Clinical characteristics.

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is characterized by two phenotypes: classic MLC and improving MLC.

• Individuals with classic MLC present with macrocephaly, often in association with seizures, gradual onset of ataxia, spasticity, and sometimes extrapyramidal findings, mild gross motor developmental delays, and late-onset cognitive deterioration. Macrocephaly, observed in most affected individuals, may be present at birth but more frequently develops during the first year of life. The degree of macrocephaly is variable, with head circumferences reaching four to six standard deviations greater than the mean. After the first year of life, head growth trajectory typically normalizes and growth follows a line parallel to, although several standard deviations above, the 98th centile. Initial mental and motor development is normal in most individuals. Walking is often unstable, followed by ataxia of the trunk and extremities, pyramidal dysfunction, and brisk deep tendon reflexes. Early-onset seizures are common, and approximately 60% of individuals have epilepsy that is typically well controlled with anti-seizure medication, but status epilepticus occurs relatively frequently. Cognitive deterioration occurs later in the course of the disease and is usually mild in severity. Overall disease severity varies, with some individuals being able to ambulate independently for only a few years from disease onset to other individuals continuing to independently walk in the fifth decade of life.
• Individuals with improving MLC have a similar initial presentation with delayed cognitive or motor development, followed by an improving clinical course: macrocephaly usually persists, but some children become normocephalic; motor function improves or normalizes; hypotonia and clumsiness may persist in some or neurologic examination may become normal. Some individuals have intellectual disability that is stable, with or without autism spectrum disorder. Epilepsy is much less frequent than in classic MLC.

Diagnosis/testing.

The diagnosis of classic MLC is established in individuals with suggestive clinical findings and characteristic abnormalities identified on brain MRI examination, including abnormal and swollen cerebral hemispheric white matter and subcortical cysts in the anterior temporal and often frontoparietal regions; and/or biallelic loss-of-function variants in MLC1 or HEPACAM or a heterozygous pathogenic variant in GPRC5B identified by molecular genetic testing.

The diagnosis of improving MLC is established in individuals with suggestive clinical findings and a heterozygous gain-of-function variant in HEPACAM or biallelic pathogenic variants in AQP4 identified by molecular genetic testing.

Management.

Treatment of manifestations: Physical therapy to improve motor function; speech therapy as needed; special education; anti-seizure medication to control seizures.

Agents/circumstances to avoid: Contact sports and other activities associated with a high risk of head trauma should be avoided.

Genetic counseling.

MLC is inherited in an autosomal recessive or an autosomal dominant manner.

• Classic MLC is most commonly caused by biallelic pathogenic variants in MLC1 or HEPACAM and is inherited in an autosomal recessive manner. If both parents of a child with MLC1-related classic MLC or biallelic HEPACAM-related classic MLC are known to be heterozygous for an MLC1 or HEPACAM pathogenic variant, each sib of an affected individual has at conception a 25% chance of inheriting biallelic pathogenic variants and being affected, a 50% chance of being heterozygous, and a 25% chance of inheriting neither of the familial pathogenic variants. Once the autosomal recessive MLC-related pathogenic variants have been identified in an affected family member, carrier testing for at-risk relatives is possible. Rarely, classic MLC occurs as an autosomal dominant disorder caused by a heterozygous GPRC5B pathogenic variant. All individuals with GPRC5B-related classic MLC reported to date have had the disorder as a result of a de novo pathogenic variant.
• Improving MLC most commonly occurs as an autosomal dominant disorder caused by an inherited or de novo pathogenic variant in HEPACAM. Approximately 20% of individuals with heterozygous HEPACAM-related improving MLC have the disorder as a result of a de novo pathogenic variant. Each child of an individual with heterozygous HEPACAM-related improving MLC has a 50% chance of inheriting the pathogenic variant. Rarely, improving MLC is due to biallelic pathogenic variants in AQP4 and is inherited in an autosomal recessive manner.

Once the MLC-related pathogenic variant(s) have been identified in an affected family member, prenatal and preimplantation genetic testing for MLC are possible.

Suggestive Findings

Two phenotypes are observed in megalencephalic leukoencephalopathy with subcortical cysts (MLC): classic MLC and improving MLC.

Classic MLC

Classic MLC should be suspected in individuals with the following clinical and brain imaging findings and family history.

Clinical findings

• Macrocephaly with onset that is either congenital or within the first year of life
• Normal or mildly delayed early development
• Slow deterioration of motor functions with cerebellar ataxia and mild spasticity
• Seizures
• Dysarthria
• Decline in cognitive function (occurs later and is typically milder than motor decline)
• Behavioral problems in some individuals
• Temporary exacerbation of signs and symptoms after minor head trauma

Imaging findings. Brain MRI features include the following abnormalities (see Figure 1):

Figure 1.

Brain images of an individual with MLC (A, C) and an unaffected individual (B, D) A. Transverse T₂-weighted image of a child age nine years with MLC, showing diffusely abnormal and mildly swollen white matter

• Diffusely abnormal and mildly swollen cerebral hemispheric white matter
• Subcortical cysts are almost invariably present in the anterior temporal region and often in the frontoparietal regions.
• Over time, white matter swelling decreases and cerebral atrophy ensues. The subcortical cysts may increase in size and number. In some individuals, the cysts become very large, occupying a large part of the frontoparietal white matter. In others, the white matter abnormalities decrease over time, and signal intensities become less abnormal.
• Diffusion-weighted imaging reveals increased diffusivity of abnormal white matter [Itoh et al 2006, van der Voorn et al 2006].

Family history. Family history is most often consistent with autosomal recessive inheritance (e.g., affected sibs and/or parental consanguinity). Rarely, a proband has a de novo pathogenic variant and represents a simplex case. Absence of a known family history does not preclude the diagnosis.

