DiagnosisClinical DiagnosisRefsum disease, also referred to as "classic Refsum disease" (CRD) or "adult Refsum disease" (ARD), is suspected in individuals with late childhood-onset retinitis pigmentosa and variable combinations (in decreasing order of frequency) of the following: Anosmia Sensory motor neuropathy Hearing loss Ataxia Ichthyosis Short metacarpals and metatarsals present from birth (~35% of individuals) Cardiac arrhythmias and cardiomyopathyIt should be noted that: (1) the full constellation of signs and symptoms is rarely seen in an affected individual; (2) most features develop with age. TestingTable 1. Comparison of Peroxisomal Metabolites in the Two Forms of Refsum Disease and AMACR DeficiencyView in own windowPhytanoyl-CoA Hydroxylase DeficiencyPTS2 Receptor DeficiencyAMACR DeficiencyNormalGene SymbolPHYHPEX7AMACR --Plasma Phytanic Acid Concentration 1>200 µmol/L ² >200 µmol/L ² >20 µmol/L<10 µmol/LPlasma Pristanic Acid Concentration<2 µmol/L<2 µmol/L>20 µmol/L<3.0 µmol/LPhytanic Acid / Pristanic Acid RatioElevatedElevatedDecreasedNormalPlasma Pipecolic Acid ConcentrationMildly elevated in 20%NormalNormalNormalErythrocyte Plasmalogen Concentration 1NormalDecreased - NormalNormalNormalDi- and Trihydroxycholestanoic AcidNormalNormalElevated ^{3NormalAMACR = α-methylacyl-CoA racemase. See Differential Diagnosis.Plasma very-long-chain fatty acids (VLCFA) are normal in all three conditions. 1. Measured by gas chromatography2. Plasma phytanic acid concentration may vary considerably because phytanic acid intake is dependent on local diet and may be deceptively low in populations with lower intakes of saturated fatty acids and cholesterol.3. Mildly elevated in 20% of individuals [Wierzbicki et al 2002]Deficiency of phytanoyl-CoA hydroxylase enzyme activity. Greater than 90% of individuals with Refsum disease have deficiency of phytanoyl-CoA hydroxylase, the enzyme encoded by PHYH that catalyzes the conversion of phytanoyl-CoA into 2-hydroxyphytanoyl-CoA, a key step in the breakdown of phytanic acid via alpha-oxidation in peroxisomes. To assess activity of this enzyme, phytanic acid alpha-oxidation is first measured in cultured fibroblasts. If alpha-oxidation is deficient, the activity of the enzyme phytanoyl-CoA hydroxylase is measured. Deficiency of the peroxisome-targeting signal type 2 (PTS2) receptor. Fewer than 10% of individuals with Refsum disease have a deficiency of the PTS2 receptor encoded by PEX7. The PTS2 receptor plays a key role in peroxisome biogenesis by catalyzing the transport across the peroxisomal membrane of proteins equipped with a peroxisome-targeting signal type 2 (like phytanoyl-CoA hydroxylase). Abnormalities in fibroblasts of such individuals include the presence of an abnormal molecular form of peroxisomal thiolase (44 kd) and a partial deficiency of alkyldihydroxyacetonephosphate synthase [van den Brink et al 2003a]. CSF protein concentration. The normal CSF protein concentration in adults ranges from 15 to 50 mg/dL. Values in individuals with Refsum disease are considerably higher. In one Arab family, CSF protein concentration was 101 mg/dL [Fertl et al 2001]. Molecular Genetic TestingGenes. Mutations in two different genes have been identified in Refsum disease. PHYH, encoding phytanoyl-CoA hydroxylase, is mutated in more than 90% of individuals with Refsum disease [Waterham & Wanders, unpublished observations]. PEX7, encoding the PTS2 receptor, is mutated in fewer than 10% of individuals with Refsum disease. Clinical testingFull gene sequencing. Sequencing of coding exons and flanking intron sequences from genomic DNA for PHYH and PEX7 detects mutations in more than 95% of individuals [Waterham & Wanders, unpublished observations]. Table 2. Summary of Molecular Genetic Testing Used in Refsum DiseaseView in own windowGene SymbolProportion of Refsum Disease Attributed to Mutations in This GeneTest MethodMutations DetectedMutation Detection Frequency by Gene and Test Method 1Test AvailabilityPHYH>90%Sequence analysisSequence variants 2>95%Clinical PEX7<10%Sequence variants 2Clinical 1. The ability of the test method used to detect a mutation that is present in the indicated gene2. Examples of mutations detected by sequence analysis may include small intragenic deletions/insertions and missense, nonsense, and splice-site mutations.Interpretation of test results. For issues to consider in interpretation of sequence analysis results, click