Intrathecal enzyme replacement therapy: successful treatment of brain disease via the cerebrospinal fluid

Abstract

Treatment of brain disease with recombinant proteins is difficult due to the blood-brain barrier. As an alternative to direct injections into the brain, we studied whether application of high concentrations of therapeutic enzymes via intrathecal (IT) injections could successfully drive uptake across the ependyma to treat brain disease. We studied IT enzyme replacement therapy with recombinant human iduronidase (rhIDU) in canine mucopolysaccharidosis I (MPS I, Hurler syndrome), a lysosomal storage disorder with brain and meningeal involvement. Monthly or quarterly IT treatment regimens with rhIDU achieved supranormal iduronidase enzyme levels in the brain, spinal cord, and spinal meninges. All regimens normalized total brain glycosaminoglycan (GAG) storage and reduced spinal meningeal GAG storage by 58-70%. The improvement in GAG storage levels persisted three months after the final IT dose. The successful use of enzyme therapy via the CSF represents a potentially useful approach for lysosomal storage disorders.

Fig. 1

IT rhIDU regimens and doses received by the dogs. Dogs ranged in age from 13 to 40 months at the end of the study and consisted of 14 males and 5 females. Gray arrows indicate IT rhIDU injection. The weekly regimen (n=4) consisted of four ~ 1 mg (270,000 units) doses of IT rhIDU at weekly intervals. The monthly regimen (n=5) consisted of four 1.38 mg (270,000 units) doses of IT rhIDU at monthly intervals. Quarterly regimen (n=4) consisted of three 1.38 mg (270,000 units) doses IT rhIDU at three month intervals. Dogs were necropsied 48 hours after the last IT rhIDU injection (black arrows). Dogs receiving low-dose IT rhIDU received 0.46 mg (90,000 units) per dose at monthly intervals, and were necropsied 48 hours (n=2) or three months (n=4) after the final IT rhIDU dose (black arrows). Dogs on monthly, quarterly, and low dose regimens received concomitant weekly 0.58 mg/kg (125,000 units/kg) IV rhIDU. During the three-month hiatus, dogs continued to receive weekly IV rhIDU until necropsied.

Fig. 2

Iduronidase levels in MPS I dogs treated with IT rhIDU. (a) Distribution of iduronidase activity in treated canine brains. Normal brain iduronidase activity is 11.9 ± 1.95 units/mg protein. In the brain, core samples in the temporoparietal regions were divided into surface and deep regions. Deep regions were at least three mm below the brain surface. All but one region had supranormal iduronidase activity (above 12 units/mg); the level in this region was 11.7 units/mg. Iduronidase levels are also shown in the spinal cord and spinal meninges. Spinal meninges sections consist of dura and arachnoid materes assayed together. Sections were taken from the cervical (C2-3), thoracic (T4-5), and lumbar (L4-5) regions. Normal spinal cord iduronidase is 11.7 ± 2.38; meninges, 15.4 ± 3.55. Values from 2 to 338 fold-normal were achieved with treatment. (b) Bar graph illustrating supranormal levels of iduronidase in total brain, spinal cord, and spinal meninges with monthly, quarterly, and low-dose IT rhIDU. As expected, lower iduronidase levels are achieved with the lower dose.

Fig. 3

Quantitative glycosaminoglycan (GAG) storage in brain, spinal cord, and spinal meninges of normal dogs, untreated MPS I dogs, and MPS I dogs treated with IT rhIDU. (a) GAG levels in the brain were normal in all treatment groups. Mean GAG storage in the spinal meninges was reduced by more than half in all treatment groups versus untreated MPS I dogs. (b) Brain GAG levels in the IT-treated dogs reached normal levels regardless of age or treatment regimen.

Fig. 4

Pathological evaluation of IT-treated MPS I dogs shows improved GAG storage in perivascular cells (top panels) and neurons (bottom panels) compared with untreated MPS I dogs. LM, light microscopy; EM, electron microscopy. Toluidine blue stain of perivascular cells (centered in each upper panel) shows lysosomal storage (arrowhead) in an untreated dog that is not apparent in that of the treated dog. The blood vessel lumen is marked with an asterisk. Electron microscopy (lower panels) shows lipid and GAG storage vesicles (arrows) in the neuron of an untreated dog that is greatly reduced in a treated dog. Treated dogs did have some neuronal storage, though it was decreased from untreated levels.

Fig. 5

Reduced meningitis with less frequent dosing and immune suppression in dogs treated with IT rhIDU. (a) A nontolerant dog treated with weekly IT rhIDU developed a moderate meningitis, as shown by hematoxylin and eosin stain of the spinal meninges. A dog receiving the quarterly regimen (b) and a dog receiving the low dose regimen (c) developed a much milder degree of meningitis. Both dogs had received immunosuppressive drugs, and the low dose dog (c) became tolerant to iduronidase. A dog treated with the low dose regimen, evaluated after a three month hiatus, also developed minimal meningitis (d). This dog was not tolerant to iduronidase. Cell differentials and pathological evaluation showed the meningitis to consist of lymphocytes, plasma cells, and monocytes/macrophages. This infiltrate was seen in the neocortical and spinal meninges, and was absent in the parenchyma of the brain and spinal cord.