L’apparato visivo nei pazienti mitocondriali: le disfunzioni ed i trattamenti

Valerio Carelli, MD, PhD

IRCCS Institute of Neurological Sciences of Bologna, Bellaria Hospital, Bologna, Italy

Neurology Unit, Department of Biomedical and Neuromotor Sciences (DIBINEM), University of Bologna, Bologna, Italy

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LCUN SAGY

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ND5 ND6

ND4

ND4L

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COIII G

ATPase6/8 K COII D

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16S

12S D-loop Cyt.b

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3243A->G MELAS CPEO DDM

tRNA-Ile mutations CARDIOPATHY

8344A->G MERRF

tRNA-Ser mutations DEAFNESS

1555A->G DEAFNESS (aminoglycosides)

cyt. b mutations MYOGLOBINURIC MYOPATHY

8993T->G NARP/MILS

3460G-A 11778G->A 14484T->C LHON

PEO, KSS, Pearson

Leber Hereditary Optic Neuropathy (mtDNA)

Dominant optic atrophy (OPA1) Recessive optic atrophy X-linked optic atrophy

Mito-pathways to RGCs loss OXPHOS deficiency (complex I) Reactive oxygen species (ROS) – calcium handling (mito-ER) Mito-dynamics-biogenesis-mitophagy Mitochondrial apoptosis

NATURAL HISTORY OF LHON Barboni et al., Ophthalmology 2010

A specific pattern of neurodegeneration

The pattern of neuro-degeneration follows the axonal diameter starting from the

smallest fibers on the temporal quadrant(papillo-macular bundle)

The smallest fibers are the most vulnerable in maintaining the energy supply because their volume is limited in accommodating

more mitochondria to compensate

CONTROL LHON/3460

A B C

D E F

Abnormal myelin maintenance

10 µm

10 µm

A B C

D E F

A B

What is the role of myelin maintenance in triggering RGCs loss or promoting RGCs sparing and function?

-Axonal swelling, -Increase of abnormal mitochondria -Thinning of myelin

-Preferential loss of smallest axons

Mouse model of OPA1 haploinsufficiency (Prof. Luca Scorrano)

COMMON THEMES LINKING RGC LOSS IN MITOCHONDRIAL OPTIC NEUROPATHIES

mtDNA mutations: -OXPHOS (complex I) deficiency -increased ROS -mito-fragmentation -activated biogenesis/mitophagy -myelin pathology -vascular changes

nDNA mutations: -OXPHOS (complex I/supercomplex) deficiency -increased ROS -mito-fragmentation -activated biogenesis/mitophagy -myelin pathology

pathologic axonal transport and ultimately APOPTOSIS of RGCs

Mitochondrial optic neuropathies are a convenient model for clinical trials: -only one tissue affected -fairly well understood pathogenesis and relatively predictable natural history -good quantitative outcome measures (visual acuity, visual fields, OCT assessments of retinal fiber layer thickness, etc.) -animal models available (OPA1 and ND6 mutants)

THERAPY FOR LHON and DOA

• Antioxidant - Idebenone and EPI-743 in affected • Activation of mitochondrial biogenesis for carriers • Anti-apoptotic drugs for acute phase • Gene therapy for LHON and NARP: allotopic

expression • Gene therapy: xenotopic expression of alternative

oxidase (S. Cervisiae single subunit NADH oxidase Ndi1)

• Gene therapy for DOA: in haploinsufficiency, AAV-dependent delivery of OPA1

Idebenone in LHON (and DOA)

New LHON trial with idebenone in UK (Dr. Patrick Yu-Wai-Mann), with high dosages and for a longer time!

EPI-743

*Eye from this patient was excluded #Eye did not experience vision loss

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Figure 1. Mean deviation, logMar visual acuity and OCT average retinal nerve fiber layer from the 1st affected eye – from baseline to 24 months of EPI-743 therapy

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Figure 3. Examples of Humphrey visual fields at baseline and after 24 months of treatment

Baseline 24 months after treatment A.