Establishing the Diagnosis

The clinical diagnosis of megalencephalic leukoencephalopathy with subcortical cysts (MLC) can be established in a proband based on the presence of clinical and imaging findings detailed in Suggestive Findings.

The molecular diagnosis of classic MLC is established in a proband with suggestive findings and biallelic pathogenic (or likely pathogenic) variants in MLC1 or HEPACAM or a heterozygous pathogenic (or likely pathogenic) variant in GPRC5B identified by molecular genetic testing.

The molecular diagnosis of improving MLC is established in a proband with suggestive findings and a heterozygous pathogenic (or likely pathogenic) variant in HEPACAM or biallelic pathogenic (or likely pathogenic) variants in AQP4 identified by molecular genetic testing.

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

Molecular genetic testing approaches can include a combination of gene-targeted testing (multigene panel) and comprehensive genomic testing (exome sequencing, genome sequencing). Gene-targeted testing requires that the clinician determine which gene(s) are likely involved (see Option 1), whereas comprehensive genomic testing does not (see Option 2).

Clinical Description

The two phenotypes observed in individuals with megalencephalic leukoencephalopathy with subcortical cysts (MLC) are classic MLC and improving MLC.

To date, approximately 500 individuals have been identified with biallelic pathogenic variants in MLC1 or HEPACAM, or heterozygous pathogenic variants in HEPACAM [Hamilton et al 2018]. To date, three individuals with heterozygous pathogenic variants in GPRC5B and classic MLC as well as two individuals with a biallelic pathogenic variant in AQP4 and improving MLC have been identified [Passchier et al 2023]. The following description of the phenotypic features associated with this spectrum is based on these reports.

Phenotype Correlations by Gene

Classic MLC. A review of 17 individuals with biallelic HEPACAM-related classic MLC revealed no phenotypic differences from individuals with MLC1-related classic MLC [Hamilton et al 2018].

The three individuals with GPRC5B-related classic MLC are phenotypically indistinguishable from individuals with classic MLC due to biallelic pathogenic variants in MLC1 or HEPACAM [Passchier et al 2023].

Improving MLC. Only two individuals (sibs) have been described to date with biallelic pathogenic variants in AQP4 [Passchier et al 2023], so it is currently unclear whether heterozygous HEPACAM-related improving MLC and AQP4-related MLC are phenotypically distinguishable.

Genotype-Phenotype Correlations

AQP4. Two sibs with biallelic AQP4 pathogenic variants have been reported with improving MLC similar to heterozygous HEPACAM-related improving MLC [Passchier et al 2023].

GPRC5B. Three unrelated individuals with de novo heterozygous pathogenic variants in GPRC5B have been described [Passchier et al 2023]. For these individuals, brain MRI presentation and clinical course are indistinguishable from MLC1-related classic MLC and biallelic HEPACAM-related classic MLC.

HEPACAM. Most known biallelic and heterozygous HEPACAM pathogenic variants affect the extracellular or transmembrane domains of the protein [López-Hernández et al 2011, Elorza-Vidal et al 2020].

• In biallelic HEPACAM-related classic MLC, pathogenic variants in HEPACAM are widely distributed over the entire extracellular and transmembrane region of the protein, whereas in heterozygous HEPACAM-related improving MLC, pathogenic variants are clustered in the first extracellular immunoglobulin-like domain [López-Hernández et al 2011, Elorza-Vidal et al 2020].
• HEPACAM pathogenic variants associated with autosomal recessive classic MLC and HEPACAM pathogenic variants associated with autosomal dominant improving MLC do not overlap, although they may affect the same residue [López-Hernández et al 2011]. At present, it is unclear why some HEPACAM pathogenic variants are associated with autosomal recessive classic MLC and others with autosomal dominant improving MLC. Notably, only missense variants in HEPACAM have been reported in improving MLC [López-Hernández et al 2011, van der Knaap et al 2012, Elorza-Vidal et al 2020].

MLC1. A review of 187 individuals with biallelic MLC1 pathogenic variants revealed that disease severity and clinical course can vary significantly in individuals from the same family [Hamilton et al 2018]. There are no known genotype-phenotype correlations to date.

Nomenclature

Subtypes of megalencephalic leukoencephalopathy with subcortical cysts (MLC) were previously named according to the genetic loci associated with each subtype:

• Classic MLC (MLC1, MLC2A, and MLC3)
  • MLC1, associated with biallelic pathogenic variants in MLC1
  • MLC2A, associated with biallelic pathogenic variants in HEPACAM
  • MLC3, associated with heterozygous pathogenic variants in GPRC5B
• Improving MLC (MLC2B and MLC4)
  • MLC2B, associated with heterozygous pathogenic variants in HEPACAM
  • MLC4, associated with biallelic pathogenic variants in AQP4

Other names previously used for MLC:

• Leukoencephalopathy with swelling and a discrepantly mild course
• Leukoencephalopathy with swelling and cysts
• Infantile leukoencephalopathy and megalencephaly
• Vacuolating leukoencephalopathy

Penetrance

Classic MLC. The penetrance of biallelic pathogenic variants in MLC1 or HEPACAM and heterozygous pathogenic variants in GPRC5B is expected to be complete.

Improving MLC. The penetrance of heterozygous pathogenic variants in HEPACAM is reduced [López-Hernández et al 2011]. The proportion of individuals with a heterozygous HEPACAM variant who exhibit or have exhibited clinical manifestations of improving MLC is unknown. The penetrance of biallelic pathogenic AQP4 variants is unknown.

Prevalence

There are several populations where founder variants in MLC-associated genes have been identified (see Table 8).

Founder variants in MLC1 have been identified in East Indian individuals from the Agrawal community [Leegwater et al 2002, Singhal et al 2003, Gorospe et al 2004], Libyan and Turkish Jewish individuals [Ben-Zeev et al 2002], Korean individuals [Choi et al 2017], and Egyptian individuals [Abdel-Salam et al 2016].

