Liver transplant as a curative treatment in a pediatric patient with classic homocystinuria: A case report.

We report a patient with homocystinuria and hyperoxaluria who was cured of homocystinuria-related disease following liver transplant. The patient was diagnosed with homocystinuria as a newborn and was treated with dietary modifications and supplements. At 22 months, he passed a calcium oxalate stone and was found to have numerous bilateral kidney stones. Genetic testing confirmed primary hyperoxaluria, type 1. He underwent preemptive liver transplant at age four to treat primary hyperoxaluria. Following transplant, his serum methionine and homocysteine levels normalized, thus, demonstrating resolution of homocystinuria. Methionine and homocysteine levels remained normal 6 years later. Homocystinuria is associated with ophthalmologic, skeletal, neurologic, and thromboembolic complications. As cystathionine beta-synthase resides in the liver, transplant was hypothesized to be an effective treatment. Primary hyperoxaluria generally progresses to chronic kidney disease and is treated with combined kidney-liver transplant at the time of end stage kidney disease. Given this patient's dual diagnoses, we proceeded with preemptive liver transplantation. Three prior cases of patients with homocystinuria treated with liver transplantation have been reported. In all cases, transplant resolved metabolic effects. However, our case represents a pediatric patient without disease-related complications prior to transplant. This case supports liver-targeted gene therapies as an effective treatment for homocystinuria.

A cautionary tale of pyridoxine toxicity in cystathionine beta-synthase deficiency detected by two-tier newborn screening highlights the need for clear pyridoxine dosing guidelines.

Classic homocystinuria is due to deficiency of cystathionine beta-synthase (CBS), a pyridoxine-dependent enzyme that, depending on the molecular variants, may be co-factor responsive. Elevated methionine is often used as the primary analyte to detect CBS deficiency (CBSD) on newborn screening (NBS), but is limited by increased detection of other biochemical disorders with less clear clinical significance such as methionine aminotransferase (MAT) I/III heterozygotes. Our state has implemented a two-tier NBS algorithm for CBSD that successfully reduced the number of MATI/III heterozygotes, yet effectively detected a mild, co-factor responsive form of CBSD. After initial diagnosis, newborns with CBSD often undergo a pyridoxine challenge with high-dose pyridoxine to determine responsiveness. Here we describe our NBS-identified patient with a mild form of pyridoxine responsive CBSD who developed respiratory failure and rhabdomyolysis consistent with pyridoxine toxicity during a pyridoxine challenge. This case highlights the need for weight-based dosing and duration recommendations for pyridoxine challenge in neonates.

The vitamin B12 processing enzyme, mmachc, is essential for zebrafish survival, growth and retinal morphology.

Cobalamin C (cblC) deficiency, the most common inborn error of intracellular cobalamin metabolism, is caused by mutations in MMACHC, a gene responsible for the processing and intracellular trafficking of vitamin B12. This recessive disorder is characterized by a failure to metabolize cobalamin into adenosyl- and methylcobalamin, which results in the biochemical perturbations of methylmalonic acidemia, hyperhomocysteinemia and hypomethioninemia caused by the impaired activity of the downstream enzymes, methylmalonyl-CoA mutase and methionine synthase. Cobalamin C deficiency can be accompanied by a wide spectrum of clinical manifestations, including progressive blindness, and, in mice, manifests with very early embryonic lethality. Because zebrafish harbor a full complement of cobalamin metabolic enzymes, we used genome editing to study the loss of mmachc function and to develop the first viable animal model of cblC deficiency. mmachc mutants survived the embryonic period but perished in early juvenile life. The mutants displayed the metabolic and clinical features of cblC deficiency including methylmalonic acidemia, severe growth retardation and lethality. Morphologic and metabolic parameters improved when the mutants were raised in water supplemented with small molecules used to treat patients, including hydroxocobalamin, methylcobalamin, methionine and betaine. Furthermore, mmachc mutants bred to express rod and/or cone fluorescent reporters, manifested a retinopathy and thin optic nerves (ON). Expression analysis using whole eye mRNA revealed the dysregulation of genes involved in phototransduction and cholesterol metabolism. Zebrafish with mmachc deficiency recapitulate the several of the phenotypic and biochemical features of the human disorder, including ocular pathology, and show a response to established treatments.

Taurine alleviates repression of betaine-homocysteine S-methyltransferase and significantly improves the efficacy of long-term betaine treatment in a mouse model of cystathionine ß-synthase-deficient homocystinuria.

