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BACKGROUND: Primary carnitine deficiency (PCD) is a rare autosomal recessive fatty acid oxidation disorder caused by variants in SLC22A5, with its prevalence and SLC22A5 gene mutation spectrum varying across races and regions. This study aimed to systematically analyze the incidence of PCD in China and delineate regional differences in the prevalence of PCD and SLC22A5 gene variants. METHODS: PubMed, Embase, Web of Science, and Chinese databases were searched up to November 2023. Following quality assessment and data extraction, a meta-analysis was performed on screening results for PCD among Chinese newborns. RESULTS: After reviewing 1,889 articles, 22 studies involving 9,958,380 newborns and 476 PCD cases were included. Of the 476 patients with PCD, 469 underwent genetic diagnosis, revealing 890 variants of 934 alleles of SLC22A5, among which 107 different variants were detected. The meta-analysis showed that the prevalence of PCD in China was 0.05 [95%CI, (0.04, 0.06)] or 1/20 000 [95%CI, (1/16 667, 1/25 000)]. Subgroup analyses revealed a higher incidence in southern China [0.07, 95%CI, (0.05, 0.08)] than in northern China [0.02, 95%CI, (0.02, 0.03)] (P < 0.001). Furthermore, the result of the meta-analysis showed that the frequency of the variant with c.1400C > G, c.51C > G, c.760C > T, c.338G > A, and c.428C > T were 45% [95%CI, (34%, 59%)], 26% [95%CI, (22%, 31%)], 14% [95%CI, (10%, 20%)], 6% [95%CI, (4%, 8%)], and 5% [95%CI, (4%, 8%)], respectively. Among the subgroup analyses, the variant frequency of c.1400C > G in southern China [39%, 95%CI, (29%, 53%)] was significantly lower than that in northern China [79, 95%CI, (47, 135)] (P < 0.05). CONCLUSIONS: This study systematically analyzed PCD prevalence and identified common SLC22A5 gene variants in the Chinese population. The findings provide valuable epidemiological insights and guidance for future PCD screening effects in newborns.
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Carnitina , Hiperamonemia , Miembro 5 de la Familia 22 de Transportadores de Solutos , Humanos , China/epidemiología , Carnitina/deficiencia , Recién Nacido , Miembro 5 de la Familia 22 de Transportadores de Solutos/genética , Hiperamonemia/genética , Hiperamonemia/epidemiología , Hiperamonemia/diagnóstico , Cardiomiopatías/genética , Cardiomiopatías/epidemiología , Enfermedades Musculares/genética , Enfermedades Musculares/epidemiología , Mutación/genética , Tamizaje Neonatal/métodos , Pueblos del Este de AsiaRESUMEN
Introduction: Primary carnitine deficiency (PCD) is a rare autosomal recessive disorder caused by loss of function mutations in the solute carrier family 22 member 5 (SLC22A5) gene that encodes a high-affinity sodium-ion-dependent organic cation transporter protein (OCTN2). Carnitine deficiency can result in acute metabolic decompensation or, in a more insidious presentation, cardiomyopathy. Cardiomyopathy associated with PCD often presents with life-threatening heart failure. This presentation also usually includes skeletal muscle myopathy. Early recognition of this disorder and treatment with carnitine can avoid life-threatening complications related to cardiomyopathy. Case Presentation: Herein, we present a 10-month-old male patient with PCD, which was diagnosed while investigating the etiology of dilated cardiomyopathy and confirmed by molecular genetic analysis. Conclusion: Homozygous c.254_265 insGGCTCGCCACC (p.I89Gfs) pathogenic variant of the SLC22A5 gene was detected. With oral L-carnitine supplementation, the free carnitine level increased up to 14 µmol/L and the symptoms disappeared. LVEF increased by 45-70%. We would like to emphasize that this problem is a combination of the metabolic decompensation and the cardiac phenotypes, which are usually separated to either phenotype.
