1. Down Syndrome: A Complex Disease
Dr Una Fairbrother

2. Down Syndrome (DS) Affects one in about 700-1000 newborns
Most significant genetic cause of mild to moderate mental retardation
Due to alterations in neural development.

3. Trisomy 21 or DS
Only autosomal aneuploidy not lethal in the fetal or early postnatal period
DS phenotypes show variable penetrance, affecting many different organs

4. Clinical features of neonates with DS
Flat facial profile, 90%
Hypotonia, 80%
Poor moro reflex, 85%
Hyperflexibility of joints, 80%
Excess skin on back of neck, 80%
Slanted palpebral fissures, 80%
Pelvic dysplasia, 70%
Anomalous ears, 60%
Dysplasia of mid joint of 5th finger, 60%
Simian crease, 45%

5. Down Syndrome – Disease Phenotype
Mental retardation,
Congenital malformations of the heart
and gastrointestinal tract,
Duodenal stenosis or atresia,
Imperforate anus,
Hirschsprung disease,
Leukemia,
Hearing loss,
Alzheimer disease
and others

6. Human Chromosome 21 Many other disorders involving Chr21 genes
165 hits on OMIM including:
Familial amyotrophic lateral sclerosis (SOD1 gene)
Bethlem myopathy (COL6A1, A2 and A3)
Acute myeloid leukemia (AML1)

7. Chromosome 21 Structure
Length of the short arm can vary greatly between individuals
First example of large genomic region that can expand or contract on a scale of many mega bases
Long arm: 225 genes originally identified (545 on chromosome 22)

8. How Does Trisomy Arise?
95% of DS arises from non-disjunction of homologous chromosomes during 1st meiotic division in oogenesis
Homogeneity means that phenotype severity cannot be predicted from the karyotype
Need to understand the pathology and identify candidate genes

9. Down Syndrome Research
Functions of most of the Chromosome 21 genes are largely unknown
As is their contribution, to the DS phenotype.

10. Trisomy and Disease
How does an extra copy of all or part of Chr21 result in the DS phenotype?
Specific case of the more general problem - how does chromosomal imbalance produce abnormalities in morphology and function?
No single mechanism explains the consequences
No simple solution to counteract its phenotypic impact.

11. Trisomy and Disease: Theories
Amplified developmental instability hypothesis:
DS phenotype due to non-specific disturbance of chr balance

12. Or… Gene dosage hypothesis:
DS due to cumulative effect of overexpression of specific chr 21 gene products
Corroborated experimentally in mouse models
Some genes undergo dosage compensation
Stage and tissue specific

13. Or Interaction effects
Through interaction of Chr21 genes products with:
each other
the whole genome
transcriptome
Proteome
Possibly through trans acting effects

14.  A proposed unifying model for the genetic mechanisms acting in autosomal trisomy
Fitzpatrick (2005) TIGS 21:

15. Mouse Models
useful to elucidate contribution of specific genes to the DS phenotype.
Strategies:
assess single-gene by overexpressing single or combinations of genes
assess effects of overexpressing large foreign DNA pieces, (YACs or BACs) in transgenic mice
mouse trisomies that carry all (usually lethal) or part of MuChr16, which has regions of conserved homology with HuChr21.

16. Down Syndrome Critical Regions (DSCRs)
Loci in the DS critical regions (DSCRs) may have major effect
Approach limited by high phenotypic variation among DS individuals
To date little evidence of association between any particular phenotypic trait and overexpression of a specific Chr21 gene.

17. Postgenomics
Even knowing the molecular defect, it is difficult to decipher the complex pathophysiology of the disease,
Developmental consequences of the trisomy
Impact on behaviour and cognitive function.
Facing a new era -
postgenomics where the goal is to identify protein function and physiological role of gene products.

