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Mouse models to study the pathophysiology of combined methylmalonic acidemia and homocystinuria, cblC type.

ABSTRACT: Combined methylmalonic acidemia and homocystinuria, cblC type, is the most common inherited disorder of cobalamin metabolism and is characterized by severe fetal developmental defects primarily impacting the central nervous system, hematopoietic system, and heart. CblC was previously shown to be due to mutations in the MMACHC gene, which encodes a protein thought to function in intracellular cobalamin trafficking and biosynthesis of adenosylcobalamin (AdoCbl) and methylcobalamin (MeCbl). These coenzymes are required for the production of succinyl-CoA and methionine, respectively. However, it is currently unclear whether additional roles for MMACHC exist outside of cobalamin metabolism. Furthermore, due to a lack of sufficient animal models, the exact pathophysiology of cblC remains unknown. Here, we report the generation and characterization of two new mouse models to study the role of MMACHC in vivo. CRISPR/Cas9 genome editing was used to develop a Mmachc floxed allele (Mmachc^flox/flox), which we validated as a conditional null. For a gain-of-function approach, we generated a transgenic mouse line that over-expresses functional Mmachc (Mmachc-OE^+/tg) capable of rescuing Mmachc homozygous mutant lethality. Surprisingly, our data also suggest that these mice may exhibit a partially penetrant maternal-effect rescue, which might have implications for in utero therapeutic interventions to treat cblC. Both the Mmachc^flox/flox and Mmachc-OE^+/tg mouse models will be valuable resources for understanding the biological roles of MMACHC in a variety of tissue contexts and allow for deeper understanding of the pathophysiology of cblC.

SUBMITTER: Chern T

PROVIDER: S-EPMC7669717 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background: Methylmalonic acidemia (MMA) can display many clinical manifestations, among which acute lymphoblastic leukemia (ALL) has not been reported, and congenital heart disease (CHD) is also rare. Case presentation: We report an MMA case with ALL and CHD in a 5.5-year-old girl. With developmental delay and local brain atrophy in MRI, she was diagnosed with cerebral palsy at 9 months old. Rehabilitation was performed since then. This time she was admitted to hospital because of weakness and widespread bleeding spots. ALL-L2 (pre-B-cell) was confirmed by bone marrow morphology and immunophenotyping. Echocardiography showed patent foramen ovale. The girl was treated with VDLD and CAML chemotherapy, during which she developed seizures, edema and renal insufficiency. Decrease of muscle strength was also found in physical examination. Screening for inherited metabolic disorders showed significantly elevated levels of methylmalonate-2, acetylcarnitine (C2), propionylcarnitine (C3), C3/C2 and homocysteine. Gene analysis revealed a compound heterozygous mutaion in MMACHC (NM_015,560): c.80A > G (p.Gln27Arg) and c.609G > A (p.Trp203*). CblC type MMA was diagnosed. Intramuscular injection of cyanocobalamin and intravenous L-carnitine treatment were applied. The edema vanished gradually, and chemotherapy of small dosage of vindesine was given intermittently when condition permitted. 2 months later, muscle strength of both lower limbs were significantly improved to nearly grade 5. The levels of methylmalonic acid and homocysteine were improved. Conclusion: Metabolic disease screening and gene analysis are very necessary for diseases with complex clinical symptoms. ALL can be a rare manifestation for MMA. Synopsis: We report a case of methylmalonic acidemia with acute lymphoblastic leukemia and congenital heart disease, which uncovered the importance of genetic testing and metabolic diseases screening in patients with multiple systemic organ involvement.

Project description:BackgroundCombined methylmalonic acidemia and homocystinuria, cobalamin C type (cblC defect) is the most common inborn error of cobalamin metabolism, and different approaches have been applied to its prenatal diagnosis. To evaluate the reliability of biochemical method for the prenatal diagnosis of cblC defect, we conducted a retrospective study of our 10-year experience at a single center.Methods248 pregnancies whose probands were diagnosed as cblC defect were referred to our center for prenatal diagnosis from January 2010 to December 2019. Prenatal data of Hcy levels determined by enzymatic cycling assay, acylcarnitine analysis using liquid chromatography tandem mass spectrometry, organic acid analysis using gas chromatography mass spectrometry, and genetic analysis by direct sequencing of 248 at-risk fetuses were retrospectively reviewed.ResultsFor 2.0 and 16.0 μmol/L levels of Hcy AF samples, the relative errors were - 2.5% and 2.8%, respectively. The respective measurement uncertainties were 13.07% and 14.20%. For the 248 at-risk fetuses, 63 fetuses were affected and 185 fetuses were unaffected. Hcy level of 13.20 (6.62-43.30) μmol/L in 63 affected fetuses was significantly higher than that in 185 unaffected fetuses of 2.70 (0.00-5.80) μmol/L, and there was no overlap between the affected and unaffected groups. The diagnostic sensitivity and specificity of Hcy were 100% and 92.05%, respectively. The positive and negative predictive values of the combination of Hcy, propionylcarnitine (C3), ratio of C3 to acetylcarnitine (C2; C3/C2), methylmalonic acid (MMA), and methylcitric acid (MCA) were both 100%. Sixteen fetuses displayed inconclusive genetic results of MMACHC variants, in which seven fetuses were determined to be affected with elevated levels of Hcy, C3, C3/C2 and MMA, and their levels were 18.50 (6.70-43.30) μmol/L, 8.53(5.02-11.91) μmol/L, 0.77 (0.52-0.97), 8.96 (6.55-40.32) mmol/mol Cr, respectively. The remaining nine fetuses were considered unaffected based on a normal amniotic fluid metabolite profile.ConclusionsHcy appears to be another characteristic biomarker for the prenatal diagnosis of cblC defect. The combination of Hcy assay with acylcarnitine and organic acid analysis is a fast, sensitive, and reliable prenatal diagnostic biochemical approach. This approach could overcome the challenge of the lack of genetic analysis for families with at-risk cblC defect fetuses.

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Mouse models to study the pathophysiology of combined methylmalonic acidemia and homocystinuria, cblC type.

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