Gene therapy for ciliopathies

Abstract

To provide vectors for treatment of ciliopathies, including Bardet-Biedl syndrome.SOLUTION: There is described a vector for treating a ciliopathy such as Bardet-Biedl syndrome, where the vector comprises a promoter operably linked to a ciliopathy gene, where the vector can provide transduction of the ciliopathy gene into multiple organs, where the promoter is a ubiquitous promoter which can provide expression of the ciliopathy gene in the transduced organs, and where the ciliopathy gene encodes the protein corresponding to a functional protein that is mutated in the ciliopathy. Also described is the use of the vector in a method of treating a ciliopathy, the method comprising administering a therapeutically effective amount of the vector to a patient suffering from a ciliopathy.SELECTED DRAWING: Figure 11

Description

[Technical Field]

[0001] The present invention relates to gene therapy vectors for the treatment of ciliopathy, including Bardet-Biedl syndrome. [Background technology]

[0002] Ciliopathy has recently emerged as a medically important class of diseases caused by the malfunction of non-motile cilia found on most cells in the body (Waters & Beales, Pediatr Nephrol (2011) 26:1039-1056). Most ciliopathy causes various diseases, including retinal degeneration. They share a common phenotype. It is estimated that there are more than 100 diseases that likely result from cilia dysfunction, with more than 30 currently documented. All of these diseases are debilitating, many are fatal, and because they are nearly all loss-of-function, most would benefit from a gene therapy approach to treatment. Ciliopathy has a collective prevalence of approximately 1 in 500 in the general population. All ciliopathy involve disruption of cilia function and overlap in resulting organ pathology.

[0003] Autosomal recessive Bardet-Biedl syndrome (BBS) is one of the best-characterized ciliopathy and is associated with early-onset blindness, severe obesity, multiple endocrine dysfunction, cognitive impairment, and renal failure. Patients born with hereditary Bardet-Biedl syndrome suffer from a range of debilitating medical problems, some of which are fatal. Affected children typically begin to lose vision by the age of 10 due to damage to the light-sensitive cells in the back of the eye (retina), ultimately resulting in complete blindness. Patients gain an abnormal amount of weight during the first year of life, and if unchecked, can progress to life-threatening obesity, diabetes, and hypertension. Many patients also develop renal failure at some point in their lives (which may require dialysis and / or kidney transplants), and most will have some form of learning difficulty. Together, these problems affect adult patients' ability to live independently, with most patients being unemployed. Even when diagnosed early, symptom-based treatment only manages non-preventable complications such as retinal degeneration and obesity that does not respond to dietary measures.

[0004] Like many ciliary diseases, BBS is an autosomal recessive genetic disorder. To date, 21 genes have been implicated. Many of these gene products interact in multisubunit complexes. For example, many of these proteins form a complex called the BBSome. The BBSome is thought to mediate protein transport to the primary cilium. Another complex, the BBS / CCT chaperonin complex, facilitates BBSome assembly and is composed of multiple BBS proteins and numerous CCT chaperonin proteins. As a result of the physical interaction of the protein products of BBS genes to perform a common function, mutations in many different genes can produce the same abnormal combination of phenotypic findings. The most commonly mutated genes in BBS patients are BBS1 (42%) and BBS10 (22%). More than 30 mutations in the BBS1 gene have been identified in individuals with Bardet-Biedl syndrome. The human BBS1 gene is located on the long (q) arm of chromosome 11 at position 13. Mutations in the BBS1 gene similarly affect the normal formation and function of cilia. Defects in these cellular structures disrupt important chemical signaling pathways during development, leading to abnormalities in sensory perception. The human BBS1 gene contains 17 exons and spans approximately 23 kb. Most BBS1 gene mutations are missense or stop mutations, with the most common mutation exchanging the amino acid methionine for the amino acid arginine at protein position 390 (Met390Arg or M390R). The M390R mutation accounts for approximately 80% of all BBS1 mutations. The human BBS10 gene is located on chromosome 12, and the BBS10 transcript contains only two exons encoding a 723-amino acid protein. Mutations found in BBS10 patients are a mixture of missense, nonsense, and frameshift mutations. The most common alteration is C91fs, occurring in approximately 50% of cases.

[0005] All current gene therapies for ciliopathies aim to treat a single damaged organ. For example, several groups have attempted to treat ocular retinopathy using subretinal injections of gene therapy vectors (e.g., Seo et al., Invest Ophthalmol Vis Sci. 54(9):6118-32 (2013)). However, the use of such approaches directly targeting a single organ means that, in multisystem disorders such as ciliopathies, different vectors must be tailored for each target organ. The need to use multiple vectors can be cost-prohibitive and require cumbersome routine procedures. The use of a single injection rather than multiple injections would be highly desirable for patients, be less invasive, require fewer outpatient visits, and reduce the risks associated with repeated procedures. Therefore, there is a need for approaches that can address defects in several or all affected organs rather than individual organs.

