Clinical characteristics.

CLCN4-related neurodevelopmental disorder (CLCN4-NDD), an X-linked disorder, is characterized in the 36 males reported to date by developmental delay or intellectual disability, behavioral/mental health issues (e.g., autism spectrum disorder, anxiety, hyperactivity, and bipolar disorder), epilepsy, and gastrointestinal dysfunction. The five heterozygous females with a de novo CLCN4 variant reported to date had findings very similar to those of affected males. Twenty-two of 25 heterozygous females identified in family studies following identification of an affected male were unaffected or had only mild specific learning difficulties and/or mental health concerns, whereas three were more severely affected.

Diagnosis/testing.

The diagnosis of CLCN4-NDD is established in a male proband with suggestive findings and a hemizygous pathogenic variant in CLCN4 identified by molecular genetic testing. The diagnosis of CLCN4-NDD is usually established in a female proband with suggestive findings and a heterozygous pathogenic variant in CLCN4 identified by molecular genetic testing; however, the phenotype in females with a pathogenic variant can range from asymptomatic to severe.

Management.

Treatment of manifestations: Treatment is supportive and often includes multidisciplinary specialists in neurology, pediatrics, mental health, physiatry, occupational and physical therapy, gastroenterology, feeding therapy, ophthalmology, audiology, and medical genetics.

Surveillance: Routine monitoring of neurologic findings (response to anti-seizure medications; emergence of new findings), development and educational progress, psychiatric/behavioral issues (response to medications; emergence of new findings), mobility and self-help skills, growth and gastrointestinal manifestations, ophthalmologic findings, hearing, and family support systems.

Genetic counseling.

CLCN4-NDD is inherited in an X-linked manner. The father of an affected male will not have the disorder nor will he be hemizygous for the CLCN4 pathogenic variant. If the mother of a proband has a CLCN4 pathogenic variant, the chance of transmitting it in each pregnancy is 50%: males who inherit the pathogenic variant will be affected; females who inherit the pathogenic variant will be heterozygotes and may be unaffected or have clinical findings ranging from mild learning difficulties and mental health concerns to severe manifestations. If the proband represents a simplex case and if the CLCN4 pathogenic variant cannot be detected in the leukocyte DNA of the mother, the risk to sibs is presumed to be low but greater than that of the general population. Once the CLCN4 pathogenic variant has been identified in an affected family member, prenatal and preimplantation genetic testing are possible.

Suggestive Findings

CLCN4-related neurodevelopmental disorder (CLCN4-NDD) should be considered in individuals with the following clinical and brain MRI findings and family history.

Clinical findings

• Developmental delay or intellectual disability
• Autism spectrum disorder
• Epilepsy
• Mental health conditions including anxiety and bipolar disorder
• Gastrointestinal dysfunction
• Unremarkable facial features. Although a subtle lengthening of the face and squaring of the chin that becomes more prominent with age has been noted in several individuals (see Figure 3 in Palmer et al [2018]; full text), these features are not likely to be distinctive enough to trigger consideration of this specific diagnosis.

Brain MRI. Minor structural brain malformations include periventricular leukomalacia, cortical atrophy/ventriculomegaly, and hypoplasia or dysplasia of the corpus callosum [Veeramah et al 2013, Palmer et al 2018, Zhou et al 2018, He et al 2021].

Family history is consistent with X-linked inheritance (e.g., no male-to-male transmission). Absence of a known family history does not preclude the diagnosis.

Establishing the Diagnosis

Male proband. The diagnosis of CLCN4-NDD is established in a male proband with suggestive findings and a hemizygous pathogenic (or likely pathogenic) variant in CLCN4 identified by molecular genetic testing (see Table 1).

Female proband. The diagnosis of CLCN4-NDD is usually established in a female proband with suggestive findings and a heterozygous de novo pathogenic (or likely pathogenic) variant in CLCN4 identified by molecular genetic testing (see Table 1).

Note: (1) Per ACMG/AMP variant interpretation guidelines, the terms "pathogenic variants" and "likely pathogenic variants" are synonymous in a clinical setting, meaning that both are considered diagnostic and both can be used for clinical decision making [Richards et al 2015]. Reference to "pathogenic variants" in this section is understood to include any likely pathogenic variants. (2) Identification of a hemizygous or heterozygous CLCN4 variant of uncertain significance does not establish or rule out a diagnosis.

Molecular genetic testing in a child with developmental delay or an older individual with intellectual disability may begin with chromosomal microarray analysis (CMA). Other options include use of a multigene panel or exome sequencing. Note: Single-gene testing (sequence analysis of CLCN4, followed by gene-targeted deletion/duplication analysis) is rarely useful and typically NOT recommended.

