Therapeutic response to intravenous infusions of glucocerebrosidase in a patient with Gaucher disease.

Enzyme replacement has been under consideration as a therapeutic strategy for patients with Gaucher disease for more than two decades. Previous studies indicated that single injections of purified glucocerebrosidase reduced the amount of storage material in the liver. It was important to determine whether administration of exogenous enzyme on a regular basis would be of clinical benefit.

Structure of the N-asparagine-linked oligosaccharide units of human placental beta-glucocerebrosidase.

The N-asparagine-linked oligosaccharide chains of homogeneous human placental beta-glucocerebrosidase were released by hydrazinolysis, and their structures were analyzed. The sequence of sugars, linkage, and anomeric configuration of the glycosidic bonds were determined. Approximately 20% of the released sugar chain was of the typical high mannose type.

Mutations of glucocerebrosidase: discrimination of neurologic and non-neurologic phenotypes of Gaucher disease.

Multiple molecular forms of beta-glucocerebrosidase that permit discrimination between neurologic and non-neurologic phenotypes of Gaucher disease have been identified radioimmunologically in fibroblasts and human brain tissue. In normal human fibroblasts these forms have been shown by NaDodSO4/polyacrylamide gel electrophoresis to have apparent Mr of 63,000 (form A1), 61,000 (form A2), and 56,000 (form B). The Mr 63,000 form may be a precursor of the Mr 56,000 form.

Delivery of hexosaminidase A to the cerebrum after osmotic modification of the blood--brain barrier.

The present studies were undertaken to evaluate the possibility that hexosaminidase A, the enzyme deficient in Tay--Sachs disease, could be effectively delivered to brain. Previous studies from our laboratory have shown that hypertonic mannitol can be used to osmotically produce reversible disruption of the blood--brain barrier in animals (rat and dog) and man without significant neurotoxicity and that such barrier modification significantly increases the delivery of cytoreductive chemotherapy agents to selected areas of brain. By using the rat model of blood--brain barrier modification and radiolabeled enzyme, increased hexosaminidase A delivery to brain has been demonstrated in more than 85 animals.

Uptake and distribution of placental glucocerebrosidase in rat hepatic cells and effects of sequential deglycosylation.

The clearance of native human placental glucocerebrosidase by rat liver shows the presence of two distinct enzyme forms with different recognition characteristics. The clearance and uptake of native enzyme by liver cells was compared to that of glucocerebrosidase sequentially treated with neuraminidase, beta-galactosidase and beta-N-acetylglucosaminidase. The initial rate of clearance of infused enzyme was increased greater than 10-fold for the asialo-, agalacto- and ahexoenzymes over that of native glucocerebrosidase.

Status of enzyme replacement therapy for Gaucher disease.

At this point in time, enzyme replacement therapy for Gaucher disease appears both biochemically effective (patients 1-3, and 10) as well as clinically promising (patients 4 and 5), although these salutary responses have not been obtained consistently (patients 6-9). The discrepancy may be due to the method employed for the isolation of the enzyme. One current opinion is that glucocerebrosidase prepared by conventional fractionation and ion exchange chromatographic procedures may have been more effective in vivo than enzyme prepared by butanol extraction and hydrophobic column chromatography, which we developed because of the difficulty in obtaining large quantities of glucocerebrosidase by the conventional method.

Enzyme replacement therapy in Gaucher's disease: large-scale purification of glucocerebrosidase suitable for human administration.

Enzyme replacement therapy for the alleviation of Gaucher's disease has been impeded because of the difficulty in preparing large amounts of glucocerebrosidase, the enzyme that is deficient in patients with this disorder. A large-scale procedure for the purification of human placental glucocerebrosidase has been developed. The method uses cholate extraction, ammonium sulfate fractionation, acid precipitation, butanol extraction, and hydrophobic chromatography; the final enzyme preparation has a specific activity of more than 10(6) units/mg of protein with an overall recovery of 30%.

A practical chromogenic procedure for the diagnosis of Krabbe's disease.

Krabbe's disease is caused by a deficiency of galactocerebrosidase in organs and tissues. Determinations of galactocerebrosidase activity had required the use of galactocerebroside labeled with radiocarbon or radiohydrogen. These materials are expensive and their use is restricted to laboratories with radioactive counting facilities.