Bone Marrow Transplantation, (1999) 24, 103–107
 1999 Stockton Press All rights reserved 0268–3369/99 $12.00
http://www.stockton-press.co.uk/bmt
Case report
Niemann–Pick disease type C (a cellular cholesterol lipidosis) treated
by bone marrow transplantation
Y-S Hsu1, W-L Hwu1, S-F Huang2, M-Y Lu1, R-L Chen1, D-T Lin1, SSF Peng3 and K-H Lin1
Departments of 1Pediatrics, 2Pathology, 3Medical Image, National Taiwan University Hospital, National Taiwan University College
of Medicine, Taiwan
Summary:
Bone marrow transplantation (BMT) has been used for
a wide variety of lysosomal storage diseases with
encouraging results. We report a 3-year 5-month-old
girl with Niemann–Pick type C disease (NPC) who
received an allogeneic BMT. The patient presented with
repeated lower respiratory tract infections, hepatosplenomegaly, failure to thrive, and developmental delay.
Chest computed tomography (CT) revealed diffuse
interstitial lung infiltration. Bone marrow and liver
biopsies revealed abundant lipid-filled foamy macrophages. Skin fibroblast sphingomyelinase assay revealed
partial deficiency. The ability of her skin fibroblasts to
esterify cholesterol was very low, and the cells stained
brightly for free cholesterol. She received BMT from a
healthy HLA-identical male sibling donor at the age of
2 year 6 months. Full engraftment was evidenced by
repeated bone marrow sex chromosome studies.
Regression of the hepatosplenomegaly, markedly
reduced foamy macrophage infiltration in bone marrow, and decreased interstitial lung infiltration was
noted 6 months after BMT. Her neurological status,
however, deteriorated. Follow-up magnetic resonance
image (MRI) revealed progressive, diffuse brain atrophy. We conclude that resolution occurred in the liver,
spleen, bone marrow and lung following successful
engraftment. Such a response is remarkable since the
underlying problem involves a membrane receptor for
cholesterol. This positive response might be due to
replacement of the monocyte–phagocytic system or it
may imply the existence of cross-correction in the NPC
membrane receptor defect by BMT approach. Since
BMT did not halt the neurological deterioraton, it is
unlikely to be an adequate treatment for NPC.
Keywords: Niemann–Pick type C disease; lysosomal
storage disease; BMT
Correspondence: Dr K-H Lin, Department of Pediatrics, National Taiwan
University Hospital, No. 7, Chung-Shan South Rd, Taipei, 100, Taiwan,
ROC
Received 23 September 1998; accepted 25 January 1999
Niemann–Pick type C disease (NPC) is an autosomal
recessive storage lipidosis in which fibroblasts exhibit a
unique biochemical lesion characterized by delayed and
impaired homeostatic responses to exogenous low-density
lipoprotein (LDL) cholesterol loading, with lysosomal
accumulations of unesterified cholesterol. The diagnosis is
established by demonstration of both impaired cholesterol
esterification in cultured fibroblasts and a characteristic pattern of intense perinuclear fluorescence after filipin staining.1–2 NPC is biochemically entirely different and phenotypically distinct from the sphingomyelin lipidoses (type
A and type B Niemann–Pick disease3), with which it has
traditionally been grouped. The cardinal pathologic features
of all forms of NPC are visceral foamy cells and neuronal
storage. Foamy cells and sea-blue histiocytes containing
accumulations of sphingomyelin and cholesterol are conspicuous in the spleen, tonsils, lymph nodes, liver and lung
and usually cause visceromegaly. Most patients with NPC
have progressive neurologic disease, although hepatic damage is prominent in certain cases.4 Both linkage and complementation analyses have shown that at least two separate
genes, NPC1 (major locus) and NPC2, induce identical
clinical and biochemical phenotypes.5 The NPC1 gene on
chromosome 18q11 was recently cloned by positional cloning and the NPC1 protein is thought to be an integral membrane protein.6 Specific treatment that will change the natural history of the disease is not yet available. Bone marrow
transplantation (BMT) and combined liver and bone marrow transplantation in the C57BL/KsJ murine model of
NPC partially reversed tissue storage of cholesterol and
sphingomyelin, but did not influence the continued neurologic deterioration. There is no previous report of BMT for
human NPC. We report a case of NPC who successfully
engrafted after allogeneic BMT.
Case report
In September 1996, a 14-month-old girl was referred for
investigation of repeated lower respiratory tract infections,
developmental delay, and persistent hepatosplenomegaly.