[0006] Williams CL et al. (Mol Ther. 25(4):904-916(2017)) reported that in Bardet-Biedl syndrome

[10] describes the recovery of peripheral olfactory impairment through intranasal administration of a gene therapy vector containing the mouse BBS gene tagged with a fluorescent protein (GFP or mCherry). Williams targets olfactory sensory neurons (OSNs) in mouse nasal tissues through intranasal administration of a gene therapy vector containing the mouse BBS gene tagged with a fluorescent protein (GFP or mCherry). Crucially, in the mouse model used by Williams, BBS protein function is defective only in mature olfactory sensory neurons (OSNs), so this mouse model can only demonstrate recovery of OSN cells. This means that, due to the mouse model used, the experiments described in Williams's literature cannot provide any information regarding BBS gene expression in other cell types or tissues and cannot demonstrate multiorgan recovery.

[0007] WO03 / 102141 describes the identification of mutated BBS1 genes and their various uses. Summary of the Invention [Problem to be solved by the invention]

[0008] In a first aspect of the present invention, there is provided a vector for treating a ciliopathies, the vector comprising a promoter operably linked to a ciliopathies gene, the vector being capable of providing transduction of the ciliopathies gene into multiple organs, the promoter being a ubiquitous promoter capable of providing expression of the ciliopathies gene in the transduced organs, and the ciliopathies gene encoding a protein corresponding to a functional protein mutated in the ciliopathies. [Means for solving the problem]

[0009] Ciliopathies are generally caused by mutations in a single gene, which leads to the dysfunction of non-motile cilia found on most cells in the body. Therefore, the introduction of a correct gene that expresses a functional protein compensates for the mutated gene and improves the effects of ciliopathies. The vector defined above provides transduction in multiple organs. Therefore, the administration of the vector via a single administration route can be used to provide gene expression in multiple organs and improve the pathology associated with ciliopathies. This means that it is not necessary to treat each affected organ or tissue individually, as has been done previously. This approach of targeting multiple organs at once has not been used in ciliopathies in the past, and it was not intended that such an approach would work.

[0010] Ciliopathies are a group of disorders associated with mutations in genes encoding defective proteins that result in abnormal cilia formation or function. Ciliopathies are therefore defined as "disorders associated with mutations in genes encoding no protein or defective proteins that result in abnormal cilia formation or function." Because cilia are components of nearly all vertebrate cells, ciliary dysfunction can characteristically manifest as a group of features including retinal degeneration, kidney disease, and cerebral abnormalities. Additional manifestations include congenital fibrocystic disease of the liver, diabetes, obesity, and skeletal dysplasia. Ciliopathies are associated with mutations in over 40 genes.

[0011] The ciliopathy that can be treated using the above-described vectors can be any ciliopathy that can be treated by expressing a protein corresponding to a functional protein mutated in the ciliopathy. Primarily, these are ciliopathy that result from a mutation that causes loss of protein function. Expression of the functional protein restores protein function and improves the abnormal formation or function of cilia. Such ciliopathy is known to those skilled in the art. The ciliopathy that can be treated may be selected from Bardet-Biedl syndrome, Meckel-Gruber syndrome, medullary polycystic kidney disease, Senior-Loken syndrome, McKusick-Kaufman syndrome, Leber congenital amaurosis, and Joubert syndrome. In some embodiments, the ciliopathy treated with the vector is selected from Bardet-Biedl syndrome, medullary polycystic kidney disease, Senior-Loken syndrome, McKusick-Kaufman syndrome, and Leber congenital amaurosis. In other embodiments, the ciliopathies treated with the vectors are selected from Bardet-Biedl syndrome, Senior-Loken syndrome, McKusick-Kaufman syndrome, and Leber congenital amaurosis. In various embodiments, the ciliopathies treated with the vectors are selected from Bardet-Biedl syndrome and McKusick-Kaufman syndrome. In particular embodiments, the ciliopathies treated with the vectors are Bardet-Biedl syndrome.

[0012] The vector contains a ciliopathic gene encoding a functional protein corresponding to the mutated protein in ciliopathic disease. In other words, the ciliopathic gene encodes a functional protein corresponding to the mutant protein that induces ciliopathic disease. The ciliopathic gene preferably encodes a human protein, e.g., a wild-type human protein. The exact ciliopathic gene depends on the ciliopathic disease being treated and the gene that is mutated and induces the ciliopathic condition. Thus, for example, if a patient has ciliopathic disease caused by a mutation in the BBS1 gene, the vector for treating the patient would contain a ciliopathic gene encoding a functional BBS1 protein.

[0013] In some embodiments, the ciliopathy gene encodes a functional protein selected from a BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, BBS11 / TRIM32, BBS12, BBS13 / MKS1, BBS14 / CEP290, BBS15 / C2ORF86, BBS16 / SDCCAG8, BBS17 / LZTFL1, BBS18 / BBIP1, BBS19 / IFT27, BBS20 / IFT74, and BBS21 / C8ORF37 protein.

[0014] In other embodiments, the ciliopathy gene encodes a functional protein selected from BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, BBS11 / TRIM32, BBS12, BBS14 / CEP290, BBS15 / C2ORF86, BBS16 / SDCCAG8, BBS17 / LZTFL1, BBS18 / BBIP1, BBS19 / IFT27, and BBS20 / IFT74 proteins.

[0015] In various embodiments, the ciliopathy gene is BBS1, BBS2, BBS3 / ARL 6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, BBS11 / TRIM32, BBS12, BBS15 / C2ORF86, BBS16 / SDCCAG8, BBS17 / LZTFL1, BBS18 / BBIP1, BBS19 / IFT27 and BBS20 / IFT74 proteins.

