CAG Expansion and Huntingtons Disease

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CAG Expansion and Huntingtons Disease -

• Could you repeat that, please?
• A research review on Huntingtons disease

Image source http//www.vh.org/Providers/Textbook
s/BrainAnatomy/Ch5Text/Section01.html
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Heres the Plan

• Introduction
• Clinical symptoms
• Pathology
• Molecular basis
• Current research
• Treatment ideas
• Future plans
• Conclusions

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Huntingtons Disease
Introduction

• Dr. George Sumner Huntington
• Published paper in 1872
• Patients of East Hampton, Long Island

http//www.uic.edu/depts/mcne/founders/page0048.ht
ml
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Check Out My Figures
Introduction

• Affects 30,000 people in the U.S.
• Frequency of about 1 in 10,000
• 150,000 may develop the disorder
• Strikes both men and women
• 10 of the cases are children

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Symptoms A Threefold Attack
Symptoms

• Three stages of progression
• Three areas affected
• Mental Dementia
• Emotional Mood swings
• Physical Chorea, unsteady gait, weight loss

http//www.nlm.nih.gov/medlineplus/ency/article/00
3199.htm
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Pathology The Wreckage of HD
Pathology

• General loss of brain mass
• Specific neuronal loss in the striatum (caudate
  nucleus and the putamen) and the cerebral cortex

DebBurman, S.K.
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Damage Report
Pathology

• Inhibitory and excitatory neurons
• Loss of inhibitory control
• Manifested in chorea

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CULPRIT Huntingtin Gene its Mutation
Molecular Basis

• Normal huntingtin protein - vital, but function
  unknown
• Gene identified as IT15
• Chromosome location 4p16.3
• Dynamic mutation
• Expanded CAG repeat

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How mutation happens Expanded CAG Repeat
Sequence
Molecular Basis
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Biological ModelMutant Huntingtin causes disease
Molecular Basis
Huntingtin Gene IT15
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Longer the repeat, earlier the onset
Molecular Basis
Persichetti et al., 1994
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Paternal Anticipation
Molecular Basis

• Longer CAG repeats are more unstable
• Expansions occur more when passed through the
  male germ line
• Meiotic transmission may cause the CAG expansion
• Age of onset decreases with the increasing number
  of generations
• Paternal anticipation is not fully understood

Decreasing Age of Onset
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Conclusions
Molecular Basis

• A dynamic mutation results in the CAG repeat
• The mutation may be caused by DNA polymerase
  slippage
• Inverse relation between length of CAG repeat
  and age at death
• Paternal Anticipation results in longer CAG
  repeat and earlier age of onset

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Let Us Demonstrate
Demonstration

• An example of a sequence specific amplification

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Gaps in Knowledge in 1997

• How does mutant huntingtin protein misfold?
• What kind of cell death takes place?
• How does misfolded huntingtin cause cell death?
• Who talks with huntingtin in health?
• How does that change in disease?
• How can we prevent HD?

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1. How does Huntingtin misfold?Discovery of
Neuronal Intranuclear Inclusions (NIIs)
Current Research

• N-terminus of htt accumulates in neuronal nuclei
• PolyQ repeat leads to the formation of insoluble
  ?-sheets or barrels
• Role of NIIs in HD pathology is presently unclear

DiFiglia, 1997
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Causal Role Of NIIs?
Current Research

• NIIs are hallmarks of polyQ diseases
• Observed in the striatum and cortex
• Appear before onset of HD
• Ubiquitinated
• Reduction NIIs decreases in vitro cell death

Saudou, 1998
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The Plot Thickens Are NIIs Helpful or
Deleterious?
Current Research

• htt induces cell death
• Reduction in NII formation shows increased death
  from HD
• NIIs a possible protection mechanism?

