Submission Details

Normal Pressure Hydrocephalus (NPH) is a neurological disorder characterized by an abnormal accumulation of cerebrospinal fluid (CSF) in the brain’s ventricles and cavities. Normally, CSF is secreted by choroid cells into the lateral ventricles, flows to the third and fourth ventricles and then to the spinal cord. If the flow of CSF throughout the brain and spinal cord is in some way disrupted and CSF accumulates, the ventricles enlarge, resulting in hydrocephalus. The difference between NPH and other forms of hydrocephalus is that even though CT or MRI brain imaging of hydrocephalus patients shows enlarged ventricles, intracranial pressure remains within normal range (PMID: 30707192). NPH is a chronic, steadily progressing disease, frequently not diagnosed until patients are 65 years old or older (PMID: 32160166). In addition to the enlargement of ventricles, NPH patients develop gait and balance disturbances as the dominant symptom, often accompanied by cognitive decline and urinary incontinence. In humans, the synchronized beat of ependymal motile cilia helps drive circulation of the CSF within brain spaces. Loss or dysfunction of cilia in the brain ventricles is associated with restrained CSF circulation, causing CSF accumulation in the ventricles (PMIDs: 35903173, 34233705).

A mutation was first reported in the CFAP43 (Cilia-and Flagella-Associated Protein 43) gene in association with Normal Pressure Hydrocephalus 1 in a Japanese family (PMID: 31004071). Whole exome sequencing of a 55 year old woman and two unaffected siblings was used to identify a heterozygous nonsense CFAP43 mutation present in both tested affected family members and absent in three unaffected siblings. The genotypes of the affected mother and a third affected sibling were not available. The specific disease entity, Normal Pressure Hydrocephalus 1, is an autosomal dominant disorder caused by monoallelic mutation in the CFAP43 gene. Per criteria outlined by the ClinGen Lumping and Splitting Working Group, we found the autosomal dominant mode of inheritance and the phenotypes seen in family members with NPH1 to be distinct from the molecular mechanism, autosomal recessive mode of inheritance, and phenotype of male patients with Spermatogenic Failure 19. We are treating Normal Pressure Hydrocephalus 1 (MONDO:0009366, OMIM:236690) and Spermatogenic Failure 19 (MONDO: 0054723, OMIM: 617592) as separate disease entities.

Of note, in addition to the usual triad of symptoms associated with NPH (gait disturbance, cognitive impairment, and urinary incontinence) two of the three affected siblings were described as having recurrent respiratory tract infections and chronic sinusitis. While these patients carry a variant in a cilia and flagella associated gene (CFAP43), they had none of the other hallmark features of Primary Ciliary Dyskinesia. There was no report of neonatal respiratory distress, no laterality defects, no bronchiectasis, no chronic ear infections, no nasal nitric oxide measurements, no studies of ciliary function, and fertility was not assessed. Per criteria outlined by the ClinGen Lumping and Splitting Working Group, we found the autosomal dominant mode of inheritance and phenotypes reported in these family members with NPH, to be distinct from those of patients with Primary Ciliary Dyskinesia. We are treating Normal Pressure Hydrocephalus 1 (MONDO:0009366, OMIM:236690) and Primary Ciliary Dyskinesia (MONDO: 0016575) as separate disease entities.

Evidence from one proband described above has been included in this curation, representing one unique nonsense variant associated with NPH1. The mechanism of pathogenicity of this CFAP43 mutation remains undefined. There are no additional cases of CFAP43 related NPH1 in the currently available public literature, hence we scored only 0.5 out of 12 possible points for genetic evidence.

The gene-disease association between CFAP43 and Normal Pressure Hydrocephalus 1 is supported by gene expression and protein localization studies. Human RNAseq expression studies show that CFAP43 is expressed in ciliated tissues, including the brain (PMID:25970244). In mice, in situ expression studies show Cfap43 transcripts in multiciliated tissues, including brain cells (ependymal epithelial cells), during embryogenesis as well as in adult mice (PMID: 31884020). Immunofluorescent studies In Tetrahymena thermophila show that Fap43p is exclusively targeted to cilia and is present along the length of the cilium (PMID: 29687140).