here.Testing Strategy To confirm/establish the diagnosis in a probandAnalysis of phytanic acid concentration in plasma or serum is the first step. Normal plasma phytanic acid levels essentially rule out the two forms of Refsum disease and AMACR deficiency (see Differential Diagnosis). Studies of enzyme activity in fibroblasts distinguish between Refsum disease caused by PHYH or PEX7 mutations and AMACR deficiency. Molecular genetic testing of PHYH, PEX7, and/or AMACR is required to establish the diagnosis.Carrier testing for at-risk relatives requires prior identification of the disease-causing mutations in the family. Plasma phytanic acid concentration cannot be used to identify carriers as it is completely normal in carriers.Note: Carriers are heterozygotes for this autosomal recessive disorder and are not at risk of developing the disorder.Prenatal diagnosis and preimplantation genetic diagnosis (PGD) for at-risk pregnancies require prior identification of the disease-causing mutations in the family.Genetically Related (Allelic) DisordersPHYH. No other phenotypes are associated with mutations in PHYH. PEX7. Mutations in PEX7 account for rhizomelic chondrodysplasia punctata type 1 (RCDP type I), a severe and often lethal disorder characterized by intellectual disability, rhizomelic shortening of the upper extremities, dwarfism, and cataract [Braverman et al 2002, Motley et al 2002].}

Natural HistoryOnset of symptoms in "classic Refsum disease" (CRD) or "adult Refsum disease" (ARD) ranges from age seven months to after age 50 years. However, because the onset is insidious, it is difficult for many individuals to know exactly when symptoms first started. A few individuals remain asymptomatic until adulthood [Skjeldal et al 1987]. Early-onset disease is not necessarily associated with a poor prognosis for life span. Retinitis pigmentosa is, in most cases, an early clinical feature. Other findings that may occur in the following ten to 15 years in decreasing order of frequency [Skjeldal et al 1987]:NeuropathyDeafnessAtaxiaIchthyosis Wierzbicki et al [2002] documented in 15 individuals the cumulative incidence of the following features over many decades: Retinitis pigmentosa (15/15) Anosmia (14/15) Neuropathy (11/15) Deafness (10/15) Ataxia (8/15)Ichthyosis (4/15) In a few instances, psychiatric disturbances have also been observed. The four cardinal features, originally described by Refsum [1946] (retinitis pigmentosa, a chronic polyneuropathy, ataxia, and raised CSF protein concentration) are rarely seen in a single individual. Some investigators distinguish between acute ARD and chronic ARD. In acute ARD, polyneuropathy, weakness, ataxia, sudden visual deterioration, and often auditory deterioration are often accompanied by ichthyosis, possibly cardiac arrhythmias, and elevated liver transaminases and bilirubin. Triggers for acute presentations include weight loss, stress, trauma, and infections. In contrast, in chronic ARD, RP is present, but the other features of ARD are subtle. Ophthalmologic findings. Retinitis pigmentosa (pigmentary retinal degeneration, tapeto-retinal degeneration) is present in all individuals with biochemical findings of Refsum disease and therefore appears to be an obligatory finding; in a series of 17 individuals, retinitis pigmentosa was present in all [Skjeldal et al 1987]. Virtually every individual ultimately diagnosed to have Refsum disease experiences visual symptoms first. If a detailed past medical history is obtained, many individuals confirm the onset of night blindness in childhood. The delay between first ophthalmologic evaluation and diagnosis ranged between one and 28 years (mean = 11 years) in one study of 23 individuals [Claridge et al 1992]. Typically, individuals with Refsum disease experience night blindness years before the progressive changes of constricted visual fields and decreased central visual acuity appear. Because night blindness can be difficult to ascertain, particularly in children, electroretinography, which shows either a reduction or a complete absence of rod and cone responses, can help support the diagnosis in early stages (see Retinitis Pigmentosa Overview). Anosmia. This is the absence of smell. Although the sense of smell and the sense of taste have their own specific receptors, they are intimately related. Both may be normal, reduced, or absent in individuals with Refsum disease. Studies by Wierzbicki et al [2002] have shown that anosmia is present in most, if not all, individuals with ARD/CRD. If smell is tested experimentally, all individuals with ARD/CRD have an abnormal smell test [Gibberd et al 2004]. Polyneuropathy. The polyneuropathy is a mixed motor and sensory neuropathy that is asymmetric, chronic, and progressive in untreated individuals. It may not be clinically apparent at the start of the illness. Initially, symptoms often wax and wane. Later, the distal lower limbs are affected with resulting muscular atrophy and weakness. Over the course of years, muscular weakness can become widespread and disabling, involving not only the limbs, but also the trunk. Almost without exception, individuals with Refsum disease have peripheral sensory disturbances, most often impairment of deep sensation, particularly perception of vibration and position-motion in the distal legs. Hearing loss. Bilaterally symmetric mild-to-profound sensorineural hearing loss affects the high frequencies or middle-to-high frequencies [Oysu et al 2001, Bamiou et al 2003]. Auditory nerve involvement (auditory neuropathy) may be evident on testing of auditory brain stem evoked responses (ABER) [Oysu et al 2001, Bamiou et al 2003]. Individuals with auditory nerve involvement may experience hearing difficulty even in the presence of a normal audiogram (see Hereditary Hearing Loss and Deafness). Ataxia. Although cerebellar dysfunction is considered to be a main clinical sign of Refsum disease, onset is nevertheless relatively late, particularly when compared with the onset of retinopathy and neuropathy. Unsteadiness of gait is the main symptom related to cerebellar dysfunction. Ataxia is thus characteristically more marked than the degree of muscular weakness and sensory loss would indicate (see Hereditary Ataxia Overview). Ichthyosis. Mild generalized scaling may occur in childhood, but usually begins in adolescence. This finding is present in a minority of affected individuals. Cardiac arrhythmias. Cardiac arrhythmia and heart failure resulting from cardiomyopathy are frequent causes of death in Refsum disease. Elevated plasma concentration of pipecolic acid. A few individuals with elevated plasma concentrations of both phytanic acid and pipecolic acid have been described [Tranchant et al 1993, Baumgartner et al 2000]. Subsequently, sequencing of the gene encoding human L-pipecolic acid oxidase [Ijlst et al 2000] in these individuals did not reveal mutations [Waterham & Wanders, unpublished results], supporting the view that the accumulation of pipecolic acid in these patients is a secondary event. Baumgartner et al [2000] described an individual with psychomotor retardation and abnormally short metatarsals and metacarpals, but no other signs of classic Refsum disease. Phytanoyl-CoA hydroxylase enzyme activity was deficient; a homozygous deletion of PHYH causing a frameshift was identified. L-pipecolic acid concentration was elevated in plasma. Microscopy of the liver showed the presence of peroxisomes, although they were reduced in number and increased in size. These abnormal liver peroxisomes lacked catalase. Moreover, in fibroblasts, a mosaic pattern of cells with and without peroxisomes was found, in contrast to the peroxisomes in fibroblasts from individuals with classic Refsum disease who cannot be distinguished from controls by catalase immunofluorescence. Most likely, this individual is affected by two distinct genetic disorders with mutations in different genes, of which one is PHYH and the other remains to be identified. The latter gene may well be one of the PEX genes.Tranchant et al [1993] described three members of a family diagnosed with Refsum disease. Two had a significant increase of pipecolic acid concentration in plasma and a fourth individual, a brother, died at age 17 years from a progressive neurologic disorder with unusual clinical and neuropathologic abnormalities. Homozygosity mapping localized the gene to chromosome 10p. Subsequently these sibs were confirmed to have phytanoyl-CoA hydroxylase deficiency in fibroblasts and disease-causing mutations in PHYH [Waterham & Wanders, unpublished observations]. Most likely, the mild accumulation of pipecolic acid in these individuals is the secondary result of the accumulation of phytanic acid. This hypothesis is supported by data of Wierzbicki et al [2002] showing elevated plasma pipecolic acid levels in 20% of individuals with Refsum disease.