18. Congenital Heart Defects and DS
40% of trisomy 21 individuals have CHDs
incidence in the euploid population, 0.8%

19. Frequency of types of CHD in Down Syndrome

20. CHD Schematic Endocardial cushion VSD Atrium Ventricle septa Heart ASD
AVSD

21. Trisomy 21 and CHD
If CHD phenotype is caused by overexpression of a gene/s on chromosome 21 then
why do not all individuals with DS have a CHD?
May be related to specific alleles and their protein products, which function well in the diploid but not in the trisomic state.

22. CHD Critical Regions on Chr21

23. How Might haplotypic diversity Cause CHD in DS?
Variants interact indirectly/ directly with other loci modulating phenotypes
Functionality of gene drops below critical levels, endocardial cushions fail to fuse
Hypothesis: combinations of mutations embedded in particular haplotypes, in trisomic individuals, disturb the supramolecular structure of a vital protein and modulate the predisposition of an individual to a single or several types of CHD.

24. Do all CHD arise from the same mechanism?
4 CHDs in individuals+ DS, related but do not occur at same time in gestation
Counter-intuitive that a single, simple mechanism is responsible for all CHDs
Variants may interact indirectly/directly with other loci to modulate varying phenotypes
(compound?) heterozygosity for rare, functionally deficient alleles, may cause analogous CHD in non-syndromic, euploid individuals.

25. Heterotrisomy, a significant contributing factor to ventricular septal defect associated with Down syndrome?
Baptista MJ, Fairbrother UL, Howard CM, Farrer MJ, Davies GE, Trikka D, Maratou K, Redington A, Greve G, Njolstad PR, Kessling AM. Hum Genet :
Heterotrisomy for a gene or genes in a narrowed region region is a contributing factor to the pathogenesis of VSD in trisomy 21
Either through the presence of three different specific alleles or through the presence of specific combinations of alleles.

26. Down syndrome congenital heart disease: a narrowed region and a candidate gene.
Barlow GM, Chen XN, Shi ZY, Lyons GE, Kurnit DM, Celle L, Spinner NB, Zackai E, Pettenati MJ, Van Riper AJ, Vekemans MJ, Mjaatvedt CH, Korenberg JR.Genet Med :
These data suggest that the presence of three copies of gene(s) from the region is sufficient for the production of subsets of DS-CHD.
Propose DSCAM as a candidate for VSD.
Encodes a cell adhesion molecule, spans more than 840 kb of the candidate region, and is expressed in the heart during cardiac development.

27. Nuclear factor of activated T cells or DSCR1
DSCR1 gene expression is dependent on NFATc1 during cardiac valve formation and colocalizes with anomalous organ development in trisomy 16 mice.
Lange AW, Molkentin JD, Yutzey KE Dev Biol ;266(2):
Nuclear factor of activated T cells or DSCR1
regulatory protein in calcineurin/NFAT signal transduction pathway.
expressed during valvuloseptal development- in the endocardium of the developing atrioventricular and semilunar valves, the muscular interventricular septum, and the ventricular myocardium.
Candidate for abnormal development of heart in DS

28. Cell type-specific over-expression of chromosome 21 genes in fibroblasts and fetal hearts with trisomy 21.
Li CM, Guo M, Salas M, Schupf N, Silverman W, Zigman WB, Husain S, Warburton D, Thaker H, Tycko B BMC Med Genet ;7:24.
Expression microarrays
Expression differed across cell/tissue types.
MX1 up in senescent +21 fibroblasts. (MX1 is an interferon target)
Interferon signaling a candidate pathway for senescence
GART up in fetal hearts with +21
Abnormal purine metabolism may have a role in cardiac defects.

29. Complex disease
non-mendelian- familial aggregation, but no clear segregation.
suggests causative agents and physiological mechanisms; evolution, development, homeostatic processes.
generally more frequent than single-gene disorders.
single-gene disorders and complex diseases have multiple genetic, developmental, and environmental factors. In single-gene disorders, one gene has a pronounced effect in producing the phenotype.
Complex diseases is the major contributor to morbidity/mortality in developed countries.
Single-gene disorders are a smaller burden of disease and death than do complex diseases.