Gene therapy of the human retina
In 2008, three independent research groups reported that patients with the rare genetic retinal disease
In retinal gene therapy, the most widely used vectors for ocular gene delivery are based on
There is still a lot of knowledge missing in regards of retina dystrophies. Detail characterization is needed in order to improve knowledge. To address this issue, creation of Registries is an attempt to grouped and characterize rare diseases. Registries help to localize, and measure all the phenotype of these conditions and therefore to provide easy follow-ups and provide a source of information to scientist community. Registry designs varies from region to region, however localization and characterization of the phenotype are the standard gold. Examples of Registries are: RetMxMap<ARVO 2009>. A Mexican and Latin-American registry created since 2009. This registry was created by Dr Adda Lízbeth Villanueva Avilés. She is a clinical-scientist gene mapping inherited retina dystrophies in Mexico and other Latin countries.
Clinical trials
Leber's congenital amaurosis
Preclinical studies in mouse models of
Following the successful clinical trials in LCA, researchers have been developing similar treatments using adeno-associated virus for
Choroideremia
In October 2011, the first clinical trial was announced for the treatment of choroideremia.[9] Dr. Robert MacLaren of the University of Oxford, who lead the trial, co-developed the treatment with Dr. Miguel Seabra of the Imperial College, London. This Phase 1/2 trial used subretinal AAV to restore the REP gene in affected patients.[10] Initial results of the trial were reported in January 2014 as promising as all six patients had better vision.[11][12]
Color blindness
Recent research has shown that AAV can successfully restore color vision to treat color blindness in adult monkeys.[13] Although this treatment has not yet entered clinical trials for humans, this work was considered a breakthrough for the ability to target cone photoreceptors.[14]
Mechanism
Physiological components in retinal gene therapy
The vertebrate neural retina composed of several layers and distinct cell types (see
In retinal gene therapy, AAV is capable of "transducing" these various cell types by entering the cells and expressing the therapeutic DNA sequence. Since the cells of the retina are non-dividing, AAV continues to persist and provide expression of the therapeutic DNA sequence over a long time period that can last several years.[15]
AAV tropism and routes of administration
AAV is capable of transducing multiple cell types within the retina. AAV serotype 2, the most well-studied type of AAV, is commonly administered in one of two routes: intravitreal or subretinal. Using the intravitreal route, AAV is injected in the
The reason that different routes of administration lead to different cell types being transfected (e.g., different
Tropism modification and novel AAV vectors
One important factor in
Initial studies with AAV in the retina have utilized AAV serotype 2. Researchers are now beginning to develop new variants of AAV, based on naturally-occurring AAV serotypes and engineered AAV variants.[20]
Several naturally-occurring serotypes of AAV have been isolated that can transduce retinal cells. Following intravitreal injection, only AAV serotypes 2 and 8 were capable of transducing retinal ganglion cells. Occasional Muller cells were transduced by AAV serotypes 2, 8, and 9. Following subretinal injection, serotypes 2, 5, 7, and 8 efficiently transduced photoreceptors, and serotypes 1, 2, 5, 7, 8, and 9 efficiently transduce RPE cells.[17]
One example of an engineered variant has recently been described that efficiently transduces Muller glia following intravitreal injection, and has been used to rescue an animal model of aggressive, autosomal-dominant retinitis pigmentosa.[21][22]
AAV and immune privilege in the retina
Importantly, the retina is immune-privileged, and thus does not experience a significant inflammation or immune-response when AAV is injected.[23] Immune response to gene therapy vectors is what has caused previous attempts at gene therapy to fail, and is considered a key advantage of gene therapy in the eye. Re-administration has been successful in large animals, indicating that no long-lasting immune response is mounted.[24]
Recent data indicates that the subretinal route may be subject to a greater degree of immune privilege compared to the intravitreal route.[25]
Promoter sequence
Expression in various retinal cell types can be determined by the promoter sequence. In order to restrict expression to a specific cell type, a tissue-specific or cell-type specific promoter can be used.
For example, in
Modulation of expression
Sometimes modulation of transgene expression may be necessary since strong constitutive expression of a therapeutic gene in retinal tissues could be deleterious for long-term retinal function. Different methods have been utilized for the expression modulation. One way is using exogenously regulatable promoter system in AAV vectors. For example, the tetracycline-inducible expression system uses a silencer/transactivator AAV2 vector and a separate inducible doxycycline-responsive coinjection.[26][27] When induction occurs by oral doxycycline, this system shows tight regulation of gene expression in both photoreceptor and RPE cells.