Genotype-Phenotype CorrelationsMore studies are needed to determine if Refsum disease caused by mutations in PHYH and Refsum disease caused by mutations in PEX7 are phenotypically different. The Refsum disease phenotype caused by mutations in PEX7 may be milder [van den Brink et al 2003b], as exemplified by the patient described by Horn et al [2007].Manifestations of Refsum disease may vary considerably among affected individuals in a family, i.e., among individuals with identical PHYH mutations. These phenotypic differences are comparable to those among affected individuals from different families. Consequently, no clear phenotype-genotype correlations have been identified as yet. Phenotypic variation may be related to the dietary intake of phytanic acid, which is thought to be the toxic compound.

Differential DiagnosisPhytanic acid is also elevated in other peroxisomal disorders including: Alpha-methylacyl-CoA racemase (AMACR) deficiency (OMIM 604489). The phenotype is adult-onset sensory motor neuropathy with or without associated pigmentary retinopathy [Ferdinandusse et al 2000]. This enzyme plays a key role in the breakdown of pristanic acid and the C27-bile acid intermediates di- and trihydroxycholestanoic acid. As a consequence of the impaired degradation of pristanic acid, both pristanic acid and phytanic acid accumulate with pristanic concentrations much more elevated than phytanic acid concentrations (see Table 1). AMACR deficiency and classic Refsum disease can be distinguished by screening peroxisome metabolites in the plasma, followed by fibroblast studies and molecular genetic testing.Peroxisomal biogenesis disorders, Zellweger syndrome spectrum. These disorders are characterized by a defect in peroxisome biogenesis and caused by mutations in one of the many PEX genes. Zellweger syndrome spectrum disorders can readily be distinguished from classic Refsum disease on clinical grounds. Furthermore, in addition to plasma phytanic acid concentration, additional peroxisomal abnormalities are found in plasma, including elevated very-long-chain fatty acids, pristanic acid concentration, and the bile acid intermediates di- and trihydroxycholestanoic acid. Rhizomelic chondrodysplasia punctata (RCDP) type 1, caused by mutations in PEX7. Classic Refsum disease can be distinguished easily from RCDP type 1, although a few individuals with a mild form of RCDP type 1 with a Refsum-like phenotype have been described [van den Brink et al 2003a] (see Molecular Genetics for more details). In these individuals plasma phytanic acid concentration is also elevated. Retinitis pigmentosa. Since visual deterioration is almost always the first symptom of Refsum disease, plasma phytanic acid concentration should be measured in any individual with retinitis pigmentosa, especially when combined with other features suggestive of Refsum syndrome, including anosmia and impaired hearing (see Retinitis Pigmentosa Overview). Retinitis pigmentosa and sensorineural hearing loss Usher syndrome type I is characterized by a congenital, bilateral, profound sensorineural hearing loss, vestibular areflexia, and adolescent-onset retinitis pigmentosa. Inheritance is autosomal recessive. Usher syndrome type II is characterized by: congenital, bilateral, sensorineural hearing loss predominantly in the higher frequencies that ranges from mild to severe; normal vestibular function; and adolescent-to-adult onset of retinitis pigmentosa. Inheritance is autosomal recessive. Usher syndrome type III is characterized by progressive sensorineural hearing loss and adolescent-onset retinitis pigmentosa. Inheritance is autosomal recessive. Alström syndrome is characterized by cone-rod dystrophy, obesity, progressive sensorineural hearing impairment, dilated cardiomyopathy, insulin resistance, and developmental delay. Inheritance is autosomal recessive. Kearns-Sayre syndrome (see Mitochondrial DNA Deletion Syndromes). Kearns-Sayre syndrome (KSS) is defined by the triad of onset before age 20 years, pigmentary retinopathy, and progressive external ophthalmoplegia (PEO). In addition, affected individuals have at least one of the following: cardiac conduction block, cerebrospinal fluid protein concentration greater than 100 mg/dL, or cerebellar ataxia. Sensorineural hearing loss is seen in almost all affected individuals. Kearns-Sayre syndrome is caused by deletion of mitochondrial DNA (mtDNA). Ataxia. Friedreich ataxia is characterized by slowly progressive ataxia with onset usually before age 25 years typically associated with depressed tendon reflexes, dysarthria, Babinski responses, and loss of position and vibration senses. Hearing loss is uncommon. Inheritance is autosomal recessive. (See also Hereditary Ataxia Overview.) Ichthyosis. Sjögren-Larsson syndrome is characterized by congenital ichthyosis and onset of ataxia in early childhood. Increased CSF protein concentration. High CSF protein concentrations can be found in a variety of conditions.