[0016] In embodiments, the ciliopathy gene encodes a functional protein selected from BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, BBS12, and BBS18 / BBIP1 proteins.

[0017] In particular embodiments, the ciliopathy gene encodes a functional protein selected from BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, and BBS12 proteins.

[0018] In a specific embodiment, the ciliopathy gene encodes a functional protein selected from BBS1 and BBS10 proteins.

[0019] In some embodiments, the ciliopathy gene encodes a functional BBS1 protein.

[0020] In other embodiments, the ciliopathy gene encodes a functional BBS10 protein.

[0021] In some embodiments, the ciliopathies being treated are Bardet-Biedl syndrome and the ciliopathies genes encode functional proteins selected from BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, BBS11 / TRIM32, BBS12, BBS13 / MKS1, BBS14 / CEP290, BBS15 / C2ORF86, BBS16 / SDCCAG8, BBS17 / LZTFL1, BBS18 / BBIP1, BBS19 / IFT27, BBS20 / IFT74, and BBS21 / C8ORF37 proteins.

[0022] In other embodiments, the ciliopathies being treated are Meckel-Gruber syndrome and the ciliopathies gene encodes a functional BBS13 / MKS1 protein.

[0023] In various embodiments, the ciliopathies being treated is medullary polycystic kidney disease and the ciliopathies gene encodes a functional BBS14 / CEP290 protein.

[0024] In a particular embodiment, the ciliopathies being treated is Senior-Loken syndrome and the ciliopathies gene encodes a functional BBS14 / CEP290 protein.

[0025] In some embodiments, the ciliopathies being treated is McKusick-Kaufman syndrome and the ciliopathies gene encodes a functional BBS6 / MKKS protein.

[0026] In other embodiments, the ciliopathies being treated is Leber's congenital amaurosis and the ciliopathies gene encodes a functional BBS14 / CEP290 protein.

[0027] In various embodiments, the ciliopathic disease to be treated is Joubert syndrome and the ciliopathic gene encodes a functional BBS14 / CEP290 protein.

[0028] The functional protein encoded by the ciliopathic gene preferably does not contain additional amino acids not found in the wild-type protein. Any additional amino acids may interfere with the normal function of the protein. For example, the functional protein preferably does not contain a fluorescent protein such as green fluorescent protein (GFP) or mCherry, or a tag such as a FLAG tag or a polyhistidine tag.

[0029] In a specific embodiment, the ciliopathy gene comprises the nucleotide sequence of SEQ ID NO:1. In some embodiments, the ciliopathy gene has the nucleotide sequence of SEQ ID NO: 1 or has at least 72% sequence identity thereto and encodes a functional BBS1 protein. and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 74% sequence identity thereto and encodes a functional BBS1 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 76% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 78% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 80% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 82% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 84% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 85% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 86% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 88% sequence identity thereto and encodes a functional BBS1 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 90% sequence identity thereto and encodes a functional BBS1 protein. In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:1 or has at least 92% sequence identity thereto and encodes a functional BBS1 protein.In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 94% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 95% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 96% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 97% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:1 or has at least 98% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:1, or has at least 99% sequence identity thereto, and encodes a functional BBS1 protein. In particular embodiments, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:1.

[0030] In a specific embodiment, the ciliopathy gene comprises the nucleotide sequence of SEQ ID NO:2 In some embodiments, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:2 or has at least 72% sequence identity thereto and encodes a functional BBS10 protein. and encodes a functional BBS10 protein. In many embodiments, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:2 or is at least 7 times different therefrom. 4% sequence identity and encodes a functional BBS10 protein. In another embodiment, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:2, or

[0023] In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 78% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 80% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 82% sequence identity thereto and encodes a functional BBS10 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 84% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 85% sequence identity thereto and encodes a functional BBS10 protein. In certain embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 86% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 88% sequence identity thereto and encodes a functional BBS10 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 90% sequence identity thereto and encodes a functional BBS10 protein. In certain embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 92% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 94% sequence identity thereto and encodes a functional BBS10 protein.In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 95% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 96% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 97% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 98% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:2 or has at least 99% sequence identity thereto and encodes a functional BBS10 protein. In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:2.

[0031] In the above embodiment, the nucleotide sequence of the ciliopathic gene may be codon-optimized to maximize expression of the protein. In codon-optimization, the amino acid sequence of the encoded protein will remain the same and therefore still be functional. It is simply a modified nucleotide sequence. SEQ ID NOs: 11 and 12 are sequences encoding B BS1 is a codon-optimized nucleotide sequence encoding SEQ ID NO: 13 and 14 are codon-optimized nucleotide sequences encoding BBS10. These sequences have been found to confer an unexpectedly large increase in gene expression.

[0032] In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:11. In some embodiments, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:11 or has at least 72% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:11 or has at least one identical sequence to it. 74% sequence identity and encodes a functional BBS1 protein. In another embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:11 or has at least 76% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 78% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 80% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 82% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 84% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 85% sequence identity thereto and encodes a functional BBS1 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 86% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 88% sequence identity thereto and encodes a functional BBS1 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 90% sequence identity thereto and encodes a functional BBS1 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 92% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:11, or has at least 94% sequence identity thereto, and encodes a functional BBS1 protein.In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 95% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 96% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 97% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 98% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:11 or has at least 99% sequence identity thereto and encodes a functional BBS1 protein. In a specific embodiment, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:11.

