Gene Therapy (2013) 20, 913–921
& 2013 Macmillan Publishers Limited All rights reserved 0969-7128/13
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ORIGINAL ARTICLE
Disease correction by combined neonatal intracranial AAV and
systemic lentiviral gene therapy in Sanfilippo Syndrome type B
mice
CD Heldermon1, EY Qin2, KK Ohlemiller3, ED Herzog4, JR Brown5, C Vogler6, W Hou7, JL Orrock8, BE Crawford5 and MS Sands2
Mucopolysaccharidosis type IIIB (MPS IIIB) or Sanfilippo Syndrome type B is a lysosomal storage disease resulting from the
deficiency of N-acetyl glucosaminidase (NAGLU) activity. We previously showed that intracranial adeno-associated virus (AAV)based gene therapy results in partial improvements of several aspects of the disease. In an attempt to further correct the disease,
MPS IIIB mice were treated at 2–4 days of age with intracranial AAV2/5-NAGLU (IC-AAV), intravenous lentiviral-NAGLU (IV-LENTI) or
the combination of both (BOTH). The BOTH group had the most complete biochemical and histological improvements of any
treatment group. Compared with untreated MPS IIIB animals, all treatments resulted in significant improvements in motor function
(rotarod) and hearing (auditory-evoked brainstem response). In addition, each treatment group had a significantly increased
median life span compared with the untreated group (322 days). The combination arm had the greatest increase (612 days),
followed by IC-AAV (463 days) and IV-LENTI (358 days). Finally, the BOTH group had nearly normal circadian rhythm measures with
improvement in time to activity onset. In summary, targeting both the systemic and central nervous system disease of MPS IIIB early
in life appears to be the most efficacious approach for this inherited metabolic disorder.
Gene Therapy (2013) 20, 913–921; doi:10.1038/gt.2013.14; published online 28 March 2013
Keywords: Sanfilippo; behavioral; MPS IIIB
INTRODUCTION
N-Acetyl-glucosaminidase deficiency (Sanfilippo Syndrome type B,
Mucopolysaccharidosis IIIB) typically causes a pediatric onset
disease characterized phenotypically, by progressive motor and
cognitive deterioration, and histologically by accumulation of
lysosomal inclusions in most tissues.1 No current treatment is
approved in humans. After the gene was identified,2,3 a murine
model of mucopolysaccharidosis type IIIB (MPS IIIB) was created.4
The MPS IIIB mouse shares many of the biochemical, histological
and clinical features with the human disease.4,5
Several groups have demonstrated the ability of either
intracranial or systemic gene therapy approaches to reduce
lysosomal distention in the brains of MPS IIIB mice.6–11 Our
group also demonstrated improvements in histology with
corresponding improvements in neurologic function and life
span using intracranial gene therapy.12 Intracranial delivery has
thus far demonstrated the most consistent improvement in
disease progression. Increases of approximately 30% in life span
have been observed with central nervous system (CNS)-directed
therapies. An intracranial gene therapy approach is now being
pursued in a larger animal model of MPS IIIB.13 Systemic-targeted
gene therapy was shown to reduce lysosomal storage in
peripheral organs. However, none of these single approaches
completely eradicates intra-cytoplasmic inclusions or normalizes
the disease phenotype. We previously attempted a combination
of CNS-directed gene therapy and bone marrow transplantation
with little or no benefit seen for the bone marrow transplantation
arms. However, the level of chimerism was relatively low and
toxicities from the radiation conditioning were evident in the
transplant arm. In other lysosomal storage disease models,
therapies to the CNS and the periphery, with enzyme
replacement, bone marrow transplantation or gene therapy,
have shown greatly improved disease correction especially
when initiated in the neo-natal period.14–17
Therefore, we hypothesized that neonatal combination therapy
directed to both the CNS and the periphery would provide better
correction of the disease, especially if higher systemic levels of
N-acetyl glucosaminidase (NAGLU) activity could be attained. We
describe here the benefits obtained from each mode of gene
therapy and the synergistic effect of combining intracranial
adeno-associated virus (AAV)-NAGLU and systemic lentiviralNAGLU gene therapy.
RESULTS
Treatments
All gene therapy injections were performed in mice pups at
2–4 days of age. Intracranial AAV-NAGLU treatment was
1
Department of Medicine, University of Florida, Gainesville, FL, USA; 2Department of Internal Medicine, Washington University, St Louis, MO, USA; 3Department of
Otorhinolaryngology, Washington University, St Louis, MO, USA; 4Department of Biology, Washington University, St Louis, MO, USA; 5Zacharon Pharmaceuticals, San Diego, CA,
USA; 6Department of Pathology, Saint Louis University, St Louis, MO, USA; 7Department of Biostatistics, University of Florida, Gainesville, FL, USA and 8Department of Zoology,
University of Wisconsin-Madison, Madison, WI, USA. Correspondence: Dr CD Heldermon, Department of Medicine, University of Florida, 1600 SW Archer Road, Box 100278,
Gainesville, FL 32610, USA. E-mail:
[email protected]fl.edu
Or Professor MS Sands, Department of Internal Medicine, Washington University in St Louis, 660 South Euclid, Campus Box 8007, St Louis, MO 63110, USA. E-mail:
Received 14 September 2011; revised 11 February 2013; accepted 21 February 2013; published online 28 March 2013
Correction of Sanfilippo Syndrome type B
CD Heldermon et al
914
performed as described previously with six direct injections of 2 ml
each into frontal, temporal and cerebellar regions of the brain of
vector at a concentration of 1.5 1012 viral particles per ml.5,12
Intravenous lentiviral-NAGLU injections were also performed as
described previously by injection of 100 ml of 1.6 108 infectious
units per ml viral aliquot into the superficial temporal vein.18
NAGLU activity
Biochemical analysis of N-acetyl-glucosaminidase activity for
various organs was determined in mice from each group at
B8 months of age and compared with untreated MPS IIIB animals
(MPS IIIB NO TX; Figure 1). The MPS IIIB animals receiving only
intravenous lentiviral vector treatment (MPS IIIB IV-LENTI)
had detectable NAGLU activity in all organs assayed (o2% of
normal in the brain (Po0.05) and kidneys (not significant (NS)),
11% in heart (Po0.01), 12% in lungs (NS), 34% in liver (Po0.001),
34% in spleen (Po0.05) and 28% in the serum (NS; data not
shown). Conversely, animals treated only with intracranial AAV
(MPS IIIB IC-AAV) had approximately 200% (Po0.05), 3% (NS) and
5% (NS) of normal activity in the brain, liver and serum (data not
shown), respectively, and little or no activity in the spleen,
heart, lungs or kidneys. Combination therapy (MPS IIIB BOTH)
yielded NAGLU activity levels of 424% for brain (Po0.01), 13.6% for
liver (Po0.001), 6.7% for heart (Po0.01), 19.8% for spleen
(Po0.01), 2.9% for lungs (Po0.05), 42% for serum (NS) and o1 (...truncated)