She was born by normal spontaneous vaginal delivery after
39 weeks of gestation with a birth weight 2170 g. Intrauterine growth retardation was noted during pregnancy. No specific problems occurred in the neonatal period except for
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104
transient hyperbilirubinemia. At 3 months of age, acute
bronchiolitis was noted. Jaundice recurred and hepatosplenomegaly was noted. Liver function tests showed: AST
190 U/l, ALT 104 U/l, total bilirubin 4.9 mg/dl, direct bilirubin 2.7 mg/dl, and alkaline phosphatase 842 U/l. Work
up for congenital infection, hepatitis, cytomegalovirus, sepsis and biliary tract disease yielded no positive results. The
jaundice subsided 2 months later without any treatment.
However, persistent hepatosplenomegaly, delayed developmental milestones and four episodes of acute bronchiolitis
without sign of respiratory distress occurred during the first
year of life. Initially, physical examination in our hospital
revealed hepatosplenomegaly and slightly coarse breathing
sounds. Her liver was palpable 4 cm below the right costal
margin and spleen 8 cm below the left costal margin.
Neurologically, she had mild mental retardation, dysarthria,
ataxia, dysmetria, bilateral lower extremity spasticity, and
vertical supranuclear ophthalmoplegia. The Babinski sign
was positive bilaterally and deep tendon reflex of all
extremities was also increased. Laboratory tests revealed:
AST 114 U/l, ALT 16 U/l, total bilirubin 0.4 mg/dl, direct
bilirubin 0.1 mg/dl, and alkaline phosphatase 374 U/l. The
cerebrospinal fluid was acellular with normal protein and
glucose levels and without oligoclonal bands. Hematopoietic parameters were within normal limits. Plain chest radiography and high-resolution chest computed tomography
(CT) revealed diffuse interstitial lung infiltration
(Figure 1a). Brain magnetic resonance image (MRI)
revealed normal myelination without obvious brain atrophy
(Figure 2a). However, electroencephalography (EEG)
showed diffuse cortical dysfunction. Bone marrow and liver
biopsies revealed abundant foamy lipid-filled macrophages
(Figure 3a), and the foamy cells were filled with secondary
lysosomes containing membranous cytoplasmic bodies of
variable sizes resembling concentric lamellated myelin
figures on electron microscopic examination (Figure 4).
The ability of cultured skin fibroblasts to esterify cholesterol was very low: 3 ⫾ 8 (pmoles cholesterol ester
formed/mg protein, normal value: 3432 ⫾ 1729, average
Niemann–Pick type C value: 90 ⫾ 140), and the cells
stained brightly for free cholesterol using the filipin stain,
performed by Dr Adele Cooney at the Developmental
Metabolic Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda,
MA, and also by Dr David A Wenger at the Division of
Medical Genetics, Jefferson Medical College, Philadelphia,
Pensylvania, USA. Sphingomyelinase assay of the cells
revealed partial deficiency (40% of normal level). Hence,
NPC was confirmed.
Allogeneic BMT with her HLA-identical sibling donor
(5-year-old brother, donor blood type O vs recipient blood
type B) was performed at 2 years 6 months of age. Evaluation 1 month before transplant showed her body weight
to be below the 3rd percentile and both body height and
head girth were around 3rd–10th percentile. Liver and
spleen were 3 cm and 8 cm below the costal margins,
respectively. Developmental regression was noted. There
had been no speech development during the past year and
even the ability and frequency of voicing repetitive consonant sounds (mama, dada) decreased. She had been able to
sit without support and to crawl at 14 months old, but both
Figure 1 High-resolution chest CT. (a) Before BMT, widespread, prominent peribronchovascular, interlobular, and centrilobular interstitial thickening are evident in both lungs. Also, segmental consolidation at bilateral
posterior aspects of the lower lobes is noted, more on the left side.(b) Ten
months after BMT, as compared with (a), the interstitial changes have
almost cleared up. Air-trapping in the posterior areas of bilateral lung
fields is seen due to acute bronchiolitis.
skills were lost before BMT. Laboratory tests revealed AST
97 U/l, ALT 21 U/l, total bilirubin 0.6 mg/dl, direct bilirubin 0.2 mg/dl, alkaline phosphatase 643 U/l, and a normal
hemogram. The conditioning regimen consisted of busulfan
4 mg/kg p.o. in divided doses daily for 4 days (total dose
16 mg/kg), followed by cyclophosphamide 60 mg once
daily i.v. for 2 days (total dose 120 mg/kg). For prophylaxis
of graft-versus-host disease (GVHD), she received cyclosporine 3 mg/kg i.v. daily starting one day before BMT
(day −1), and methotrexate (MTX) 15 mg/m2 on day +1,
and MTX 10 mg/m2 on days +3, +6 and +11. A generalized
tonic–clonic seizure occurred on day −3. After a series of
investigations including brain CT, electrolytes, electrocardiography, and cerebrospinal fluid studies, no specific etiology was found, and busulfan was considered to be responsible. The seizure was controlled with phenytoin. However,
acute neurologic deterioration was noted during this period.