[0033] In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:12. In some embodiments, the ciliopathy gene has the nucleotide sequence of, or has at least 70% sequence identity with, SEQ ID NO: 12 and encodes a functional BBS1 protein. In some embodiments, the ciliopathy gene has the nucleotide sequence of, or has at least 72% sequence identity with, SEQ ID NO: 13. In many embodiments, the ciliopathy gene has the nucleotide sequence of SEQ ID NO:12 or has at least one identical sequence to it. 74% sequence identity and encodes a functional BBS1 protein. In another embodiment, the ciliopathy gene has the nucleotide sequence of SEQ ID NO: 12 or has at least 76% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 78% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 80% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 82% sequence identity thereto and encodes a functional BBS1 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 84% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 85% sequence identity thereto and encodes a functional BBS1 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 86% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 88% sequence identity thereto and encodes a functional BBS1 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 90% sequence identity thereto and encodes a functional BBS1 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 92% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:12, or has at least 94% sequence identity thereto, and encodes a functional BBS1 protein.In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 95% sequence identity thereto and encodes a functional BBS1 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 96% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 97% sequence identity thereto and encodes a functional BBS1 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 98% sequence identity thereto and encodes a functional BBS1 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:12 or has at least 99% sequence identity thereto and encodes a functional BBS1 protein. In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:12.

[0034] In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:13. In some embodiments, the ciliopathy gene has the sequence of, or at least 70% sequence identity to, the BBS10 protein and encodes a functional BBS10 protein. having the nucleotide sequence of NO:13 or at least 72% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathic gene has the nucleotide sequence of, or at least and encodes a functional BBS10 protein. In another embodiment, the ciliopathy gene has the nucleotide sequence of SEQ ID NO: 13. or has at least 76% sequence identity thereto and encodes a functional BBS10 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 78% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 80% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 82% sequence identity thereto and encodes a functional BBS10 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 84% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 85% sequence identity thereto and encodes a functional BBS10 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 86% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 88% sequence identity thereto and encodes a functional BBS10 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 90% sequence identity thereto and encodes a functional BBS10 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 92% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:13, or has at least 94% sequence identity thereto, and encodes a functional BBS10 protein.In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 95% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 96% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 97% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 98% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:13 or has at least 99% sequence identity thereto and encodes a functional BBS10 protein. In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:13.

[0035] In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:14. In some embodiments, the ciliopathy gene has the sequence of, or at least 70% sequence identity to, the BBS10 protein and encodes a functional BBS10 protein. having the nucleotide sequence of NO:14 or at least 72% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathic gene has the nucleotide sequence of, or at least and encodes a functional BBS10 protein. In another embodiment, the ciliopathy gene has the nucleotide sequence of SEQ ID NO: 14. or has at least 76% sequence identity thereto and encodes a functional BBS10 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 78% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 80% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 82% sequence identity thereto and encodes a functional BBS10 protein. In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 84% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 85% sequence identity thereto and encodes a functional BBS10 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 86% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 88% sequence identity thereto and encodes a functional BBS10 protein. In other embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 90% sequence identity thereto and encodes a functional BBS10 protein. In specific embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 92% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:14, or at least 94% sequence identity thereto, and encodes a functional BBS10 protein.In various embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 95% sequence identity thereto and encodes a functional BBS10 protein. In particular embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 96% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 97% sequence identity thereto and encodes a functional BBS10 protein. In many embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 98% sequence identity thereto and encodes a functional BBS10 protein. In some embodiments, the ciliopathies gene has the nucleotide sequence of SEQ ID NO:14 or has at least 99% sequence identity thereto and encodes a functional BBS10 protein. In a specific embodiment, the ciliopathic gene has the nucleotide sequence of SEQ ID NO:14.

[0036] In various embodiments, the ciliopathy gene has the protein sequence of SEQ ID NO:9. In some embodiments, the functional BBS1 protein encodes a functional BBS1 protein having the protein sequence of SEQ ID NO:9 or at least 80% sequence identity thereto. In some embodiments, the functional BBS1 protein has the protein sequence of SEQ ID NO:9 or at least 85% sequence identity thereto. In one embodiment, the functional BBS1 protein has the protein sequence of SEQ ID NO:9. In many embodiments, the functional BBS1 protein has the protein sequence of SEQ ID NO:9, or at least 90% sequence identity thereto. In a particular embodiment, the functional BBS1 protein has the protein sequence of SEQ ID NO:9.

[0037] In another embodiment, the ciliopathy gene is the protein sequence of SEQ ID NO:10. The functional BBS10 protein encodes a functional BBS10 protein having the protein sequence of SEQ ID NO:10 or at least 80% sequence identity thereto. In some embodiments, the functional BBS10 protein has the protein sequence of SEQ ID NO:10 or at least 85% sequence identity thereto. In various embodiments, the functional BBS10 protein has the protein sequence of SEQ ID NO:10 or at least 90% sequence identity thereto. In many embodiments, the functional BBS10 protein has the protein sequence of SEQ ID NO:10 or at least 95% sequence identity thereto. In particular embodiments, the functional BBS10 protein has the protein sequence of SEQ ID NO:10.