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BACKGROUND: Primary carnitine deficiency (PCD) denotes low carnitine levels with an autosomal recessive pattern of inheritance. Cardiomyopathy is the most common cardiac symptom in patients with PCD, and early diagnosis can prevent complications. Next-generation sequencing can identify genetic variants attributable to PCD efficiently. OBJECTIVE: We aimed to detect the genetic cause of the early manifestations of hypertrophic cardiomyopathy and metabolic abnormalities in an Iranian family. METHODS: We herein describe an 8-year-old boy with symptoms of weakness and lethargy diagnosed with PCD through clinical evaluations, lab tests, echocardiography, and cardiac magnetic resonance imaging. The candidate variant was confirmed through whole-exome sequencing, polymerase chain reaction, and direct Sanger sequencing. The binding efficacy of normal and mutant protein-ligand complexes were evaluated via structural modeling and docking studies. RESULTS: Clinical evaluations, echocardiography, and cardiac magnetic resonance imaging findings revealed hypertrophic cardiomyopathy as a clinical presentation of PCD. Whole-exome sequencing identified a new homozygous variant, SLC22A5 (NM_003060.4), c.821G > A: p.Trp274Ter, associated with carnitine transport. Docking analysis highlighted the impact of the variant on carnitine transport, further indicating its potential role in PCD development. CONCLUSIONS: The c.821G > A: p.Trp274Ter variant in SLC22A5 potentially acted as a pathogenic factor by reducing the binding affinity of organic carnitine transporter type 2 proteins for carnitine. So, the c.821G > A variant may be associated with carnitine deficiency, metabolic abnormalities, and cardiomyopathic characteristics.
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Cardiomiopatías , Cardiomiopatía Hipertrófica , Hiperamonemia , Enfermedades Musculares , Masculino , Humanos , Niño , Enfermedades Musculares/diagnóstico , Enfermedades Musculares/genética , Carnitina/genética , Carnitina/metabolismo , Irán , Miembro 5 de la Familia 22 de Transportadores de Solutos/genética , Hiperamonemia/diagnóstico , Hiperamonemia/genética , Hiperamonemia/complicaciones , Cardiomiopatías/diagnóstico por imagen , Cardiomiopatías/genética , Cardiomiopatía Hipertrófica/complicaciones , MutaciónRESUMEN
Primary carnitine deficiency (PCD) caused by pathogenic variants in the solute carrier family 22 member 5 (SLC22A5) gene is a rare autosomal recessive disease that results in defective fatty acid oxidation. PCD can be detected through tandem mass spectrometry (MS/MS), but transplacental transport of free carnitine from mothers may cause false negatives or positives during newborn screening (NBS). This study aimed to analyze the genetic characteristics of SLC22A5 and estimate the prevalence of PCD in the Chinese population, providing useful information for NBS and genetic counseling. We manually curated SLC22A5 pathogenic or likely pathogenic (P/LP) variants according to the American College of Medical Genetics and Genomics (ACMG) guidelines and identified 128 P/LP variants. Based on the China Neonatal Genomes Project (CNGP), the estimated PCD prevalence was 1:17,456, which was higher than that in other populations. The genotype-phenotype association analysis showed that patients carrying homozygous c.760C>T and c.844C>T were more likely to present cardiomyopathy, whereas those carrying homozygous c.1400C>G were more likely to be asymptomatic (all p-values < 0.05). We found that there was no significant difference in initial C0 concentrations between patients and carriers, but there was a significant difference in the second-tier screening of C0 concentration between them (p-value < 0.05). We established a cost-effective variant panel containing 10 high-frequency sites and developed a screening algorithm incorporating gene panels with MS/MS, which could rescue one more patient who was undetected from MS/MS. In conclusion, the prevalence of PCD in the Chinese population is relatively high. The combination of conventional NBS with genetic sequencing is suggested for early diagnosis of PCD.
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Introduction: To investigate the clinical characteristics and disease outcomes of a pedigree with compound heterozygous mutations in the SLC22A5 gene. Methods: Serum acylcarnitine profiles of patients were analyzed using tandem mass spectrometry. DNA samples isolated from patients and their first-degree relatives were subjected to high-throughput sequencing, and mutations were validated using Sanger sequencing. Results: The proband, a 4-month-old girl, presented with seizure episodes and mild cardiac hypertrophy and was diagnosed with primary carnitine deficiency (PCD), with carnitine levels of 5.165â mol/L. Her brother, a 6-year-and 4-month-old boy, was also diagnosed with PCD with serum-free carnitine levels of 1.014â mol/L (reference values 10-60â mol/L). Compound heterozygous mutations (c.760C > T [p.R254X] and c.825G > A [p.W275X]) were detected in the SLC22A5 gene in both patients and were inherited from the mother and father, respectively. Oral L-carnitine significantly improved or resolved the clinical symptoms. Conclusion: Children with compound mutations in SLC22A5 may present different clinical manifestations, particularly at different ages. Early clinical manifestations have a greater impact on the organs and may cause irreversible damage. PCD can cause epilepsy and dilated cardiomyopathy. Tandem mass spectrometry and high-throughput sequencing are recommended to confirm the diagnosis. Early L-carnitine supplementation can improve symptoms and reverse organ damage in some children. Tandem mass spectrometry should be used to screen for newborns with a family history of PCD.
