Clinical manifestations

Presentation and course

Cystathionine beta-synthase deficiency represents a multisystem disorder with involvement of the eyes, integument, skeleton, vascular system, and nervous system. The most comprehensive overview of clinical manifestations in cystathionine beta-synthase deficiency was given by Mudd and colleagues in 1985 when they investigated the natural course of the disease in 629 patients by an international questionnaire survey (65).

Eyes. Ectopia lentis is a striking and readily recognizable manifestation of the disease. It may be the only manifestation, and by 38 years of age only 3% of patients have both lenses in place. Dislocation is usually present by 10 years of age. The dislocation is usually downward--the opposite of the situation in Marfan syndrome. Other ocular abnormalities include myopia, glaucoma, cataracts, retinal detachment, and, rarely, optic atrophy.

Skin and hair. The skin and hair may show a decrease in pigment. A pronounced malar flush was first recognized in Ireland (12). The skin may otherwise have blotchy erythema and pallor. Livido reticularis is particularly common in the distal extremities, which may be cold and show other evidence of vascular instability (26).

Skeletal system. Generalized osteoporosis is the most common skeletal finding, and 50% of patients have osteoporotic changes in the radiograph of the spine at the age of 16 years (65). Low bone mineral density was found in 38% of patients when investigated by DEXA scan (84). There are a large number of other skeletal abnormalities, including genu valgum, everted feet, pectus excavatum or carinatum, scoliosis, biconcave (“codfish”) vertebrae, large metaphyses, and epiphyses. Notable among these is dolichostenomelia (thinning and lengthening of the long bones), which produces tall and thin individuals who are often considered to have a marfanoid appearance, but true arachnodactyly is rare (74; 10).

Vascular system. Thromboembolism is the major cause of morbidity and the most frequent cause of death in cystathionine beta-synthase deficiency. Vascular occlusion can occur in any vessel and at any age, including infancy (39). By the end of the third decade, 50% of untreated patients have had one or more thromboembolic events (65). The international survey of Mudd and colleagues revealed that 158 (of 629) patients were reported to have had a total of 253 thromboembolic events. Among these events, 51% involved peripheral veins (of which about a quarter resulted in pulmonary embolism), 32% were cerebrovascular accidents, 11% affected peripheral arteries, 4% produced myocardial infarctions, and 2% fell into none of these categories (65).

Central nervous system. Intellectual disability of varying severity is the most common manifestation of central nervous system involvement in the disease. Presenting as developmental delay during the first years of life, intellectual disability is often the first recognized sign of cystathionine beta-synthase deficiency. The cognitive capability of untreated patients varies widely. IQ scores range from 10 to 138, with a median of approximately 64 (65). IQ scores among affected siblings were similar (01). Seizures occur in about 20% of untreated patients (65). The most common seizure type is generalized tonic-clonic. Typical strokes or focal neurologic signs suggest the presence of a cerebrovascular occlusion. Other neurologic features have included abnormal electroencephalograms and extrapyramidal disturbances, such as dystonia (01; 38). Diffuse leukoencephalopathy, observed in 3 cases to date, is an extremely rare finding in classical homocystinuria. In all the cases the leukoencephalopathy was reversible, and it was complicated by severe hypermethioninemia (greater than 1000 µmol/L). Thus, brain edema rather than demyelination was the most likely cause for leukoencephalopathy (88; 16; 82). Psychiatric abnormalities have been reported in about a half of one series of 63 patients. Four diagnostic categories predominated: episodic depression (10%), chronic disorders of behavior (17%), chronic obsessive-compulsive disorder (5%), and personality disorders (19%) (01). Another series investigating neuropsychiatric outcomes noted psychological symptoms in 16 of 25 adult patients (64%), including a high prevalence of anxiety (32%) and depression (32%) that correlated with IQ values below 85 (03).

Risk for development of Alzheimer diseases is likely greater based on the observation that increased plasma homocysteine represents an independent risk factor. With a plasma homocysteine level greater than 14 µmol per liter, the risk of Alzheimer disease nearly doubles (76).

All of the above-mentioned clinical manifestations except for the skeletal manifestation are commonly more pronounced or occur earlier in pyridoxine nonresponsive patients as compared to those who are pyridoxine responsive. The skeletal manifestations do not appear to depend on the pyridoxine responsiveness of patients. In a more granular analysis regarding pyridoxine responsiveness, 328 patients with CBS deficiency were classified into 4 groups of pyridoxine responsivity: nonresponders, partial, full, and extreme responders (41). Lens dislocation was common in all groups except extreme responders, but the age of dislocation increased with increasing responsiveness. Developmental delay was commonest in the nonresponder group, whereas no extreme responder patient had cognitive impairment. Thromboembolism was the commonest presenting feature in extreme responder patients, whereas it was least likely at presentation in the nonresponder group. This probably is due to the differences in ages at presentation. All groups had a similar number of thromboembolic events per 1000 patient-years. Clinical severity of CBS deficiency depends on the degree of pyridoxine responsiveness. Along those lines, Ochoa-Ferraro and colleagues reported cerebral venous thrombosis as the first manifestation of homocystinuria in adults (67).