[0038] In the preceding description, the term "identity" is used to refer to the similarity between two sequences. For the purposes of the present invention, identity is defined herein as aligning two sequences for optimal comparison purposes to determine their percent identity (e.g., gaps can be introduced into the first sequence for optimal alignment with the second amino acid or nucleic acid sequence). The nucleotides / amino acid residues at each position are then compared. If a position in the first sequence is occupied by the same amino acid or nucleotide residue as the corresponding position in the second sequence, the molecules are identical at that position. The percent identity between two sequences is a function of the number of identical positions shared by the sequences (i.e., % identity = number of identical positions / total number of positions (i.e., overlapping positions) × 100). Generally, the two sequences are the same length. Sequence comparison is typically performed over the entire length of the two sequences being compared.

[0039] Those skilled in the art will recognize that several different computer programs are available for determining the identity between two sequences. For example, the alignment of two sequences and the determination of percent identity can be accomplished using a mathematical algorithm. In a preferred embodiment, the percent identity between two nucleic acid sequences is determined using the sequence alignment software Clone Manager 9 (Sci-Ed software - www.scied.com) using global DNA alignment, parameters: both strands, scoring matrix: linear (mismatch 2, OpenGap 4, ExtGap 1).

[0040] Alternatively, the percent identity between two amino acid or nucleic acid sequences can be determined utilizing the Needleman and Wunsch (1970) algorithm as implemented in the GAP program within the Accelrys GCG software package (available from http: / / www.accelrys.com / products / gcg / ), using either a Blosum 62 matrix or a PAM250 matrix, and gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6. Another method for assessing the percent identity between two amino acid or nucleic acid sequences can be using the BLAST sequence comparison tool available from the National Center for Biotechnology Information (NCBI) website (www.blast.ncbi.nlm.nih.gov), e.g., BLASTn for nucleotide sequences or BLASTp for amino acid sequences using default parameters.

[0041] The ciliopathies gene encodes a "functional" protein. This means that the protein, when expressed, has the same function and activity as the wild-type human protein. This can be easily determined by one of ordinary skill in the art. The protein encoded by the ciliopathies gene may be a wild-type human protein. The wild-type human sequences of the various proteins discussed above are well known to those of ordinary skill in the art. For example, they can be found in the publicly accessible databases of the National Center for Biotechnology Information. Furthermore, these The nucleotide sequence encoding the protein (and contained within the vector) can be readily discovered or determined by one of skill in the art, for example, using the genetic code, which correlates particular nucleotide codons with particular amino acids.

[0042] The promoter contained within the vector is a ubiquitous promoter operably linked to the ciliopathic gene, such that the promoter directs expression of the ciliopathic gene in the transduced organ. A ubiquitous promoter is one that has strong activity in a wide range of cells and tissues and provides constitutive expression. Suitable ubiquitous promoters are readily available to those skilled in the art. It is well known that ubiquitous promoters are not tissue specific; they provide expression in multiple tissues / organs. The ubiquitous promoters result in expression of the ciliopathies gene in the transduced organ, and the expressed protein ameliorates the pathology associated with ciliopathies.

[0043] Suitable ubiquitous promoters include the short elongation factor promoter (EFS), CAG promoter, cytomegalovirus immediate early promoter (CMV), ubiquitin C promoter (UBC), phosphoglycerate kinase promoter (PGK), and beta-actin promoters, such as chicken beta-actin promoter (CBA). These promoters are well known to those skilled in the art. Examples of sequences of these promoters are SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45, SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, S EQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8 and SEQ I D NO:45. Thus, in some embodiments, the promoter SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8 and SEQ ID It has a sequence selected from NO:45.

[0044] In particular embodiments, the promoter is SEQ ID NO:3 or SEQ ID NO:1 The EFS promoter may have the nucleotide sequence of D NO:46.

[0045] In some embodiments, the promoter is SEQ ID NO:4 or SEQ The CAG promoter may have the nucleotide sequence of ID NO:47.

[0046] In various embodiments, the promoter is SEQ ID NO:6 or SEQ ID NO:1 A CMV promoter may have the nucleotide sequence of D NO:45.

[0047] In certain embodiments, the promoter is SEQ ID NO:5 or SEQ ID NO:1 The UBC promoter may have the nucleotide sequence of D NO:48.

[0048] In many embodiments, the promoter comprises the nucleotide sequence of SEQ ID NO:7. The PGK promoter may have the sequence:

[0049] In some embodiments, the promoter is selected from the group consisting of SEQ ID NO:8 and SEQ ID NO:10. A beta-actin promoter may have the nucleotide sequence of D NO:49.

[0050] Regarding a particular combination of elements, the vector may comprise an EFS promoter operably linked to the BBS1 gene, and the vector is for the treatment of Bardet-Biedl syndrome. The BBS1 gene is represented by SEQ ID NOs: 1, 11 and 12. For example, the vector may be selected from SEQ ID NOs: 15, 16 and 17. Alternatively, the vector may comprise an EFS promoter operably linked to the BBS10 gene, and the vector is for the treatment of Bardet-Biedl syndrome. The BBS10 gene may comprise one of the sequences of SEQ ID NO:2. , 13 and 14. For example, the vector may be selected from SEQ ID NO: 30 , 31 and 32.