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l-carnitine is indispensable for transfer of fatty acids to mitochondria for the process of ß-oxidation, a process, whose significance in cancer has drawn attention in recent years. In humans majority of carnitine is delivered by diet and enters the cell due to activity of solute carriers (SLCs), mainly by ubiquitously expressed organic cation/carnitine transporter (OCTN2/SLC22A5). In control and cancer human breast epithelial cell lines the major fraction of OCTN2 is present as a not matured non-glycosylated form. Studies on overexpressed OCTN2 demonstrated an exclusive interaction with SEC24C, as the cargo-recognizing subunit of coatomer II in transporter exit from endoplasmic reticulum. Co-transfection with SEC24C dominant negative mutant completely abolished presence of the mature form of OCTN2, pointing to a possibility of trafficking regulation. SEC24C was previously shown to be phosphorylated by serine/threonine kinase AKT, known to be activated in cancer. Further studies on breast cell lines showed that inhibition of AKT with MK-2206 in control and cancer lines decreased level of OCTN2 mature form. Proximity ligation assay showed that phosphorylation of OCTN2 on threonine was significantly abolished by AKT inhibition with MK-2206. Carnitine transport was positively correlated with the level of OCTN2 phosphorylated by AKT on threonine moiety. The observed regulation of OCTN2 by AKT places this kinase in the center of metabolic control. This points to both proteins, AKT and OCTN2, as druggable targets, in particular in a combination therapy of breast cancer.
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Neoplasias de la Mama , Proteínas de Transporte de Catión Orgánico , Humanos , Femenino , Proteínas de Transporte de Catión Orgánico/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Carnitina/metabolismo , Miembro 5 de la Familia 22 de Transportadores de Solutos/metabolismo , Cationes/metabolismoRESUMEN
BACKGROUND: Primary carnitine deficiency (PCD) is screened by expanded newborn screening (NBS) using tandem mass spectrometry (MS/MS) that can detect both affected neonates and mothers. This study aimed to delineate the clinical, biochemical, and molecular findings of Thai PCD patients. METHODS: Expanded NBS using MS/MS was implemented in Bangkok and 146,757 neonates were screened between 2014 and 2018. PCD was screened by low free carnitine (C0) levels in dried blood spots. Plasma C0 levels and C0 clearance values were measured in neonates and their mothers with positive screening results. Clinically diagnosed cases were described. The coding regions and intron-exon boundaries of the SLC22A5 gene were sequenced in all cases with low plasma C0 levels. RESULTS: There were 14 cases with confirmed PCD: two clinically diagnosed cases, and 12 cases identified through NBS including five newborns, six mothers, and one older sibling. Thus, the incidence of PCD in neonates was 1:29,351. All affected neonates and mothers were asymptomatic except one mother with dilated cardiomyopathy. SLC22A5 gene sequencing identified biallelic causative variants in all cases, comprising 10 different variants of which four were novel. c.51C > G (p.Phe17Leu) and c.760C > T (p.Arg254Ter) were the most prevalent variants in this study. Cases with significant clinical features tended to have higher C0 clearance values. CONCLUSIONS: Primary carnitine deficiency is a common inherited metabolic disorder (IMD) in Thailand. Our findings broaden the spectrum of SLC22A5 variants. The future national NBS program will shed more light on PCD and other IMDs in Thailand.
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Cardiomiopatías , Miembro 5 de la Familia 22 de Transportadores de Solutos , Espectrometría de Masas en Tándem , Femenino , Humanos , Recién Nacido , Cardiomiopatías/diagnóstico , Cardiomiopatías/genética , Carnitina/metabolismo , Mutación , Tamizaje Neonatal/métodos , Miembro 5 de la Familia 22 de Transportadores de Solutos/genética , Pueblos del Sudeste Asiático/genética , Tailandia/epidemiologíaRESUMEN
Background: Primary carnitine deficiency (PCD) is an autosomal recessive disease caused by mutations in the SLC22A5 gene, which encodes the organic cation transporter 2 (OCTN2). Patients with PCD may be at risk of skeletal or cardiac myopathy, metabolic decompensation, and even sudden death. This study aimed to analyze the biochemical, clinical, and genetic characteristics of PCD patients identified by newborn screening (NBS) in Shanghai. Methods: Dried blood spot (DBS) samples of newborns were analyzed through tandem mass spectrometry (MS/MS) from January 2003 to December 2021. Newborns with low free carnitine (C0) levels were recalled. Mutation in the SLC22A5 gene was analyzed on suspected positive newborns with low C0 levels after recall. Results: 1,247,274 newborns were screened by MS/MS and 40 newborns were diagnosed with PCD, therefore the incidence of PCD in Shanghai was approximately 1:31,200. The mean C0 level in newborns with PCD was 5.37 ± 1.79 µmol/L before treatment and increased to 24.45 ± 10.87 µmol/L after treatment with L-carnitine. Twenty-three different variants were identified in the SLC22A5 gene, including 8 novel variants, of which c.51C>G (p.F17L) was the most frequent (27.27%, 18/66), followed by c.1400C>G (p.S467C) (25.76%, 17/66). Almost all the screened PCD patients were asymptomatic. Conclusion: NBS via MS/MS was a quick and efficient method for the early diagnosis of PCD. The incidence of PCD in Shanghai was 1:31,200. Eight novel variants were identified, which greatly expanded the variant spectrum of SLC22A5. MS/MS combined with genetic testing could effectively improve the diagnostic accuracy of PCD.
