2. Diseases caused by mtDNA mutations
There are more than 700 known disease-associated mtDNA mutations
(mitomap.org):
- 285 tRNA/rRNA
- 266 protein coding and control region point mutations;
- 131 deletions
Acquired, age related - neurodegenerative diseases, Parkinson, heart
diseases, diabetes, cancer
Inherited - neuropathy, encephalopathy, cardiomyopathy, myopathy,
diabetes, metabolic syndromes
Up to 4,000 children are born in the United States every year with inherited
mtDNA syndromes
2
3. Complex nature of mtDNA genetics and inheritance
Leber’s hereditary optic neuropathy (LHON)
44% 25%
2% 85% 15% 52%
0% 2%
I
II
III
IV
3
4. Inherited mtDNA diseases
mtDNA is maternally inherited - through the egg
Complex, unpredictable pattern of inheritance
These diseases are fatal or severely debilitating
No cure for mtDNA disease
Ultimate goal is to prevent transmission of mtDNA disorder
s by replacement of mutated genes in eggs
4
5. Mitochondrial Gene Replacement in Oocytes
Complete replacement of entire mtDNA
Applicable to any mtDNA mutation type
Eliminates entire spectrum of mtDNA disease
Genetic corrections will be heritable and passed on to
later generations
Prevents the need for repeated therapy generation after gene
ration
5
6. mtDNA replacement in oocytes
Feasibility and efficacy of MII spindle-chromosome
complex transfer (ST)
Developmental Potential
Mutated mtDNA carryover
Nuclear/Mitochondrial genome compatibility?
6
7. Mitochondrial gene replacement in oocytes
Spindle imaging
Separated chromosomes (nuclear DNA) and
mitochondrial DNA
Distribution of mitochondria
in mature oocytes
Spindle removal 7
12. Normal growth and development of monkey offspring
following mtDNA replacement
Tachibana et al., Nature 2013
12
13. • 7 egg donors
• A total of 106
mature MII oocytes
used for ST or
served as controls
13
14. mtDNA replacement by Spindle Transfer (ST) in human
oocytes: efficacy, fertilization and embryo development
Tachibana et al., Nature 2013
14
15. Fertilization outcomes in human zygotes following mtDNA
replacement
15
Tachibana et al., Nature 2013
16. ESC lines from human ST and control embryos
5 ESC lines from 13 human ST blastocysts (38%)
contained normal euploid karyotypes
mtDNA carryover 1% or lower
1 ESC line from 6 abnormally fertilized ST blastocysts
(17%) was triploid
9 ESC lines from 16 control blastocysts (56%), 2 cell lines
were also karyotypically abnormal (XYY or X0)
16
17. Conclusions
Entire cytoplasm containing mtDNA in human oocytes can be
efficiently replaced by ST
Use of mt genome from donor egg (not recombinant)
Applicable to any mtDNA mutation type
ST is feasible with cryopreserved eggs
A portion of manipulated oocytes displayed abnormal fertilization
Normally fertilized zygotes develop to blastocysts and produce
karyotypically normal ESCs at rates similar to controls
Thorough screening for abnormal fertilization is critical for selecting
ST embryos for transfers
17
18. Clinical Trials
Current efficiency allows generation of several (3-4) healthy embryos
by ST suitable for embryo transfers for each cycle
Recruit families –carriers of early onset mtDNA diseases (at least one
affected child, living or deceased)
Recruit healthy mtDNA egg donors
Conduct ST followed by PGD and/or prenatal diagnosis to ensure
complete mtDNA replacement and chromosomal normalcy
Follow up with birth and development of healthy children
18