[0051] The vector may comprise a UBC promoter operably linked to a BBS1 gene, and the vector is for treating Bardet-Biedl syndrome. The BBS1 gene may be selected from SEQ ID NOs: 1, 11, and 12. For example, the vector The vector may comprise one of the sequences of SEQ ID NOs: 18, 19, and 20. Alternatively, the vector may contain a UBC promoter operably linked to the BBS10 gene. The vector may comprise a BBS10 gene selected from SEQ ID NOs: 2, 13, and 14, and the vector is for the treatment of Bardet-Biedl syndrome. For example, the vector may comprise one of the sequences of SEQ ID NOs: 33, 34, and 35. may also include:

[0052] The vector may comprise a CMV promoter operably linked to a BBS1 gene, and the vector is for the treatment of Bardet-Biedl syndrome. The BBS1 gene may be selected from SEQ ID NOs: 1, 11, and 12. For example, the vector The vector may contain one of the sequences of SEQ ID NOs: 21, 22, and 23. Alternatively, the vector may comprise a CMV promoter operably linked to the BBS10 gene, and the vector is for treating Bardet-Biedl syndrome. The BBS10 gene may be selected from SEQ ID NOs: 2, 13, and 14. For example, the vector may comprise one of SEQ ID NOs: 36, 37, and 38.

[0053] The vector may comprise a CBA promoter operably linked to a BBS1 gene, and the vector is for treating Bardet-Biedl syndrome. The BBS1 gene may be selected from SEQ ID NOs: 1, 11, and 12. For example, the vector The vector may contain one of the sequences of SEQ ID NOs: 24, 25, and 26. Alternatively, the vector may comprise a CBA promoter operably linked to the BBS10 gene, and the vector is for treating Bardet-Biedl syndrome. The BBS10 gene may be selected from SEQ ID NOs: 2, 13, and 14. For example, the vector may comprise one of SEQ ID NOs: 39, 40, and 41.

[0054] The vector may comprise a CAG promoter operably linked to a BBS1 gene, and the vector is for the treatment of Bardet-Biedl syndrome. The BBS1 gene may be selected from SEQ ID NOs: 1, 11, and 12. For example, the vector The vector may comprise one of SEQ ID NOs: 27, 28, and 29. Alternatively, the vector may comprise a CAG promoter operably linked to the BBS10 gene, and the vector is for treating Bardet-Biedl syndrome. The BBS10 gene may be selected from SEQ ID NOs: 2, 13, and 14. For example, the vector may comprise one of SEQ ID NOs: 42, 43, and 44.

[0055] The vector may comprise a PGK promoter operably linked to a BBS1 gene, and the vector is for the treatment of Bardet-Biedl syndrome. The BBS1 gene may be selected from SEQ ID NOs: 1, 11, and 12. The vector may comprise a PGK promoter operably linked to a BBS10 gene, and the vector is for treating Bardet-Biedl syndrome. The BBS10 gene may be selected from SEQ ID NOs: 2, 13, and 14.

[0056] The vectors described above can provide transduction of ciliopathies genes into multiple organs. This can be any suitable vector, and such vectors are well known to those skilled in the art. In particular embodiments, the vector can cross the blood-brain barrier. This allows transduction within the brain and nervous system, including the eye, as well as in the visceral organs and muscular system. Therefore, a single vector can be used to provide gene expression in multiple organs to improve ciliopathies-related pathologies. This gene expression can occur in multiple sites throughout the body and can be systemic. Furthermore, vector administration via a limited number of routes can be used to provide systemic gene expression to improve ciliopathies-related pathologies throughout the body. This means that it is not necessary to treat each affected tissue individually. This approach of targeting multiple organs at once has not been used in ciliopathies before, and it was not anticipated that such an approach would work.

[0057] Organs that can be transduced with the previously described vectors may be selected from the central nervous system, eye (e.g., retinal photoreceptors and retinal pigment epithelium), heart, liver, muscle, pancreas, spleen, lung, and kidney. Thus, in some embodiments, the vector provides for transduction of a ciliopathic gene into multiple organs selected from the central nervous system, eye, heart, liver, muscle, pancreas, spleen, lung, and kidney. In other embodiments, the vector provides for transduction of a ciliopathic gene into at least three organs selected from the central nervous system, eye, heart, liver, muscle, pancreas, spleen, lung, and kidney. In various embodiments, the vector provides for transduction of a ciliopathic gene into at least four of the mentioned organs. In many embodiments, the vector provides for transduction of a ciliopathic gene into at least five of the mentioned organs. In some embodiments, the vector provides for transduction of a ciliopathic gene into at least six of the mentioned organs. In other embodiments, the vector provides for transduction of a ciliopathic gene into at least seven of the mentioned organs. In various embodiments, the vector provides for transduction of the ciliopathic disease gene into at least eight of the mentioned organs. In particular embodiments, the vector provides for transduction of the ciliopathic disease gene into the central nervous system, eyes, heart, liver, muscle, pancreas, spleen, lungs, and kidneys. In particular embodiments, the vector provides for transduction of the ciliopathic disease gene into at least the central nervous system (e.g., brain) and eyes. In various embodiments, the vector provides for transduction of the ciliopathic disease gene into at least the central nervous system (e.g., brain), eyes, and one of the liver, kidneys, and spleen. In some embodiments, the vector provides for transduction of the ciliopathic disease gene into at least the central nervous system (e.g., brain), eyes, and liver. In particular embodiments, the vector provides for transduction of the ciliopathic disease gene into at least the central nervous system (e.g., brain), eyes, liver, kidneys, and spleen.