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Figure 4. Examples of color optic disc photographs at baseline and after 24 months of treatment.

Baseline 24 months after treatment A.

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RNFL thickness between treated and untreated LHON patients

Pretreatment 24-months 18-months 12-months

EPI-743

Untreated affected LHON

patients

97.8 64.3 65.0 67.8 50.4

Not statistically significant

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�Marked improvement � Mild improvement � No improvement

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24 months after treatment

RNFL swelling as a negative prognostic marker for LHON patients on therapy

*Definition of baseline: 6 months from vision loss

The greater the initial RNFL swelling, the worse the final outcome for the 1st eye after 24 months with EPI-743 treatment

THERAPY FOR LHON and DOA

• Antioxidant - Idebenone and EPI-743 in affected • Activation of mitochondrial biogenesis for carriers • Anti-apoptotic drugs for acute phase • Gene therapy for LHON and NARP: allotopic

expression • Gene therapy: xenotopic expression of alternative

oxidase (S. Cervisiae single subunit NADH oxidase Ndi1)

• Gene therapy for DOA: in haploinsufficiency, AAV-dependent delivery of OPA1

Females are partially protected in LHON

Oestrogens activate mito-biogenesis

Conclusions Oestrogens present a multilayer effect on complex I defective LHON cybrids, leading to: -reduced ROS production

-small but significant improvement in cybrids energetic competence -partial rescue of cell viability in galactose medium by restoring membrane potential and limiting apoptotic cell death -coordinated activation of mitochondrial biogenesis

THERAPY FOR LHON and DOA

• Antioxidant - Idebenone and EPI-743 in affected • Activation of mitochondrial biogenesis for carriers • Anti-apoptotic drugs for acute phase • Gene therapy for LHON and NARP: allotopic

expression • Gene therapy: xenotopic expression of alternative

oxidase (S. Cervisiae single subunit NADH oxidase Ndi1)

• Gene therapy for DOA: in haploinsufficiency, AAV-dependent delivery of OPA1

Dubbi sull’efficienza della metodica

Ancora risultati controversi

• We took advantage of cell models harboring the homoplasmic disruptive m.3571insC MTND1 mutation. We thereby demonstrated that allotopic expression of corrected mtDNA-encoded ND1 subunit complements and allows reassembly of human respiratory complex I along with recovery of bioenergetic deficiency, hence providing definitive proof that the allotopic expression is a feasible approach for gene therapy of mitochondrial diseases.

Strategie per la LHON • Terapia preventiva in fase pre-sintomatica:

attivatori della biogenesi mitocondriale nei carriers di mutazione non affetti e terapia genica

• Terapia in fase acuta: combinazione di farmaci

antiapoptotici (ciclosporina etc..?) e antiossidanti (idebenone e EPI-743) o terapia genica

• Terapia in fase cronica: antiossidanti (idebenone e

EPI-743) e attivatori della biogenesi mitocondriale e/o terapia genica

Thanks for the attention and thanks to the many collaborators: • Alfredo A. Sadun, USC, Los Angeles • Rubens Belfort Jr, UNIFESP, Sao Paolo • Carla Giordano, University “La Sapienza”, Rome • Palmiro Cantatore, University of Bari, Bari • Anna Ghelli, University of Bologna, Bologna • Pio D’Adamo, University of Trieste, Trieste • Vamsi Mootha, Harvard, Boston • Massimo Zeviani, MRC, Cambridge • Patrick Chinnery and Patrick Yu-Wai-Man, University of Newcastle, Newcastle • Piero Barboni, San Raffaele, Milano

• Lab of Neurogenetics: Alessandra Maresca, Leonardo Caporali, Sabrina Farne’, Chiara La Morgia, Maria Lucia Valentino

Melanopsin expressing retinal ganglion cells (mRGCs) survive in LHON and DOA

Melanopsin RGCs and conventional RGCs are loaded with mitochondria

mRGC

mRGC

RGC

RGC

RGC

RGC

Courtesy Fred Ross Cisneros and Carla Giordano