Gene therapy for color blindness

The retina of the human eye contains photoreceptive cells called cones that allow color vision. A normal trichromat individual possesses three different types of cones to distinguish different colors within the visible spectrum from 380 nm to 740 nm.[1] The three types of cones are designated L, M, and S cones, and each type is sensitive to a certain range of wavelength of light depending on what photopigment it contains. More specifically, the L cone absorbs around 560 nm, the M cone absorbs near 530 nm, and the S cone absorbs near 420 nm.[1] Contrary to popular belief, the peak absorption frequency for L, M, and S cones do not exactly correspond to red, green, and blue wavelength. Rather, the peak frequency for the L cone is orange, yellowish green in M cones, and blue-violet in S cones. These cones transduce the absorbed light into electrical information to be relayed to neurons in the retina such as retinal bipolar cells and retinal ganglion cells, before reaching the brain.[1]

The signals from different cones are added or subtracted from each other to process the color of incoming light. For instance, the color red stimulate L cones more than M cones, whereas the color green stimulates the L and M cones more than the S cones.[1] The colors are perceived in an opponent process, such that red and green are perceived in opposition, as are blue and yellow, black and white.[1]

The gene loci coding for the photopigments: M-opsin and L-opsin are located in close proximity within the X chromosome and are highly polymorphic.[1] Among the population, some have a deleted gene for the M photopigment in the X chromosome (such as in deuteranopia), whereas others have a mutated form of the gene (such as in deuteranomaly). Individuals who can express only two types of opsins in the cones are called dichromats. Because males have only one copy of the X chromosome, dichromatism is much more prevalent among men.[1] With only two types of cones, dichromats are less capable of distinguishing between two colors. In the most common form of color blindness, deuteranopes have difficulty discriminating between red and green color.[1] This is shown by their poor performance in Ishihara test. Although dichromatism poses little problem for daily life, dichromats may find some color-coded diagrams and maps difficult to read.

Less common forms of dichromacy include protoanopia (lack of L-cones), and tritanopia (lack of S-cones). If a person lacks two types of photopigments, they are considered monochromats. People lacking the three types of photopigments are said to have complete color blindness or achromatopsia. Color blindness can also result from damages to the visual cortex in the brain.[1]

Experiments using a variety of mammals (including primates) demonstrated that it is possible to confer color vision to animals by introducing an opsin gene that the animal previously lacked. Using a replication-defective recombinant adeno-associated virus (rAAV) as a vector, the cDNA of the opsin gene found in the L or M cones can be delivered to some fraction of the cones within the retina via subretinal injection. Upon gaining the gene, the cone begins to express the new photopigment. The effect of therapy lasts until the cones die or the inserted DNA is lost within the cones.

While gene therapy for humans has been ongoing with some success, a gene therapy for humans to gain color vision has not been attempted to date. However, demonstrations using several mammals (including primates such as a squirrel monkey) suggest that the therapy should be feasible for humans as well. It is also theoretically possible for trichromats to be "upgraded" to tetrachromats by introducing new opsin genes.

The goal of the gene therapy is to make some of the cones in the retina of a dichromat individual to express the missing photopigment. Although partial color blindness is considered to be a mild disability and even an advantage under certain circumstances (such as spotting camouflaged objects), it can pose challenges for many occupational fields such as law enforcement, aviation, railroad, and military service.[2] More generally, color codes in maps and figures may be difficult to read for individuals with color blindness.

Because only a single gene codes for a photopigment and the gene is only expressed in the retina, it is a relatively easy condition to treat using gene therapy compared to other genetic diseases. However, there remains the question of whether the therapy is worthwhile, for an individual to undergo an invasive subretinal injection to temporarily treat a condition that is more of an inconvenience than a disorder.

However, complete color blindness, or achromatopsia, is very rare but more severe. Indeed, achromats cannot see any color, have a strong photophobia (blindness in full sun), and a reduced visual acuity (generally 20/200 after correction).

Moreover, the research may have strong implications toward genetic therapy of other cone diseases. Other cone diseases such as Leber's congenital amaurosis, cone-rod dystrophy, and certain types of maculopathies may be treatable using the same techniques as the gene therapy used for color blindness.[3][4]

Experimental treatments for Leber's congenital amaurosis, a genetic disorder in photoreceptors that can lead to vision loss and blindness have been performed. These treatments use AAV vector and is delivered in much the same way as the gene therapy for color blindness.[5][6]

Human L-cone photopigment have been introduced into mice. Since the mice possess only S cones and M cones, they are dichromats.[7] M-opsin was replaced with a cDNA of L-opsin in the X chromosome of some mice. By breeding these "knock-in" transgenic mice, they generated heterozygous females with both an M cone and an L cone. These mice had improved range of color vision and have gained trichromacy, as tested by electroretinogram and behavioral tests. However, this is more difficult to apply in the form of gene therapy.

Recombinant AAV vector was to introduce the green fluorescent protein (GFP) gene in the cones of gerbils.[8] The genetic insert was designed to only be expressed in S or M cones, and the expression of GFP in vivo was observed over time. Gene expression could stabilize if a sufficiently high dose of the viral vector is given.

Adult dichromat squirrel monkeys was converted into trichromats using gene therapy.[9] New world monkeys such as squirrel monkeys lack the L-opsin gene and are incapable of discriminating between certain shades of red and green.[9] Recombinant AAV vector was used to deliver a human L-opsin gene into the monkey’s retina. Cones that gained the missing genes began expressing the new photopigment.[9]

If the therapy worked — the monkeys would either remain dichromatic with greater sensitivity for longer wavelength of light, or they would become trichromats.[9] Electroretinogram recordings demonstrated that they are able to discriminate blue-green from red-violet, and have indeed gained trichromacy.[9] The treated monkeys were also more successful when their color vision was tested with a modified Ishihara test.[9]

Gene therapy was to restore some of the sight of mice with achromatopsia. The results were positive for 80% of the mice treated.[10]

Gene therapy for a form of achromatopsia was performed in dogs. Cone function and day vision have been restored for at least 33 months in two young dogs with achromatopsia. However, this therapy was less efficient for older dogs.[11]

According to research by David H. Hubel and Torsten Wiesel, suturing shut one eye of monkeys at an early age resulted in an irreversible loss of vision in that eye, even after the suture was removed.[1][12] The study concluded that the neural circuitry for vision is wired during a "critical period" in childhood, after which the visual circuitry can no longer be rewired to process new sensory input. Contrary to this finding, Mancuso et al.’s success in conferring trichromacy to adult squirrel monkeys suggests that it is possible to adapt the preexisting circuit to allow greater acuity in color vision. The researchers concluded that integrating the stimulus from the new photopigment as an adult was not analogous to vision loss following visual deprivation.[9]

It is yet unknown how the animals that gain a new photopigment are perceiving the new color. While the article by Mancuso et al. states that the monkey has indeed gained trichromacy and gained the ability to discriminate between red and green, they claim no knowledge of how the animal internally perceives the sensation.[9]

While red/green color blindness among deuteranopes can be treated by introducing M-opsin genes, rarer forms of color blindness such as tritanopia can in principle be treated as well. For tritanopia, the S-opsin gene must be introduced instead of M-opsin gene.

Despite the success in animals, there still remain challenges to conducting gene therapy on humans for treating color blindness.

• Color vision
• Achromatopsia