Biological basis

Etiology and pathogenesis

Homocystinuria is an autosomal recessive inherited disorder due to deficiency of cystathionine beta-synthase (EC 4.2.1.22, McKusick 236200). The gene has been cloned and mapped to chromosome 21 in the subtelomeric area q22.3 (66). The human cystathionine beta-synthase cDNA sequence and structure was published in 1993 (43). More than 320 individual homocystinuric alleles have been studied, and 100 mutations were detected. Most of these mutations are missense mutations (42). Testing for pathogenicity revealed that mutations p.M173del, p.I278S, p.D281N, and p.D321V showed null activity in all conditions tested, whereas mutations p.49L, p.P200L, and p.A446S retained different degrees of activity and response to stimulation (15). In a series of 84 cystathionine beta-synthase alleles previously sequenced from patients with homocystinuria, 37% of the alleles were partially functional or could be rescued by cofactor supplementation (55). Deficient regulation of cystathionine beta-synthase by its activator S-adenosylmethionine seems to be a frequently found mechanism in 20% of individuals with cystathionine beta-synthase deficiency (57).

Cystathionine beta-synthase activity can be found in many tissues, including liver, brain, pancreas, and cultured fibroblasts (79). The enzyme is a tetramer of identical 63-kDa subunits but can also exist as a more active dimer of 48-kDa subunits (44). In addition to the cofactor pyridoxal phosphate, cystathionine beta-synthase also binds to other ligands: the activator S-adenosylmethionine and 1 mole of heme per subunit (40; 23).

Fibroblasts from patients with homocystinuria due to cystathionine beta-synthase deficiency can be classified into 3 groups: those exhibiting no residual activity, those exhibiting reduced activity with normal affinity for pyridoxal phosphate, and those exhibiting residual activity with markedly reduced affinity for pyridoxal phosphate (22). The in vitro properties do not necessarily correspond to in vivo responsiveness to pyridoxine, but in almost all pyridoxine responders there is at least a small amount of residual enzymatic activity detectable in fibroblasts. Nevertheless, there is ample evidence that the pyridoxine-induced response in patients is due to its action as a cofactor of cystathionine beta-synthase. Approximately equal proportions of patients are pyridoxine responsive and nonresponsive (65; 14).

Not all of the pyridoxine-responsive patients reveal complete normalization of homocysteine after pyridoxine treatment. Some patients who are responsive to pyridoxine continue to have slight elevations of homocysteine in plasma and urine. Brenton and Cusworth defined 3 classes of pyridoxine responsiveness, including a group intermediate between those who display little or no response and those with clear response (11). Approximately 13% of patients may show such intermediate responses (65).

Kožich and colleagues have analyzed clinical and laboratory data at the time of diagnosis in 328 patients with CBS deficiency from the E-HOD (European network and registry for Homocystinurias and methylation Defects) registry and developed comprehensive criteria to classify patients into 4 groups of pyridoxine responsivity: nonresponders, partial, full, and extreme responders (41). All groups showed overlapping concentrations of plasma total homocysteine, whereas pyridoxine responsiveness inversely correlated with plasma/serum methionine concentrations. The full and extreme responder groups had a later age of onset and diagnosis and a longer diagnostic delay than nonresponder and partial responder patients.

Deficiency of cystathionine beta-synthase leads to tissue accumulation of methionine, homocysteine, and their S-adenosyl derivatives, with lack of cystathionine and low levels of cysteine. Thus, cysteine becomes an essential amino acid in cystathionine beta-synthase deficiency. The –SH group of homocysteine can easily react with –SH groups of other molecules, leading to the formation of a number of disulfide compounds, such as homocystine, homocysteine-cysteine mixed disulfide, or protein-bound homocysteine (06).

The pathophysiology of cystathionine beta-synthase deficiency has not yet been completely elucidated, but accumulation of homocysteine probably plays a major role in determining some of the most relevant clinical manifestations, including generalized vascular damage and thromboembolic complications. This view is supported by the observation that patients with cystathionine beta-synthase deficiency due to defects of remethylation, but without accumulation of methionine, show similar lesions in blood vessels (08).

