ASSAYS FOR DIAGNOSING AND EVALUATING TREATMENT OPTIONS FOR FABRY DISEASE
Provided are in vitro and in vivo methods for determining whether a patient with Fabry disease will respond to treatment with a specific pharmacological chaperone.
1 . A method for determining whether a patient having a deficiency in activity of a protein will respond to treatment with a specific pharmacological chaperone for the protein, which method comprises
a. contacting cells from a patient an individual with a pharmacological chaperone specific for the protein; and
b. comparing protein activity in cells not contacted with the specific pharmacological chaperone, with protein activity in cells contacted with the specific pharmacological chaperone
wherein an increase in protein activity in cells contacted with the specific pharmacological chaperone over activity in cells not contacted with the specific pharmacological chaperone indicates that the patient will respond to treatment with the specific pharmacological chaperone.
2 . The method of claim 1 , wherein the deficiency of activity is caused by a missense mutation in a gene encoding the protein.
3 . The method of claim 1 , wherein the protein is an enzyme.
4 . The method of claim 3 , wherein the enzyme is a lysosomal enzyme.
5 . The method of claim 4 , wherein the patient has been diagnosed with a lysosomal storage disorder.
6 . The method of claim 5 , wherein the lysosomal enzyme is α-GAL and the lysosomal storage disorder is Fabry disease.
7 . The method of claim 5 , wherein the specific pharmacological chaperone is 1-deoxygalactonojirimycin.
8 . The method of claim 7 , wherein the cells are white blood cells and the contact with the specific pharmacological chaperone occurs in vivo.
9 .- 16 . (canceled)
17 . The method of claim 8 , wherein the patient is administered 1-deoxygalactonojirimycin daily for about 2 weeks.
18 . The method of claim 17 , wherein the administration is oral.
19 . The method of claim 17 , wherein the 1-deoxygalactonojirimycin is administered at a dose of about 50-500 mg/day.
20 . The method of claim 19 , wherein the dose is about 100-250 mg/day.
21 . The method of claim 20 , wherein the dose is about 150 mg/day.
22 . The method of claim 19 , wherein the 1-deoxygalactonojirimycin is administered once a day.
23 . The method of claim 17 , further comprising collecting a blood sample at the end of two weeks and separating the white blood cells.
24 . The method of claim 17 wherein α-GAL activity is determined using a fluorometric assay that quantifies hydrolysis of substrate in lysates from the white blood cells.
25 . The method of claim 24 wherein the sufficient increase in activity in the lysates in the presence of the 1-deoxygalactonojirimycin which indicates whether the patient will respond is measured according to the following criteria:
i) If baseline activity is less than 1% of normal, the activity following culture or following treatment with SPC must be at least 2% of normal;
ii) If baseline activity is between 1% but less than 5% of normal then the activity following culture or treatment with SPC must be at least 2 times the baseline level;
iii) If baseline activity is between 5% but less than 10% of normal, then the activity following culture or treatment with SPC must be at least 5% of normal higher the baseline level of normal;
iv) if baseline activity is 10% of normal or more, then activity following culture or treatment with SPC must be at least 1.5× the baseline level.
26 . A kit comprising:
a. at least one T cell stimulatory agent;
b. a specific pharmacological chaperon
c. a labeled substrate for the chaperone;
d. GalNAc; and
e. instructions for performing a protein enhancement assay.
27 . The kit of claim 26 , wherein the T-cell stimulatory agent is a mitogen.
28 . The kit of claim 27 , wherein the mitogen is PHA.
29 . The kit of claim 26 , wherein the stimulatory agent is a cytokine.
30 . The kit of claim 29 , wherein the cytokine is IL-2.
31 . The kit of claim 26 , wherein the pharmacological chaperone is 1-deoxygalactonojirimycin.
32 . The kit of claim 26 , further comprising one or more a blood collection tubes, centrifuge tubes, and cryotubes.
33 . The kit of claim 26 , wherein the protein is an enzyme.
34 . The kit of claim 33 , wherein the enzyme is α-galactosidase A.