She became totally bed ridden, unable to voice repetitive
consonant sounds (mama, dada) and spasticity increased,
but she could still swallow well, keep her head erect, sit in
a chair with her trunk erect, and no head lag. She received
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Y-S Hsu et al
105
Figure 2 Head MRI, T2WI. (a) Before BMT, myelination was normal and grossly no obvious brain atrophy is depicted. (b) Six months after BMT,
sulci and cisterns are diffusely dilated. The lateral and third ventricles are diffusely prominent. Myelination is still normal. As compared with the previous
MRI, brain atrophy is evident.
8 × 108 nucleated cells/kg recipient weight and 8.16 × 106
CD34+ nucleated cells/kg recipient weight. The post-BMT
course was smooth, without acute GVHD.
Bone marrow aspiration 3 weeks later showed full
engraftment as evidenced by sex chromosome studies (all
cells were 46 XY male donor type) although many foamy
cells could still be seen. One month after BMT, she was
discharged in stable clinical condition but with persistent
hepatosplenomegaly, and bed-ridden.
Six months after transplantation, her liver and spleen
were impalpable. Grade I GVHD developed. Bone marrow
aspiration and biopsy still showed complete engraftment
with normocellularity. The most striking finding was that
the lipid-filled foamy macrophages seen in previous
biopsies were not found in most areas (Figure 3b). Chest
CT at 6 and 9 months revealed almost complete resolution
of the previous lung infiltrate (Figure 1b). However, developmental regression progressed after the transplant. She
could vocalize when talked to or sung to but there was
no ability to imitate speech sounds or repeat consonants.
Difficulty with swallowing occurred during these months.
She could not keep her head erect or raise her body or
maintain a prone position at 45°. She became unable to sit
in a chair with her trunk erect. Head lag became obvious
during traction pulling to sit. In addition, horizontal supranuclear ophthalmoplegia appeared. The developmental quotient of gross motor, language, fine motor, and personalsocial skills decreased significantly both before and after
BMT. The estimated developmental age using the Denver
Development Screening Test II was 10–14 months, 10–14
months, 5–8 months, 3–5 months, 2–4 months at chronological ages of 1 year 8 months (1y8m), 2y5m, 2y7m,
2y11m, 3y3m, respectively. EEG showed markedly diffuse
cortical dysfunction. Although she had no obvious brain
atrophy at the time of BMT, brain MRI 6 months later
showed obvious diffuse brain atrophy with decreased thickness of the white matter (Figure 2b).
Discussion
NPC has heterogeneous clinical manifestations. The classic
phenotype is characterized by variable hepatosplenomegaly, vertical supranuclear ophthalmoplegia, progressive
ataxia, dystonia, and dementia. The onset is usually in late
childhood, and death occurs in the second decade. Other
phenotypes include fatal neonatal liver disease, early infantile onset with hypotonia and delayed motor development,
and adult variants where psychosis and dementia predominate.7 There is still no specific treatment for NPC. Treatment strategies to reduce intracellular cholesterol accumulation have been formulated based on the hypothesis that
cholesterol is an offending metabolite in NPC. BALB/C
mutant mice fed with a low cholesterol diet live twice as
long as those given a 2% cholesterol diet.4 The combination
of cholesterol lowering agents including cholestyramine,
lovastatin, and nicotinic acid lowered cholesterol by 20%
in the liver and blood of NPC patients with minimal sideeffects, but any influence on the rate of neurologic progression is doubtful.8 Dimethyl sulfoxide (DMSO) has been
proved to be an activator of acid sphingomyelinase and to
accelerate the intracellular mobilization of LDL-derived
cholesterol. It has been known to correct a partial
deficiency of acid sphingomyelinase activity in NPC fibroblasts and oral administration of DMSO has brought about
clinical improvement including reduction in hepatosplenomegaly and seizure frequency but the progressive clinical
course did not change although it appeared to slow down.9–11
DMSO may be worth trying in NPC patients.
In the C57BL/KsJ spm/spm murine model of NPC, BMT
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Y-S Hsu et al
106
Figure 4 Electron microscopic examination for liver biopsy. The macrophage (Kupffer cells) in the sinusoid spaces are enlarged and filled with
abundant secondary lysosomes of variable sizes, which contain membranous cytoplasmic bodies (arrows) resembling concentric lamellated myeline figures (H, hepatocytes; K, Kuppfer cell; original magnification
×12 000).