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(1) Background: Many transporters of the SLC22 family (e.g., OAT1, OAT3, OCT2, URAT1, and OCTN2) are highly expressed in the kidney. They transport drugs, metabolites, signaling molecules, antioxidants, nutrients, and gut microbiome products. According to the Remote Sensing and Signaling Theory, SLC22 transporters play a critical role in small molecule communication between organelles, cells and organs as well as between the body and the gut microbiome. This raises the question about the potential role of SLC22 transporters in cancer biology and treatment. (2) Results: In two renal cell carcinoma RNA-seq datasets found in TCGA, KIRC and KIRP, there were multiple differentially expressed (DE) SLC22 transporter genes compared to normal kidney. These included SLC22A6, SLC22A7, SLC22A8, SLC22A12, and SLC22A13. The patients with disease had an association between overall survival and expression for most of these DE genes. In KIRC, the stratification of patient data by pathological tumor characteristics revealed the importance of SLC22A2, SLC22A6, and SLC22A12 in disease progression. Interaction networks combining the SLC22 with ADME genes supported the centrality of SLC22 transporters and other transporters (ABCG2, SLC47A1) in disease progression. (3) Implications: The fact that many of these genes are uric acid transporters is interesting because altered uric acid levels have been associated with kidney cancer. Moreover, these genes play key roles in processing metabolites and chemotherapeutic compounds, thus making them potential therapeutic targets. Finally, our analyses raise the possibility that current approaches may undertreat certain kidney cancer patients with low SLC22 expression and only localized disease while possibly overtreating more advanced disease in patients with higher SLC22 expression. Clinical studies are needed to investigate these possibilities.
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OCTN2 (SLC22A5) is a carnitine transporter whose main function is the active transport of carnitine into cells. In skeletal muscle and other organs, the regulation of the SLC22A5 gene transcription has been shown to depend on the nuclear transcription factor PPAR-α. Due to the observation that the muscle OCTN2 mRNA level is maintained in PPAR-α knock-out mice and that PGC-1α overexpression in C2C12 myoblasts increases OCTN2 mRNA expression, we suspected additional regulatory pathways for SLC22A5 gene transcription. Indeed, we detected several binding sites of the myocyte-enhancing factor MEF2 in the upstream region of the SLC22A5 gene, and MEF2C/MEF2D stimulated the activity of the OCTN2 promoter in gene reporter assays. This stimulation was increased by PGC-1α and was blunted for a SLC22A5 promoter fragment with a mutated MEF2 binding site. Further, we demonstrated the specific binding of MEF2 to the SLC22A5 gene promoter, and a supershift of the MEF2/DNA complex in electrophoretic mobility shift assays. In immunoprecipitation experiments, we could demonstrate the interaction between PGC-1α and MEF2. In addition, SB203580, a specific inhibitor of p38 MAPK, blocked and interferon-γ stimulated the transcriptional activity of the SLC22A5 gene promoter. Finally, mice with muscle-specific overexpression of OCTN2 showed an increase in OCTN2 mRNA and protein expression in skeletal muscle. In conclusion, we detected and characterized a second stimulatory pathway of SLC22A5 gene transcription in skeletal muscle, which involves the nuclear transcription factor MEF2 and co-stimulation by PGC-1α and which is controlled by the p38 MAPK signaling cascade.
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Carnitina , Receptores Activados del Proliferador del Peroxisoma , Ratones , Animales , Carnitina/metabolismo , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Interferón gamma/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Músculo Esquelético/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Factores de Transcripción MEF2/genética , Factores de Transcripción MEF2/metabolismoRESUMEN
Due to lack of nuclei and de novo protein synthesis, post-translational modification (PTM) is imperative for erythrocytes to regulate oxygen (O2) delivery and combat tissue hypoxia. Here, we report that erythrocyte transglutminase-2 (eTG2)-mediated PTM is essential to trigger O2 delivery by promoting bisphosphoglycerate mutase proteostasis and the Rapoport-Luebering glycolytic shunt for adaptation to hypoxia, in healthy humans ascending to high altitude and in two distinct murine models of hypoxia. In a pathological hypoxia model with chronic kidney disease (CKD), eTG2 is critical to combat renal hypoxia-induced reduction of Slc22a5 transcription and OCNT2 protein levels via HIF-1α-PPARα signaling to maintain carnitine homeostasis. Carnitine supplementation is an effective and safe therapeutic approach to counteract hypertension and progression of CKD by enhancing erythrocyte O2 delivery. Altogether, we reveal eTG2 as an erythrocyte protein stabilizer orchestrating O2 delivery and tissue adaptive metabolic reprogramming and identify carnitine-based therapy to mitigate hypoxia and CKD progression.
