Abstract
High consanguinity rates, poor access to accurate diagnostic tests, and costly therapies are the main causes of increased burden of lysosomal storage disorders (LSDs) in developing countries. Therefore, there is a major unmet need for accurate and economical diagnostic tests to facilitate diagnosis and consideration of therapies before irreversible complications occur. In cross-country study, we utilized dried blood spots (DBS) of 1,033 patients clinically suspected to harbor LSDs for enzymatic diagnosis using modified fluorometric assays from March 2013 through May 2015. Results were validated by demonstrating reproducibility, testing in different sample types (leukocytes/plasma/skin fibroblast), mutation study, or measuring specific biomarkers. Thirty percent (307/1,033) were confirmed to have one of the LSDs tested. Reference intervals established unambiguously identified affected patients. Correlation of DBS results with other biological samples (nā=ā172) and mutation studies (nā=ā74) demonstrated 100% concordance in Gaucher, Fabry, Tay Sachs, Sandhoff, Niemann-Pick, GM1, Neuronal ceroid lipofuscinosis (NCL), Fucosidosis, Mannosidosis, Mucopolysaccharidosis (MPS) II, IIIb, IVa, VI, VII, and I-Cell diseases, and 91.4% and 88% concordance in Pompe and MPS-I, respectively. Gaucher and Pompe are the most common LSDs in India and Pakistan, followed by MPS-I in both India and Sri Lanka. Study demonstrates utility of DBS for reliable diagnosis of LSDs. Diagnostic accuracy (97.6%) confirms veracity of enzyme assays. Adoption of DBS will overcome significant hurdles in blood sample transportation from remote regions. DBS enzymatic and molecular diagnosis should become the standard of care for LSDs to make timely diagnosis, develop personalized treatment/monitoring plan, and facilitate genetic counseling.
Competing interests: None declared
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Abbreviations
- CK:
-
Creatine phosphokinase
- DBS:
-
Dried blood spot
- EQAS:
-
External Quality Assurance Scheme
- ERNDIM:
-
European Research Network for evaluation and improvement of screening, diagnosis, and treatment of inborn errors of metabolism
- GAGs:
-
Glycosaminoglycans
- LSDs:
-
Lysosomal storage disorders
- MPS:
-
Mucopolysaccharidosis
References
Bostick WD, Dlnsmore SR, Mrochek JE, Waalkes TP (1978) Separation and analysis of aryl sulfatase isoenzymes in body fluids of man. Clin Chem 24(8):1305ā1316
Burin M, Dutra-Filho C, Brum J, Mauricio T, Amorim M, Giugliani R (2000) Effect of collection, transport, processing and storage of blood specimens on the activity of lysosomal enzymes in plasma and leukocytes. Braz J Med Biol Res 33:1003ā1013
Camelier MV, Burin MG, De Mari J, Vieira TA, Marasca G, Giugliani R (2011) Practical and reliable enzyme test for the detection of Mucopolysaccharidosis IVA (Morquio Syndrome type A) in dried blood samples. Clin Chim Acta 412:1805ā1808
Ceci R, de Francesco PN, Mucci JM, Cancelarich LN, Fossati CA, Rozenfeld PA (2011) Reliability of enzyme assays in dried blood spots for diagnosis of 4 lysosomal storage disorders. Adv Biol Chem 1:58ā64
Chace DH, Kalas TA, Naylor EW (2003) Use of tandem mass spectrometry for multianalyte screening of dried blood specimens from newborns. Clin Chem 49:1797ā1817
Chamoles NA, Blanco M, Gaggioli D (2001) Fabry disease: enzymatic diagnosis in dried blood spots on filter paper. Clin Chim Acta 308:195ā196
Chamoles NA, Blanco M, Gaggioli D, Casentini C (2002a) Gaucher and Niemann-Pick diseases- enzymatic diagnosis in dried blood spots on filter paper: retrospective diagnosis in newborn- screening cards. Clin Chim Acta 317:191ā197
