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US-12622965-B2 - Therapeutical tools and methods using temperature-sensitive receptors for treating blindness

US12622965B2US 12622965 B2US12622965 B2US 12622965B2US-12622965-B2

Abstract

The present inventions relates to a method for vision restoration comprising the steps of expressing a temperature-sensitive transient receptor potential (TRP) channel having an extracellular tag in the retina of a subject and of contacting said retina with a nanomaterial conjugated to a molecule specifically binding to said extracellular tag, wherein said nanomaterial generates heat by absorbing radiations of a specific wavelength. And reagents therefor.

Inventors

  • Dasha NELIDOVA
  • Botond Roska

Assignees

  • FRIEDRICH MIESCHER INSTITUTE FOR BIOMEDICAL RESEARCH
  • INSTITUTE OF MOLECULAR AND CLINICAL OPHTHALMOLOGY BASEL

Dates

Publication Date
20260512
Application Date
20210218
Priority Date
20200219

Claims (10)

  1. 1 . An isolated nucleic acid molecule comprising a nucleotide sequence coding for a temperature-sensitive transient receptor potential (TRP) channel having an extracellular epitope tag, for use in a method for vision restoration comprising the steps of expressing said temperature-sensitive TRP channel having an extracellular tag in the retina of a subject and of contacting said retina with a nanomaterial conjugated to a molecule specifically binding to said extracellular tag, wherein said nanomaterial generates heat by absorbing radiations of a specific wavelength, wherein said isolated nucleic acid molecule further comprises a promoter operatively linked to said temperature-sensitive TRP channel having an extracellular tag and leading to the specific expression of said temperature-sensitive TRP channel having an extracellular tag in at least one specific cell population of said retina, and wherein said promoter leads to the specific expression of said temperature-sensitive TRP channel having an extracellular tag in at least one photoreceptor type.
  2. 2 . The isolated nucleic acid molecule of claim 1 , wherein said temperature-sensitive TRP channel is selected from the group consisting of: TRPM, TRPA, TRPV, and homologs thereof.
  3. 3 . The isolated nucleic acid molecule of claim 1 , wherein said epitope tag is selected from the group consisting of: OLLAS, AviTag, C-tag, Calmodulin-tag, polyglutamate tag, E-tag, FLAG-tag, HA-tag, His-tag, Myc-tag, NE-tag, Rho1D4-tag, S-tag, SBP-tag, Softag 1, Softag 3, Spot-tag, Strep-tag, TC tag, Ty tag, V5 tag, VSV-tag, Xpress tag, Isopeptag, SpyTag, SnoopTag, SnoopTagJr, DogTag, SdyTag, BCCP (Biotin Carboxyl Carrier Protein), Glutathione-S-transferase-tag, Green fluorescent protein (GFP)-tag, HaloTag, SNAP-tag, CLIP-tag, Maltose binding protein-tag, Nus-tag, Thioredoxin-tag, Fc-tag, and Carbohydrate Recognition Domain or CRDSAT-tag.
  4. 4 . The isolated nucleic acid molecule of claim 1 , wherein said tag is selected from OLLAS and His-tag.
  5. 5 . A vector comprising the nucleic acid molecule of claim 1 .
  6. 6 . The vector of claim 5 , wherein said vector is a viral vector.
  7. 7 . The vector of claim 6 , wherein said vector is an AAV vector, a PRV vector or a lentivirus vector.
  8. 8 . The isolated nucleic acid molecule of claim 2 , wherein said temperature-sensitive TRP channel is TRPV1, TRPA1, or homologs thereof.
  9. 9 . The isolated nucleic acid of claim 1 , wherein said promoter leads to the specific expression of said temperature-sensitive TRP channel having an extracellular tag in cones.
  10. 10 . The vector of claim 7 , wherein said vector is an AAV vector.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS This application is a U.S. National Stage Application under 35 U.S.C. § 371 of International Application No. PCT/IB2021/051384, filed on Feb. 18, 2021, which claims priority to, and the benefit of, European application No. 20158285.5, filed on Feb. 19, 2020, the contents of each of which are hereby incorporated by reference in their entireties. FIELD OF THE INVENTION The present invention relates to methods of treating blindness using temperature sensitive receptors. The present invention also relates to reagents for use in treating blindness, as well as their use in the manufacture of a medicament for treating blindness. BACKGROUND