IOO Genetics

LCA is an severe, early-onset form of retinal degeneration in which peripheral and night vision is lost from the first decade of life, followed in most cases by central, daytime vision. Although it is a rare condition, affecting around 1/80,000 people, LCA is thought to account for around 1/5 cases of childhood blindness and there is no treatment currently available.

We have developed gene therapy for LCA caused by damage to the RPE65 gene, which is found in the retinal pigment epithelium cells that support the light-sensitive photoreceptor cells of the retina. Mutations in RPE65 are thought to cause around 8% of LCA cases.

With proof-of-concept for RPE65 gene therapy established in models of disease, we are testing this therapy in our first-in-man clinical trial of gene therapy for LCA caused by mutations in the RPE65 gene. Results from this trial indicate that gene delivery using viruses is well tolerated and is able to improve vision (Bainbridge et al. NEJM 2011).

Further trials will help determine whether improvements to vector design can enhance the beneficial effect of RPE65 gene therapy on sight.

• Current clinical trial: Phase I/II trial to assess safety and efficacy of viral RPE65 gene therapy

• Key publications: Bainbridge et al. NEJM 2011, Annear et al. Gene Therapy 2010

  • More publications on gene and cell therapy for inherited sight loss
• Collaborator: Simon Petersen-Jones
• EyeTherapy information hub: Gene therapy clinical trial for LCA caused by damage to the RPE65 gene

LCA is an severe, early-onset form of retinal degeneration in which peripheral and night vision is lost from the first decade of life, followed in most cases by central, daytime vision. Although it is a rare condition, affecting around 1/80,000 people, LCA is thought to account for around 1/5 cases of childhood blindness and there is no treatment currently available.

We have shown that treating mice with sight loss caused by damage to the AIPL1 gene, by delivering a working copy of AIPL1, restores structure and function to both rod and cone photoreceptor cells. We have also demonstrated that this gene replacement therapy can prevent the loss of photoreceptor cells from the retina - this means that gene replacement therapy for AIPL1 is one of the most robust therapies to be tested in models of inherited retinal degenerations.

We are planning to start a clinical trial for patients with LCA caused by damage to the AIPL1 gene.

We are planning a clinical trial for patients with the AIPL1 form of LCA (changes to the retina in AIPL1 LCA are shown in the left panel), having shown that gene delivery using AAV restores vision in a model of disease (right panel shows that gene delivery restores AIPL1, coloured green, to the light-sensitive photoreceptor cells - which also correctly house the machinery needed to detect light, coloured red here)

• Selected publications: Tan et al Hum Mol Genet 2009, Sun et al Gene Therapy 2010, Tan et al PLoS One 2012

  • More publications on gene and cell therapy for inherited sight loss
• Collaborators: Tiansen Li, Dr Yvan Arsenijevic, Dr Alberto Auricchio

Achromatopsia is a rare, recessively inherited condition affecting around 1/30,000 people in which colour vision is completely lost, central detailed vision is distorted and patients experience extreme light sensitivity. Currently there is no treatment available although some symptoms may be lessened by wearing red contact lenses and dark wrap-around glasses.

Damage to the CNGB3 gene is associated with around 50% of achromatopsia cases. We have demonstrated that delivering a working copy of the CNGB3 gene can restore vision in a model of achromatopsia lacking CNGB3 - our study showed long-term restoration of sight and included improvements to the ability of treated mice to track a visual stimulus.

AAV gene therapy improves cone photoreceptor cell survival in model of achromatopsia caused by mutation in CNGB3. Compared with untreated (top right, bottom right), AAV.CNGB3 gene therapy restores CNGB3 protein (top left) and the light-sensitive opsin pigment (bottom left) to cone photoreceptor cells

We are planning to conduct a clinical trial testing the delivery of the CNGB3 gene to patients with achromatopsia using an AAV vector.

• Proof-of-concept publication: Carvalho et al Hum Mol Genet 2011

  • More publications on gene and cell therapy for inherited sight loss
• Collaborator: Xi-Qin Ding

Ongoing research shows that gene therapy in a mouse that lacks RDH12 can restore the biochemical pathway that is responsible for sight loss in this form of LCA.

