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Sam Berns & Progeria: What a Rare Disease Teaches Us About a Universal Condition
Francis S. Collins, M.D., Ph.D.Director, National Institutes of Health
Thomas E. Cone Jr. History Lecture, 2015 AAP NCEOctober 24, 2015
Clinical Picture of Hutchinson-Gilford Progeria Syndrome General:
– Short stature– Weight for height
Skin: Subcutaneous fat Hair: Alopecia Head:
– Micrognathia– Prominent scalp veins
Teeth: Delayed and misaligned Limbs:
– Coxa valga– Joints prominent and stiff
Bones: Distal resorption (clavicles, phalanges) Cardiovascular: severe atherosclerosis
A “Silent” Mutation Causes Progeria
Eriksson, M. et al., Nature (2003)
Splice donor consensus TGG A
GA G T
Normal LMNA sequence
Mutation GGC=>GGT(G608G)
G G T G G G C
G G T G G G T
3’UTR12
Normal splice: lamin A
Exon 11
progerin
Exon 10
*
*Glycine
N1 H1 N2 H2 N3 N4 HeLa no-RNA
3’UTR12
Normal: lamin A
Exon 11
progerin
Exon1--10
lamin Aprogerin
Forward primer
Reverse primer
N1-4: normal H1-2: HGPS
RT-PCR Demonstrates Abnormal Splice
Goldman, R.D. et al., Proc. Natl. Acad. Sci. USA (2004)
Lamin AProgerinLamin C
Phenotypes in Interphase HGPS Cells
Capell, B.C. et al., Proc. Natl. Acad. Sci. USA (2005)
Patient Fibroblasts: In vitro 72 Hour FTI Treatment
Mouse Model of Progeria:A G608G BAC Transgenic
G608G
*P RAB25P (c1orf5)SSR2 P (FLJ12287)
164.4 kb
25.4
13.5
22.2 (24.1)
LMNA44.3
9.33.717.59.5 (11.9)
15.2
lumen
Wild type
Progeriatransgenic
Progeriatransgenic
+ FTI
G608G BACtransgenic
showssevere lossof vascular
smooth muscle cellsin aorta at
nine months
lumen
Farnesyl-transferaseinhibitorsprevent damage
lumen
Therapeutic Strategies for Progeriaand Other Laminopathies
Small Molecule Drugs– Reduce progerin production
• Farnesyltransferase inhibitors
– Increase progerin clearance• mTOR inhibitors
RNA Therapy– Block abnormal splice
• Morpholinos
Therapeutic Strategies for Progeriaand Other Laminopathies
Small Molecule Drugs– Reduce progerin production
• Farnesyltransferase inhibitors
– Increase progerin clearance• mTOR inhibitors
RNA Therapy– Block abnormal splice
• Morpholinos
Rapamycin improves nuclear blebbing in HGPS cells
24
What about other laminopathies?
Cao et al., Science Translational Med (2011)
Next Steps for mTOR Inhibition Strategy
Trial in G608G mouse model vs. wt controls– Everolimus (mTOR inhibitor)– Lonafarnib (farnesyltransferase inhibitor)– Both drugs– Neither drug
Cross breed progeria mouse with mTOR ∆/∆ Consider human trial of lonafarnib plus everolimus
Therapeutic Strategies for Progeriaand Other Laminopathies
Small Molecule Drugs– Reduce progerin production
• Farnesyltransferase inhibitors
– Increase progerin clearance• mTOR inhibitors
RNA Therapy– Block abnormal splice
• Morpholinos
Can you deliver morpholinos to vascular smooth muscle? Sarepta eGFP tester mouse
27
Transgene only produces eGFP if morpholinoreaches tissue and blocks intronic splice donor
Morpholinoblocks splice donor
SA SD
Intravenous morpholino achieves good delivery to vascular smooth muscle cells of mouse aorta
28
vehicle only morpholino
Therapeutic Strategies for Progeriaand Other Laminopathies
Small Molecule Drugs– Reduce progerin production
• Farnesyltransferase inhibitors
– Increase progerin clearance• mTOR inhibitors
RNA Therapy– Block abnormal splice
• Morpholinos
Lots More Ideas!!
N1 H1 N2 H2 N3 N4 HeLa no-RNA
3’UTR12
Normal: lamin A
Exon 11
progerin
Exon1--10
lamin Aprogerin
Forward primer
Reverse primer
N1-4: normal H1-2: HGPS
RT-PCR Demonstrates Abnormal Splice
Major questions about role of progerin in cellular senescence
What else do we know about molecular events that lead to senescence?– Shortening of telomeres at the end of chromosomes
Does shortening of telomeres correlate with activation of progerin prodution?– YES
If telomeres are prevented from shortening (by activating telomerase), does progerin production cease?– YES
If telomeres are “uncapped” in early passage cells, does that activate progerin production?
TRF2 is a dominant negative shelterinprotein
Progerin production is upregulated in normal human cells with uncapped telomeres
Next Steps for pf/pf Mice
Detailed assessment of phenotype Assess potential for extended longevity Passage MEFs and/or skin fibroblasts repeatedly to
assess timing of senescence Compare pf/pf and wt gene expression in multiple
tissues Determine if pf/pf MEFs are protected from senescence
after telomere uncapping Determine if pf/pf background alters phenotype of
telomere KO mice
Nature is nowhere accustomed more openly to display her secret mysteries than in cases where she shows traces of her workings apart from the beaten path; nor is there any better way to advance the proper practice of medicine than to give our minds to the discovery of the usual law of nature, by the careful investigation of cases of rarer forms of disease.
~ William Harvey, Letter IX, to John Vlackveld, 24 Apr 1657
Rare Diseases: Window on Nature?
Collins LaboratoryKan CaoCecie BlairBrian CapellAmanda DuboseMike ErdosMaria ErikssonDina FaddahStephen LichtensteinNoreen PetrashUrraca TavarezMelissa TruongRenee Varga
SareptaRyszard KoleDan MourichStacy CrumleyVictoria Philbin
Progeria Res. Fndn.Leslie GordonAudrey Gordon
Dana-FarberMark Kieran
Boston Children’sMonica Kleinman
Columbia UniversityKarima Djabali
Northwestern Univ.Bob Goldman
Mass GeneralDimitri Krainc
Progeria Research Team