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Carnitina , Insuficiencia Renal Crónica , Animales , Carnitina/metabolismo , Eritrocitos/metabolismo , Eritrocitos/patología , Homeostasis , Humanos , Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Ratones , Oxígeno/metabolismo , Insuficiencia Renal Crónica/patología , Miembro 5 de la Familia 22 de Transportadores de Solutos/metabolismo , Transglutaminasas/metabolismoRESUMEN
BACKGROUND: Primary carnitine deficiency (PCD) is an autosomal recessive disorder of carnitine transportation that leads to impaired fatty acid oxidation. Large-scale studies on newborn screening (NBS) for PCD are limited. This study aimed to investigate the biochemical and genetic characteristics of patients with PCD detected through NBS. RESULTS: A total of 548 247 newborns were screened for PCD between January 2014 and June 2021; 1714 newborns with low free carnitine (C0) levels were called back and 49 patients were diagnosed with PCD. The latest incidence rate in Quanzhou, China, was estimated to be 1 in 11 189 newborns. NBS results showed that the 49 patients had varying degrees of decreased C0 levels, whereas seven patients exhibited normal C0 levels during the recall review. All patients harbored biallelic pathogenic variants of the SLC22A5 gene. Nineteen distinct SLC22A5 variants were detected in these 49 patients, and most of the detected variants were clustered in exons 1, 4, and 7. The top eight variants had an allele frequency of 86.73%. The most common variant was c.760C > T (p.R254*) with an allele frequency of 31.63%, followed by c.51C > G (p.F17L) (17.35%) and c.1400C > G (p.S467C) (16.33%). The C0 level of patients with the N/N genotype was significantly lower than that of the M/M group. The C0 levels of patients with genotypes of R254*/R254* and R254*/F17L were far lower than those of patients with the R254*/S467C genotype. CONCLUSIONS: This study presented more than 500,000 NBS data with the latest incidence of 1:11 189 in the Quanzhou area. The SLC22A5 variant spectrum in the selected southern Chinese population has been updated. Patients with null variants were associated with low C0 levels. Combining NBS with genetic testing is critical to improve screening efficiency because patients with PCD may have normal C0 levels during NBS and recall review.
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Hiperamonemia , Enfermedades Musculares , Cardiomiopatías , Carnitina/deficiencia , Humanos , Hiperamonemia/diagnóstico , Hiperamonemia/genética , Recién Nacido , Enfermedades Musculares/diagnóstico , Enfermedades Musculares/genética , Mutación/genética , Tamizaje Neonatal , Miembro 5 de la Familia 22 de Transportadores de Solutos/genéticaRESUMEN
Primary carnitine deficiency (PCD) is an autosomal recessive disorder that could result in sudden death. It is caused by a defect in the carnitine transporter encoded by SLC22A5 (Solute Carrier Family 22 Member 5, MIM:603377). Currently, a number of variants in SLC22A5 have been identified, however, the PCD prevalence and its variants in Ningbo area are unclear. In this study, we screened 265,524 newborns by using tandem mass spectrometry. Variants in SLC22A5 were further detected by next-generation sequencing in individuals with abnormal free carnitine levels (C0). We identified 53 newborns with abnormal C0 levels and 26 with variants in SLC22A5. Among them, 16 with compound heterozygous or homozygous variants in SLC22A5 were diagnosed with PCD, suggesting the PCD birth prevalence in Ningbo city was 1/16,595. Moreover, the C0 level was significantly (P = 0.013) higher in PCD patients than in those with one variant. Besides, the c.1400C > G (p. S467C) and c.51C > G (p. F17L) variants were the most frequent and six novel variants are all predicted to be damaging. This study reports the largest PCD patients in Ningbo area by newborn screening and expands the variant spectrum of SLC22A5. Our findings demonstrate the clinical value of combining NBS program results with DNA analysis for the diagnosis of PCD.