We are planning a clinical trial to test the safety and efficacy of delivering the RDH12 gene to patients with LCA.

• Collaborator: Debra A. Thompson

X-linked RP is caused by damage to one of two genes; the form associated with mutations in the RPGR gene causes 14% of RP cases making it the commonest cause of retinitis pigmentosa. Patients usually experience loss of peripheral vision and reduced central vision by the age of 20, accompanied by significant changes to pigmentation in the retina that are characteristic of RP.

Damage to the RPGR gene causes X-linked RP, causing pigmentary changes in the retina (top panel) and loss of sight from an early age. AAV gene therapy rescues cone photoreceptors in a model of X-linked RP lacking RPGR, restoring RPGR protein (middle right panel) and cone opsin pigment (bottom right panel) compared with untreated (middle and bottom left panels)

With proof-of-concept for gene replacement being established in animal models of disease, we are now planning clinical trials to test the safety and efficacy of RPGR gene delivery using AAV in patients.

• Collaborators: Prof. Tiansen Li, Prof. Alan Wright
• Publications on gene and cell therapy for inherited sight loss

LCA is an severe, early-onset form of retinal degeneration in which peripheral and night vision is lost from the first decade of life, followed in most cases by central, daytime vision. Although it is a rare condition, affecting around 1/80,000 people, LCA is thought to account for around 1/5 cases of childhood blindness and there is no treatment currently available.

We have demonstrated long-term improvements in vision following the delivery of the RetGC-1 gene (also known as GUCY2D) using AAV in a model of LCA. Treatment leads to increased survival of cone photoreceptor cells and improvements in electrical activity of the retina as measured by ERG.

Gene therapy restores expression of RetGC-1 protein, shown here in green, in a model of LCA in which RetGC-1 is missing

We are in the early stages of planning a clinical trial for this condition, which would test the safety and efficacy of delivering the RetGC-1 gene to patients with LCA caused by damage to that gene.

• Proof-of-concept publication: Mihelec et al Hum Gen Ther 2011

  • More publications on gene and cell therapy for inherited sight loss

A disease such as RP is inherited dominantly when damage to just one copy of a gene is enough to significantly alter the function of cells and cause disease. In such cases gene replacement therapy is unlikely to provide benefit – we need to silence the mutant gene, and in most cases would need to supply a working copy of the gene that isn’t silenced alongside – this approach is known as ‘suppression-replacement therapy.’

RNA interference (RNAi) is a naturally-occurring process in which small RNA molecules silence specific genes by binding to them and initiating their breakdown by the cell.

We have shown that RNAi delivered by rAAV vectors can effectively silence the expression of Peripherin-2,  which is damaged in both autosomal dominant retinitis pigmentosa and dominant maculopathies.

RNA interference can silence the Peripherin-2 gene (shown in the right panel with a reduction in green staining for the Peripherin protein), which is damaged in dominant retinitis pigmentosa (left panel shows the changes to pigmentation in the retina of a patient with dominant RP)

We are now applying a suppression-replacement approach to models of disease in which mutations in Peripherin-2 cause dominant retinal degeneration, and investigating how RNA interference could be applied to other models of dominant disease.

• Proof-of-concept publication: Georgiadis et al Gene Therapy 2010

  • More publications on gene and cell therapy for inherited sight loss

Gene supplementation strategies

Our programme of gene therapy research includes many more proof-of-concept studies, showing that several other forms of inherited retinal degeneration may benefit from gene supplementation therapy - delivering a working copy of a gene that is damaged in disease.

We have shown that retinal cell structure and function is improved following viral gene delivery in models of inherited sight loss caused by mutations in the following genes:

• Prph2 (Ali RR et al Nature Genetics 2000), the first study showing rescue of inherited retinal degeneration using an AAV vector)
• MerTK (improvements to RPE cell function following gene delivery both AAV [Smith AJ et al Molecular Therapy 2003] and Lentivirus [Tschernutter et al Gene Therapy 2005] vectors)
• RPGRIP1 (Pawlyk BS et al IOVS 2005, convincing long-term reversal of photoreceptor cell loss and retinal function following AAV gene delivery)

Neuroprotection - preventing retinal cell loss

We have studied the delivery of genes encoding growth factors to photoreceptor cells, in a strategy called neuroprotection. This strategy aims to prevent photoreceptor cells from being lost, and is designed to be used as a more general therapy alongside specific gene replacement treatments.

