“Cytopenias and proteinuria as key diagnostic features of Imerslund-Gräsbeck Syndrome: a case series of four Argentine patients”

“Cytopenias and proteinuria as key diagnostic features of Imerslund-Gräsbeck Syndrome: a case series of four Argentine patients” | Authorea try { document.documentElement.classList.add('js'); } catch (e) { } var _gaq = _gaq || []; _gaq.push(['_setAccount', 'G-8VDV14Y67G']); _gaq.push(['_trackPageview']); (function() { var ga = document.createElement('script'); ga.type = 'text/javascript'; ga.async = true; ga.src = ('https:' == document.location.protocol ? 'https://ssl' : 'http://www') + '.google-analytics.com/ga.js'; var s = document.getElementsByTagName('script')[0]; s.parentNode.insertBefore(ga, s); })(); Skip to main content Preprints Collections Wiley Open Research IET Open Research Ecological Society of Japan All Collections About About Authorea FAQs Contact Us Quick Search anywhere Search for preprint articles, keywords, etc. Search Search ADVANCED SEARCH SCROLL This is a preprint and has not been peer reviewed. Data may be preliminary. 25 March 2025 V1 Latest version Share on “Cytopenias and proteinuria as key diagnostic features of Imerslund-Gräsbeck Syndrome: a case series of four Argentine patients” Authors : Verónica Bindi 0000-0002-5872-199X [email protected] , Crespo Carolina , Gravina L. , Hernán Eiroa , and Rossetti E. Authors Info & Affiliations https://doi.org/10.22541/au.174291335.58828505/v1 276 views 112 downloads Contents Abstract Supplementary Material Information & Authors Metrics & Citations View Options References Figures Tables Media Share Abstract Imerslund-Gräsbeck syndrome (IGS) is a rare autosomal recessive disorder caused by pathogenic variants in CUBN or AMN genes, leading to selective vitamin B12 malabsorption. Symptoms include failure to thrive, cytopenia, and proteinuria. This study aims to analyze clinical, biochemical, and molecular genetic profiles of 4 patients diagnosed with IGS at a single center in Argentina. Macrocytic anemia and neutropenia were present in three cases, two with thrombocytopenia. All had proteinuria. A novel CUBN : c.2450G>A; p.(Cys817Tyr) variant was found in three patients, while the fourth carried AMN : c.1006+34_1007-31del in homozygous state. Early and lifelong parenteral vitamin B12 ensures an excellent prognosis. Title: “Cytopenias and proteinuria as key diagnostic features of Imerslund-Gräsbeck Syndrome: a case series of four Argentine patients” Authors: Bindi V. 1 Crespo C. 2 Gravina L. 2 Eiroa H. 1 Rossetti E. 3 Filliations of authors 1. Department of Inborn Errors of Metabolism, Hospital de Pediatría J. P. Garrahan, Buenos Aires, Argentina. 2. Molecular Biology Laboratory - Genetics, Hospital de Pediatría J. P. Garrahan, Buenos Aires, Argentina. 3. Pediatric Hematology and Oncology Department, Hospital de Pediatría Juan P. Garrahan, Buenos Aires, Argentina Corresponding author: Verónica Bindi Department of Inborn Errors of metabolism, Hospital de Pediatría Garrahan Combate de los Pozos 1881 (C 1245) Buenos Aires, Argentina e-mail: [email protected] Telephone: (+54-11) 4122-6309 Word Count for: a) Abstract 100 b) Main Text 1198 –The number of Tables: one –Short running title “Cytopenia and Proteinuria in Imerslund-Gräsbeck Syndrome” – Keywords Imerslund-Gräsbeck syndrome (IGS), Macrocytic anemia, Vitamin B12 malabsorption, Proteinuria. -Abbreviations key [1]¿p1 [1]¿m1 CUBN Cubilin AMN Amnioless Cbl Vitamin B12, cobalamin IF Intrinsic factor IM OHcbl Intramuscular hydroxocobalamin MMAU Urinary Methylmalonic Acid NA not available Hb hemoglobin LDH lactate dehydrogenase MCV mean corpuscular volume NR normal range Abstract Imerslund-Gräsbeck syndrome (IGS) is a rare autosomal recessive disorder caused by pathogenic variants in CUBN or AMN genes, leading to selective vitamin B12 malabsorption. Symptoms include failure to thrive, cytopenia, and proteinuria. This study aims to analyze clinical, biochemical, and molecular genetic profiles of 4 patients diagnosed with IGS at a single center in Argentina. Macrocytic anemia and neutropenia were present in three cases, two with thrombocytopenia. All had proteinuria. A novel CUBN : c.2450G>A; p.