Chamoles NA, Blanco M, Gaggioli D, Casentini C (2002b) Tay-Sachs and Sandhoff diseases: enzymatic diagnosis in dried blood spots on filter paper: retrospective diagnosis in newborn-screening cards. Clin Chim Acta 318(1ā2):133ā137
Civallero G, Michelin K, de Mari J et al (2006) Twelve different enzyme assays on dried-blood filter paper samples for detection of patients with selected inherited lysosomal storage diseases. Clin Chim Acta 372:98ā102
Cobos PN, Steqlich C, Santer R, Lukacs Z, Gal A (2015) Dried blood spots allow targeted screening to diagnose mucopolysaccharidosis and mucolipidosis. JIMD Rep 15:123ā132
Coelho J, Guigliani R (2000) Fibroblasts of skin fragments as a tool for the investigation of genetics diseases: technical recommendations. Genet Mol Biol 23:269ā271
Filocamo M, Morrone A (2011) Lysosomal storage disorders: molecular basis and laboratory testing. Hum Genomics 5:156ā169
Hollak CE, Van Weely S, van Oers MH, Aerts JM (1994) Marked elevation of plasma chitotriosidase activity. A novel hallmark of Gaucher disease. J Clin Invest 93:1288ā1292
Hopwood JJ, Muller V, Smithson A, Baggett N (1979) A fluorometric assay using 4 methylumbelliferyl alpha-L-iduronide for the estimation of alpha-L-iduronidase activity and the detection of Hurler & Scheie syndromes. Clin Chim Acta 92:257ā265
Kaminsky P, Lidove O (2014) Current therapeutic strategies in lysosomal disorders. Presse Med 43:1174ā1184
Kelly S (1977) Biochemical methods in medical genetics, vol 1008, 18th edn, Harvard American lecture series. Thomas, Springfield
Lukacs Z, Santavuori P, Keil A, Steinfeld R, Kohlschutter A (2003) Rapid and simple assay for the determination of tripeptidyl peptidase and palmitoyl protein thioesterase activities in dried blood spots. Clin Chem 49:509ā511
Marsh J, Fensom AH (1985) 4 Methylumbelliferyl Ī±-N-acetyl glucosaminidase activity for diagnosis of Sanfilippo B disease. Clin Genet 27:258ā262
Mistri M, Oza N, Sheth F, Sheth J (2014) Prenatal diagnosis of lysosomal storage disorders: our experience in 120 cases. Mol Cytogenet 7(Suppl 1):P126
Muller KB, Rodrigues MD, Pereira VG, Martins AM, DāAlmeida V (2010) Reference values for lysosomal enzymes activities using dried blood spots samples- a Brazilian experience. Diagn Pathol 5:65ā72
Sheth J, Mistri M, Sheth F et al (2014) Burden of lysosomal storage disorders in India: experience of 387 affected children from a single diagnostic facility. JIMD Rep 12:51ā63
van Diggelen OP, Keulemans JL, Winchester B et al (1999) A rapid fluorogenic palmitoyl-protein thioesterase assay: pre- and postnatal diagnosis of INCL. Mol Genet Metab 66:240ā244
van Diggelen OP, Voznyi YV, Keulemans JLM et al (2005) A new fluorometric enzyme assay for the diagnosis of NiemannāPick A/B, with specificity of natural sphingomyelinase substrate. J Inherit Metab Dis 28:733ā741
Verma PK, Ranganath P, Dalal AB, Phadke SR (2012) Spectrum of lysosomal storage disorders at a medical genetics center in northern India. Indian Pediatr 49:799ā804
Verma J, Thomas DC, Sharma S et al (2015) Inherited metabolic disorders: quality management for laboratory diagnosis. Clin Chim Acta 447:1ā7
Voznyi YV, Keulemans JLM, van Diggelen OP (2001) A fluorimetric enzyme assay for the diagnosis of MPS II (Hunter disease). J Inherit Metab Dis 24:675ā680
Willey AM, Carter TP, Kally S et al (1982) Clinical genetics: problems in diagnoses and counseling. Harcourt Brace/Jovanovich, New York/London, Page 41
Winchester B, Bali D, Bodamer OA et al (2008) Methods for a prompt and reliable laboratory diagnosis of Pompe disease: report from an international consensus meeting. Mol Genet Metab 93:275ā281
Acknowledgement
