (13 cases), and notably, CNVs were responsible for three cases (23.1%). The remaining genes in which mutations were detected included one case each of ERCC2, DSG4, LRP6, and LIPH.

Table 1 Descriptive statistics of positive ectodermal dysplasia cases (n = 20)Cases with EDA/EDAR mutations

EDA and EDAR mutations were detected in 16 cases, including 15 male patients and 1 female patient, who tested positive for EDAR. The average patient age was 15.5 years. In positive cases, mutation sites within EDA and EDAR were annotated in the functional domains. (Fig. 1). Missense mutations were localized toward the termini of the established furin and TNF domains, whereas loss-of-function (LOF) mutations were predominantly clustered within the COL domain. Concerning EDAR mutations, three alterations were detected in the death domain. Notably, a novel missense mutation in EDAR gene (NM_022336 c.1043T > C, p.L348P) was confirmed as a de novo mutation based on parental testing results (Supplementary Figure S1).

Fig. 1

Genetic variants of EDA/EDAR in positive ectodermal dysplasia cases. Among 16 cases, 15 sequence variants were identified (p.G201Rfs*39 was detected in both ED10 and ED20). Three EDA CNVs are shown at the top left. Among the four frameshift mutations, three were located in the COL domain of the EDA gene, two missense mutations were in the furin domain, and three were at the end of the TNF domain. Three missense EDAR mutations were identified: two in the C-terminal end of the DD and one slightly upstream of the DD domain. COL, collagen-like domain; TNF, tumor necrosis factor; DD, death domain

Cases with mutations in genes other than EDA/EDAR

ED6 had biallelic mutations on ERCC2. He is a 5-year-old boy who exhibited classic symptoms including hypotrichosis and hypohidrosis. Hypodontia was not prominent; instead, dysmorphic teeth were observed. Notably, he had a history of frequent febrile episodes, along with the presence of micropenis accompanied by underdeveloped scrotum, and adrenal hemorrhage during the neonatal period (Table 1). ERCC2 is associated with xeroderma pigmentosum group D (MIM: 278730) in an autosomal recessive manner. Moreover, biallelic mutations in ERCC2 can also lead to photosensitive trichothiodystrophy. Two mutations in ERCC2 were identified: a frameshift mutation and a missense mutation that was initially categorized as VUS. However, subsequent parental testing led to the segregation and reclassification of these two mutations as likely pathogenic, thereby confirming the genetic basis of the disease.

ED9 had a homozygous mutation, c.574T > C, p.Ser192Pro, in the DSG4 gene. DSG4 is the causal gene of autosomal recessive hypotrichosis 6 (MIM: 607903), which is inherited in an autosomal recessive manner. Interestingly, ED9 exhibited hypohidrosis, whereas typically DSG4-mutation positive patients show normal sweating.

ED16, a 4-year-old boy, presented with a normal complement of 20 deciduous teeth; however, routine X-rays revealed multiple missing permanent teeth (more than half absent). A de novo mutation in LRP6 was detected. This variant is a heterozygous pathogenic nonsense mutation (c.94 C > T, p.Arg32*) in the LRP6 gene. LRP6 is associated with the autosomal dominant tooth agenesis-selective 7 (MIM: 616724).

Lastly, ED23, a 7-year-old boy, exhibited hypotrichosis, although hypohidrosis was not observed. Hypodontia could not be confirmed due to delayed dentition. ED23 patient harbored a pathogenic mutation (c.736T > A, p.Cys246Ser, heterozygous) and a likely pathogenic frameshift variant (c.928del, p.Asp310Ilefs*4, heterozygous) in the LIPH gene. LIPH is associated with autosomal recessive hypotrichosis 7 (MIM: 604379).

Analysis of negative cases with ED symptoms

Table 2 displays 7 negative cases. Among these, all seven patients were male, with dental issues evident in six cases (85.7%), hypohidrosis/anhidrosis in two cases (28.6%), and hypotrichosis in three cases (42.9%). Furthermore, three patients exhibited symptoms unrelated to the ectodermal origin, such as optic neuropathy and failure to thrive. Novel VUS were identified in four patients. Specifically, WNT10A variants were observed in 2 cases, and EDA and KDF1 were present in 1 case. Except for the WNT10A c.511 C > T variant, the remaining three variants were predicted to be deleterious based on all three in silico prediction tools. The sequence variants identified in EDA and KDF1 are not annotated in the GnomAD database of human variations. Parental testing was only feasible for the KDF1 case, where the mother was found to be a carrier and had few permanent teeth.

Table 2 Descriptive statistics of negative ectodermal dysplasia cases (n = 7)Correlation between EDA/EDAR mutation detection and classic symptom presentation

Table 3 illustrates the mutation detection rates of EDA and EDAR based on the presence of hair, skin, or dental symptoms. Notably, of patients exhibiting all three symptoms, 94.1% had EDA/EDAR mutations; of those who did not show these three symptoms, none had EDA/EDAR mutations.

Table 3 Detection rate of EDA/EDAR mutations according to symptoms