ManagementEvaluations Following Initial DiagnosisTo establish the extent of disease in an individual diagnosed with Refsum disease, the following evaluations are recommended:Ophthalmology. Examination for retinitis pigmentosa / miosis, cataract and visual fields [Claridge et al 1992] Anosmia testing using the procedure described by Gibberd et al [2004] Neurology. Ataxia, neuropathy, myography, and electrophysiologic assessment Audiology. Pure tone audiometry and possibly otoacoustic emission testing and auditory brain stem evoked response (ABER) testing if hearing difficulties are suspected but not identified on pure tone audiometry [Bamiou et al 2003] Radiology. Physical examination of hands, feet, and knees; radiologic assessment of hands and feet Cardiology. Cardiac evaluation including electrocardiography (ECG) and cardiac ultrasound examination Treatment of ManifestationsChronic Treatment Dietary restriction of phytanic acid intake Avoidance of sudden weight loss Lifelong treatment with hydrating creams Regular care by a cardiologist for cardiac arrhythmias and cardiomyopathy in order to treat signs and symptoms properly with anti-arrhythmic and cardiogenic supportive drugs Because the pupils do not dilate well if at all, other measures, such as use of iris hooks, may be necessary to allow sufficient pupillary enlargement during cataract surgery. In addition, an anterior chamber lens with iris fixation may be necessary because the brittleness of the zonular fibers holding the lens capsule may not allow positioning of an intra-ocular lens in the capsular bag after cataract removal, a complication observed in one patient [BP Leroy 2007, personal communication].Treatment of Acute Presentation Many acute features such as polyneuropathy, ataxia, ichthyosis, and cardiac arrhythmias resolve with reduction in plasma phytanic acid concentration (see Prevention of Primary Manifestations).Plasmapheresis or lipapheresis can be used in the event of acute arrhythmias or extreme weakness because phytanic acid is transported on lipoproteins [Wierzbicki et al 1999]. During plasmapheresis, cardiac monitoring should be continuous and plasma glucose concentration should be kept high to prevent onset or exacerbation of arrhythmias. A low phytanic acid diet can be given orally or by nasogastric tube. If oral intake is restricted, appropriate parenteral nutrition and fluid therapies are needed to maintain plasma glucose concentrations and prevent ketosis. Prevention of Primary ManifestationsNo curative therapy currently exists for Refsum disease. By restricting dietary intake of phytanic acid or eliminating phytanic acid by plasmapheresis or lipid apheresis, plasma phytanic acid concentrations can be reduced by 50% to 70%, typically to about 100 to 300 µmol/L. This reduction in plasma phytanic acid concentration successfully resolves symptoms of ichthyosis, sensory neuropathy, and ataxia in approximately that order. However, it is uncertain whether treatment affects the progression of the retinitis pigmentosa, anosmia, and deafness [Gibberd & Wierzbicki 2000]. A high-calorie diet is necessary to avoid mobilization of stored lipids, including phytanic acid, into the plasma. Postoperative care requires parenteral nutrition with solutions that do not contain phytanic acid, e.g., Intralipid^® available in 10%, 20%, and 30% concentrations, which are all based on soybean oil and egg yolk phospholipid.Prevention of Secondary Complications Treatment of hypertension may help in delaying cardiomyopathy, which inevitably leads to arrhythmias. It is probably better to avoid amiodarone as an anti-arrhythmic drug, because of the risk of hyperthyroidism, which results in catabolism and increased phytanic acid release from tissues if not recognized in time. This complication was observed in one individual [BP Leroy 2007, personal communication].Once cardiomyopathy has become difficult to treat, cardiac transplantation can be life-saving. One individual with PHYH-related Refsum disease and one with