Roscoe Brady, M.D.
Scientist Emeritus
National Institutes of Health
Bethesda, Maryland USA
CURRENT APPROACHES TO TREAT PATIENTS WITH
HEREDITARY METABOLIC STORAGE DISORDERS
1. ENZYME REPLACEMENT THERAPY
2. SUBSTRATE DEPLETION
3. CHEMICAL CHAPERONES
4. GENE THERAPY
Can Additional Strategies Be Developed?
For many years it was believed that alterations of
the normal sequence of amino acids caused a
reduction in the ability of glucocerebrosidase to
degrade glucocerebroside that accumulates in
patients with Gaucher disease.
My colleagues and I discovered that the decrease
of glucocerebrosidase activity is due, at least in
part, to a reduction of the amount of this enzyme
in the cells and tissues of patients with Gaucher
disease.
(Lu J, et al. Proc Natl Acad Sci USA 2010; 107: 21665)
Alterations of the amino acid sequence of
glucocerebrosidase cause abnormal folding of
the mutated enzymes within cells.
Cells cannot tolerate mis-folded proteins. An
enzyme complex called the proteaosome
destroys mis-folded proteins.
We investigated proteasomal degradation of
mutated glucocerebrosidase in skin fibroblasts
derived from patients with Gaucher disease with
the N370S and L444P mutations.
The quantity of glucocerebrosidase is reduced
in cells of patients with Gaucher disease.
% of normal
Type 1 Adult GD (N370S/N370S)
Type 2 Acute neuronopathic GD (L444P/L444P)
Type 3 Chronic neuronopathic GD (L444P/L444P)
43.
11.
22.
These changes cause a comparable
reduction of the catalytic activity of
glucocerebrosidase in cells of patients
with Gaucher disease
% of normal
Type 1 GD (N370S/N370S)
Type 2 GD (L444P/L444P)
Type 3 GD (L444P/L444P)
42.
10.
25.
Why does this happen?
Cellular strategy of protein quality control
Biosynthesis, Folding and Transport of Glucocerebrosidase to Lysosomes
messenger RNA
Hsp70 prevents nascent peptides from
aggregating and being rendered nonfunctional
The TCP1 ring complex consists of two identical stacked rings, each
containing eight different proteins. Unfolded polypeptides enter the
central cavity of the complex and are folded in an ATP-dependent manner.
Biosynthesis, Mis-folding and Degradation of Mutated Glucocerebrosidase
messenger RNA
Hsp90 is essential for the creation, maintenance
and destruction of proteins. Its normal function
is critical to maintaining the health of cells
Quantification of the increase in binding
of N370S and L444P glucocerebrosidase
mutants to the Hsp90 complex that
facilitates degradation.
Quantification of the reduction of binding
of N370S and L444P glucocerebrosidase
mutants to Hsp70 and TCP1 required for
correct folding of nascent proteins.
The catalytic activity of the residual
glucocerebrosidase in fibroblasts from
patients with the N370S and L444P
mutations was found to be normal.
Conclusions
1. Missense mutations destabilize glucocerebrosidase
2. Mutated glucocerebrosidase is partially degraded
3. Lysosomal localization is reduced
4. Rather than a decrease of intrinsic catalytic
activity, proteasomal degradation underlies
the reduction of enzymatic activity in patients with
Gaucher disease caused by missense mutations
Would suppression of proteasomal degradation of
mutated glucocerebrosidase provide an additional
treatment option for patients with Gaucher disease?
Can proteostasis regulators promote the binding of N370S
and L444P mutated glucoerebrosidases to chaperonins
and reduce their degradation?
Histones
Alkaline proteins in cell nuclei that package DNA into
structural units
Histone deacetylases
Enzymes that remove acetic acid residues from histones
Histone deacetylase inhibitors
Decrease proteoasomal degradation thereby restoring
the function of mis-folded proteins
Histone deacetylase inhibitors (HDACi) have been
found to correct aberrant protein folding in:
1. Type 2 diabetes
2. Cystic fibrosis
3. Fibroblasts derived from patients with Type C
Niemann-Pick disease
Can histone deacetylase inhibitors
(HDACi) reduce proteasomal degradation
of mutated enzymes in hereditary
enzyme deficiency disorders such as
Gaucher disease?
My colleagues and I discovered that histone
deacetylase inhibitors prevent the degradation
and restore glucocerebrosidase activity in Gaucher
disease.
(Lu J et al. Proc Natl Acad Sci USA 2011;108:21200)
Improved lysosomal trafficking of mutated glucocerebrosidase by HDACi
Histone deacetylase inhibitor approved for the treatment
of patients with cutaneous T cell lymphoma
o
I
N
o
I
H
Suberoylanilide Hydroxamic acid (SAHA)
LB-205
Longer lasting histone deacetylase inhibitor
Lixte Biotechnology, East Setauket, NY
Effect of histone deacetylase inhibitors on the
quantity of glucocerebrosidase
(Percent of Normal)
Mutation
Untreated
Treated with SAHA Treated with LB-205
N370S/N370S
38
68
81
L444P/L444P
12
43
41
Effect of histone deacetylase inhibitors on the
catalytic activity of mutated glucocerebrosidase
(Percent of Normal)
Mutation
Untreated
Treated with SAHA Treated with LB-205
N370S/N370S
28
63
76
L444P/L444P
16
35
40
Reported toxic effects of SAHA
Nausea, vomiting, diarrhea, reduced appetite,
headache, fatigue and indigestion that resolved
during a week of rest.
A safety and dose-response trial with LB-205
is under consideration
OVERVIEW
Considerable efforts are underway concerning the development
of innovative therapies for Gaucher disease and other hereditary
metabolic disorders.
The use of histone deacetylase inhibitors appears to be a
promising therapeutic strategy for the following reasons:
1. They are administered orally thereby eliminating the necessity
for intravenous infusions.
2. They cross the blood-brain barrier and are expected to be
beneficial for patients with central nervous system
involvement.
3. Because they are small molecules, it is presumed that their
cost will be less than enzyme replacement and other
therapies.
How do we proceed from here?
1. Determine the effect of HDAC inhibitors in murine
models of Gaucher disease.
2. If positive results are obtained, carry out clinical
studies in patients with Type 1 and neuronopathic
forms of Gaucher disease.
Thank you