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PXE Research Update Patrick F. Terry

President PXE International, Inc.

January 14, 2009 1

Webinar Instructions   Slides are Numbered. 1, 2, 3…

  Talk: ~ 45 Minutes   Questions: Ask questions by typing your

question “Questions Box” during the presentation and it will be answered as I go.

  Concerns: Use the “Questions Box”

  Questions & Answers: We will have time at the end for a discussion.

  No Medical Advice. I’m Not a Doctor!!!   More Info at the End. Stay on the line.

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Presentation   Background & Overview   Basic Genetics   Genetics of PXE   PXE Animal Models   Next Steps Toward an Intervention:

  Diagnostic Test   Current Treatments   Therapy Approach   Drug Development

  Human Clinical Trials 3

1995 4

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TM

Genomic Health, Inc. Redwood City California

www.GenomicHealth.com NASDAQ: GHDX

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Patient Managed Research Foundation Not for Profit Patient Support Organization

501(c)3 Foundation

Fir

ew

all

PXE International

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PXE International BioBank

Founded, Supported, & Managed Since 1995 9

PXE Gene

Therapeutic Intervention

Genomic SNP/rSNPs

Proteomics

Regulomics

Cellular Modeling

Model Organisms Protein

Production Rational

Intervention Design Experimental

Assays

In vivo Studies

Delivery Vectors

Protein Structure/Function

Molecular Pathway Genotype / Phenotype

Associations

Accurate Correlative

Linkage

Phenotype Severity

Prediction

Epidemiological Research

Sequence Analysis

Code Sequence Cause Mutations Diagnostics

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PXE Around the World = PXE Patients

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PXE ASSOCIATED RESEARCH

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Benefit Sharing - Academics

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Bio-informatics

Technical Equipment

Training / Staffing & Consumables

Funds & Publicatio

ns

Genetics

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Male Genome 17

Haploid Genome 18

PXE International

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PXE International

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Recessive Disease

ATTAGATCTATGCTATAGGCGT

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PXE Gene Found

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About 10 years ago, several groups had used a candidate gene approach in an attempt to identify gene mutations responsible for the PXE phenotype. The failure of this approach prompted several investigators to consider a systematic genome search for the PXE gene (Struk et al., 1997, van Soest et al.,1997). These studies both reported a single locus at 16p13.1. 5cM region. The locus was refined to an 820-kb region containing 6 candidate genes. (1998) The exclusion of five of these genes and the identification of the first mutations responsible for the development of PXE in a gene encoding a protein associated with multi-drug resistance-associated Protein (ABCC6). (2000)

Pseudoxanthoma Elasticum

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PXE International 24

Complex Inheritance Patterns Even though the first description of the disease was performed over a century ago (Darier, 1896), most cases of PXE have been reported in just the last two decades. In the majority of cases, PXE appears to be inherited as an autosomal recessive (AR) or sporadic means. Mild manifestations of PXE have also been found in known carriers in recessive families (Bachelli et al, 1990) suggesting that heterozygote (haplo-insufficiency) individuals might be at risk particularly for the development of certain signs. The overall incidence of PXE in North America and Europe has been reported to be between 1:25,000 to 1:100,000 live births.

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ABC Transporters

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ATP-binding cassette transporters (ABC-transporter)

Atp Binding Cassette = ABC

DNA super-family that is the largest and most ancient family with examples from prokaryotes (simple bacteria), plants, to humans.

These are transmembrane proteins that function in the transport of a wide variety of substances across extra- and intracellular membranes, including metabolic products, lipids, sterols, and drugs.

ABC transporters are involved in tumor drug resistance, cystic fibrosis, bacterial multidrug resistance, pseudoxanthoma elasticum, and a wide range of other inherited human diseases.