Thromboembolism has been suggested to be the end-point of homocysteine-induced abnormalities of platelets, endothelial cells, and coagulation factors. Homocysteine may also cause abnormal cross-linking of collagen (71), leading to abnormalities of the skin, joints, and skeleton in patients (51). Impaired cartilage differentiation has been demonstrated in cystathionine beta-synthase deficient mice (73). These mechanisms seem unlikely to cause damage of the non-collagenous zonular fibers of the lens; instead, disturbed fibrillin structure has been proposed (63; 86). Investigations on the potential of homocysteine to modify structural and functional properties of recombinant human fibrillin-1 fragments strongly suggest that structural and functional modifications as well as degradation processes of fibrillin-1 in the connective tissues of patients with homocystinuria play a major role in the pathogenesis of this disorder (34; 33).

It has not yet been elucidated how cystathionine beta-synthase deficiency impairs cognitive capabilities. Increased homocysteine is considered a risk factor for vascular dementia and stroke. Research results in murine models suggest that homocysteine causes arterial remodeling, at least in part, by increasing the collagen/elastin ratio. This increases vascular resistance that leads to a decrease in carotid artery blood flow (46). Other studies have attempted to clarify the mechanisms involved in the complex neuropathological features associated with abnormal homocysteine metabolism. Investigations on the effect of elevated homocysteine on the blood-brain barrier suggest an important toxic effect of elevated homocysteine on brain microvessels and implicate homocysteine in the disruption of the blood-brain barrier (36). Investigations on the regional and cellular distribution of cystathionine beta-synthase in the adult and developing mouse brain suggest that the enzyme plays a crucial role in the development and maintenance of the CNS and that enzyme deficiency might cause radial glia or astrocyte dysfunction (17).

Epidemiology

The estimated prevalence of cystathionine beta-synthase deficiency ranges from 1 in 1800 newborns in the state of Qatar and over 1 in 50,000 in Ireland and Kuwait to 1 in 1 million in Japan (91; 04). The overall frequency has been reported to be between 1 in 200,000 and 1 in 335,000 (64). Moorthie and colleagues performed a systematic review and metaanalysis and found a worldwide prevalence based on diagnosis of symptomatic individuals of 0.82 in 100,000, whereas the prevalence based on neonatal screening by MS/MS was 1.01 in 100,000 newborns (58). The minimum worldwide prevalence of homocystinuria based on the frequency of the 25 most common pathogenic alleles of homocystinuria in a large genomic database (gnomAD) was estimated to be about 0.38 in 100,000 and was higher in non-Finnish Europeans (approximately 0.72:100,000) and Latin Americans (approximately 0.45:100,000), and lower in Africans (approximately 0.20:100,000) and Asians (approximately 0.02:100,000) (85).

Data for the frequency of cystathionine beta-synthase deficiency are derived from the number of patients detected by newborn screening for cystathionine beta-synthase deficiency, though a measurement of methionine is likely an underestimate of the true rate of occurrence of cystathionine beta-synthase deficiency. Particularly, the pyridoxine-responsive cystathionine beta-synthase deficiency, the most readily treatable form, is being preferentially missed by newborn screening. This is corroborated by some studies on allele frequencies. A DNA-based screening of newborns in Denmark showed 1.4% of them to be heterozygous for the I278T mutation (28). This value corresponds to a homozygote frequency of approximately 1:20,000. In the Czech population, the frequency of the c.1105C>T mutation, which typically causes a milder phenotype, was 0.005. This predicts a birth prevalence of homocystinuria of 1:40,000, which increased to 1:15,500 in a model that included 10 additional mutations (35). Skovby and colleagues concluded that the predominant portion of such homozygotes are clinically unaffected or may be identified due to thromboembolic events that occur no sooner than the third decade of life (78).

Prevention

From observation in patients in whom homocystinuria was detected early and treated, some evidence indicates that presymptomatic initiation of treatment is able to prevent cognitive impairment, lens dislocation, and thromboembolic events (65). This is especially important in pyridoxine-responsive forms. Newborn screening for cystathionine beta-synthase deficiency would represent the tool for the required timely identification of affected patients. Although it is not possible to draw any conclusions based on controlled studies, there are uncontrolled case series that support the efficacy of newborn screening for homocystinuria and its early treatment (83). Newborn screening programs searching for the disease by methionine measurement may fail because of their insufficient diagnostic sensitivity of methionine, which is not dependent on the screening technology (75). To overcome the lack of sensitivity of methionine screening, total homocysteine has been implemented successfully as a first tear test in Qatar (29); however, low prevalence of the disease in most parts of the world may not justify the incremental cost that would come with homocysteine as a first tear test. Instead, lowering the methionine cut-off combined with additional ratios, like the methionine/phenylalanine ratio, or the combination with total homocysteine as a secondary tear test may present cheaper alternatives to improve specificity and sensitivity of newborn screening for homocystinuria (37).