We thank all patients and normal subjects whose blood was used as a control/reference material to generate the data for this study. We extend our thanks to Genzyme (India) who has chosen our center as a referral laboratory for lysosomal enzymes testing. We express our gratitude to doctors from India and neighboring countries for referring patients to our genetic center. We appreciate Genzyme (India), a Sanofi company for extending support to patients for testing and treatment. We also thank Chairman and Director Administration, Sir Ganga Ram Hospital, for their cooperation in setting up genetic testing facility.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Additional information
Communicated by: Gajja Salomons
Appendices
Take Home Message
Dried blood spots (DBS) have potential to substitute the established liquid bio-matrices (plasma/leukocytes/cultured fibroblasts) for reliable, accurate, and timely diagnosis of lysosomal storage disorders (LSDs) as demonstrated by 100% sensitivity and 95.2% specificity obtained after biochemical (using liquid biological samples) and molecular confirmation of the DBS-based enzymatic diagnoses to facilitate prognostication and genetic counseling and its implementation will also overcome challenges in cross-country transportation of conventional biological samples.
Compliance with Ethics Guidelines
Authorsā contribution
Concept, design, and method standardization: Jyotsna Verma
Experimentation/Acquisition of data/analysis/interpretation: Jyotsna Verma, Divya C. Thomas, Sandeepika Sharma, Pramod K. Mistry, David C. Kasper
Patientsā management/clinical assessment: I. C. Verma, Sunita Bijarnia, Ratna D. Puri
Drafting article or revising or reviewing it critically for important content: Jyotsna Verma, Divya C. Thomas, Pramod K. Mistry, David C. Kasper, I. C. Verma, Ratna D. Puri
Final approval for publication: All authors
Conflict of Interest
Jyotsna Verma and David C. Kasper received travel grant from Genzyme, a Sanofi company. Same company also supported I. C. Verma for organization of continuing medical education (CME). Divya C. Thomas, Sandeepika Sharma, Ratna D. Puri, Sunita Bijarnia-Mahay, and Pramod K. Mistry declare that they have no conflict of interest.
Informed Consent
All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2000 (5) of the World Medical Association. Informed consent was obtained from all patients/normal subjects for being included in the study.
Funding
Patients for enzymatic testing of four disorders (Pompe, Gaucher, Fabry, and MPS1) were sponsored by Genzyme (India), a Sanofi company via doctors from India and other neighboring countries to help in providing treatment on compassionate ground under their āIndia Compassionate Access Program.ā Genzyme has under no circumstance extended funds to us under any project head/grant number nor has it provided us with any financial support to facilitate the functioning of our laboratory so as to influence the results of our study.
Rights and permissions
Copyright information
Ā© 2016 SSIEM and Springer-Verlag Berlin Heidelberg
About this chapter
Cite this chapter
Verma, J. et al. (2016). Inherited Metabolic Disorders: Efficacy of Enzyme Assays on Dried Blood Spots for the Diagnosis of Lysosomal Storage Disorders. In: Morava, E., Baumgartner, M., Patterson, M., Rahman, S., Zschocke, J., Peters, V. (eds) JIMD Reports, Volume 31. JIMD Reports, vol 31. Springer, Berlin, Heidelberg. https://doi.org/10.1007/8904_2016_548
Download citation
DOI: https://doi.org/10.1007/8904_2016_548
Received:
Revised:
Accepted:
Published:
Publisher Name: Springer, Berlin, Heidelberg
Print ISBN: 978-3-662-54118-0
Online ISBN: 978-3-662-54119-7
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)