Figure 3 Photomicrography of bone marrow biopsies (hematoxylin
eosin, original magnification ×200). (a) Before BMT, many lipid-filled
foamy cells are seen (arrows) disseminated among the hematopoietic cells.
(b) Six months after BMT, a normal cellular marrow with no foamy cells
is shown.
decreased the accumulation of sphingomyelin and cholesterol quantitatively in the spleen, and the sphingomyelin
deposited in the bone marrow was also reduced on histology. On the other hand, neurological involvement was
not improved by the graft.12 Combined liver and bone marrow transplantation in mice partially reversed tissue storage
of cholesterol and sphingomyelin, and ameliorated hepatosplenomegaly but did not influence the continued neurologic deterioration and life span. Foamy cells disappeared
from bone marrow, liver, spleen, thymus, and lymph nodes,
but depletion of Purkinje cells was not prevented.13
However, in BALB/c mice with cholesterol-storage diseases, transplantation of fetal liver cells corrects accumulation of lipids in tissues and prevents fatal neuropathy. Tremors disappeared and normal weight was almost regained.
Deposition of lipids in tissues was decreased, neuropathy
was prevented, and survival was significantly prolonged.14
Nevertheless, liver transplantation in a 7-year-old girl with
Niemann–Pick type C disease and liver cirrhosis was successful in restoring hepatic function but failed to slow down
neurologic progression.15
The results of BMT in our patient were similar to the
C57BL/KsJ spm/spm mouse receiving BMT. Acute accelerated then continuous neurological deterioration was noted
after BMT. Acute accelerated neurological deterioration
may be due to a side-effect of busulfan. MRI shows obvious atrophic changes of the brain 1/2 month before and 6
months after BMT (Figure 2a and b) and provides evidence
for the failure of BMT to prevent CNS deterioration. However, long-term CNS follow-up will still be undertaken in
this patient. In addition, rectal biopsy will also be considered in this patient and other new cases, since it may
allow insight into the effects of the engrafted normal marrow on ganglion cells as neuronal storage also occurs in
the Schwann’s cells and axons.16
Over the decade, BMT has been used to treat many storage diseases including adrenoleukodystrophy, metachromatic leukodystrophy, globoid cell leukodystrophy, Gaucher’s disease and Hurler syndrome, etc. Engraftment and the
resulting enzymatic reconstitution are concordant, and positive changes occur in the central nervous system (CNS)
following long-term engraftment. After engraftment, significant improvement in the clinical course of each of these
diseases occurs. Earlier diagnosis will allow BMT to be
performed in the presymptomatic stage, providing enhancement of the positive effects of such treatment.17 It is not
clear whether in NPC it will be helpful if BMT is performed earlier, before the symptomatic stage.
In this report, we have shown that a correction occurred
in the liver, spleen, bone marrow and lung following successful engraftment in human NPC. Such a therapeutic
response is remarkable since the underlying hypothesis is
that of a membrane receptor problem for cholesterol. This
positive response may be directly due to the replacement
of foamy histiocytes in the viscera with donor cells, but
cross-correction in the integral membrane protein defect by
BMT is still a possibility. In vitro studies have revealed
that NPC1/NPC2 somatic cell hybrids exhibit a normal
phenotype. Suprisingly, co-cultivation of NPC1 and NPC2
BMT for Niemann–Pick type C disease
Y-S Hsu et al
fibroblasts also corrects the phenotype18 to a small degree.
It is possible that a soluble factor which enhances cholesterol esterification in NPC-1 fibroblasts is released into the
medium by NPC-2 cells or exchanged by cellular junctions,
or perhaps a correcting factor is secreted, which acts downstream of NPC1.19
Although the BMT result in this case is not encouraging,
diagnosis in fetal life in families where an older sibling
is affected or by clinical judgement (prolonged cholestatic
jaundice in newborn period) may allow an immediate postnatal transplant or other interventions before the symptomatic stage. Earlier diagnosis and intervention are
important and are most likely to provide an answer concerning whether or not neurologic disease can be prevented.
Acknowledgements
The authors want to thank Dr Adele Cooney at the Developmental
Metabolic Branch, National Institute of Neurological Disorders
and Stroke, National Institutes of Health, and Dr David A Wenger
at the Division of Medical Genetics, Jefferson Medical College,
Philadelphia, Pensylvania, USA for assisting us in the diagnosis
of Niemann–Pick type C disease in the patient.
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