OF THE INVENTION Blindness is a major health problem that disables millions of people worldwide. The most common cause of blindness is the disfunction of the retina. The three most common forms of retinal blindness are retinitis pigmentosa (RP), macular degeneration (MD) and glaucoma (G). In RP and MD the primary problem is the degeneration of photoreceptors and the consequent loss of photosensitivity. There is thus a need to be able to obviate the problems associated with such degeneration of photoreceptors. One approach has been to develop a retinal prosthesis, a “seeing eye” chip with as many as 1,000 tiny electrodes to be implanted in the eye. This would have the potential to help people who have lost their sight to regain enough vision to function independently, but the numbers of electrodes is simply insufficient to provide a high degree or level of sight to be obtained. Moreover, there are problems associated with inserting foreign bodies into the eye. Recently a number of genes has been isolated and/or manipulated that when expressed can make cells light sensitive. In some cases additional non-genetic factors are also needed to make cells light sensitive. This is complex and problems are likely to arise if the channel is delivered to the wrong type of retinal cell. Bi et al., (Neuron, 50, 2006, p 23-33) discloses the use of microbial-type rhodopsin to restore visual responses in mice with photoreceptor degeneration. However, the expression of the rhodopsin gene is likely to have occurred in a variety of types of cell in the eye which is potentially undesirable and/or problematic. It also appears that the threshold light intensity required for producing responses is much higher than for normal rod and cone photoreceptors, but there is no teaching of how this may be addressed in, for example, low light environments. An alternative method has been described by some of the present inventors in WO-A-2008/022772, wherein e.g. channelrhodopsin-2 is targeted to e.g. ON-cells. This method has however the disadvantage of being sub-optimal with OFF-cells. Enabling the detection of near-infrared (NIR) light at wavelengths (>900 nm) far away from the human visible spectrum (390-700 nm) could provide a way of supplementing or restoring vision in affected retinal regions, without interfering with remaining vision. Upconversion nanoparticles applied to the eye allow the perception of NIR light after conversion of NIR back to visible light (Cell 177, 243-255 e215 (2019).). However, they rely on functional photoreceptors. Photoreceptor dysfunction rules out upconversion as a technology to enable NIR light detection in the blind retina. It is amongst the objects of the present invention to obviate and/or mitigate at least one of the aforementioned disadvantages. SUMMARY OF THE INVENTION To address this need, the present inventors considered the use of temperature-sensitive transient receptor potential (TRP) channels. A few species such as boas, pythons and pit vipers can detect infrared light (1-30 μm) using temperature sensitive TRP channels expressed in a specialized organ (Nature 464, 1006-1011 (2010)). Thermal and visual images superimpose within the snake's brain, presumably enabling the snake to react to the environment with greater precision than with use of a single image only (Science 213, 789-791 (1981)). In order to restore vision, TRP channels could potentially be targeted to retinal cell types, as they are sensitive to infrared radiation. However, heat transfer to ectopically-expressed TRP channels via direct near-infrared illumination is inefficient, and requires high intensities that would damage the retina. In order to develop a more efficient, NIR detector for retinal cell types, the present inventors engineered a two-component system consisting of a genetic and a nanomaterial component. The genetic component consisted of temperature sensitive TRP channels, engineered to incorporate an extracellular epitope recognizable by a specific antibody (Science 336, 604-608 (2012)). The nanomaterial component consisted of gold nanorods conjugated to an antibody against this epitope (Bioconjug Chem 24, 878-888 (2013)). This system uses surface plasmon resonance for heat transfer (Chem Soc Rev 41, 1191-1217 (2012)): gold nanorods capture NIR lig