Classical cystathionine ß-synthase-deficient homocystinuria (HCU) is a life-threatening inborn error of sulfur metabolism. Treatment for pyridoxine-nonresponsive HCU involves lowering homocysteine (Hcy) with a methionine (Met)-restricted diet and betaine supplementation. Betaine treatment efficacy diminishes significantly over time due to impairment of betaine-Hcy S-methyltransferase (BHMT) function. Little is known regarding the regulation of BHMT in HCU. Using a betaine-responsive preclinical mouse model of HCU, we observed that this condition induces a 75% repression of BHMT mRNA, protein and enzyme activity, and significant depletion of hepatic betaine levels. BHMT repression was proportional to plasma Hcy levels but was not observed in mouse models of homocystinuria due to either methylenetetrahydrofolate reductase or Met synthase deficiency. Both Met supplementation and chemically induced glutathione depletion exacerbated hepatic BHMT repression in HCU mice but not wild-type (WT) controls. Conversely, cysteine treatment normalized hepatic BHMT expression in HCU mice but had no effect in WT control animals. Taurine treatment induced BHMT expression in HCU mice by 5-fold and restored maximal lowering of Hcy levels during long-term betaine treatment with a concomitant normalization of inflammatory cytokine expression and a significantly improved coagulative phenotype. Collectively, our findings indicate that adjuvantial taurine treatment has the potential to significantly improve clinical outcomes in HCU.-Maclean, K. N., Jiang, H, Phinney, W. N., Keating, A. K., Hurt, K. J., Stabler, S. P. Taurine alleviates repression of betaine-homocysteine S-methyltransferase and significantly improves the efficacy of long-term betaine treatment in a mouse model of cystathionine ß-synthase-deficient homocystinuria.

High dietary folate in pregnant mice leads to pseudo-MTHFR deficiency and altered methyl metabolism, with embryonic growth delay and short-term memory impairment in offspring.

Methylenetetrahydrofolate reductase (MTHFR) generates methyltetrahydrofolate for methylation reactions. Severe MTHFR deficiency results in homocystinuria and neurologic impairment. Mild MTHFR deficiency (677C > T polymorphism) increases risk for complex traits, including neuropsychiatric disorders. Although low dietary folate impacts brain development, recent concerns have focused on high folate intake following food fortification and increased vitamin use. Our goal was to determine whether high dietary folate during pregnancy affects brain development in murine offspring. Female mice were placed on control diet (CD) or folic acid-supplemented diet (FASD) throughout mating, pregnancy and lactation. Three-week-old male pups were evaluated for motor and cognitive function. Tissues from E17.5 embryos, pups and dams were collected for choline/methyl metabolite measurements, immunoblotting or gene expression of relevant enzymes. Brains were examined for morphology of hippocampus and cortex. Pups of FASD mothers displayed short-term memory impairment, decreased hippocampal size and decreased thickness of the dentate gyrus. MTHFR protein levels were reduced in FASD pup livers, with lower concentrations of phosphocholine and glycerophosphocholine in liver and hippocampus, respectively. FASD pup brains showed evidence of altered acetylcholine availability and Dnmt3a mRNA was reduced in cortex and hippocampus. E17.5 embryos and placentas from FASD dams were smaller. MTHFR protein and mRNA were reduced in embryonic liver, with lower concentrations of choline, betaine and phosphocholine. Embryonic brain displayed altered development of cortical layers. In summary, high folate intake during pregnancy leads to pseudo-MTHFR deficiency, disturbed choline/methyl metabolism, embryonic growth delay and memory impairment in offspring. These findings highlight the unintended negative consequences of supplemental folic acid.

High folic acid consumption leads to pseudo-MTHFR deficiency, altered lipid metabolism, and liver injury in mice.

BACKGROUND: Increased consumption of folic acid is prevalent, leading to concerns about negative consequences. The effects of folic acid on the liver, the primary organ for folate metabolism, are largely unknown. Methylenetetrahydrofolate reductase (MTHFR) provides methyl donors for S-adenosylmethionine (SAM) synthesis and methylation reactions. OBJECTIVE: Our goal was to investigate the impact of high folic acid intake on liver disease and methyl metabolism. DESIGN: Folic acid-supplemented diet (FASD, 10-fold higher than recommended) and control diet were fed to male Mthfr(+/+) and Mthfr(+/-) mice for 6 mo to assess gene-nutrient interactions. Liver pathology, folate and choline metabolites, and gene expression in folate and lipid pathways were examined. RESULTS: Liver and spleen weights were higher and hematologic profiles were altered in FASD-fed mice. Liver histology revealed unusually large, degenerating cells in FASD Mthfr(+/-) mice, consistent with nonalcoholic fatty liver disease. High folic acid inhibited MTHFR activity in vitro, and MTHFR protein was reduced in FASD-fed mice. 5-Methyltetrahydrofolate, SAM, and SAM/S-adenosylhomocysteine ratios were lower in FASD and Mthfr(+/-) livers. Choline metabolites, including phosphatidylcholine, were reduced due to genotype and/or diet in an attempt to restore methylation capacity through choline/betaine-dependent SAM synthesis. Expression changes in genes of one-carbon and lipid metabolism were particularly significant in FASD Mthfr(+/-) mice. The latter changes, which included higher nuclear sterol regulatory element-binding protein 1, higher Srepb2 messenger RNA (mRNA), lower farnesoid X receptor (Nr1h4) mRNA, and lower Cyp7a1 mRNA, would lead to greater lipogenesis and reduced cholesterol catabolism into bile. CONCLUSIONS: We suggest that high folic acid consumption reduces MTHFR protein and activity levels, creating a pseudo-MTHFR deficiency. This deficiency results in hepatocyte degeneration, suggesting a 2-hit mechanism whereby mutant hepatocytes cannot accommodate the lipid disturbances and altered membrane integrity arising from changes in phospholipid/lipid metabolism. These preliminary findings may have clinical implications for individuals consuming high-dose folic acid supplements, particularly those who are MTHFR deficient.

