2013 Research Update

APBDRFH. Orhan AKMAN, PhD

Dr. DiMauro’s Laboratory

Columbia University Medical Center

Department of Neurology

New York

Adult Polyglucosan Body DiseaseDiagnosis, and Development of tools for Research and Treatment

Mouse model of APBD

Possible Pharmacological Treatments of APBD

Diagnosis of Manifesting Heterozygote Patients

Mouse model of APBD with Y329S mutation

Y329S TAT to TCTTyr Ser

FRT

LoxP

5’ ARM 2.1 kb PGK- Neomycin 3’ARM 6.5 kb Exon 7

Exon 75’ ARM 3’ARM

Generation of mouse model of APBD:

Breeding scheme and expected genotypes

Y329S TAT to TCTTyr Ser

FRT

LoxP

5’ ARM 2.1 kb PGK-Neomycin 3’ARM 6.5 kb Exon 7

Exon 75’ ARM 3’ARM

1 2 1

Expected & other genotypes

5’ PGK-Neomycin 3’Exon 7

5’ PGK-Neomycin 3’Exon 7

Y329S-Neo

WT-Neo

Exon 75’ 3’

Y329S

Exon 75’ 3’

WT-lox

5’ 3’

Exon-7 del or D

Polyglucosan can be used in the liver during fasting

Polyglucosan can be used in the liver during fasting

Difference between Gbe1Y329S and Gbe1neo/neo mice

Gbe1+/+ Gbe1neo/neoPercent

changeGbe1ys/ys

Percent

change

Brain 230 ± 17 25 ± 8 11% 41± 9 n=4 17.8%

Heart 457 ± 56 72 ± 34 16% 92± 21 n=4 20.1%

Liver 855 ± 43 115 ± 42 13% 210± 35 n=4 24.6%

Muscle 251 ± 23 38 ± 18 15% ND n=4 ND

Gbe1Y329S mice live longer than Gbe1neo/neo mice

Gbe1Y329S mice has polyglucosan in the liver

PAS Diastase-PAS

Muscle

PAS Diastase-PAS

Brain30 days old mice

PAS Diastase-PAS

Cerebellum30 days old mice

PAS Diastase-PAS

Brain 4 month old mice

PAS Diastase-PAS

Brain 4 month old mice Gbe1ys-neo

PAS Diastase-PAS

High throughput screening

We have tested 1700 FDA approved compounds.

Analysis has yield 15 compounds decreasing PGB content in tissue culture

Validation of 15 HTS hits

0.0

1.0

2.0

3.0

4.0

5.01 2 3 4 5 6 7 8 9

10

11

12

13

14

15

Co

ntr

ol

Re

lati

ve G

lyco

gen

Co

nte

nt

(mic

rogr

am/m

g p

rote

in)

Treatment with Guaiacol

YS untreated

YS treated

PAS Diastase-PAS

Quantification of glycogen in tissues

Diagnosis of APBD

GBE enzyme assay

Histologic examination of sural nerve biopsy

Sequencing genomic DNA and messenger RNA.

Sequencing mRNA Proof of second genetic change

Search for the genetic change that eliminates the normal allele

1 to 6 5 to 10 9 to 14

12 to 15 14 to 16

C P1 P2 P3 C

C P1 P2 P3 C

THANK YOUMichio Hirano, MD Sergey Pompou, MDValentina Emmanuelle, MD Carmen Paradas, MD

Salvatore DiMauro, MDSindu Krishna, PhDPablo Abreu