• Chromosomal microarray analysis (CMA) uses oligonucleotide or SNP arrays to detect genome-wide large deletions/duplications (including CLCN4) that cannot be detected by sequence analysis.
• An intellectual disability and/or epilepsy and/or autism spectrum disorder multigene panel that includes CLCN4 and other genes of interest (see Differential Diagnosis) is most likely to identify the genetic cause of the condition while limiting identification of variants of uncertain significance and pathogenic variants in genes that do not explain the underlying phenotype. Note: (1) The genes included in the panel and the diagnostic sensitivity of the testing used for each gene vary by laboratory and are likely to change over time. (2) Some multigene panels may include genes not associated with the condition discussed in this GeneReview. (3) In some laboratories, panel options may include a custom laboratory-designed panel and/or custom phenotype-focused exome analysis that includes genes specified by the clinician. (4) Methods used in a panel may include sequence analysis, deletion/duplication analysis, and/or other non-sequencing-based tests.
  For an introduction to multigene panels click here. More detailed information for clinicians ordering genetic tests can be found here.
• Comprehensive genomic testing does not require the clinician to determine which gene is likely involved. Exome sequencing is most commonly used; genome sequencing is also possible.
  For an introduction to comprehensive genomic testing click here. More detailed information for clinicians ordering genomic testing can be found here.

Clinical Description

CLCN4-related neurodevelopmental disorder (CLCN4-NDD), an X-linked disorder, is characterized in the 36 males reported to date by developmental delay or intellectual disability, behavioral/mental health issues (e.g., autism spectrum disorder, anxiety, hyperactivity, and bipolar disorder), epilepsy, and gastrointestinal dysfunction. The five heterozygous females with a de novo CLCN4 variant reported to date had findings very similar to those of affected males. Twenty-two of 25 heterozygous females identified in family studies following identification of an affected male were unaffected or had only mild specific learning difficulties and/or mental health concerns; three were more severely affected.

The findings of affected individuals reported to date are summarized in Table 2.

Genotype-Phenotype Correlations

No genotype-phenotype correlations have been identified to date.

Significant phenotypic variability has been observed both between individuals from different families with a recurrent variant and among individuals from a single family who have the same variant [Palmer et al 2018, He et al 2021].

The phenotypes of males with haploinsufficiency (due to exon deletions or frameshift or nonsense variants) were noted to be relatively milder than the phenotypes of individuals with missense variants [Palmer et al 2018, He et al 2021].

Nomenclature

CLCN4-related neurodevelopmental disorder may also be referred to as Raynaud-Claes syndrome, based on two of the families reported by Hu et al [2016], who had been reported as families MRX15 [Raynaud et al 1996] and MRX49 [Claes et al 1997] before identification of the causative gene.

Prevalence

CLCN4-NDD is rare, and (likely) pathogenic variants have, to date, only been reported in 66 individuals, 22 of whom are asymptomatic or only very mildly affected heterozygous females [Veeramah et al 2013, Hu et al 2016, Palmer et al 2018, Zhou et al 2018, Guo et al 2021, He et al 2021, Xu et al 2021].

CLCN4-NDD is not known to be more prevalent in any specific populations, and no founder variants are known.

Evaluations Following Initial Diagnosis

To establish the extent of disease and needs in an individual diagnosed with CLCN4-related neurodevelopmental disorder (CLCN4-NDD), the evaluations summarized in Table 3 (if not performed as part of the evaluation that led to the diagnosis) are recommended.

Treatment of Manifestations

Treatment is supportive and often includes multidisciplinary specialists in neurology, pediatrics, mental health, physiatry, occupational and physical therapy, gastroenterology, feeding therapy, ophthalmology, audiology, and medical genetics.

Surveillance

Evaluation of Relatives at Risk

See Genetic Counseling for issues related to testing of at-risk relatives for genetic counseling purposes.

Therapies Under Investigation

Search ClinicalTrials.gov in the US and EU Clinical Trials Register in Europe for access to information on clinical studies for a wide range of diseases and conditions. Note: There may not be clinical trials for this disorder.

Mode of Inheritance

CLCN4-related neurodevelopmental disorder (CLCN4-NDD) is inherited in an X-linked manner.