Saudou, 1998
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Presently the role of NIIs in HD is unclear
Current Research
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2. How Do Neurons Die?
Current Research

• Apoptosis - programed cell death
• Non-apoptotic model - role for NIIs
• Excitotoxicity - death by over-excitement

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A. Role of Caspases in apoptosis
Current Research

• Degrade apoptotic cells
• Triggers activation of other caspases
• Overactivation of caspases in HD

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B. A Non-Apoptotic Model
Current Research
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C. Excitotoxicity
Current Research

• High glutamate levels excite some neurons to
  death
• Expected increased cell death in HD mice
• Surprise! HD mice are resistant to the addition
  of quinolinic acid

Hansson et al, 1999
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So Which One is It?
Current Research

• No one knows!
• Contradictory evidence from different
  laboratories and many unanswered questions
• Still investigating each mechanism

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Who does huntingtin talk with?Healthy
pathogenic Protein Interactions
Current Research

• Molecular Chaperones Transcription Factors

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Chaperones Fold With Me
Current Research

• Yeast model
• Different levels of polyQ
• Ubiquitination?
• Still aggregation

http//www.letterset.com/ufo.htm
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Keep Foldin, Foldin, Foldin
Current Research

• Overexpression or deletion of HSPs
• Increased HSPs, increased aggregation
• Deletion of HSP104 - NO AGGREGATION

http//www.ccc.nottingham.ac.uk/mbzspd/tutorials/
CSC.htm
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Additional Folding Models
Current Research

• Mammalian model
• HSPs increase aggregation
• Worm model
• _ NIIs move to cytoplasm with more Hsps
• Fly model
• Hsp70 and Hsp40 together Inhibit fibril
  (aggregate) formation

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Transcribe thisconnecting with p53
Current Research

• htt and p53
• p53 aggregation
• Repressed transcription
• Leads to Apoptosis?

Perutz, 1994
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What in the CA150??
Current Research

• Aggregation leads to
• Interfered transcription
• Essential protein synthesis disruption
• Cell death

• CA150
• Transcriptional co-activator
• Rich in Glu-Ala
• Found with ubiquitinated htt in inclusions
• binds preferentially with htt

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So
Current Research

• HSPs are essential in htt misfolding
• Possibility that the pathology involved htt
  aggregrating with transcription factors, which
  leads to cell death

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4. Therapeutic Possibilities
Treatment Ideas

• Addition of Minocycline
• Turn off promoter to prevent htt translation
• Fetal striatal tissue transplant

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A. Minocycline Treatment
Treatment Ideas

• An antibiotic that can cross BBB
• Will inhibit caspases
• Delays disease progression
• No inhibitory effect on the nuclear inclusions

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B. Inhibiting transcription halts HD in
mice
Treatment Ideas

• Transgenic mice developed with the ability to
  inhibit htt expression with addition of
  doxycycline (DOX)

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Lose Your Inhibitions
Treatment Ideas

• Inhibition of htt expression allows for neuronal
  recovery
• Gene off mice show caudate regeneration

Yamamoto, 2000
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C. Transplanting the Seeds of Hope... with fetal
transplants
Treatment Ideas
Freeman, et al., 2000

• Fetal striatal transplants by Freeman et al.
• Used human subject
• Only 18 months of data collected
• Transplanted neurons unaffected

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The Underlying Theme
Conclusion

• The field of HD has become increasingly
  controversial at the molecular level
• Mechanisms are unknown even though progress has
  been substantial
• HD does not follow suit with the rest of the
  trinucleotide repeat diseases

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What Next?
Future Plans

• Combine treatments to target the different
  proposed mechanisms
• Continued research to determine a definitive
  mechanism
• Re-evaluate other trinucleotide repeat diseases

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We Wish We Knew...
Future Plans

• How are the symptoms and pathology interrelated?
• What is the exact mechanism of HD?
• The role of NIIs is it cause or effect?
• What causes paternal genetic anticipation?

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We Would Like to Thank
Acknowledgments

• Dr D
• Aash Bhatt
• Becky Bielang
• the TA staff
• Our Moms

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