Evaluations Following Initial Diagnosis

To establish the extent of disease and needs in an individual diagnosed with megalencephalic leukoencephalopathy with subcortical cysts (MLC), the evaluations summarized in Table 4 (if not performed as part of the evaluation that led to the diagnosis) are recommended.

Treatment of Manifestations

There is no cure for MLC.

Supportive care to improve quality of life, maximize function, and reduce complications is recommended. This ideally involves multidisciplinary care by specialists in relevant fields (see Table 5).

Surveillance

To monitor existing manifestations, the individual's response to supportive care, and the emergence of new manifestations, the evaluations summarized in Table 6 are recommended.

Agents/Circumstances to Avoid

In individuals with classic MLC, minor head trauma may lead to temporary motor deterioration, seizures, or (rarely) coma. For this reason, contact sports and other activities with a high risk of head trauma should be avoided in affected individuals. Wearing a helmet should also be considered for situations associated with increased risk of head trauma.

Evaluation of Relatives at Risk

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

Pregnancy Management

Potential teratogenic effects of anti-seizure medication should be discussed with affected women of childbearing age, ideally prior to conception.

See MotherToBaby for further information on medication use during pregnancy.

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

Classic megalencephalic leukoencephalopathy with subcortical cysts (MLC) is most commonly caused by biallelic pathogenic variants in MLC1 or HEPACAM and inherited in an autosomal recessive manner. Rarely, classic MLC occurs as an autosomal dominant disorder caused by a de novo heterozygous GPRC5B pathogenic variant.

Improving MLC most commonly occurs as an autosomal dominant disorder caused by an inherited or de novo heterozygous pathogenic variant in HEPACAM. Rarely, improving MLC is caused by biallelic pathogenic variants in AQP4 and inherited in an autosomal recessive manner.

Note: HEPACAM pathogenic variants associated with classic MLC and HEPACAM pathogenic variants associated with improving MLC do not overlap, although they may affect the same residue [López-Hernández et al 2011]. In a few individuals with a pathogenic variant associated with autosomal dominant HEPACAM-related improving MLC on one allele and a pathogenic variant associated with autosomal recessive HEPACAM-related classic MLC on the other allele, classic MLC was observed [Authors, personal communication]. In these individuals, risk to offspring depends on the transmitted pathogenic variant: offspring who inherit the allele associated with heterozygous HEPACAM-related improving MLC may be affected and offspring who inherit the allele associated with biallelic HEPACAM-related classic MLC will be asymptomatic carriers.

Autosomal Recessive Inheritance – Risk to Family Members

Parents of a proband

• The parents of a child with autosomal recessive MLC (i.e., a child with MLC1-related classic MLC, biallelic HEPACAM-related classic MLC, or AQP4-related improving MLC) are typically heterozygous for an autosomal recessive MLC-related pathogenic variant.
• If a molecular diagnosis has been established in the proband, molecular genetic testing is recommended for the parents of the proband to confirm that both parents are heterozygous for an autosomal recessive MLC-related pathogenic variant and to allow reliable recurrence risk assessment.
• If a pathogenic variant is detected in only one parent and parental identity testing has confirmed biological maternity and paternity, it is possible that one of the pathogenic variants identified in the proband occurred as a de novo event in the proband [López-Hernández et al 2011] or as a postzygotic de novo event in a mosaic parent. If the proband appears to have homozygous pathogenic variants (i.e., the same two pathogenic variants), additional possibilities to consider include:
  • A single- or multiexon deletion in the proband that was not detected by sequence analysis and that resulted in the artifactual appearance of homozygosity;
  • Uniparental isodisomy for the parental chromosome with the pathogenic variant that resulted in homozygosity for the pathogenic variant in the proband.
• Individuals who are heterozygous for an MLC1 pathogenic variant are typically asymptomatic.
• Individuals who are heterozygous for a HEPACAM pathogenic variant associated with classic MLC are asymptomatic. HEPACAM pathogenic variants associated with autosomal dominant improving MLC do not overlap with HEPACAM pathogenic variants associated with autosomal recessive classic MLC (see Genotype-Phenotype Correlations; also see Note under Mode of Inheritance).
• Individuals who are heterozygous for an AQP4 pathogenic variant are asymptomatic to date.

Sibs of a proband

• If both parents are known to be heterozygous for a pathogenic variant associated with autosomal recessive MLC, each sib of an affected individual has at conception a 25% chance of inheriting biallelic pathogenic variants and being affected, a 50% chance of being heterozygous, and a 25% chance of inheriting neither of the familial pathogenic variants.
• Intrafamilial variability has been observed in MLC1-related classic MLC. A review of 187 individuals with biallelic MLC1 pathogenic variants revealed that in individuals from the same family, disease severity and clinical course can vary significantly [Hamilton et al 2018].
• Individuals who are heterozygous for an MLC1 pathogenic variant are typically asymptomatic.
• Individuals who are heterozygous for a HEPACAM pathogenic variant associated with classic MLC are asymptomatic. HEPACAM pathogenic variants associated with autosomal dominant improving MLC do not overlap with HEPACAM pathogenic variants associated with autosomal recessive classic MLC (see Genotype-Phenotype Correlations; also see Note under Mode of Inheritance).
• Individuals who are heterozygous for an AQP4 pathogenic variant are asymptomatic to date.

Offspring of a proband. The offspring of an individual with autosomal recessive MLC (i.e., MLC1-related classic MLC, biallelic HEPACAM-related classic MLC, or AQP4-related improving MLC) are obligate heterozygotes (carriers) for a pathogenic variant.

Other family members. Each sib of the proband's parents is at a 50% risk of being a carrier of a pathogenic variant associated with autosomal recessive MLC.