(Cys817Tyr) variant was found in three patients, while the fourth carried AMN : c.1006+34_1007-31del in homozygous state. Early and lifelong parenteral vitamin B12 ensures an excellent prognosis. Keywords: Imerslund-Gräsbeck syndrome (IGS), Macrocytic anemia, Vitamin B12 malabsorption, Proteinuria. Introduction Vitamin B12 (cobalamin, Cbl) is the most structurally complex vitamin and is indispensable for critical physiological processes, including growth, reproduction, DNA synthesis, methylation, and mitochondrial metabolism [1]. Imerslund-Gräsbeck syndrome (IGS, OMIM 261100/618882) is a rare autosomal recessive disorder characterized by selective Cbl malabsorption and proteinuria, with an estimated prevalence of 6 per 1,000,000 individuals. Untreated, IGS manifests with non-specific symptoms, including failure to thrive, pallor, fatigue, recurrent infections, and neurological deficits. The hallmark megaloblastic anemia results from impaired hematopoietic cell division due to defective DNA replication, affecting other rapidly dividing cells [2]. IGS results from homozygous or compound heterozygous mutations in CUBN (10p12.1) or AMN (14q32), encoding cubilin and amnionless, which form the cubilin-amnionless (cubam) complex. This receptor is essential for intestinal absorption of the Cbl-intrinsic factor (IF) complex and renal protein reabsorption [3,4]. IGS typically appears between four months and five years, distinguishing it from transcobalamin II deficiency, which presents at birth [5]. Diagnosis relies on clinical features and biochemical evidence of vitamin B12 malabsorption unresponsive to intrinsic factor supplementation. Biochemical assessment shows reduced serum cobalamin with elevated homocysteine and methylmalonic acid. Molecular analysis confirming AMN or CUBN mutations provides definitive diagnosis [2-4]. Lifelong management involves parenteral vitamin B12 supplementation, predominantly via intramuscular injections, which effectively address clinical and biochemical abnormalities while mitigating complications, despite persistent proteinuria. This study presents an analysis of the clinical, biochemical, and molecular characteristics of four Argentine patients diagnosed with IGS at our center. Case Descriptions We describe four unrelated IGS patients in our center from September 2008 to January 2025. One of them was from a consanguineous family. Three of these patients were males and one a female ( Table 1 ). The median age of symptom onset was 21.5 months (16-72m). The median age at diagnosis was 112.5 months (84-192m), with a median diagnostic delay of 93.5 months. Laboratory tests revealed megaloblastic anemia in 3/4 patients. One patient had normocytic anemia, with normal iron deposits. Three patients presented with mild to moderate neutropenia, and one with mild thrombocytopenia. All patients had a previous history of anemia and vitamin B12 deficiency, two of them having transfusion requirements. Consultation in all patients was because of fatigue, pallor, failure to thrive and gastrointestinal manifestations including diarrhea and vomiting. All patients had hemolytic parameters, with a median lactate dehydrogenase of 4295 UI/L (1919-6406). Severe vitamin B12 deficiency (<125 pg/mL; normal range: 180–914 pg/mL) and proteinuria with a median of 0.78 g/L (0.4-1.2) (normal range: <0.12 g/L) were observed in all patients. Renal ultrasound showed no structural anomalies of the kidneys or urinary tract. Gastrointestinal symptoms (diarrhea, vomiting, failure to thrive) were noted in all patients, with mucosal lesions (sore tongue, oral ulcers) in one patient. Dermatological findings were observed in three patients, with non-specific findings (hyperpigmentation, hypopigmentation, eczema). Neurological symptoms were observed in only one patient (P1), who presented with axial hypotonia and motor skill regression, which fully resolved after treatment initiation. Currently, at 9 years of age, the patient exhibits mild intellectual disability (IQ 58). Genetic screening for AMN and CUBN mutations was performed, identifying a novel missense variant, c.2450G>A; p.