PEX7-related Refsum disease have successfully received a donor heart [BP Leroy 2007, personal communication].Surveillance The following are appropriate:Measurement of plasma phytanic acid levels every three to six months and during any illness that may lead to a catabolic stateAnnual ophthalmologic examination to identify vision loss resulting from cataracts, which are treatable Annual cardiac examination to identify cardiomyopathy and concomitant arrhythmiasAgents/Circumstances to AvoidAvoid the following:All food products containing phytanic acid such as ruminant (cow, sheep, and goat) products and certain fish (cod) products. some nuts should also be avoided: see Brown et al [1993]. Fasting because stored lipids, including phytanic acid, are mobilized into the plasma Ibuprofen because it is metabolized by AMACR and may interfere with the metabolism of phytanic acid Evaluation of Relatives at RiskIt is appropriate to evaluate the sibs of a proband by measuring plasma phytanic acid concentration before symptoms of Refsum disease occur in order to institute early treatment to reduce plasma phytanic acid concentration. See Genetic Counseling for issues related to testing of at-risk relatives for genetic counseling purposes. Therapies Under InvestigationAt present, the potential of enzyme replacement therapy (ERT) similar to that for lysosomal storage diseases (e.g., Hurler syndrome [see MPS I], Fabry disease, and Gaucher disease) is under investigation. This may eventually replace dietary restrictions and plasma- or lipapheresis. In the long run, gene therapy may be the treatment of choice, but many issues need to be resolved before it can be applied. Search ClinicalTrials.gov for access to information on clinical studies for a wide range of diseases and conditions.

Molecular GeneticsInformation in the Molecular Genetics and OMIM tables may differ from that elsewhere in the GeneReview: tables may contain more recent information. —ED.Table A. Refsum Disease: Genes and DatabasesView in own windowGene SymbolChromosomal LocusProtein NameLocus SpecificHGMDPHYH10p13Phytanoyl-CoA dioxygenase, peroxisomalPHYH homepage - Mendelian genesPHYHPEX76q23​.3Peroxisomal targeting signal 2 receptordbPEX, PEX7 Gene Database
PEX7 homepage - Mendelian genesPEX7Data are compiled from the following standard references: gene symbol from HGNC; chromosomal locus, locus name, critical region, complementation group from OMIM; protein name from UniProt. For a description of databases (Locus Specific, HGMD) to which links are provided, click here.Table B. OMIM Entries for Refsum Disease (View All in OMIM) View in own window 266500REFSUM DISEASE, CLASSIC 601757PEROXISOME BIOGENESIS FACTOR 7; PEX7 602026PHYTANOYL-CoA HYDROXYLASE; PHYHMolecular Genetic PathogenesisMutations in PHYH and PEX7 are known to cause Refsum disease by interfering with the alpha-oxidation of phytanic acid.Alpha-oxidation of phytanic acid. Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) is derived from dietary sources only, mainly from dairy and ruminant fats. Phytanic acid is a 3-methyl branched chain fatty acid, which cannot undergo straightforward beta-oxidation like other fatty acids since the presence of the methyl group at the 3 position blocks beta-oxidation. Nature has resolved this problem by creating an alpha-oxidation mechanism in which the terminal carboxyl group is released as CO₂. Accordingly, phytanic acid first undergoes alpha-oxidative chain shortening to produce pristanic acid (2,4,6,10-tetramethylpentadecanoic acid) and CO₂. All steps from phytanoyl-CoA to pristanic acid occur in peroxisomes [Wanders et al 2001]: Activation of phytanic acid to phytanoyl-CoA Hydroxylation of phytanoyl-CoA to 2-hydroxyphytanoyl-CoA by the enzyme phytanoyl-CoA hydroxylase, encoded by PHYH. Phytanoyl-CoA hydroxylase is equipped with a PTS2 signal, and is targeted to the peroxisome by means of the PTS2 receptor, encoded by PEX7. ¹ Conversion of 2-hydroxyphytanoyl-CoA into pristanal and formyl-CoA via the enzyme 2-hydroxyphytanoyl-CoA