Autosomal Dominant Inheritance – Risk to Family Members

Parents of a proband

• Many individuals with heterozygous HEPACAM-related improving MLC have a parent with macrocephaly or a history of childhood macrocephaly. Given the neurologic improvement that is observed in this phenotype, the parent is not at risk of neurologic regression. Whether having a heterozygous gain-of-function HEPACAM pathogenic variant leads to other manifestations late in life has not been investigated; however, to date no evidence suggests disease progression later in life.
• A proband with heterozygous HEPACAM-related improving MLC may have the disorder as the result of a de novo HEPACAM pathogenic variant. The proportion of individuals with HEPACAM-related improving MLC caused by a de novo HEPACAM pathogenic variant is approximately 20%.
• In the three individuals reported to date with GPRC5B-related classic MLC, the heterozygous pathogenic GPRC5B variant arose de novo.
• If the proband appears to be the only affected family member (i.e., a simplex case), molecular genetic testing is recommended for the parents of the proband to confirm their genetic status and to allow reliable recurrence risk counseling.
• If the pathogenic variant identified in the proband is not identified in either parent and parental identity testing has confirmed biological maternity and paternity, the following possibilities should be considered:
  • The proband has a de novo pathogenic variant.
  • The proband inherited a pathogenic variant from a parent with germline (or somatic and germline) mosaicism. Note: Testing of parental leukocyte DNA may not detect all instances of somatic mosaicism and will not detect a pathogenic variant that is present in the germ cells only.
• The family history of some individuals diagnosed with heterozygous HEPACAM-related improving MLC may appear to be negative because of failure to recognize the disorder in family members, reduced penetrance, or milder phenotypic presentation. Therefore, an apparently negative family history cannot be confirmed unless molecular genetic testing has demonstrated that neither parent is heterozygous for the pathogenic variant identified in the proband.

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

• If a parent of the proband is affected and/or is known to have the autosomal dominant MLC-related pathogenic variant identified in the proband, the risk to the sibs of inheriting the variant is 50%.
• If the heterozygous HEPACAM or GPRC5B pathogenic variant found in the proband cannot be detected in the leukocyte DNA of either parent, the recurrence risk to sibs is estimated to be 1% because of the theoretic possibility of parental germline mosaicism [Rahbari et al 2016].
• If the parents have not been tested for the heterozygous HEPACAM or GPRC5B pathogenic variant but are clinically unaffected, the sibs are still at increased risk for heterozygous HEPACAM-related improving MLC or GPRC5B-related classic MLC because of the possibility of reduced penetrance in a heterozygous parent or the theoretic possibility of parental germline mosaicism.

Offspring of a proband. Each child of an individual with autosomal dominant MLC (i.e., heterozygous HEPACAM-related improving MLC or GPRC5B-related classic MLC) has a 50% chance of inheriting the pathogenic variant.

Other family members. The risk to other family members depends on the status of the proband's parents: if a parent has the heterozygous HEPACAM or GPRC5B pathogenic variant, the parent's family members may be at risk.

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 carriers (or are at risk of being carriers) of MLC-related pathogenic variants.

DNA banking. Because it is likely that testing methodology and our understanding of genes, pathogenic mechanisms, and diseases will improve in the future, consideration should be given to banking DNA from probands in whom a molecular diagnosis has not been confirmed (i.e., the causative pathogenic mechanism is unknown). For more information, see Huang et al [2022].

Prenatal Testing and Preimplantation Genetic Testing

Once the pathogenic variant(s) have been identified in an affected family member, prenatal and preimplantation genetic testing for MLC 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

MLC1. Membrane protein MLC1 (MLC1) is highly expressed in the human brain, as well as in leukocytes. Within the human brain, MLC1 is almost exclusively expressed in astrocytic endfeet in the perivascular, subependymal, and subpial regions. The localization of the protein, combined with the clinical phenotype of megalencephalic leukoencephalopathy with subcortical cysts (MLC), suggests a role for MLC1 in brain ion and water homeostasis [van der Knaap et al 2012]. MLC1 shares its endfeet localization with members of the dystrophin glycoprotein-associated complex, including aquaporin-4 [Boor et al 2007]. MLC1 has been shown to interact with a variety of proteins potentially involved in brain ion and water homeostasis. These include the Na⁺/K⁺-ATPase, TRPV4, caveolin-1, and Kir4.1, among others [Brignone et al 2015].

MLC1 pathogenic variants show a low steady-state expression of MLC1 with a consequent reduction in surface expression [Teijido et al 2004, Montagna et al 2006]. Electrophysiologic studies reveal a reduction of volume-regulated anion channel (VRAC) activity in lymphoblasts from individuals with biallelic MLC1 pathogenic variants. This results in impaired regulatory volume decrease upon hypotonic cell swelling [Ridder et al 2011]. Cell lines overexpressing normal MLC1 show enhanced VRAC activity, which is absent upon overexpression of mutated MLC1, supporting the role of MLC1 in VRAC regulation [Ridder et al 2011].

HEPACAM encodes hepatic and glial cell adhesion molecule (called either hepaCAM or glialCAM), a single transmembrane protein with two extracellular immunoglobulin domains that is predominantly expressed in the brain. GlialCAM colocalizes with MLC1 in astrocyte endfeet. However, glialCAM is also expressed in axons, on the outside of myelin sheaths, and in oligodendrocytes [Favre-Kontula et al 2008, López-Hernández et al 2011]. GlialCAM acts as a chaperone for MLC1, ensuring the localization of both proteins at astrocyte-astrocyte junctions in endfeet and in cultured astrocytes [López-Hernández et al 2011, Capdevila-Nortes et al 2013]. GlialCAM also acts as an auxiliary subunit for the chloride channel ClC-2. It increases ClC-2 mediated currents, changes their functional properties, and is necessary for ClC-2 localization to astrocyte-astrocyte junctions [Jeworutzki et al 2012, Hoegg-Beiler et al 2014]. GlialCAM also interacts with the gap junction subunit connexin 43 [Wu et al 2016].