The implementation of a feasible method to measure total homocysteine in the newborn screening sample should overcome the problem (25). Indeed, various laboratories have reported the successful analysis of total homocysteine with liquid chromatography–mass spectrometry methods (24). A review of newborn screening practice for homocystinuria recommends the revision of decision limits considering population median, the combination of relevant screening markers, and tHcy as second-tier marker (37).

The risk for thromboembolic events seems to be directly correlated with the plasma homocysteine concentration. Normalization of plasma homocysteine at any time seems to prevent the occurrence of new thromboembolic events, but even when normalization cannot be reached by treatment, lowering the homocysteine concentration lowers the risk for such an event. Thus, lowering homocysteine as much as possible represents the most important measure in homocystinuric patients. Otherwise, the protection for vascular occlusions must be carried out by anticoagulants.

Differential diagnosis

The differential diagnosis for homocystinuria due to cystathionine beta-synthase deficiency is as follows:

Diagnostic workup

Determination of plasma total homocysteine and of plasma amino acids allows diagnosis of cystathionine beta-synthase deficiency. Total homocysteine in cystathionine beta-synthase deficiency is often higher than 200 µmol/L (reference, less than 15 µmol/L). Methionine is clearly increased in most cases (approximately 500 to 800 µmol/L, reference less than 35 µmol/L). Urinary analysis reveals increased homocystine and homocysteine-cysteine excretion. The cyanide nitroprusside test is positive. Confirmation analysis consists of cystathionine beta-synthase measurement in fibroblasts or plasma (02) and mutational analysis in the cystathionine beta-synthase gene. In vitro pyridoxine responsiveness of residual cystathionine beta-synthase enzyme activity in fibroblasts can be estimated by enzyme assay with and without addition of pyridoxine. However, the in vitro response does not reflect the in vivo response to pyridoxine in all patients. Before initiation of treatment, the in vivo response to pyridoxine has to be examined by administration of vitamin B6 (100 mg twice a day, over 2 weeks). After measuring total homocysteine, folate must be added (5 mg, over next 2 weeks). After 4 weeks, the patients can be considered to be pyridoxine-responsive when total homocysteine is less than 60 µmol/L, as pyridoxine-partially responsive when total homocysteine decreases but is greater than 60 µmol/L, and as pyridoxine-non-responsive when total homocysteine remains unchanged.

Prenatal diagnosis of cystathionine beta-synthase deficiency is feasible in chorionic villi and cultured amniotic cells.

A review from Morris and colleagues provides guidance for the diagnosis and management of homocystinuria (59).

Management

The goal of treatment in cystathionine beta-synthase deficiency is normalization of plasma total homocysteine. The target concentration is less than 20 µmol/L. Whenever normalization is not accessible, the total homocysteine concentration must be kept in the lowest range possible.

In addition, the total cysteine content in plasma is crucial. Plasma cysteine concentrations below 150 µmol/L counteract the efforts to lower plasma homocysteine. An exogenous supply of 40 mg cysteine/kg per day might be sufficient to maintain the glutathione synthesis in erythrocytes (48; 68; 80)

Several strategies used to lower homocysteine levels include:

(1) Enhancing transsulphuration by using pharmacological doses of vitamin B6

(2) Increasing remethylation by folate, vitamin B12, and betaine

(3) Reducing methionine load by a low protein diet combined with a methionine-free amino acid mixture, containing supplemented cysteine.

About half of all cystathionine beta-synthase deficiency patients are responsive to pharmacological doses of vitamin B6, and this treatment alone will substantially reduce plasma homocysteine levels. With treatment, all of these patients will eventually become folate depleted, and probably also B12 depleted, and these vitamins are essential. Occasionally, patients are partially responsive to pyridoxine. Most pyridoxine-responsive patients cannot achieve a normal level of homocysteine on pyridoxine, folate, and B12 treatment alone, although the levels obtained evidently result in a greatly improved outcome. Addition of a low protein diet and methionine-free amino acid supplement, if tolerated, will result in near-normal total homocysteine levels in most patients. Pyridoxine-nonresponsive patients need betaine in addition to folate, vitamins B12 and B6, and a low-protein diet with a methionine-free amino acid supplement. Usually, only patients diagnosed as neonates are fully compliant with diet and the amino acid supplement.