(Cys817Tyr) in the CUBN gene in homozygous state in three patients, and an homozygous intronic deletion, c.1006+34_1007-31del, in the AMN gene in one patient. DNA samples from the parents of two patients with the CUBN variant were available, confirming them as carriers. Parenteral vitamin B12 supplementation was initiated, leading to normalization of vitamin B12 levels, resolution of anemia, neutropenia and thrombocytopenia, and improvement in associated symptoms. Discussion Imerslund-Gräsbeck syndrome (IGS) is an autosomal recessive disorder caused by mutations in the CUBN or AMN genes, which encode the cubam receptor subunits: cubilin and amnionless. Cubilin is a multifunctional protein involved in the uptake of vitamin B12 by binding to the IF-vitamin B12 complex. Additionally, these proteins play a role in renal tubular reabsorption of proteins. Symptoms of IGS typically appear between 1 and 5 years of age, once fetal vitamin B12 stores are depleted, as seen in all our patients. However, although vitamin B12 deficiency was diagnosed in all at initial consultation, deficiency was interpreted as secondary to malnutrition, and the diagnostic delay of IGS had a median of 7,7 years. As described by Grasbeck in his review of IGS [3], megaloblastic anemia is a typical manifestation in IGS. It was seen in 3/4 of our patients. Neurocognitive symptoms associated with vitamin B12 deficiency are linked to impaired myelination and neurotoxicity from elevated homocysteine and methylmalonic acid. These symptoms are usually mild and non-specific, including developmental delays, learning difficulties, or psychological disturbances [6]. Frequency of this manifestation was 16% according to the review by Kingma et al [4], similar to that described in our series, with only one patient affected. The pathophysiology of proteinuria in IGS is not fully understood but is likely linked to defects in cubilin ( CUBN) and amnionless ( AMN ), which are essential for the reabsorption of filtered plasma proteins in the proximal renal tubules. Their absence leads to impaired protein reabsorption and proteinuria. Despite persistent proteinuria in IGS, renal function does not appear to progress in patients treated with vitamin B12 supplementation, though the exact reasons remain unclear. Although not always present, Kingma et al describe a high incidence with 89% of patients presenting with proteinuria. When present it is highly suggestive of IGS. In our case series, all patients presented with proteinuria, with no affection of renal function [2-4]. A definitive diagnosis of IGS requires molecular analysis of the CUBN and AMN genes. Over 100 different homozygous and compound heterozygous variants have been described in both genes. We found a novel homozygous variant, c.2450G>A in the CUBN gene in three patients. This rare variant, located in exon 19, leads to the substitution of cysteine with tyrosine at codon 817 of the cubilin protein (p.Tyr121His). The missense variant was classified as of unknown significance (VUS) according to the ACMG guidelines[7]. Variants affecting the amino-terminal third of cubilin (exons 1-20) have been associated with both Cbl deficiency and proteinuria occurring simultaneously [8], as in the three patients in this study. On the other hand, c.1006+34_1007-31del in the AMN gene, also known as c.1006+34_48del15bp, has been previously described in patients with IGS, and it has been proved that the intronic deletion of 15 bp causes the complete loss of exon 9 in the resulting mRNA [8]. Initially, vitamin B12 deficiency in IGS patients was treated with monthly intramuscular injections due to malabsorption, and oral treatment was considered ineffective. However, high doses of oral vitamin B12 can be effective, as approximately 1% is absorbed through passive diffusion, even in IGS patients. With sufficiently high doses, this limited absorption can correct the deficiency. Lifelong vitamin B12 supplementation is required for