All MLC-causing variants in HEPACAM affect the extracellular region of glialCAM. In cultured rat primary astrocytes, mutated glialCAM with either a dominant or a recessive pathogenic variant disrupts localization of MLC1 and glialCAM at astrocyte-astrocyte junctions. Coexpression of wild type glialCAM rescues the detrimental effect of glialCAM with a recessive variant on MLC1 localization, but it does not rescue the effect of the dominant variant of glialCAM [López-Hernández et al 2011]. Biochemical analysis shows that some dominant and recessive pathogenic variants disrupt the ability of glialCAM to homo-oligomerize [Arnedo et al 2014]. However, what causes a HEPACAM variant to be dominant vs recessive is still unclear.

GPRC5B encodes G-protein coupled receptor (GPCR) family C group 5 member B (GPRC5B). In the brain, GPRC5B is highly expressed in perivascular astrocyte endfeet, a location shared with MLC1 and glialCAM [Alonso-Gardón et al 2021, Passchier et al 2023]. GPCRs can be activated by different types of chemical or mechanical stimuli, or by interaction with other proteins. GPRC5B is considered an orphan receptor since its ligand has not been identified. An interaction of GPRC5B with MLC1 and glialCAM has been described [Alonso-Gardón et al 2021]. Volume regulation in lymphoblasts from GPRC5B patients is disrupted [Passchier et al 2023]. Knockdown of GPRC5B in astrocytes reduces VRAC activity [Alonso-Gardón et al 2021], while overexpression in astrocytoma cells increases VRAC activity [Passchier et al 2023]. Therefore, expression patterns and cellular mechanisms linked to GPRC5B overlap with those of MLC1 and glialCAM. The two dominant pathogenic variants that have been identified in GPRC5B are duplications of two neighboring amino acids in the fourth transmembrane region of the receptor (p.Ile176dup and p.Ala177dup) [Passchier et al 2023]. How these variants alter the function of the receptor is currently unknown.

AQP4 encodes the ubiquitous brain water channel aquaporin-4. Like the other proteins involved in MLC, aquaporin-4 is highly expressed in perivascular astrocyte endfeet and at other brain-fluid interfaces. At these locations, aquaporin-4, similar to MLC1, is part of the dystrophin glycoprotein-associated complex [Boor et al 2007]. The presence of large numbers of water channels at astrocyte endfeet enables fast water exchange between the extracellular space and the interconnected network of astrocytes and other glial cells [Nagelhus & Ottersen 2013]. These combined features increase the ability of astrocytes to rapidly adjust cell volume and increases their capacity for ion and water homeostasis, but simultaneously makes them more prone to pathologic swelling in situations of acute edema formation. Modulation of aquaporin-4 function or localization is being investigated as potential therapeutic strategy for the treatment of acute brain edema [Kitchen et al 2020].

The recessive pathogenic variant p.Ala215Thr in aquaporin-4 alters the second of two highly conserved Asn-Pro-Ala (NPA) motifs that form the narrow central part of the water channel. These motifs are crucial for selective water permeability and for localization of the channel to the plasma membrane [King et al 2004]. The p.Ala215Thr variant leads to protein misfolding, resulting in loss of membrane localization and water channel function [Passchier et al 2023].

Mechanism of disease causation

• Loss of function. MLC1, biallelic HEPACAM (classic MLC), and AQP4 pathogenic variants
• Gain of function or dominant negative. Heterozygous HEPACAM (improving MLC) and GPRC5B pathogenic variants

Author Notes

The authors are happy to offer cDNA analysis of MLC1 on lymphoblast cell lines of individuals with MLC without an identified molecular etiology in any of the known genes. If a molecular etiology is not identified, the authors are also willing to examine brain MRIs of these individuals to help determine whether the MRI is diagnostic of MLC and, if so, offer further genetic analysis to identify the genetic basis.

Author History

Truus EM Abbink, PhD (2018-present)

Rogier Min, PhD (2018-present)

JC Pronk, PhD; Vrije Universiteit Medical Center, Amsterdam (2003-2008)

Gert C Scheper, PhD; Vrije Universiteit Medical Center, Amsterdam (2008-2018)

Marjo S van der Knaap, MD, PhD (2003-present)

Revision History

• 27 July 2023 (gm) Comprehensive update posted live
• 29 March 2018 (sw) Comprehensive update posted live
• 3 November 2011 (me) Comprehensive update posted live
• 29 July 2008 (me) Comprehensive update posted live
• 29 November 2005 (me) Comprehensive update posted live
• 11 August 2003 (me) Review posted live
• 12 June 2003 (mvdk) Original submission