lyase ² encoded by 2-HPCL. Conversion of pristanal into pristanic acid via an ill-defined aldehyde dehydrogenase [Jansen et al 2001]. Conversion of pristanic acid into pristanoyl-CoA via the peroxisomal very-long-chain acyl-CoA synthetase ², which has its catalytic site facing the lumen of the peroxisome [Smith et al 2000]. Note: (1) In case of a deficiency at the level of the PTS2 receptor, phytanoyl-CoA hydroxylase cannot be properly imported into the peroxisome, leading to its functional deficiency. (2) The other enzymes from the phytanic acid alpha-oxidation route, including 2-hydroxyphytanoyl-CoA lyase and very-long-chain acyl-CoA synthetase, are PTS1 proteins, which are targeted to the peroxisome by means of the PTS1 receptor, encoded by PEX5. Beta-oxidation. Once pristanic acid has been activated to pristanoyl-CoA, it undergoes three cycles of beta-oxidation in the peroxisomes to generate 4,8-dimethylnonanoyl-CoA, which then is transported to mitochondria for final oxidation to CO₂ and H₂O in the form of its carnitine ester. Note that unimpaired breakdown of phytanic acid is only possible if pristanic acid, the product generated from phytanic acid by alpha-oxidation, also undergoes unimpaired degradation. If pristanic acid oxidation is blocked, as in α-methylacyl-CoA racemase (AMACR) deficiency caused by mutations in AMACR, phytanic acid degradation is partially blocked, leading to elevated plasma phytanic acid concentration.PHYH Normal allelic variants. Jansen et al [2004] have described one sequence variant (c.636A>G) with an incidence of around 10% in 93 control individuals that causes no amino acid change. Pathologic allelic variants. Sequence analysis of PHYH has now been performed in 31 unrelated families and has revealed 29 different variants of which 18 (62.1%) are unique to one patient or family. Of these 29 variants, Sixteen (55.2%) are missense mutations. Four (13.8%) are deletions. All deletions are found in the first half of the coding sequence and all cause a frameshift. Two are insertions (6.8%). One causes a frameshift; the other, a three nucleotide insertion, causes the insertion of a single amino acid into the PhyH protein.Six (20.7%) are splice-site mutations. Two splice-site variants, c.135-2A>G and c.135-1G>C, lead to skipping of exon 3, which consists of 111 nucleotides. Either change causes an in-frame deletion of 37 internal amino acids, and an altered protein (p.Tyr46_Arg82del), which, when heterozygously expressed in S. cerevisiae, is clearly detectable by Western blot analysis, but completely lacks enzymatic activity [Jansen et al 2000]. Four splice-site variants – c.497-2A>G, c.678+2T>G, c.678+5G>T, and c.679-1G>T – cause skipping of exon 6, which results in a frameshift and a premature stop codon (p.Lys167Glyfs*3). One (3.4%) is a non-disease-causing variant that is also present in about 10% of a control population. Table 3. Selected PHYH Allelic Variants View in own windowClass of Variant AlleleDNA Nucleotide ChangeProtein Amino Acid Change
(Alias ¹)Reference SequencesNormalc.636A>Gp.=NM_006214​.3
NP_006205​.1Pathologicc.135-2A>Gp.Tyr46_Arg82delc.135-1G>Cp.Tyr46_Arg82delc.497-2A>Gp.Lys167Glyfs*3
(A166fs*3)c.678+2T>Gp.Lys167Glyfs*3
(A166fs*3)c.678+5G>Tp.Lys167Glyfs*3
(A166fs*3)c.679-1G>Tp.Lys167Glyfs*3
(A166fs*3)c.524A>Gp.His175Argc.526C>Ap.Gln176Lysc.530A>Gp.Asp177Glyc.610G>Ap.Gly204Serc.805A>Cp.Asn269HisSee Quick Reference for an explanation of nomenclature. GeneReviews follows the standard naming conventions of the Human Genome Variation Society (www​.hgvs.org).1. Variant designation that does not conform to current naming conventionsNormal gene product. See Molecular Genetic Pathogenesis. Abnormal gene product. The impact of any mutation in the phytanoyl-CoA hydroxylase gene can be assessed by evaluating the consequences of certain mutations on the stability and the catalytic activity of the hydroxylase upon expression. Interestingly, 15 of the 17 missense mutations identified in the phytanoyl-CoA hydroxylase are located