Treating colour blindness with Gene Therapy

Recent research has demonstrated that colour blindness may be capable of rescue by a simple sub-retinal injection of the genetic sequence for the missing photopigment. A research team, based at the University of Washington, have comprehensively shown that animals, previously documented to be colour-blind, are capable of colour discrimination within 20 weeks of treatment. The research not only adds optimism to the field of gene therapy for many other retinal disorders but also suggests an encouraging level of plasticity in how the brain manages new information.

Colour vision
Colour vision is both a fascinating and complex process. Fascinating because interpretation of the world around us through the capacity of colour vision almost defines the "human" experience; complex because the "sensation" of colour and fine acuity vision involves an array of highly differentiated and specialised cell types communicating with the cerebral cortex to create an output that continues to elude our detailed understanding.

To understand how an eye sees colour, click here.

The wavelengths of light visible to the human eye range between approximately 400nM and 700nM allowing humans to distinguish over a million different colours. This impressive feat is achieved through the processing of signals from three types of cone photoreceptor distinguished by their sensitivity to varying wavelengths: "S" (short) with a maximal sensitivity at about 430nM; "M" (medium) with a maximal sensitivity at about 530nM and finally; "L" (long) with a maximal sensitivity at about 560nM. The sensitivities however, accommodate broad "tuning" capabilities such that each type can respond to wavelengths across the visible light spectrum. The sensitivity of any particular photoreceptors are determined by the type of opsin expressed which, in turn, is determined by the sequence of amino acids that make up an opsin protein. Changes in the sequence of amino acids can change the spectral sensitivity for example, changes in 2 out of the approximate 350 amino acids in the L- and M- opsins in humans account for most of the 30nm difference in their peak wavelength sensitivities. Red-green colour blindness is a condition brought about through a disruption of either the long L- or the middle M- wavelength sensitive visual photopigments found in cone photoreceptors. Although the condition has been recognised and studied for over 200 years, the present research is the first report on the use of genetic technology to correct a deficit of colour vision in a mammalian species.

Colour blindness
Red-green colour blindness is among the most common genetic disorders found in humans. The incidence is known to vary with ethnicity (about 8% in Caucasian men, 4% in Japanese men and 3% in African men, and about 6-8% in Indians).

To understand how colour blindness affects sight, click here.

As the L- and M- wavelength sensitive visual photopigment genes ("OPN1LW"-opsin 1 long wave sensitive and "OPN1MW"-opsin 1 medium wave sensitive) are found concatenated head-to-tail along the X chromosome, this in part explains why the condition affects 3-8% of males but only 1% of females. Heterozygous carrier females are estimated at about 15% of the Caucasian population. In their research into correcting the colour deficit the University of Washington research team chose the New World squirrel monkey (Saimiri sciureus) as an experimental model. All male and some female squirrel monkeys are colour-blind "dichromats" (the three different types of cone photoreceptor make humans "trichromatic" whereas most other mammals in the animal kingdom have only two types of cone and are referred to as "dichromatic"). Dichromatic squirrel monkeys have S- cones and M- cones and the idea to deliver the L- photopigment gene sequence would allow the researchers, if successful, to demonstrate a change from dichromatic to trichromatic vision.

Insightful research
The research, led by Professors Jay and Maureen Neitz, was aimed not so much at developing a gene based therapeutic for the treatment of human colour blindness but more to demonstrate the principle of gene therapy for correcting a genetic fault in the retina. While the technology could be developed further and used to treat the condition in humans, it is likely that regulatory authorities would prefer to observe the use of gene therapy for more severe ocular disorders before approving such technology for use in an otherwise healthy retina.

The research group genetically engineered a copy of the human L-opsin gene (OPN1LW) under the control of the L/M opsin enhancer and promoter and packaged the transcript into the recombinant adeno-associated viral (AAV) genome (serotype 2, capsid 5). High-titre infectious particles were prepared and injected in batches of 100uL. Genetic regulatory elements were chosen to direct expression in M- rather than S- cones. Researchers treated colour-blind adult squirrel monkeys, colour blind from birth, with three sequential sub-retinal injections in different areas of the retina, each injection comprising a volume of approximately 100uL and in total containing an estimated 2.7 X 1013 virus particles. Prior to treatment, animals were trained to perform a computer based colour vision test (the Cambridge Colour Test) and control baseline results were built up from over a year's worth of testing.

Twenty weeks after administration, the results clearly demonstrated a change in the spectral sensitivity of a subset of the cone cell population as detected using a custom built wide-field colour multifocal electroretinogram system (mf-ERG). Following treatment, animals tested on the Cambridge Colour Test showed an improved threshold for blue-green and red-violet wavelengths and this improvement coincided with robust levels of transgenic gene expression previously reported for similarly treated squirrel monkeys. In short, the animals had gained trichromatic vision as soon as the new gene was producing opsin protein. So far the researchers have reported that the improvement in colour vision in treated animals has remained stable for more than 2 years. Plans are scheduled to continue testing to allow for long-term evaluation of the technology.

Teaching old monkeys new tricks
The signals for colour are processed through post-receptoral cells in the retina and brain and part of the processing includes computations in specific ganglion cells that subtract the signals received from different types of cone photoreceptor. Such computation, it was thought, develops specifically from birth contingent on the number and types of photoreceptors present. Adding a "new" signal to an established system was thought unlikely to work as the established system would not have developed the pathway required for that particular signal type. The current research from the University of Washington has changed that idea. As soon as the new photopigment is expressed in the retina, there is a simultaneous ability to process new wavelengths of light. Previously dichromatic monkeys acquired the capability for performing tasks of colour discrimination as proficiently as trichromats. This suggests a level of plasticity previously thought to be unlikely. Neural connections, it was thought, established during development would be unlikely to efficiently process "new inputs" (such as that delivered by the gene therapy). As both Prof. Jay and Maureen Neitz comment, "classic visual deprivation experiments [dating back to the 1960s] have led to the expectation that neural connections established during development would not appropriately process an input that was not present from birth. Therefore, it was believed that the treatment of congenital vision disorders would be ineffective unless administered to the very young". Cleary the results from the recent research suggest otherwise and the observations, reported in the journal Nature (Vol. 461, pp784-788), provide encouragement that gene delivery to the eye in the context of adult onset diseases may have a real prospect of success.

Next steps
The team are now looking at another retinal disorder - achromatopsia - and are planning to restore the missing or defective photoreceptor components to the healthy retina and thereby treat the disease in humans. Achomatopsia, meaning "without colour", is a disorder in which the individual is unable to distinguish colour due to a deficient cone mediated eletroretinogram and typically sufferers will have a severely compromised visual acuity. Approximately 1 in 30,000 individuals are affected by the disorder that can cause permanent central vision loss and for which no effective medical therapies exist. The University of Washington team is now testing a gene therapy approach in a mouse model of achromatopsia in an effort to reproduce the success demonstrated in the correction of colour-blindness.

- From Euretina