Literature Cited

• Abdel-Salam GM, Abdel-Hamid MS, Ismail SI, Hosny H, Omar T, Effat L, Aglan MS, Temtamy SA, Zaki MS. Megalencephalic leukoencephalopathy with cysts in twelve Egyptian patients: novel mutations in MLC1 and HEPACAM and a founder effect. Metab Brain Dis. 2016;31:1171-9. [PubMed: 27389245]
• Alonso-Gardón M, Elorza-Vidal X, Castellanos A, La Sala G, Armand-Ugon M, Gilbert A, Di Pietro C, Pla-Casillanis A, Ciruela F, Gasull X, Nunes V, Martínez A, Schulte U, Cohen-Salmon M, Marazziti D, Estévez R. Identification of the GlialCAM interactome: the G protein-coupled receptors GPRC5B and GPR37L1 modulate megalencephalic leukoencephalopathy proteins. Hum Mol Genet. 2021;30:1649-65. [PMC free article: PMC8369841] [PubMed: 34100078]
• Arnedo T, López-Hernández T, Jeworutzki E, Capdevila-Nortes X, Sirisi S, Pusch M, Estévez R. Functional analyses of mutations in HEPACAM causing megalencephalic leukoencephalopathy. Hum Mutat. 2014;35:1175-8. [PubMed: 25044933]
• Balestri M, Crisafulli C, Donato L, Giegling I, Calati R, Antypa N, Schneider B, Marusic D, Tarozzi ME, Marusic D, Paragi M, Hartmann AM, Konte B, Marsano A, Serretti A, Rujescu D. Nine differentially expressed genes from a post mortem study and their association with suicidal status in a sample of suicide completers, attempters and controls. J Psychiatr Res. 2017;91:98-104. [PubMed: 28327445]
• Ben-Zeev B, Levy-Nissenbaum E, Lahat H, Anikster Y, Shinar Y, Brand N, Gross-Tzur V, MacGregor D, Sidi R, Kleta R, Frydman M, Pras E. Megalencephalic leukoencephalopathy with subcortical cysts; a founder effect in Israeli patients and a higher than expected carrier rate among Libyan Jews. Hum Genet. 2002;111:214-8. [PubMed: 12189496]
• Boor I, Nagtegaal M, Kamphorst W, van der Valk P, Pronk JC, van Horssen J, Dinopoulos A, Bove KE, Pascual-Castroviejo I, Muntoni F, Estévez R, Scheper GC, van der Knaap MS. MLC1 is associated with the dystrophin-glycoprotein complex at astrocytic endfeet. Acta Neuropathol. 2007;114:403-10. [PMC free article: PMC2039857] [PubMed: 17628813]
• Brignone MS, Lanciotti A, Camerini S, De Nuccio C, Petrucci TC, Visentin S, Ambrosini E. MLC1 protein: a likely link between leukodystrophies and brain channelopathies. Front Cell Neurosci. 2015;9:66. [PMC free article: PMC4381631] [PubMed: 25883547]
• Bugiani M, Moroni I, Bizzi A, Nardocci N, Bettecken T, Gärtner J, Uziel G. Consciousness disturbances in megalencephalic leukoencephalopathy with subcortical cysts. Neuropediatrics. 2003;34:211-4. [PubMed: 12973663]
• Capdevila-Nortes X, López-Hernández T, Apaja PM, López de Heredia M, Sirisi S, Callejo G, Arnedo T, Nunes V, Lukacs GL, Gasull X, Estévez R. Insights into MLC pathogenesis: GlialCAM is an MLC1 chaperone required for proper activation of volume-regulated anion currents. Hum Mol Genet. 2013;22:4405-16. [PubMed: 23793458]
• Choi SA, Kim SY, Yoon J, Choi J, Park SS, Seong MW, Kim H, Hwang H, Choi JE, Chae JH, Kim KJ, Kim S, Lee YJ, Nam SO, Lim BC. A unique mutational spectrum of MLC1 in Korean patients with megalencephalic leukoencephalopathy with subcortical cysts. Ann Lab Med. 2017; 37:516-21. [PMC free article: PMC5587825] [PubMed: 28840990]
• Devaney JM, Donarum EA, Brown KM, Meyer J, Stöber G, Lesch KP, Nestadt G, Stephan DA, Pulver AE. No missense mutation of WKL1 in a subgroup of probands with schizophrenia. Mol Psychiatry. 2002;7:419-23 [PubMed: 11986987]
• Duarri A, Lopez de Heredia M, Capdevila-Nortes X, Ridder MC, Montolio M, López-Hernández T, Boor I, Lien CF, Hagemann T, Messing A, Gorecki DC, Scheper GC, Martínez A, Nunes V, van der Knaap MS, Estévez R. Knockdown of MLC1 in primary astrocytes causes cell vacuolation: a MLC disease cell model. Neurobiol Dis. 2011;43:228-38. [PMC free article: PMC3885813] [PubMed: 21440627]
• Dubey M, Brouwers E, Hamilton EMC, Stiedl O, Bugiani M, Koch H, Kole MHP, Boschert U, Wykes RC, Mansvelder HD, van der Knaap MS, Min R. Seizures and disturbed brain potassium dynamics in the leukodystrophy megalencephalic leukoencephalopathy with subcortical cysts. Ann Neurol. 2018; 83:636-49. [PMC free article: PMC5900999] [PubMed: 29466841]
• Elorza-Vidal X, Xicoy-Espaulella E, Pla-Casillanis A, Alonso-Gardón M, Gaitán-Peñas H, Engel-Pizcueta C, Fernández-Recio J, Estévez R. Structural basis for the dominant or recessive character of GLIALCAM mutations found in leukodystrophies. Hum Mol Genet. 2020;29:1107-20. [PMC free article: PMC7206653] [PubMed: 31960914]
• Favre-Kontula L, Rolland A, Bernasconi L, Karmirantzou M, Power C, Antonsson B, Boschert U. GlialCAM, an immunoglobulin-like cell adhesion molecule is expressed in glial cells of the central nervous system. Glia. 2008;56:633-45. [PubMed: 18293412]
• Fujino S, Yoshihashi H, Takeda R, Ihara S, Miyama S. White matter abnormality in Jacobsen syndrome assessed by serial MRI. Brain Dev. 2020;42:621-5. [PubMed: 32507665]