all IGS patients [4,9]. Three of our patients receive intramuscular supplementation and one oral supplementation, all achieving complete remission of hematological manifestations and normal homocysteine, methylmalonic acid and vitamin B12 levels. The presence of macrocytic anemia or the coexistence of anemia with other cytopenias should prompt suspicion of vitamin B12 deficiency. Similarly, the presence of proteinuria or a history of similar symptoms should always heighten awareness of IGS as a differential diagnosis. Early initiation of vitamin B12 therapy can prevent potential complications arising from this condition. Acknowledgments: The authors would like to thank the multidisciplinary team at the Inborn Errors of Metabolism, Hematology and Molecular Biology Laboratory in Garrahan Hospital with the diagnosis and assistance of the patients. Conflicts of Interest: The authors declare no conflicts of interest. References 1. Bindi V, Eiroa H, Díaz L. Trastornos adquiridos y congénitos de la deficiencia de vitamina B12 en niños. Salud(i)Ciencia. 2023;25:217-223. 2. Watkins D, Rosenblatt DS, Fowler B. Disorders of cobalamin and folate transport and metabolism. In: Saudubray J-M, Baumgartner MR, Walter JH, eds. Inborn Metabolic Diseases. 6th ed. Springer; 2016:385-399. 3. Gräsbeck R. Imerslund-Gräsbeck syndrome (selective vitamin B12 malabsorption with proteinuria). Orphanet J Rare Dis. 2006;1(1):1-6. 4. Kingma SD, Neven J, Bael A, et al. Imerslund-Gräsbeck syndrome: a comprehensive review of reported cases. Orphanet J Rare Dis. 2023;18:291. doi:10.1186/s13023-023-02889-x 5. Bindi V, Crespo C, Lochner N, et al. New data supporting that early diagnosis and treatment are possible and necessary in intracellular cobalamin depletion: the case of transcobalamin II deficiency. J Pediatr Endocrinol Metab. 2024;37(10):930. Published September 11, 2024. doi:10.1515/jpem-2024-040. 6. Kalayci Ö, Cetin M, Kirel B, Özdirim E, Yetgin S, Aysun S, Gürgey A. Neurological findings of vitamin B12 deficiency: presentation of seven cases. Turk J Pediatr. 1996;38:67-72. 7. Richards S, Aziz N, Bale S, et al. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med. 2015;17(5):405-424. doi:10.1038/gim.2015.30 8. Tanner SM, Sturm AC, Baack EC, et al. Inherited cobalamin malabsorption: mutations in three genes reveal functional and ethnic patterns. Orphanet J Rare Dis. 2012;7:56. doi:10.1186/1750-1172-7-56 9. Broch H, Imerslund O, Monn E, Hovig T, Seip M. Imerslund-Gräsbeck anemia: a long-term follow-up study. Acta Paediatr. 1984;73:248-253. Supplementary Material File (table 1 igs 2025 final.docx) Download 20.90 KB Information & Authors Information Version history V1 Version 1 25 March 2025 Copyright This work is licensed under a Non Exclusive No Reuse License. Keywords all anemias homocysteine molecular diagnosis & therapy neutropenia thrombocytopenia Authors Affiliations Verónica Bindi 0000-0002-5872-199X [email protected] Hospital de Pediatria Prof Dr Juan P Garrahan View all articles by this author Crespo Carolina Hospital de Pediatria Prof Dr Juan P Garrahan View all articles by this author Gravina L. Hospital de Pediatria Prof Dr Juan P Garrahan View all articles by this author Hernán Eiroa Hospital de Pediatria Prof Dr Juan P Garrahan View all articles by this author Rossetti E. Hospital de Pediatria Prof Dr Juan P Garrahan Servicio de Hematologia y Oncologia View all articles by this author Metrics & Citations Metrics Article Usage 276 views 112 downloads .FvxKWukQNSOunydq8rnd { width: 100px; } Citations Download citation Verónica Bindi, Crespo Carolina, Gravina L., et al. “Cytopenias and proteinuria as key diagnostic features of Imerslund-Gräsbeck Syndrome: a case series of four Argentine patients”. Authorea . 25 March 2025. DOI: https://doi.org/10.22541/au.174291335.58828505/v1 If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. Simply select your manager software from the list below and click Download. For more information or tips please see 'Downloading to a citation manager' in the Help menu . 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