Examples and animal models
Targeting RPE
One study that was done by
The protein RPE65 is used in the retinoid cycle where the all-trans-retinol within the rod outer segment is isomerized to its 11-cis form and oxidized to 11-cis retinal before it goes back to the photoreceptor and joins with opsin molecule to form functional rhodopsin.[29] In animal knockout model (RPE65-/-), gene transfer experiment shows that early intraocular delivery of human RPE65 vector on embryonic day 14 shows efficient transduction of retinal pigment epithelium in the RPE65-/- knockout mice and rescues visual functions. This shows successful gene therapy can be attributed to early intraocular deliver to the diseased animal.
Targeting of photoreceptors
Juvenile
Specifically the AAV 5 vector containing the wild-type human RSI cDNA driven by a mouse opsin promoter showed long-term retinal functional and structural recovery. Also the retinal structural reliability improved greatly after the treatment, characterized by an increase in the outer nuclear layer thickness.[26]
Retinitis pigmentosa
Different types of inheritance can attribute to this disease; autosomal recessive, autosomal dominant, X-linked type, etc. The main function of rhodopsin is initiating the
The way this system operates was shown in animal model that have a mutant rhodopsin gene. The injected AAV-ribozymes were optimized in vitro and used to cleave the mutant mRNA transcript of P23H (where most mutation occur) in vivo.[26]
Another mutation in the rhodopsin structural protein, specifically peripherin 2 which is a membrane glycoprotein involved in the formation of photoreceptor outersegment disk, can lead to recessive RP and macular degeneration in human[30] (19). In a mouse experiment, AAV2 carrying a wild-type peripherin 2 gene driven by a rhodopsin promoter was delivered to the mice by subretinal injection. The result showed improvement in photoreceptor structure and function which was detected by ERG (electroretinogram). The result showed improvement of photoreceptor structure and function which was detected by ERG. Also peripherin 2 was detected at the outer segment layer of the retina 2 weeks after injection and therapeutic effects were noted as soon as 3 weeks after injection. A well-defined outer segment containing both peripherin2 and rhodopsin was present 9-month after injection.[26]
Since
AAV-based treatment for retinal neovascular diseases
Ocular neovascularization (NV) is the abnormal formation of new capillaries from already existing blood vessels in the eye, and this is a characteristics for ocular diseases such as diabetic retinopathy (DR), retinopathy of prematurity (ROP) and (wet form) age-related macular degeneration (AMD). One of the main players in these diseases is VEGF (Vascular endothelial growth factor) which is known to induce vessel leakage and which is also known to be angiogenic.[26] In normal tissues VEGF stimulates endothelial cell proliferation in a dose dependent manner, but such activity is lost with other angiogenic factors.[36]
Many angiostatic factors have been shown to counteract the effect of increasing local VEGF. The naturally occurring form of soluble Flt-1 has been shown to reverse neovascularization in rats, mice, and monkeys.[37][38][39][40]
Pigment epithelium-derived factor (
The finding suggests that the AAV-mediated expression of angiostatic factors can be implemented to treat NV.[43][44] This approach could be useful as an alternative to frequent injections of recombinant protein into the eye. In addition, PEDF and sFlt-1 may be able to diffuse through sclera tissue,[45] allowing for the potential to be relatively independent of the intraocular site of administration.
See also
References
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- ^ "Safety and Efficacy Study of rAAV.sFlt-1 in Patients With Exudative Age-Related Macular Degeneration (AMD)". U. S. National Institutes of Health. Retrieved 1 June 2012.
- ^ "Safety and Tolerability Study of AAV2-sFLT01 in Patients With Neovascular Age-Related Macular Degeneration (AMD)". U. S. National Institutes of Health. Retrieved 1 June 2012.
- ^ "First Patient Treated in Choroideremia Gene Therapy Clinical Trial in U.K". Foundation Fighting Blindness. 28 October 2011. Retrieved 1 June 2012.
- ^ "Gene Therapy for Blindness Caused by Choroideremia". U. S. National Institutes of Health. Retrieved 1 June 2012.
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