• Gorospe JR, Singhal BS, Kainu T, Wu F, Stephan D, Trent J, Hoffman EP, Naidu S. Indian Agarwal megalencephalic leukodystrophy with cysts is caused by a common MLC1 mutation. Neurology. 2004;62:878-82. [PubMed: 15037685]
• Hamilton EMC, Tekturk P, Cialdella F, van Rappard DF, Wolf NI, Yalcinkaya C, Çetinçelik Ü, Rajaee A, Kariminejad A, Paprocka J, Yapici Z, Bošnjak VM, van der Knaap MS, et al. Megalencephalic leukoencephalopathy with subcortical cysts: characterization of disease variants. Neurology. 2018;90:e1395-403. [PMC free article: PMC5902784] [PubMed: 29661901]
• Hoegg-Beiler MB, Sirisi S, Orozco IJ, Ferrer I, Hohensee S, Auberson M, Gödde K, Vilches C, de Heredia ML, Nunes V, Estévez R, Jentsch TJ. Disrupting MLC1 and GlialCAM and ClC-2 interactions in leukodystrophy entails glial chloride channel dysfunction. Nat Commun. 2014;5:3475. [PubMed: 24647135]
• Huang SJ, Amendola LM, Sternen DL. Variation among DNA banking consent forms: points for clinicians to bank on. J Community Genet. 2022;13:389-97. [PMC free article: PMC9314484] [PubMed: 35834113]
• Ilja Boor PK, de Groot K, Mejaski-Bosnjak V, Brenner C, van der Knaap MS, Scheper GC, Pronk JC. Megalencephalic leukoencephalopathy with subcortical cysts: an update and extended mutation analysis of MLC1. Hum Mutat. 2006;27:505-12. [PubMed: 16652334]
• Itoh N, Maeda M, Naito Y, Narita Y, Kuzuhara S. An adult case of megalencephalic leukoencephalopathy with subcortical cysts with S93L mutation in MLC1 gene: a case report and diffusion MRI. Eur Neurol. 2006;56:243-5. [PubMed: 17077634]
• Jeworutzki E, López-Hernández T, Capdevila-Nortes X, Sirisi S, Bengtsson L, Montolio M, Zifarelli G, Arnedo T, Müller CS, Schulte U, Nunes V, Martínez A, Jentsch TJ, Gasull X, Pusch M, Estévez R. GlialCAM, a protein defective in a leukodystrophy, serves as a ClC-2 Cl(-) channel auxiliary subunit. Neuron. 2012;73:951-61. [PMC free article: PMC3334819] [PubMed: 22405205]
• Kaganovich M, Peretz A, Ritsner M, Bening Abu-Shach U, Attali B, Navon R. Is the WKL1 gene associated with schizophrenia? Am J Med Genet B Neuropsychiatr Genet. 2004;125B:31-7. [PubMed: 14755440]
• Kariminejad A, Rajaee A, Ashrafi MR, Alizadeh H, Tonekaboni SH, Malamiri RA, Ghofrani M, Karimzadeh P, Mohammadi MM, Baghalshooshtari A, Bozorgmehr B, Kariminejad MH, Postma N, Abbink TE, van der Knaap MS. Eight novel mutations in MLC1 from 18 Iranian patients with megalencephalic leukoencephalopathy with subcortical cysts. Eur J Med Genet. 2015;58:71-4. [PubMed: 25497041]
• King LS, Kozono D, Agre P. From structure to disease: the evolving tale of aquaporin biology. Nat Rev Mol Cell Biol. 2004;5:687-98. [PubMed: 15340377]
• Kitchen P, Salman MM, Halsey AM, Clarke-Bland C, MacDonald JA, Ishida H, Vogel HJ, Almutiri S, Logan A, Kreida S, Al-Jubair T, Winkel Missel J, Gourdon P, Törnroth-Horsefield S, Conner MT, Ahmed Z, Conner AC, Bill RM. Targeting aquaporin-4 subcellular localization to treat central nervous system edema. Cell. 2020;181:784-99.e19. [PMC free article: PMC7242911] [PubMed: 32413299]
• Leegwater PA, Boor PK, Yuan BQ, van der Steen J, Visser A, Konst AA, Oudejans CB, Schutgens RB, Pronk JC, van der Knaap MS. Identification of novel mutations in MLC1 responsible for megalencephalic leukoencephalopathy with subcortical cysts. Hum Genet. 2002;110:279-83. [PubMed: 11935341]
• Leegwater PA, Yuan BQ, van der Steen J, Mulders J, Konst AA, Boor PK, Mejaski-Bosnjak V, van der Maarel SM, Frants RR, Oudejans CB, Schutgens RB, Pronk JC, van der Knaap MS. Mutations of MLC1 (KIAA0027), encoding a putative membrane protein, cause megalencephalic leukoencephalopathy with subcortical cysts. Am J Hum Genet. 2001;68:831-8. [PMC free article: PMC1275636] [PubMed: 11254442]
• López-Hernández T, Ridder MC, Montolio M, Capdevila-Nortes X, Polder E, Sirisi S, Duarri A, Schulte U, Fakler B, Nunes V, Scheper GC, Martínez A, Estévez R, van der Knaap MS. Mutant GlialCAM causes megalencephalic leukoencephalopathy with subcortical cysts, benign familial macrocephaly, and macrocephaly with retardation and autism. Am J Hum Genet. 2011;88:422-32. [PMC free article: PMC3071909] [PubMed: 21419380]
• Mattina T, Perrotta CS, Grossfeld P. Jacobsen syndrome. Orphanet J Rare Dis. 2009;4:9. [PMC free article: PMC2670819] [PubMed: 19267933]
• Meyer J, Huberth A, Ortega G, Syagailo YV, Jatzke S, Mossner R, Strom TM, Ulzheimer-Teuber I, Stober G, Schmitt A, Lesch KP. A missense mutation in a novel gene encoding a putative cation channel is associated with catatonic schizophrenia in a large pedigree. Mol Psychiatry. 2001;6:302-6. [PubMed: 11326298]
• Montagna G, Teijido O, Eymard-Pierre E, Muraki K, Cohen B, Loizzo A, Grosso P, Tedeschi G, Palacín M, Boespflug-Tanguy O, Bertini E, Santorelli FM, Estévez R. Vacuolating megalencephalic leukoencephalopathy with subcortical cysts: functional studies of novel variants in MLC1. Hum Mutat. 2006;27:292. [PubMed: 16470554]