[0058] Suitable vectors include adeno-associated virus-8 (AAV8) and adeno-associated virus-9 (AVV9), as well as other AAVs (e.g., AAV2) pseudotyped with capsid proteins from AAV8 or AAV9. Such vectors are described in WO2005 / 033321. Other suitable vectors include AAV-PHP.A and AAVPHP.B (Nature Biotechnology 34, 204-209 (2016)), AAV9 .47 (Hum Gene Ther. 2016 Jul;27(7):497-508), AAV-B1 (Mol. Ther. 24, 1247-1257), AAV8(Y733F) (Mol Ther 2009;17:463-471), and AAV2-TT (described in WO2015 / 121501). Lentiviral vectors can also be used, for example, as described in Trends in Molecular Medicine, April 2016, Vol. 22, No. 4 and Ther Deliv. 2010 October; 1(4): 517-534.

[0059] In some embodiments, the vector is an AAV vector such as AAV8, AAV9, an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9, AAV-PHP.A, AAV-PHP.B, AAV9.47, AAV-B1, AAV8(Y733F), or AAV2-TT. In other embodiments, the vector is selected from AAV8, AAV9, an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9, AAV-PHP.A, AAV-PHP.B, AAV9.47, and AAV-B1. In various embodiments, the vector is selected from AAV8, AAV9, an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9, AAV-PHP.A, and AAV-PHP.B. In many embodiments, the vector is selected from AAV8, AAV9, an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9, and AAV-PHP.B. In particular embodiments, the vector is selected from AAV8, AAV9, an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9. In some embodiments, the vector is selected from AAV8 and an AAV vector pseudotyped with capsid proteins from AAV8 (e.g., AAV2 pseudotyped with capsid proteins from AAV8 (AAV2 / 8)). In other embodiments, the vector is selected from AAV9 and an AAV vector pseudotyped with capsid proteins from AAV9 (e.g., AAV2 pseudotyped with capsid proteins from AAV9 (AAV2 / 9)).

[0060] The adeno-associated virus vector may be a recombinant adeno-associated virus (rAAV) vector. AAV is a parvoviridae family described in Kenneth I. Berns, "Parvoviridae: The Viruses and Their Replication," Chapter 69 in Fields Virology (3d Ed. 1996).

[0061] The genomic organization of all known AAV serotypes is very similar. The AAV genome is a linear, single-stranded DNA molecule less than approximately 5,000 nucleotides (nt) in length. Inverted terminal repeats (ITRs) flank unique coding nucleotide sequences for nonstructural replication (Rep) proteins and structural (VP) proteins. The VP proteins (VP1, -2, and -3) form the capsid. The terminal 145 nt are self-complementary and organized to allow the formation of energetically stable intramolecular duplexes that form T-shaped hairpins. These hairpin structures serve as origins of viral DNA replication and as primers for the intracellular DNA polymerase complex. Following wild-type (wt) AAV infection in mammalian cells, the Rep genes (i.e., encoding the Rep78 and Rep52 proteins) are expressed from the P5 and P19 promoters, respectively, and both Rep proteins function in viral genome replication. Splicing events in the Rep ORF result in the expression of four Rep proteins (i.e., Rep78, Rep68, Rep52, and Rep40). However, it has been shown that the unspliced mRNAs encoding the Rep78 and Rep52 proteins in mammalian cells are sufficient for AAV vector production. Similarly, in insect cells, the Rep78 and Rep52 proteins are sufficient for AAV vector production.

[0062] In AAVs suitable for use as gene therapy vectors, the vector genome typically contains a packaged nucleic acid (e.g., a ciliopathogenic gene) for delivery to a target cell. According to this particular embodiment, the heterologous nucleotide sequence is located between viral ITRs at either end of the vector genome. In a further preferred embodiment, the parvovirus (e.g., AAV) cap gene and parvovirus (e.g., AAV) rep gene are deleted from the template genome (and thereby from the virion DNA produced therefrom). This structure maximizes the size of the nucleic acid sequence(s) that can be carried by the parvovirus capsid.

[0063] According to this particular embodiment, the nucleic acid is located between viral ITRs at either end of the substrate. A parvovirus genome can function with only one ITR. Thus, in parvovirus-based gene therapy vectors, the vector genome is flanked by at least one ITR, but more typically by two AAV ITRs (generally on either side of the vector genome, i.e., one at the 5' end and one at the 3' end). There may be intervening sequences between the nucleic acid in the vector genome and one or more of the ITRs.

[0064] Generally, a ciliopathic gene (i.e., a nucleotide sequence encoding a protein corresponding to the functional protein mutated in a ciliopathic disease (for expression in mammalian cells)) is integrated into the parvovirus genome, either placed between two normal ITRs or on either side of two D-region engineered ITRs.

[0065] In one aspect, the present invention provides a pharmaceutical composition comprising the vector described above and one or more pharmaceutically acceptable excipients, including carriers, diluents and / or other medicinal, pharmaceutical or adjuvant agents.

[0066] The present invention also provides a method for treating a ciliopathic disease, comprising administering a therapeutically effective amount of a vector as described above to a patient suffering from a ciliopathic disease, preferably a human.