Some patients respond on higher doses of vitamin B6; however, doses greater than 400 mg daily have been associated with peripheral neuropathy in some individuals (09). Pyridoxine toxicity (respiratory failure and rhabdomyolysis) has been observed in one neonate receiving 200 mg pyridoxine (54 mg/kg/day) (05).

Cerebral edema has been observed in a child with cystathionine beta-synthase deficiency 4 to 6 weeks after starting betaine therapy (16). The cerebral edema might have been provoked by a betaine-caused rapid increase of methionine. High plasma levels (greater than 1500 µmol/L) may possibly be associated with cerebral edema, although this is uncertain (60).

Aspirin (100 mg daily) is indicated if there are other thrombophilic factors present, such as factor V Leyden, or if there has been a thromboembolic event. Vitamin C has been shown to improve the impairment of nitric-oxide-dependent vasodilatation that occurs in cystathionine beta-synthase deficiency patients (72). Supplementation with the amino acid taurine improved preexisting reduced endothelial function in CBS-deficient patients (81).

The first case of liver transplantation intended to cure homocystinuria has been published, and the results are promising (50). A 24-year-old man with homocystinuria diagnosed and treated from birth received a deceased whole liver transplant. Because of poor adherence to the conventional diet-restrictive therapy, liver transplantation was considered when he developed hypertension and multiple infarctions over the right cerebellum early in the second decade of his life. Plasma and urine homocysteine and plasma methionine normalized, and he was completely disease free without dietary or nutritional control.

Chemical chaperone therapy might be a promising new approach in the future. Some studies have shown that proline, glycerol, sorbitol, ethanol, dimethyl sulfoxide, or trimethylamine-N-oxide can restore significant cystathionine beta-synthase enzymatic activity. The ratio of Hsp70 and Hsp26 determines whether misfolded human cystathionine beta-synthase will either be refolded or degraded. Heme arginate, for example, rescued catalytic CBS activity of 5 out of 7 human mutations in mammalian CHO-K1 cells (77; 53; 52; 56). Along those lines, proteostasis modulators may be useful in treating certain types of genetic diseases caused by missense mutations as they can restore significant enzymatic activity to misfolded cystathionine beta-synthase enzymes in mutant mice (30).

Attempts toward gene therapy indicate the efficacy and feasibility of gene delivery and long-term functional correction of cystathionine β-synthase deficiency in mice by adeno-associated viral gene therapy (70; 47).

Also, in mice, enzyme replacement therapy with PEGylated cystathionine beta synthase resulted in reduced and sustained plasma homocysteine concentration and normalized plasma cysteine, corrected liver glucose, and lipid metabolism and prevented or reversed facial alopecia, fragile and lean phenotype, and low bone mass (54).

Outcomes

Extreme variability of prognosis is a consequence of whether or not there are thromboembolic events and, if there are, which areas of the body suffer infarction. Variability also depends on whether the patients are pyridoxine-responsive or not and when the treatment was started. The outcome in 158 patients treated for up to 18 years has been reported (89). Those patients responsive to pyridoxine maintained total homocysteine levels of less than 60 µmol/L (reference less than 15 µmol/L), whereas pyridoxine-non-responsive patients had levels usually greater than 80 µmol/L. Treatment regimens varied somewhat. There was a substantial decrease in thromboembolic episodes from the number expected in untreated patients. In a subset of patients whose treatment was standardized and similar to that described below, the same benefit from this treatment regimen was seen (87). In patients with neonatal diagnosis and treatment, there is also evidence of improved outcome with avoidance of intellectual deficit and dislocation of the lens (90).

There is an increased risk for thromboembolic events from any surgery or immobilization and for acute glaucoma due to pupillary block by the dislocated lens.

Special considerations

Pregnancy

Pregnancy in cystathionine beta-synthase deficiency may well confer increased risk for maternal thromboembolism, especially postpartum, but there seems to be only limited teratogenic potential. Thromboembolic events have been reported previously in a few pregnancies, all occurring postpartum. Recommendation for anticoagulation during delivery is still controversial. Maternal cystathionine beta-synthase deficiency might predispose pregnant women to preeclampsia. Although only small numbers of pregnancies in cystathionine beta-synthase deficiency have been reported, most pregnancies have progressed without obstetric complications (49), and healthy pregnancies are feasible. That can be achieved by proactive management through a motivated clinical team and good patient engagement (31).

Anesthesia

Nitrous oxide might have adverse effects on increasing homocysteine levels because it irreversibly deactivates methionine synthase; therefore, it should not be used as an anesthetic agent.