Risk to Family Members

Parents of a male proband

• The father of an affected male will not have the disorder nor will he be hemizygous for the CLCN4 pathogenic variant; therefore, he does not require further evaluation/testing.
• In a family with more than one affected individual, the mother of an affected male is an obligate heterozygote. Note: If a woman has more than one affected child and no other affected relatives and if the CLCN4 pathogenic variant cannot be detected in her leukocyte DNA, she most likely has germline mosaicism.
• If a male is the only affected family member (i.e., a simplex case), the mother may be a heterozygote, the affected male may have a de novo CLCN4 pathogenic variant (in which case the mother is not a heterozygote), or the mother may have somatic/germline mosaicism.
  About 16% of affected males have the disorder as the result of a de novo pathogenic variant [Veeramah et al 2013, Palmer et al 2018, Zhou et al 2018, He et al 2021].
• Molecular genetic testing of the mother is recommended to confirm her genetic status and to allow reliable recurrence risk assessment.

Sibs of a male proband. The risk to sibs depends on the genetic status of the mother:

• If the mother of the proband has a CLCN4 pathogenic variant, the chance of the mother transmitting it in each pregnancy is 50%.
  • Males who inherit the pathogenic variant will be affected.
  • Females who inherit the pathogenic variant will be heterozygotes and may be unaffected or have clinical findings ranging from mild learning difficulties and mental health concerns to severe manifestations (see Clinical Description, Heterozygous Females). X-chromosome inactivation status has not been found to be helpful in assessing degree of severity of features in female heterozygotes [Palmer et al 2018].
• If the proband represents a simplex case and if the CLCN4 pathogenic variant cannot be detected in the leukocyte DNA of the mother, the risk to sibs is presumed to be low but greater than that of the general population because of the possibility of maternal germline mosaicism. To date, sib recurrence due to parental germline mosaicism has not been reported; however, somatic mosaicism has been reported in one female proband and one severely affected male [Palmer et al 2018].

Parents of a female proband

• A female proband may have inherited the CLCN4 pathogenic variant from either her mother or her father, or the pathogenic variant may be de novo. Five of the eight females reported to date with manifestations of CLCN4-NDD have the disorder as the result of a de novo pathogenic variant (see Clinical Description) [Palmer et al 2018].
• Detailed evaluation of the parents and review of the extended family history may help distinguish probands with a de novo pathogenic variant from those with an inherited pathogenic variant. Molecular genetic testing of the mother (and possibly the father, or subsequently the father) can help to determine if the pathogenic variant was inherited.

Sibs of a female proband. The risk to sibs depends on the genetic status of the parents:

• If the mother of the proband has a CLCN4 pathogenic variant, the chance of the mother transmitting it in each pregnancy is 50%.
  • Males who inherit the pathogenic variant will be affected.
  • Females who inherit the pathogenic variant will be heterozygotes and may be unaffected or have clinical findings ranging from mild learning difficulties and mental health concerns to severe manifestations (see Clinical Description, Heterozygous Females).
  • The manifestations of CLCN4-NDD in a female sib with a maternally-inherited pathogenic variant cannot be predicted on the basis of the phenotype in the heterozygous mother. It is possible for a female who inherits a CLCN4 pathogenic variant from an asymptomatic or very mildly affected mother to have a severe neurodevelopmental phenotype [Palmer et al 2018, Xu et al 2021].
  • X-chromosome inactivation status has not been found to be helpful in assessing degree of severity of features in female heterozygotes [Palmer et al 2018].
• If the father of the proband has a CLCN4 pathogenic variant, he will transmit it to all his daughters and none of his sons.
• If the proband represents a simplex case and if the CLCN4 pathogenic variant cannot be detected in the leukocyte DNA of either parent, the risk to sibs is greater than that of the general population because of the possibility of parental germline mosaicism.

Offspring of a male proband. Affected males transmit the CLCN4 pathogenic variant to all their daughters and none of their sons.

Offspring of a female proband. Women with a CLCN4 pathogenic variant have a 50% chance of transmitting the pathogenic variant to each child.

Other family members. The risk to other family members depends on the status of the proband's parents: if a parent has the CLCN4 pathogenic variant, the parent's family members may be at risk.

Note: Molecular genetic testing may be able to identify the family member in whom a de novo pathogenic variant arose, information that could help determine genetic risk status of members of the extended family.

Heterozygote Detection

Identification of female heterozygotes requires prior identification of the familial CLCN4 pathogenic variant.

Note: Females who inherit the pathogenic variant will be heterozygotes and may be unaffected or have clinical findings ranging from mild learning difficulties and mental health concerns to severe manifestations (see Clinical Description, Heterozygous Females). X-chromosome inactivation status has not been found to be helpful in assessing degree of severity of features in female heterozygotes [Palmer et al 2018].