in exon 6 and 7. Structure-function analysis has demonstrated that these exons code in part for the conserved beta-barrel core, which suggests that this structurally important element in the protein is susceptible to changes that directly cause loss of enzymatic activity. The effect of only very few of the missense mutations has been tested by expressing the mutant proteins and testing protein stability and enzyme activity. Mukherji et al [2001] have studied a few mutants including the p.His175Arg, p.Gln176Lys, and p.Asp177Gly mutants. The amino acid triad His175, Gln176, Asp177 forms the iron-binding motif and mutations in any of these amino acids have been shown to cause a fully dysfunctional enzyme since the hydroxylase is completely dependent upon Fe²⁺ for the conversion of phytanoyl-CoA to 2-hydroxyphytanoyl-CoA. Two missense mutations, p.Gly204Ser and p.Asn269His, cause the peculiar effect of uncoupling the hydroxylation of phytanoyl-CoA from the conversion of 2-oxoglutarate into succinate and CO₂. As demonstrated in expression studies, Mukherji et al [2001] found that no phytanoyl-CoA was hydroxylated while decarboxylation of 2-oxoglutarate to succinate and CO₂ still took place, although at a much-reduced rate. This uncoupling is also observed in the case of the p.Gln176Lys substitution, which is also associated with a change in the iron-binding motif. PEX7 Normal allelic variants. PEX7 contains ten exons that span 91 kb of genomic DNA. Pathologic allelic variants. So far, three individuals have been identified with mild mutations in PEX7. The two individuals described by van den Brink et al [2003a] who were both compound heterozygous for mutations in PEX7 had a p.Tyr40X nonsense mutation that introduces a premature stop codon in the N-terminal region of the protein. This mutation has also been found in classic RCDP type 1 with a severe clinical presentation [Motley et al 2002]. In patient 1, the second allele was p.Gly7Valfs*51, resulting from a 7-nucleotide duplication, predicted to cause a frame-shift leading to a premature stop codon at amino acid position 51. However, the duplication occurs between two in-frame initiation codons, which would suggest that use of the second ATG may produce a protein that lacks the first ten amino acids but retains partial peroxin 7 transport functions. In patient 2, the second allele was p.Thr14Pro, which is thought either to interfere with folding of the protein or to reduce the affinity for its binding partners [Braverman et al 2002]. Table 4. Selected PEX7 Pathologic Allelic VariantsView in own windowDNA Nucleotide Change
(Alias ¹)Protein Amino Acid ChangeReference Sequencesc.12_18dup
(7-bp duplication)p.GLy7Valfs*51NM_000288​.3
NP_000279​.1c.40A>Cp.Thr14Proc.120C>Gp.Tyr40XSee Quick Reference for an explanation of nomenclature. GeneReviews follows the standard naming conventions of the Human Genome Variation Society (www​.hgvs.org).1. Variant designation that does not conform to current naming conventionsNormal gene product. The normal product is a 323-amino acid protein with six WD40 repeats. Each WD-repeat is composed of four beta-strands, which together make up one blade of a propeller. The overall structural appearance of a WD-repeat protein such as PEX7 with six such repeats, resembles that of a propeller with as many blades as the number of WD-repeats. Peroxisomal targeting signal 2 receptor, the protein encoded by PEX7, is a receptor for a subclass of peroxisomal matrix enzymes and targets these enzymes to the peroxisomal membrane. Abnormal gene product. Defects in peroxisomal targeting signal 2 receptor result in deficient activity of all PTS2 enzymes, but other peroxisomal functions remain intact. Fibroblast assays show that PTS2 proteins remain cytosolic in individuals with RCDP1 and are likely degraded, but PTS1 proteins are imported into peroxisomes normally. Peroxisome morphology is normal in fibroblasts, but abnormal in liver, according to several case reports [Braverman et al 2002, van den Brink et al 2003b].