ABCC6 / MRP6

ATP ATP

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The WAVE® System

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NH2

COOH

ABCC6 Mutations Hundreds of Alterations in the Gene/Protein

Extracellular

Intracellular

R1114C T1130M G1133A R1138Q, R1138W A1139T

V1298F G1302R A1303P R1314W R1314Q L1335Q R1339H R1339C

I1342T Q1347H

A78T

E125K

A950T L946I T944I

G663C E669D Q698P L726P M751K G755R R760W R765Q D777N V787I

F568S

G992R

R391G K392N

R1221C, R1221H

L1226I

L463H

L495H

G1501S

G1481S

R600C

R881S

S535P

T364R

T811M

R382W

S317R

Membrane

R1164Q

R518Q

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FUNCTION

P. Terry PXE Int.

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Two

Piston

Pump

PXE International

ABCC10

ABCC11

ABCC12

ABCC13

ABCC6

Sub Family C

3 Billion Years of Evolution

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ABC - Human Diseases

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Disease Transporter Cancer ABCB1 (MDR1), ABCC1 (MRP1),

ABCG2 (MXR) Stargardt disease & AMD ABCA4 (ABCR)

Cystic fibrosis ABCC7 (CFTR) Tangier Disease and Familial HDL deficiency ABCA1 (ABC1) Progressive familial intrahepatic cholestasis ABCB11 (SPGP), ABCB4 (MDR2) Dubin-Johnson syndrome ABCC2 (MRP2) Pseudoxanthoma elasticum ABCC6 (MRP6) Persistent hypoglycemia of infancy ABCC8 (SUR1), ABCC9 (SUR2) Sideroblastic anemia and ataxia ABCB7 (ABC7) Adrenoleukodystrophy ABCD1 (ALD) Sitosterolemia ABCG5, ABCG8 Immune deficiency ABCB2 (Tap1), ABCB3 (Tap2) Lamellar Icthyosis ABCA12

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Genotype Phenotype

Project

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Results Thus Far… 1.  Gene Discovery 2.  Mutational Database 3.  Biorepository 4.  Patient Registry 5.  Genotype-Phenotype Program 6.  Rigorous Participant Protections 7.  IPR - Patent and Licensing 8.  Clinical Diagnostics 9.  Animal Models 10.  Functional Screening Assay 11. Clinical Biomarker Program 12.  Drug Discovery Program 13.  Human Clinical Trials 14.  Drug/Treatment Development 34

Research

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Human & Animal Disease 36

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Blood

Bile / Liver

ECM

To Blood ??? Peptides Enzymes Elastase Proteinase inhibitors

ABCC6

LIVER

FUNCTION

Elastin & Collagen Fibrils

Mineralization

ABCC6

Stieger, et al. ‘House Keeping Function’ Basal layer expression of ABCC6 is incorrect. Personal Communication

Peptide ?

PXE International

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Metabolism &

Liver Function

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Diagnostic Testing

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Human & Animal Intervention Approaches

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Drug Research

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Eye Intervention Treatments

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NIH Clinical Center

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Questions… •  Natural function & transported substrate? •  Liver function & metabolism •  ABCC6 tissue-based function characterization? •  Variable expressivity and penetrance (Animal & Human) •  Extreme & intermediate phenotypes •  Pathogenesis & secondary consequences? •  Correlations - genotype/phenotype, protein structure

modifications, gene modifiers / environmental factors •  Possible early disease interventions •  Human clinical intervention trials •  Clinical evidence for Avastin & Lucentis treatment

effectiveness 45

Current Projects •  Validated functional assay for natural compounds,

drugs & small molecule screening •  Non-coding & functional element analysis •  Additional gene(s) involvement •  Full mutational spectrum database •  Mouse model & clinical workup •  Fish model & clinical workup •  Liver transplant in mouse •  Liver transplant patient workup •  Systematic clinical phenotype workups •  Human intervention trial at NIH (2009)

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Exploratory Research

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•  PXE Specific Mineralization –  Phosphate –  Magnesium –  Vitamin K

•  ??? Bio-activity & Prohibition of Mineralization •  ??? Absorption and Transport (Liver & Kidney) •  ??? Tissue Stores and Distribution •  ??? Proximal & Distal w/Circulatory Exposure •  ??? Liver Metabolism Cycle

Dynamic Interplay

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Questions?