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Primary carnitine deficiency (PCD) is a rare autosomal recessive disorder caused by loss of function mutations in the solute carrier family 22 member 5 (SLC22A5) gene that encodes a high-affinity sodium-ion-dependent organic cation transporter protein (OCTN2). Reduced carnitine transport results in diminished fatty acid oxidation in heart and skeletal muscle and carnitine wasting in urine. We present a case of PCD diagnosed in an adult female after a positive newborn screen (NBS) for PCD that was not confirmed on follow-up testing. The mother was referred for evaluation of persistent fatigue and possible hypothyroidism even though all measurements of thyroid-stimulating hormone were well within the range of 0.4 to 2.5 mIU/L expected for reproductive-age women. She was found to have unequivocally low levels of both total carnitine and carnitine esters, and genetic testing revealed compound heterozygosity for 2 SLC22A5 mutations. One mutation (c.34G>A [p.Gly12Ser]) is a known missense mutation with partial OCTN2 activity, but the other mutation (c.41G>A [p.Trp14Ter]) is previously unreported and results in a premature stop codon and truncated OCTN2. This case illustrates that some maternal inborn errors of metabolism can be identified by NBS and that maternal carnitine levels should be checked after a positive NBS test for PCD.
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Hiperamonemia , Adulto , Cardiomiopatías , Carnitina/deficiencia , Femenino , Humanos , Hiperamonemia/diagnóstico , Hiperamonemia/genética , Recién Nacido , Enfermedades Musculares , Mutación , Miembro 5 de la Familia 22 de Transportadores de Solutos/genéticaRESUMEN
BACKGROUND: Rare studies focused on the tandem mass spectrometry (MS/MS) findings for the primary carnitine deficiency (PCD) in the neonates in China mainland. In this study, we aim to analyze the gene mutation spectrum of PCD in Fujian Province in China mainland. METHODS: Primary carnitine deficiency (PCD) samples used in this study were selected from 95,453 cases underwent neonatal screening between May 2015 and February 2020. SLC22A5 gene sequencing was performed on the neonates and their parents with C0 level of less than 8.8 µmol/L. RESULTS: Ten patients (male: 7; female: 3) were finally included in this study. Among these patients, nine were neonates, and one was maternal decline of C0 of less than 8.8 µmol/L. The maternal case showed two types of mutations of SLC22A5 including c.760C>T(p.R254*) and c.1400C>G(p.S467C). The other nine neonates showed compound mutations involving nine types in 18 sites, among which two mutations [i.e., c.37G>T(p.E13*) and c.694A>G(p.T232A)] were novel that had never been reported before. Bioinformatic analysis indicated that c.37G>T(p.E13*) was a pathogenic mutation, while the c.694A>G (p.T232A) was considered to be likely pathogenic. CONCLUSION: MS/MS screening on PCD contributed to the early diagnosis and screening. In addition, SLC22A5 gene mutation analysis contributed to the PCD screening.
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Cardiomiopatías/genética , Carnitina/deficiencia , Hiperamonemia/genética , Enfermedades Musculares/genética , Fenotipo , Adulto , Cardiomiopatías/sangre , Cardiomiopatías/diagnóstico , Carnitina/análogos & derivados , Carnitina/sangre , Carnitina/genética , Femenino , Frecuencia de los Genes , Humanos , Hiperamonemia/sangre , Hiperamonemia/diagnóstico , Lactante , Masculino , Enfermedades Musculares/sangre , Enfermedades Musculares/diagnóstico , Mutación , Miembro 5 de la Familia 22 de Transportadores de Solutos/genéticaRESUMEN
Gliomas are the most common primary malignant brain tumor in adults, but current treatment for glioblastoma multiforme (GBM) is insufficient. Even though glucose is the primary energetic substrate of glioma cells, they are capable of using fatty acids to generate energy. Fatty acid oxidation (FAO) in mitochondria requires L-carnitine for the formation of acylcarnitines by carnitine palmitoylotransferase 1 (CPT1) and further transport of acyl carnitine esters to mitochondrial matrix. Carnitine can be delivered to the cell by an organic cation/carnitine transporter-SLC22A5/OCTN2. In this study, we show that SLC22A5 is up-regulated in glioma cells and that they vary in the amount of SLC22A5 in the plasma membrane. Research on glioma cells (lines U87MG, LN229, T98G) with various expression levels of SLC22A5 demonstrated a correlation between the FAO rate, the level of the transporter, and the carnitine transport. Inhibition of carnitine transport by chemotherapeutics, such as vinorelbine and vincristine, led to inhibition of FAO, which was further intensified by etomoxir-a CPT1 inhibitor. This led to reduced viability and increased apoptosis in glioma cells. Modulation of SLC22A5 level by either silencing or up-regulation of SLC22A5 also affected glioma cell survival in a FAO-dependent way. These observations suggest that the survival of glioma cells is heavily reliant on both FAO and SLC22A5 activity, as well as that CPT1 and SLC22A5 might be possible drug targets.
