Diagnostic Strategies

Diagnostic Testing

Biochemical

Routine laboratory profiles including standard blood biochemistry, plasma lipids, urinalysis and cerebrospinal fluid metabolites are generally normal in NPC except in patients with hypersplenism or cholestatic jaundice. A definitive diagnosis of NPC requires demonstration of abnormal intracellular cholesterol homeostasis in cultures of fibroblasts taken from the patient.³ In NPC-positive cultures, these cells show a reduced ability to esterify cholesterol after loading with exogenously derived LDL-cholesterol. Only certain specialist laboratories perform specific biochemical testing for NPC. These can be located/contacted through various NPC resources via specialist care centres.²

Filipin staining is employed to identify any fluorescence concentrated around the nucleus, which indicates the accumulation of unesterified cholesterol in perinuclear lysosomes. Most patients with a ‘classic biochemical phenotype’ of NPC have zero or very low esterification levels with a classic, intense dotted filipin staining pattern. However, approximately 15% of individuals display the ‘variant biochemical phenotype’, showing intermediate levels of cholesterol esterification.⁴ Filipin staining is more sensitive in detecting such patients, although the variant phenotype is also associated with a less distinctive fibroblast staining pattern.^1,2 Alternative methods exist to characterise such patients more precisely,⁵ but are not yet routinely available.

Studies suggest that measurements of plasma chitotriosidase activities could be useful as a marker for lysosomal storage diseases including Gaucher disease and Niemann–Pick disease.^6–8 Plasma chitotriosidase is markedly elevated in Gaucher disease, has consistently been shown to decrease with therapy,⁹ and is recommended as a screening procedure in children in some countries.^7,8Laboratory studies suggest that the same might also apply in NPC,⁸ although patients typically show more biochemical variability in this disease.¹⁰

Histologic

Prior to the recognition of the primary biochemical defect in NPC³ and the development of direct biochemical testing for abnormal intracellular cholesterol homeostasis, histopathologic analysis of tissue biopsies and/ or tissue lipid analysis was essential to confirm a diagnosis of NPC. However, such invasive tests are now only required in rare cases.

Histologic analyses include examination of bone marrow, spleen and liver for the presence and extent of foam cells (lipid-laden macrophages) as well as the presence of sea-blue histiocytes, which may be seen in bone marrow. However, these features are considered non-specific, as failure to detect them does not rule out a diagnosis of NPC.¹Electron microscopic analyses of skin, rectal neurones, liver or brain can also be employed, and can show polymorphous cytoplasmic bodies.^2,11

Genetic

Molecular diagnosis is not considered a suitable tool for primary diagnosis,¹ but is of use in prenatal diagnosis and the identification of heterozygotes in probands’ families – especially as regards family planning, and in the confirmation of diagnosis in individuals with a variant biochemical phenotype. The majority of individuals with NPC harbour mutations in the NPC1 gene while, it is assumed, the remaining individuals with NPC have mutations in the NPC2 gene.

Imaging

Magnetic resonance imaging (MRI) or computed tomography (CT) scans of the brain are usually normal until the late stages of NPC, and imaging is considered non-specific.¹ However, cerebral atrophy and in particular, marked atrophy of the superior/anterior cerebellar vermis may be seen, usually in patients with slowly progressive disease. Other abnormalities can include thinning of the corpus callosum and increased signal in the periatrial white matter, reflecting secondary demyelination. Proton magnetic resonance spectroscopy (H-MRSI) may be a more sensitive imaging technique in NPC than standard MRI.¹²

Figure 8. MRI and H-MRSI in control and NPC brain (reproduced with permission from Tedeschi et al.¹²)
Proton

Click image for larger version.

Proton magnetic resonance spectroscopic imaging (H-MRSI) and corresponding MRI scans from (A) a control and (B) a patient with NPC. H-MRSI permits the simultaneous measurement of N-acetyl-aspartate (NA), choline-containing compounds (Cho), creatine plus phosphocreatine (Cre) and lactate (Lac) signal intensities, and identification of regions of interest. In patients with NPC, NA/Cre was significantly reduced in the frontal and parietal cortices, centrum semiovale and caudate nucleus; Cho/Cre was increased in the frontal cortex and centrum semiovale. These changes were correlated with clinical staging score.¹²

References
1. Patterson MC, Vanier MT, Suzuki K et al. Niemann–Pick disease, type C: a lipid trafficking disorder. In: Scriver CR, Beaudet AL, Sly WS, Valle D, Childs B, Vogelstein B (eds) The Metabolic and Molecular Bases of Inherited Disease, 8th ed, 2001. New York: McGraw-Hill, Ch 145, pp 3611–33.
2. Patterson MC. Niemann–Pick disease Type C. Gene Reviews 2007a (updated 9 July). Accessible at: www.geneclinics.org. Accessed 28^th May 2009.
3. Pentchev PG, Comly ME, Kruth HS et al. A defect in cholesterol esterification in Niemann–Pick disease (type C) patients. Proc Natl Acad Sci USA 1985;82:8247–51.
4. Vanier MT, Rodriguez-Lafrasse C, Rousson R et al. Type C Niemann–Pick disease: biochemical aspects and phenotypic heterogeneity. Dev Neurosci 1991;13:307–14.
5. Sun X, Marks DL, Park WD et al. Niemann–Pick C variant detection by altered sphingolipid trafficking and correlation with mutations within a specifi c domain of NPC1. Am J Hum Genet 2001;68:1361–72.
6. Hollak CE, van Weely S, van Oers MH, Aerts JM. Marked elevation of plasma chitotriosidase activity. A novel hallmark of Gaucher disease. J Clin Invest 1994;93:1288–92.
7. Wajner A, Michelin K, Burin MG et al. Biochemical characterization of chitotriosidase enzyme: comparison between normal individuals and patients with Gaucher and with Niemann–Pick diseases. Clin Biochem 2004;37:893–7.
8. Ries M, Schaefer E, Lührs T et al. Critical assessment of chitotriosidase analysis in the rational laboratory diagnosis of children with Gaucher disease and Niemann–Pick disease type A/B and C. J Inherit Metab Dis 2006;29:647–52.
9. Cox T, Lachmann R, Hollak C et al. Novel oral treatment of Gaucher’s disease with N-butyldeoxynojirimycin (OGT 918) to decrease substrate biosynthesis. Lancet 2000;355:1481–5.
10. Chien YH, Lee C, Tsai LK et al. Treatment of Niemann–Pick disease type C in two children with miglustat: Initial responses and maintenance of effects over 1 year. J Inherit Metab Dis 2007, June 21; [Epub ahead of print].
11. Boustany RN, Kaye E, Alroy J. Ultrastructural findings in skin from patients with Niemann–Pick disease, type C. Pediatr Neurol 1990;6:177–83.
12. Tedeschi G, Bonavita S, Barton NW et al. Proton magnetic resonance spectroscopic imaging in the clinical evaluation of patients with Niemann–Pick type C disease. J Neurol Neurosurg Psychiatry 1998;65:72–9.

Niemann-Pick Knowledge Centre
(Type C)