Genetics of Vascular Diseases Jake Lusis 3-730 MRL [email protected].

Genetics of Vascular Diseases

Jake Lusis3-730 MRL

[email protected]

210

410 200 210 180 430

420190460490

Carl Miller, ~1938, Oslo County Hospital, Norway

Inheritance of:

Cholesterol level (mg/dl):

Tendon xanthomas:

Early coronary artery disease (under 55 years of age):

330 210

Tendon Xanthomas

Cosegregation of triad of xanthomas, high cholesterol and early heart disease

210190 400

210 430 490420 180

170420

Inheritance of:

Cholesterol level (mg/dl):

Tendon xanthomas:

Early coronary artery disease (under 55 years of age):

210 230

810 420

This individual died from severe CAD at age 7

Lecture delivered in Sweden Dec 9, 1985 when they received the Nobel Prize in Physiology. Published in

Science, 232, 34-47, 1986

Experiments with 125I-LDL and skin fibroblasts

skin fibroblasts from normal and FH individuals

add labeled 125I-LDL ( )

incubate to allow binding

wash away unbound LDL and follow label

- 125I-LDL on surface can be displaced from cells by anionic polymers

- 125I-LDL that has been internalized cannot be displaced from cells

- 125I-LDL that has been degraded to amino acids is soluble in tricholoroacetic acid

FH homozygote

Normal

Experiments with fibroblasts from normal and FH individuals

Sequential steps in LDL receptor pathway: Discovery of receptor mediated endocytosis

Degradation of LDL by lymphocytes and following injection

Cloning of the Human LDL Receptor

cultured skin fibroblastsisolate plasma membrane

purify such that a single protein remains upon electrophoresis

extract proteins with detergent and subject to chromatography

125I-LDL bound

digest the protein and sequence the resulting

peptides

-ASN-PHE-THR-SER-based on the amino acid

sequence, synthesize corresponding oligonucleotide

probes

….CACTCGACAGCGAAC...

probe cDNA and genomic libraries.

Confirm identity of clones by sequencing

and expression studies

Domain structure of the LDL receptor

Five classes of LDL receptor mutations

Development of drugs to treat hypercholesterolemia based on concepts learned from FH studies

Combined heart-liver transplant in an individual with homozygous FH

Lessons from familial hypercholesterolemia

• Studies of an uncommon disease (FH) can have important implications for a common disease (coronary artery disease)

• Genetics studies of clinical traits can lead to an understanding of basic biologic mechanisms (receptor mediated endocytosis, cholesterol homeostasis) as well as new therapies (statins).

Francis Collins (1995)

Schematic of the approach generally utilized for positional cloningSchematic of the approach generally utilized for positional cloning

FamiliesFamilies PhysicalPhysical

MappingMapping

andand

CloningCloning

FinerFiner

GeneticGenetic

MappingMappingLinkageLinkage

AnalysisAnalysis++

TranscriptTranscript

IdentificationIdentification

CandidateCandidatecDNAscDNAs

MutationMutation

SearchSearch

..A A C T C..A A C T C

..A A G T C..A A G T C

BACsBACs

NormalNormal

MutationMutation

GeneticGeneticMarkersMarkers

Marfan Syndrome -autosomal dominant disorder characterized by weakened connective tissue -most life-treatening is aortic dissection (an aneurysm of the aorta) -other symptoms include mitral valve prolapse, emphysema-like lung disease, and ocular and skin disorders -patients tend to be tall and lanky, with very long limbs, fingers and toes

Fibrillin 1 Gene Mutated in Marfan Syndrome Patients(A)Single-strand conformational polymorphism screen(B) DNA sequence

Letter

Nature Genetics  33, 407 - 411 (2003) Published online: 24 February 2003;

Dysregulation of TGF-   activation contributes to pathogenesis in Marfan syndrome

Enid R. Neptune1, 2, Pamela A. Frischmeyer2, Dan E. Arking2, Loretha Myers2, Tracie E. Bunton3, Barbara Gayraud4, Francesco Ramirez4, Lynn Y. Sakai5 & Harry C. Dietz2, 6

Lung histopathology and morphometry of mice deficient in fibrillin-1.

                Figure 3

Active TGF-B expression and signaling in lung tissue of mice deficient in fibrillin-1.

Neutralizing antibody to TGF- rescues lung maturation in mice deficient in fibrillin-1.

Mice deficient in fibrillin-1 have preserved lung cell proliferation but greater apoptosis.

Model

Fibrillin deficiency

TGF not sequestered

TGFB activated

Apoptosis in lung, bone overgrowth, etc.

A lesson: Structural matrix elements serve crucial regulatory roles in cytokine activation and signalling

Identification of Mendelian and complex trait genes 1980-2002

0

200

400

600

800

1000

1200

1400

1600

1800

1980 1985 1990 1995 2000

Year

Number of Mendelian genes

0

10

20

30

40

50

60

70

80

90

100

Number of complex trait genes

Mendelian traits

All complex traits

Human complex traits

Glazer, et al. (2002) Science

•Ian

Nature Genetics  34, 383 - 394 (2003)

VEGF is a modifier of amyotrophic lateral sclerosis in mice and humans and protects motoneurons against ischemic deathDiether Lambrechts1, Erik Storkebaum1, Masafumi Morimoto1, Jurgen Del-Favero2, Frederik Desmet1, Stefan L Marklund3, Sabine Wyns1, Vincent Thijs1, Jörgen Andersson3, Ingrid van Marion4, Ammar Al-Chalabi5, Stephanie Bornes6, Rhiannon Musson4, Valerie Hansen5, Lars Beckman3, Rolf Adolfsson7, Hardev Singh Pall4, Hervé Prats6, Severine Vermeire1, Paul Rutgeerts1, Shigehiro Katayama8, Takuya Awata8, Nigel Leigh5, Loïc Lang-Lazdunski9, Mieke Dewerchin1, Christopher Shaw5, Lieve Moons1, Robert Vlietinck1, 10, Karen E Morrison4, Wim Robberecht1, Christine Van Broeckhoven2, Désiré Collen1, Peter M Andersen3 & Peter Carmeliet1

VEGF Polymorphisms

Lesson

Basic vascular biology studies (regulation of VEGF expression in response to hypoxia) can lead to an understanding of a complex disease