Transcript SMN2 - ASEM Catalunya

Phenylbutyrate increases SMN gene expression
in vitro and in vivo
Christina Brahe
Università Cattolica del S.Cuore, Roma
SMA is caused by homozygous absence of the
SMN1 gene
SMN2
cen
SMN1
tel
SMA patients have one or more SMN2 genes which
modulate the disease severity
SMN2
severe form
SMN2
SMN2
mild form
These genes produce an insufficient level of SMN protein
Hypothesis
Particular drugs may activate the SMN2 genes
to produce more SMN protein
SMN
SMN
improvement of muscle strength
There are drugs that open up the DNA to allow the
expression of genes at a higher level
inactive gene
HDAC
active gene
HAT
condensed chromatin
accessible chromatin
Phenylbutyrate belongs to this class of drugs
• FDA approved
for the treatment of another pediatric disease
• is generally well tolerated
• crosses the blood-brain barrier
Study of the effect of phenylbutyrate on SMN2 expression in
cell cultures from SMA patients
Results
• Increase in SMN transcripts (precursor of protein)
in most but not all cultures
• the increase in transcripts ranged from 50%-400%
• increase in SMN protein
Maximum increase in SMN2-fl transcript levels
% increase in SMN2-fl transcripts
500
450
400
350
300
250
200
150
100
50
1
2
3
SMA I
4
5
6
7
8
SMA II
9
10
11 12 13 14 15 16
SMA III
patient
Increase in SMN protein
120
% increase in SMN protein
16 h
100
24 h
80
60
40
20
SMA I
SMA II
SMA III
Study of the effect of phenylbutyrate on
SMN2 expression in leukocytes of SMA patients
• Phenylbutyrate was administered to 6 patients
(4 type II and 2 type III) and 3 parents for 7 days
• blood samples were taken before, during and after the trial
• leukocyte SMN2 transcripts were measured
SMN-fl transcripts (%) relative to T0
PB increases SMN expression in leukocytes
900
900
800
800
500
500
400
400
300
300
200
200
100
100
0
0
-100
1
2
3
4
patients
5
6
M2
M3
F6
parents
-100
T1
T2
T3
T4
T7
Mean
1
2
3
4
5
healthy untreated
controls
Brahe et al., EJHG 2005
Open pilot trial
• 10 patients with SMA II (age: 30 months-12 years)
• Treatment with phenylbutyrate for 13 weeks
(1 week on, 1 week off)
• Efficacy was evaluated by using a functional motor scale
Functional motor scale
Three point scoring system:
2 scores for unaided, 1 for assistance and 0 for inability
20 items
Score 2 points
Touch one hand to head
Score 1 point
flexes head to hand
Score 0
Score
unable
( )
score
Results on the functional scale
35
before
30
3 weeks
25
9 weeks
20
15
10
5
0
2.6
3.5
3.6
3.8
4.1
6.1
6.3
7.7
9.7
12.7
Age (years)
Mercuri et al., 2004
Mean scores in treated patients and control group
22
20
18
16
14
12
10
8
6
4
2
T0
T1
T2
treated patients
T0
T3 (6 months)
untreated controls
Mercuri et al., 2004
Conclusions of the open trial
• improvement of functional ability
• children <5 years had a more obvious improvement
• great variability of response (as previously observed
in vitro)
Large randomised, double-blind, placebo controlled trial
Enrolled : 106 patients SMA II or non-ambulant type III
Age:
30 months -12 years
Drop out: 14 (13%)
Schedule: intermittent (1 week on, 1 week off) for 13 weeks
Assessments:
• Hammersmith functional motor scale
• myometry (>5 years)
• FVC (>5 years)
Problems to be solved
• More reliable outcome measures
(more sensitive functional motor scale and quantitative muscle
testing, etc)
• protocol design to achieve the maximum response
(dosage, continuous or pulsed administration, duration)
• gain information on the mechanisms underlying the
difference in response
Istituto di Genetica Medica
Università Cattolica, Roma
Danilo Tiziano
Carla Angelozzi
Federica Borgo
Anna Maria Pinto
Daria Darelli
Tiziana Vitali
Giovanni Neri
Istituto di Neurologia
Università Cattolica, Roma
Eugenio Mercuri
Ospedale Bambino Gesù
Rome
Enrico Bertini
Acknowledgements:
Patients and parents
FSMA USA
FSMA Italy
A.S.A.M.S.I.
Telethon - Italy