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Carnitina/metabolismo , Ácidos Grasos/metabolismo , Glioma/metabolismo , Miembro 5 de la Familia 22 de Transportadores de Solutos/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Glioma/patología , Humanos , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/fisiología , ARN Interferente Pequeño/farmacologíaRESUMEN
BACKGROUND AND AIMS: Primary carnitine deficiency (PCD) is an autosomal recessive disease caused by functional defects in the carnitine transporter OCTN2 due to mutations in SLC22A5. Here, we aimed to understand the incidence, clinical, biochemical, and molecular features of PCD in Quanzhou, China. MATERIALS AND METHODS: Newborn screening (NBS) was performed through tandem mass spectrometry (MS/MS) to detect genetic metabolic diseases. Next-generation sequencing was used to detect SLC22A5 mutations in patients with suspected PCD. RESULTS: From 364,545 newborns screened, 36 were diagnosed with PCD, in addition to five mothers. The incidence of PCD in children in the Quanzhou area was 1:10126. Eighteen SLC22A5 variants were found, with five novel ones. The most prevalent variant in neonatal and maternal patients was c.760C > T (p.R254*). Twenty-five neonatal patients received L-carnitine supplementation; however, one patient discontinued treatment and sudden death occurred. One sibling presented repeated fatigue, hypoglycemia, and coma, but the symptoms disappeared after treatment. Two mothers with PCD claimed to feel weak and easily fatigued. CONCLUSION: The incidence of PCD is relatively high in the Quanzhou area. Five novel variants were found, broadening the mutation spectrum of SLC22A5. NBS is effective in identifying PCD, and sudden death may be prevented with timely treatment.
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Cardiomiopatías , Tamizaje Neonatal , Carnitina/deficiencia , Niño , China/epidemiología , Femenino , Humanos , Hiperamonemia , Recién Nacido , Enfermedades Musculares , Mutación , Miembro 5 de la Familia 22 de Transportadores de Solutos/genética , Espectrometría de Masas en TándemRESUMEN
OBJECTIVES: Primary carnitine deficiency (PCD) is a rare fatty acid metabolism disorder that can cause neonatal death. This study aims to analyze carnitine levels and detect SLC22A5 gene in newborns with carnitine deficiency, to provide a basis for early diagnosis of PCD, and to explore the relationship between carnitine in blood and SLC22A5 genotype. METHODS: A total of 40 neonates with low free carnitine (C0<10 µmol/L) in blood were the subjects of the study. SLC22A5 gene was detected by Sanger sequencing to analyze the value of carnitine, the results of gene test and their relationship. RESULTS: A total of 15 variants of SLC22A5 gene were detected, including 11 pathogenic or likely pathogenic variants and 4 variants of uncertain significance. There were 5 new mutations: c.288delG (p.G96fsX33), c.744_745insTCG (p.M258_L259insS), c.752A>G (p.Y251C), c.495 C>A (p.R165E), and c.1298T>C (p.M433T). We found 14 PCD patients including 2 homozygous mutations and 12 heterozygous mutations, 14 with 1 mutation, and 12 with no mutation among 40 children. The C0 concentration of children with SLC22A5 gene homozygous or complex heterozygous mutations was (4.95±1.62) µmol/L in the initial screening, and (3.90±1.33) µmol/L in the second screening. The C0 concentration of children with no mutation was (7.04±2.05) µmol/L in the initial screening, and (8.02±2.87) µmol/L in the second screening. There were significant differences between children with homozygous or compound heterozygous mutations and with no mutation in C0 concentration of the initial and the second screening (both P<0.05), as well as between children with truncated mutation and with untruncated mutation in C0 concentration of the initial screening (P=0.022). CONCLUSIONS: There are 5 new mutations which enriched the mutation spectrum of SLC22A5 gene. C0<5 µmol/L is highly correlated with SLC22A5 gene homozygous or compound heterozygous mutations. Children with truncated mutation may have lower C0 concentration than that with untruncated mutation in the initial screening.