Our studies show that although neuroprotection (the delivery of growth factor genes) can prevent cell loss in models of disease, in some cases it can also cause damage to photoreceptor cell function (this is particularly the case with a gene called CNTF). We also showed that an alternative growth factor, GDNF, can enhance the effects of gene replacement therapy in models of inherited retinal degeneration without causing the damaging side effects seen with CNTF.

• Proof-of-concept publication: Buch et al. Mol Ther 2006

More publications on gene and cell therapy for inherited sight loss

In a landmark study we showed that immature photoreceptor cells integrate with the host retina when transplanted into models of inherited retinal degeneration, provided they are at the correct stage of development.

If donor cells are taken from the right stage of retinal development, they can repair a damaged retina when transplanted. They show all the features of mature photoreceptor cells (left panel), including the machinery needed to detect light and connections with other retinal cells (right panel)

Cells that are too early in development, or adult cells, fail to integrate as efficiently - this means that immature photoreceptor cells are the best source of donor cells for transplantation.

We have also shown that manipulating the retina that is to receive such cell transplants can improve the number of integrated cells

• Proof-of-concept publication: MacLaren, Pearson et al Nature 2006

  • More publications on gene and cell therapy for inherited sight loss
• Collaborator: Jane Sowden
• Channel 4 News report on retinal transplantation breakthrough

Having shown that cell transplants can repair the degenerating retina, we demonstrated the restoration of vision following transplants - this breakthrough study provides the first evidence that vision-guided behaviour can be effectively restored using the transplantation of immature photoreceptor cells.

• Proof-of-concept publication: Pearson et al Nature 2012

  • More publications on gene and cell therapy for inherited sight loss
• Collaborator: Jane Sowden
• BBC News: Scientists restore sight in blind mice

The efficient transplantation of photoreceptor cells requires donor cells to be at a precise stage in retinal development. It would not be possible to obtain such cells from humans as this stage of development corresponds to the second trimester of pregnancy. Therefore the challenge is to generate sufficient cells that are suitable for transplant in a dish, and the best sources for obtaining such cells are embryonic stem cells or adult cells reprogrammed to become stem cells (induced pluripotent stem, or iPS, cells).

Following the discovery that embryonic stem cells can spontaneously develop into retinal cells if placed in the right conditions in a three-dimensional culture system, we have shown that we can recreate eye development and produce retinal cells for transplantation.

Retinal cells spontaneously develop from embryonic stem cells in 3D culture. Four panels show cells grown from embryonic stem cells expressing different proteins found in photoreceptor cells

A similar strategy is being used by Advanced Cell Technologies (ACT) to produce retinal pigment epithelium cells from human embryonic stem cells, which we are transplanting into patients with Stargardt macular dystrophy in Europe's first safety trial of embryonic stem cell-derived cells.

• Publication: West et al Stem Cells 2012

  • More publications on gene and cell therapy for inherited sight loss
• Collaborator: Masayo Takahashi

The photoreceptor cell transplantation studies carried out to date have mostly used immature rod photoreceptor cells, as theses are the easiest to study in a mouse system.

For a clinical photoreceptor transplant treatment to be effective, it is important to develop ways to transplant cone photoreceptor cells too. We have shown that when a mix of immature rod and cone photoreceptors are transplanted, we can see mature cone cells integrating into the recipient retina (Lakowski et al Hum Mol Genet 2010).

Cone photoreceptors that are genetically labelled with Green Fluourescent Protein (GFP, top panel) can integrate with host retina when isolated and transplanted - they also produce proteins found in mature cone photoreceptors cells (bottom panel)

• More publications on gene and cell therapy for inherited sight loss