Three animal models have been used to support the gene-disease relationship between CFAP43 and Normal Pressure Hydrocephalus 1. CRISPR/Cas9 technology was used in mice to generate a frameshift Cfap43 mutation similar to the nonsense CFAP43 mutation seen in human NPH1 patients (PMID: 31004071), probably in the hopes of replicating the human phenotype and providing insight into the molecular mechanism of disease. Cfap43-/- mice developed hydrocephalus. Authors also reported abnormalities in ciliary protein composition in ventricle and tracheal epithelia, ultrastructural defects (8+2 or 10+2 DMT axonemal structure) and compound cilia in mice tracheal cells, as well as sperm abnormalities in testis and epididymal tissues. The significance of these data is unclear as they are suggestive of several different types of defective ciliary phenotypes. In addition there is no information on the frequency of these abnormalities in Cfap43-/- mice so it is difficult to ascertain the strength of the data. The occasional observation of compound cilia, for example, is not uncommon and may be insignificant. Heterozygote Cfap43+/- mice showed no differences from wild-type mice. The hydrocephalus seen in this mouse model is consistent with the phenotype seen in NPH1 patients, but there is little data from this model to support a link between CFAP43 and brain ciliary dysfunction. The autosomal recessive inheritance pattern of the mouse model compared to the autosomal dominant inheritance suggested in human cases is also problematic and makes this, mechanistically, a weak animal model. A second mouse model using Cre/Lox technology also generated a Cfap43 knockout (PMID: 31884020). This mouse model confirms a role for Cfap43 in airway epithelia mucus clearance, brain ventricle development and function, and sperm development and function, all processes requiring motile cilia function. A loss of normal Cfap43 function in mice increased tracheal ciliary beat frequency and reduced tracheal cilia generated flow. Knockout mice retained motile ependymal cilia but developed enlarged ventricles and hydrocephalus. The autosomal recessive inheritance pattern of this mouse model compared to the autosomal dominant inheritance seen in human NPH1 cases is again, mechanistically, problematic and weakens the model. A Tetrahymena thermophila FAP43 knockout showed signs of impaired ciliary motility and had a reduced cilia beat amplitude (PMID: 29687140). Together, all three of these animal models confirm the plausibility that a mutation in human CFAP43 could impair ciliary motility and affect cerebrospinal fluid movement by ependymal cilia, leading to hydrocephalus.

Tetrahymena orthologs of human CFAP43 and CFAP44, Fap43p and Fap44p, were shown to be interacting proteins by two methods: Co-immunoprecipitation and a BirA* proximity labeling assay of Tetrahymena cell extracts, followed by mass spectrometry (PMID: 29687140). Authors of this study propose that a Fap43p-Fap44p complex might be involved in the regulation of ciliary beat.

Genetic and biochemical approaches combined with cryo-electron tomography identified Tetrahymena orthologs of CFAP43 and CFAP44, FAP43 and FAP44 as dimerized components of the T/TH (Tether/Tether Head) complex (PMID: 29514928). The T/TH complex is structurally linked to the inner dynein arm motor domains as well as to other axonemal complexes that may be involved in regulating ciliary motility. A change in ciliary function is consistent with a defective CFAP43 altering CSF flow in the brain.

In summary, due to the presence of only one CFAP43 variant associated with NPH1, despite strong experimental evidence for CFAP43 playing a role in motile ciliary function in other organisms, there is limited evidence supporting a gene-disease relationship between variants in CFAP43 and Normal Pressure Hydrocephalus 1. This classification was approved by the ClinGen Motile Ciliopathy GCEP on October 13, 2022 (SOP Version 9).