Asunto(s)
Hiperamonemia , Enfermedades Musculares , Cardiomiopatías , Carnitina/deficiencia , Niño , Humanos , Hiperamonemia/genética , Recién Nacido , Enfermedades Musculares/genética , Mutación , Miembro 5 de la Familia 22 de Transportadores de Solutos/genéticaRESUMEN
Objective: To evaluate and improve the performance of the newborn screening program for primary carnitine deficiency (PCD) based on tandem mass spectrometry and to investigate the incidence of PCD and molecular characteristics of SLC22A5 gene in Guangzhou. Methods: A total of 200 180 neonates born in Guangzhou from 2015 to 2019 were enrolled into the newborn screening program for PCD by tandem mass spectrometry at Guangzhou Newborn Screening Center. The positive results of screening for PCD was defined as free carnitine (C0) less than 10 µmol/L with decreased acylcarnitine species in dried blood spots of three to seven days after birth. Screen-positive newborns and their mothers were recalled for another blood spot sample. The diagnosis was confirmed based on decreased levels of C0 and acylcarnitine species in recalled blood spots and genetic analysis in SLC22A5 gene sequencing. The utility of using the sum of propionylcarnitine and palmitoylcarnitine (C3+C16) as a biomarker for acylcarnitine species in newborn screening was retrospectively evaluated. The levels of C0 and (C3+C16) at first screening were compared between newborns with PCD and newborns born to mothers with PCD by independent t test. The variant spectrum and known pathogenic variants carrier rate of SLC22A5 in 2 395 healthy children in Guangzhou Women and Children's Medical Center through whole exon sequencing were analyzed. Results: Among 200 180 neonates, 239 (0.12%) cases were screen-positive for PCD. A total of 37 patients including 15 newborns and 22 mothers had confirmed PCD. The incidence of PCD was 1/13 345 in newborns and 1/9 099 in mothers, respectively. The positive predictive value of this program was 15.5%. Taking cutoff values of C0<8.5 µmol/L or C0 8.5~9.9 µmol/L with (C3+C16)<2 µmol/L, the number of screen-positive cases would be reduced from 810 to 224 without additional false negative case, when compared with cutoff value C0<10 µmol/L only. Both levels of C0 and (C3+C16) at first screening were not significant difference between newborns with PCD and newborns born to mothers with PCD ((6.2±2.4) vs. (5.0±1.8) µmol/L, (1.4±0.4) vs. (1.2±0.5) µmol/L, t=3.826, 0.326; P=0.058, 0.572). Seven PCD mothers experienced moderate fatigue and dizziness in the morning. One of them presented with cardiomyopathy in pregnancy. Genetic analysis of the SLC22A5 gene showed that p.S467C, p.F17L, p.R254X were the three most common variants in newborns with PCD. In PCD mothers and healthy children, the p.S467C, p.F17L and R399W were the three most common whereas the severe variant p.R254X was rare. The population carrier rate for pathogenic variants was 1 in 65 and the estimated incidence of PCD was about 1/16 500. Conclusions: Newborn screening can detect PCD both in newborns and mothers. Adding a quantitative biomarker (C3+C16) <2 µmol/L into the newborn screening program can improve the PCD screen performance. The severe variant p.R253X was common in PCD newborns but rare in PCD mothers and healthy children, indicating that the current screening program maybe failed to detect all PCD newborns and under-estimated the incidence rate of PCD in Guangzhou.
Asunto(s)
Cardiomiopatías/genética , Carnitina/sangre , Carnitina/deficiencia , Hiperamonemia/diagnóstico , Enfermedades Musculares/diagnóstico , Tamizaje Neonatal/métodos , Miembro 5 de la Familia 22 de Transportadores de Solutos/genética , Cardiomiopatías/diagnóstico , Cardiomiopatías/metabolismo , Carnitina/genética , Niño , Femenino , Predisposición Genética a la Enfermedad , Humanos , Hiperamonemia/genética , Recién Nacido , Enfermedades Musculares/genética , Embarazo , Estudios Retrospectivos , Espectrometría de Masas en TándemRESUMEN
Testing for primary carnitine deficiency (PCD) has been implemented in many newborn screening (NBS) programs, but few large-scale studies on NBS for PCD have been reported in China. This study aimed to assess the incidence and biochemical, clinical, and genetic characteristics of PCD discovered by NBS. Dried blood spots from newborns were analyzed by tandem mass spectrometry (MS/MS) and suspected positive patients were further tested using molecular genetic analysis. Infants who carried two variants in SLC22A5 or those with extremely low free carnitine levels during recall were referred for follow-up and treatment. Over 3.4 million newborns were screened and 113 newborns were diagnosed with PCD, yielding a positive predictive value of 1.93%. In addition, 63 mothers with PCD were identified. The incidence of PCD in newborns and mothers in Zhejiang was 1:30,182 and 1:54,137, respectively. Thirty-seven distinct variants were identified in SLC22A5 of which 10 were novel. c.1400C > G (p.S467C) was the most prevalent variant in both newborns and mothers with PCD, while c.760C > T (p.R254*), which is reportedly common in other Chinese regions, was rarely detected in maternal PCD patients. This study reports the largest series of patients with PCD detected by NBS and identifies 10 novel variants, expanding the variant spectrum of SLC22A5.