Related Genetic Counseling Issues

Family planning

• The optimal time for determination of genetic risk and discussion of the availability of prenatal/preimplantation genetic testing is before pregnancy.
• It is appropriate to offer genetic counseling (including discussion of potential risks to offspring and reproductive options) to young adults who are affected or are heterozygotes or who are at increased risk of being heterozygotes.

Prenatal Testing and Preimplantation Genetic Testing

Once the CLCN4 pathogenic variant has been identified in an affected family member, prenatal and preimplantation genetic testing are possible.

Differences in perspective may exist among medical professionals and within families regarding the use of prenatal testing. While most centers would consider use of prenatal testing to be a personal decision, discussion of these issues may be helpful.

Molecular Pathogenesis

CLCN4/ClC-4 is broadly expressed in many tissues, with highest expression in brain and skeletal muscle. ClC-4 is a strongly outwardly rectifying, electrogenic 2Cl-/H+ exchanger, predominantly expressed in membranes of the endolysosomal system. Its biological function remains largely unknown but may be related to ion homeostasis [Jentsch & Pusch 2018].

ClC-4 forms heterodimers with its close homolog ClC-3, encoded by CLCN3 [Weinert et al 2020]. Of note, homozygous and heterozygous pathogenic variants in CLCN3 are associated with developmental delay / intellectual disability, mood or behavioral disorders, dysmorphic features, structural brain abnormalities, and seizures [Duncan et al 2021].

It is currently unknown how genetic alterations in CLCN4 cause CLCN4-NDD. This is an area of active research.

Mechanism of disease causation. All CLCN4 missense variants published to date that were tested functionally in Xenopus oocytes by electrophysiologic studies showed impaired protein function (i.e., loss of function).

Author Notes

Dr Vera Kalscheuer (PhD) is a scientist at the Max Planck Institute for Molecular Genetics in Berlin, Germany. She obtained her degree and PhD in biochemistry from the Free University in Berlin, Germany and conducted postdoctoral research at the Radboud University Nijmegen Medical Centre, the Netherlands. Her work focuses on the identification of novel genes linked to neurodevelopmental disabilities, and on the molecular and functional characterization of selected genes and proteins in order to better understand the underlying molecular and pathogenic mechanisms. With her research, she has discovered and contributed to the identification of numerous genes associated with various forms of neurodevelopmental disabilities, including CLCN4, thereby establishing a molecular diagnosis for patients and their families. In order to better understand CLCN4-related neurodevelopmental disorder, she and Dr Emma Palmer together with other collaborators conducted a large study on the genetic, clinical, and functional effect of newly identified CLCN4 variants. She and Dr Palmer are currently studying an even larger group of patients with CLCN4 defects.
Email: ed.gpm.neglom@uehcslak
Website: www.molgen.mpg.de/Vera-Kalscheuer

Dr Emma Palmer (PhD, MBBS, FRACP, BA (Hons I) Oxon) is a clinician scientist at Sydney Children's Hospital Network & University of New South Wales in Sydney, Australia. She has extensive experience at the interface of clinical and research genetics leading multidisciplinary teams and establishing international collaborations to discover new genetic conditions. She has led five international projects delineating the following novel genetic conditions:

• ARV1-developmental and epileptic encephalopathy, inherited in an autosomal recessive manner
• ATN1-congenital hypotonia, epilepsy, developmental delay, digital anomalies (CHEDDA); caused by de novo heterozygous variants
• KCNT2-developmental epileptic encephalopathy; caused by de novo heterozygous variants
• RLIM duplication; Tonne-Kalscheuer syndrome; inherited in an X-linked manner
• ZSWIM6-neurodevelopmental disorder with movement abnormalities, abnormal gait, and autistic features (NEDMAGA) acromelic frontonasal dysplasia

She works closely with rare genetic disease advocacy and consumer reference groups and aims to translate genomic discoveries to improved education and management for patients and families. She was the first author on a publication describing the effect of CLCN4 changes in 52 individuals, moderates the CLCN4 gene pages on the Human Disease Gene Webseries along with Dr Vera Kalscheuer, and is leading the clinical aspects of an international study to better understand the genetic and clinical spectrum of CLCN4-related neurodevelopmental disorder.
Email: ua.ude.wsnu@remlap.htebazile
Website: research.unsw.edu.au/people/dr-elizabeth-emma-palmer

Acknowledgments

We thank the Cure CLCN4 patient support group and Simons Foundation for their efforts supporting families with CLCN4-related neurodevelopmental disorder. We thank the families, their clinicians and clinical geneticists, and our collaborators.

Revision History

• 16 December 2021 (bp) Review posted live
• 27 August 2021 (ep) Original submission

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