• Nagelhus EA, Ottersen OP. Physiological roles of aquaporin-4 in the brain. Physiol Rev. 2013;93:1543-62. [PMC free article: PMC3858210] [PubMed: 24137016]
• Passchier EMJ, Kerst S, Brouwers E, Hamilton EMC, Bisseling Q, Bugiani M, Waisfisz Q, Kitchen P, Unger L, Breur M, Hoogterp L, de Vries SI, Abbink TEM, Kole MHP, Leurs R, Vischer HF, Brignone MS, Ambrosini E, Feillet F, Born AP, Epstein LG, Mansvelder HD, Min R, van der Knaap MS. Aquaporin-4 and GPRC5B: old and new players in controlling brain oedema. Brain. 2023. Epub ahead of print. [PMC free article: PMC10393393] [PubMed: 37143309]
• Rahbari R, Wuster A, Lindsay SJ, Hardwick RJ, Alexandrov LB, Turki SA, Dominiczak A, Morris A, Porteous D, Smith B, Stratton MR, Hurles ME, et al. Timing, rates and spectra of human germline mutation. Nat Genet. 2016;48:126-33. [PMC free article: PMC4731925] [PubMed: 26656846]
• Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, Grody WW, Hegde M, Lyon E, Spector E, Voelkerding K, Rehm HL, et al. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med. 2015;17:405-24. [PMC free article: PMC4544753] [PubMed: 25741868]
• Ridder MC, Boor I, Lodder JC, Postma NL, Capdevila-Nortes X, Duarri A, Brussaard AB, Estévez R, Scheper GC, Mansvelder HD, van der Knaap MS. Megalencephalic leucoencephalopathy with cysts: defect in chloride currents and cell volume regulation. Brain. 2011;134:3342-54. [PubMed: 22006981]
• Rubie C, Lichtner P, Gartner J, Siekiera M, Uziel G, Kohlmann B, Kohlschutter A, Meitinger T, Stober G, Bettecken T. Sequence diversity of KIAA0027/MLC1: are megalencephalic leukoencephalopathy and schizophrenia allelic disorders? Hum Mutat. 2003;21:45-52. [PubMed: 12497630]
• Selch S, Strobel A, Haderlein J, Meyer J, Jacob CP, Schmitt A, Lesch KP, Reif A. MLC1 polymorphisms are specifically associated with periodic catatonia, a subgroup of chronic schizophrenia. Biol Psychiatry. 2007;61:1211-4. [PubMed: 17210142]
• Shimada S, Shimojima K, Masuda T, Nakayama Y, Kohji T, Tsukamoto H, Matsubasa T, Oka A, Yamamoto T. MLC1 mutations in Japanese patients with megalencephalic leukoencephalopathy with subcortical cysts. Hum Genome Var. 2014;1:14019. [PMC free article: PMC4785519] [PubMed: 27081509]
• Singhal BS, Gorospe JR, Naidu S. Megalencephalic leukoencephalopathy with subcortical cysts. J Child Neurol. 2003;18:646-52. [PubMed: 14572144]
• Spijker S, Van Zanten JS, De Jong S, Penninx BW, van Dyck R, Zitman FG, Smit JH, Ylstra B, Smit AB, Hoogendijk WJ. Stimulated gene expression profiles as a blood marker of major depressive disorder. Biol Psychiatry. 2010;68:179-86. [PubMed: 20471630]
• Stenson PD, Mort M, Ball EV, Chapman M, Evans K, Azevedo L, Hayden M, Heywood S, Millar DS, Phillips AD, Cooper DN. The Human Gene Mutation Database (HGMD®): optimizing its use in a clinical diagnostic or research setting. Hum Genet. 2020;139:1197-207. [PMC free article: PMC7497289] [PubMed: 32596782]
• Sugiura C, Shiota M, Maegaki Y, Yoshida K, Koeda T, Kitahara T, Ohno K. Late-onset neuropsychological symptoms in a Japanese patient with megalencephalic leukoencephalopathy with subcortical cysts. Neuropediatrics. 2006;37:286-90. [PubMed: 17236107]
• Teijido O, Martínez A, Pusch M, Zorzano A, Soriano E, Del Río JA, Palacín M, Estévez R. Localization and functional analyses of the MLC1 protein involved in megalencephalic leukoencephalopathy with subcortical cysts. Hum Mol Genet. 2004;13:2581-94. [PubMed: 15367490]
• van der Knaap MS, Barth PG, Vrensen GF, Valk J. Histopathology of an infantile-onset spongiform leukoencephalopathy with a discrepantly mild clinical course. Acta Neuropathol (Berl). 1996;92:206-12. [PubMed: 8841668]
• van der Knaap MS, Boor I, Estévez R. Megalencephalic leukoencephalopathy with subcortical cysts: chronic white matter oedema due to a defect in brain ion and water homoeostasis. Lancet Neurol. 2012;11:973-85. [PubMed: 23079554]
• van der Voorn JP, Pouwels PJW, Hart AA, Serrarens J, Willemsen MA, Kremer HP, Barkhof F, van der Knaap MS. Childhood white matter disorders: quantitative MR imaging and spectroscopy. Radiology. 2006;241:510-7. [PubMed: 17057071]
• Verma R, Mukerji M, Grover D, B-Rao C, Das SK, Kubendran S, Jain S, Brahmachari SK. MLC1 gene is associated with schizophrenia and bipolar disorder in Southern India. Biol Psychiatry. 2005;58:16-22. [PubMed: 15992519]
• Wu M, Moh MC, Schwarz H. HepaCAM associates with connexin 43 and enhances its localization in cellular junctions. Sci Rep. 2016;6:36218. [PMC free article: PMC5098153] [PubMed: 27819278]
• Yamamoto T, Shimada S, Shimojima K, Sangu N, Ninomiya S, Kubota M. Leukoencephalopathy associated with 11q24 deletion involving the gene encoding hepatic and glial cell adhesion molecule in two patients. Eur J Med Genet. 2015;58:492-6. [PubMed: 26193381]