Dietary practices in pyridoxine non-responsive homocystinuria: a European survey.

BACKGROUND: Within Europe, the management of pyridoxine (B6) non-responsive homocystinuria (HCU) may vary but there is limited knowledge about treatment practice. AIM: A comparison of dietetic management practices of patients with B6 non-responsive HCU in European centres. METHODS: A cross-sectional audit by questionnaire was completed by 29 inherited metabolic disorder (IMD) centres: (14 UK, 5 Germany, 3 Netherlands, 2 Switzerland, 2 Portugal, 1 France, 1 Norway, 1 Belgium). RESULTS: 181 patients (73% >16 years of age) with HCU were identified. The majority (66%; n=119) were on dietary treatment (1-10 years, 90%; 11-16 years, 82%; and >16 years, 58%) with or without betaine and 34% (n=62) were on betaine alone. The median natural protein intake (g/day) on diet only was, by age: 1-10 years, 12 g; 11-16 years, 11 g; and >16 years, 45 g. With diet and betaine, median natural protein intake (g/day) by age was: 1-10 years, 13 g; 11-16 years, 20 g; and >16 years, 38 g. Fifty-two percent (n=15) of centres allocated natural protein by calculating methionine rather than a protein exchange system. A methionine-free l-amino acid supplement was prescribed for 86% of diet treated patients. Fifty-two percent of centres recommended cystine supplements for low plasma concentrations. Target treatment concentrations for homocystine/homocysteine (free/total) and frequency of biochemical monitoring varied. CONCLUSION: In B6 non-responsive HCU the prescription of dietary restriction by IMD centres declined with age, potentially associated with poor adherence in older patients. Inconsistencies in biochemical monitoring and treatment indicate the need for international consensus guidelines.

Experimental evidence of oxidative stress in plasma of homocystinuric patients: a possible role for homocysteine.

Homocystinuria is an inherited disorder biochemically characterized by high urinary excretion of homocystine and increased levels of homocysteine (Hcy) and methionine in biological fluids. Affected patients usually have a variety of clinical and pathologic manifestations. Previous experimental data have shown a relationship between Hcy and oxidative stress, although very little was reported on this process in patients with homocystinuria. Therefore, in the present study we evaluated parameters of oxidative stress, namely carbonyl formation, malondialdehyde (MDA) levels, sulfhydryl content and total antioxidant status (TAS) in patients with homocystinuria at diagnosis and under treatment with a protein restricted diet supplemented by pyridoxine, folate, betaine, and vitamin B(12). We also correlated plasma Hcy and methionine concentrations with the oxidative stress parameters examined. We found a significant increase of MDA levels and carbonyl formation, as well as a reduction of sulfhydryl groups and TAS in plasma of homocystinuric patients at diagnosis relatively to healthy individuals (controls). We also verified that Hcy levels were negatively correlated with sulfhydryl content and positively with MDA levels. Furthermore, patients under treatment presented a significant reduction of the content of MDA, Hcy and methionine concentrations relatively to patients at diagnosis. Taken together, the present data indicate that lipid and protein oxidative damages are increased and the antioxidant defenses diminished in plasma of homocystinuric patients, probably due to increased reactive species elicited by Hcy. It is therefore presumed that oxidative stress participates at least in part in the pathogenesis of homocystinuria.

Insidious peripheral neuropathy occurring under treatment in infantile MTHFR deficiency.

5,10-Methylenetetrahydrofolate reductase (MTHFR) deficiency was diagnosed in a 1-month-old baby with signs of cerebral distress. Under a classic treatment using methionine supplementation, methyl donor (betaine) folinic acid, vitamin B(6) and vitamin B(12), the neuromotor development was satisfactory. At 15 years of age, however, despite no clear modification of the biochemical markers in body fluids, she developed a clinically overt peripheral axonal neuropathy. Only partial clinical improvement was obtained after reinforcement of betaine doses. Surveillance of the peripheral nerve is indicated in MTHFR deficiency, including in the infantile form with a good therapeutic compliance.

Comprehensive dental treatment under general anesthesia for patients with homocystinuria.

Continued investigation in genetic biology has helped the medical profession to prevent complications in patients with this defect. Moreover, early diagnosis and treatment are recommended, in order to prevent mental retardation and visual deterioration. Refinement of techniques concerning preparation of the patient for general anesthesia and postoperative care merit further investigation. Simple measures undertaken in the two patients described made the completion of dental treatment possible without serious complications.