[0067] When a ciliopathic disease is "treated" by the above methods, this means that one or more symptoms of the ciliopathic disease are improved. It does not mean that the symptoms are completely cured so that they are no longer present in the patient, although this may be the case in some methods. The treatment method results in one or more symptoms of the ciliopathic disease being less severe than before treatment. The treatment method may result in multiple symptoms of the ciliopathic disease being less severe than before treatment. Transduction and gene expression in multiple organs results in improvement of the symptoms in multiple organs.

[0068] A "therapeutically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve a desired therapeutic result, such as increasing functional protein levels in a subject (to a level sufficient to ameliorate the symptoms of a ciliopathic disorder).

[0069] The method of treatment induces an increase in the level of functional protein in a subject. In some embodiments, the method of treatment results in an increase in functional protein level to about normal levels (i.e., levels found in normal, healthy subjects). In one embodiment, the method of treatment induces an increase in functional protein level to most normal levels.

[0070] The vector may be administered in any suitable manner to allow expression of the ciliopathies gene in multiple organs. In particular embodiments, a single administration of the vector can be used to provide gene expression and ameliorate symptoms associated with ciliopathies. Administration of the vector may provide systemic gene expression and ameliorate symptoms associated with ciliopathies throughout the body. The vector may be administered intravenously or intracranially. In particular embodiments, the vector is administered intravenously. In some embodiments, the vector is administered intracranially. In various embodiments, the vector is administered intravenously and intracranially.

[0071] The vector may be administered intrathecally, which may be alone or in addition to intravenous and / or intracranial administration.

[0072] Intracranial administration is the direct delivery of a vector to a specific region of the brain using stereotactic injection. Intracranial administration does not include subretinal administration, e.g., subretinal injection.

[0073] Furthermore, the vector should preferably not be administered intranasally. The intranasal administration route may limit the expression of the vector to a small fraction of intranasal cells, and the vector cannot target the main affected tissues in other parts of the body. In addition, the intranasal route does not allow the long-term persistent expression of the transgene due to the rapid replacement of a small fraction of intranasal cells.

[0074] When the vector is administered by multiple routes, for example, intravenously and intracranially, the vector is administered to both sites on the same day. In some embodiments, the multiple administrations are given within 6 hours, 4 hours, or 2 hours apart. In some embodiments, the multiple administrations are given simultaneously.

[0075] The vector may be administered at one time, for example, a single injection may be given. If the vector is administered via multiple routes, for example, intravenously and intracranially, the vector is administered to both sites only once (and on at least the same day, as described above). Then, no further administration is given.

[0076] The present invention further provides the above vectors for use in therapy, for example in the treatment of ciliopathy.

[0077] Additionally, the present invention provides the use of the above vector in the manufacture of a medicament for treating a ciliopathic disease.

[0078] All patent and literature references cited herein are incorporated by reference in their entirety. [Brief explanation of the drawings]

[0079] The invention will now be described in more detail by way of example only with reference to the following figures. [Figure 1]Restriction digestion. Plasmid DNA of AAV2 / 8, helper virus, and BBS1 digested with Xho1. Gel electrophoresis of a 1% agarose gel showing the plasmid DNA of each helper plasmid (pHGTI, lane 2), AAV2 / 8 plasmid (pLT-AAV2 / 8, lane 3), and BBS1 plasmid (pAV-EFS-BBS1, lane 4) digested with the restriction enzyme Xho1. The HyperLadder is visible in lane 1. The plasmids were used to generate hAAV2 / 8-EFS-BBS1. The band sizes for the helper plasmid were 6318 bp and 11549 bp (lane 2), and for the AAV2 / 8 plasmid were 186 bp, 2109 bp, and 4844 bp (lane 3), respectively. For pAV-EFS-BBS1, the band sizes were 236 bp, 643 bp, 1195 bp, and 5078 bp (lane 4). [Figure 2] Human BBS1 transfection of 293T cells. Protein and mRNA expression after transfection of 293T with the EFS-BBS1 plasmid. A) 2% agarose gel after reverse transcription-PCR from mRNA extracted from transfected and non-transfected 293T cells. Lane 1 marker and lanes 2, 4, and 6 show transfected cDNA; lanes 3, 5, and 7 show transfected cells without reverse transcription; lanes 8, 10, and 12 show non-transfected cDNA, and lanes 9, 11, and 13 show non-transfected, non-reverse transcriptase controls. Lane 14 shows a ddH20 sample for PCR as a negative control. B) BBS1 protein expression after transfection was visible in transfected 293T cells. Lanes 2, 3, and 4 are transfected cells, and lanes 5, 6, and 7 are non-transfected cells. Protein expression of HSP90β and GAPDH was used as internal controls. [Figure 3]Expression of human BBS1 in the retina and CNS (central nervous system). RT-PCR demonstrating transgene human BBS1 expression in P35 animals after intracranial and systemic delivery. Intracranial injection (IC) of AAV2 / 8-EFS-BBS1 shows clear expression of human BBS1 alongside mouse BBS1 in the brain and eye (black arrows), indicating successful transduction and expression. Clear expression of human BBS1 is observed in the eye of an intravenously (systemically) injected animal (IV) (white arrow). Specific primers were designed to distinguish between mouse and human BBS1: mBbs1 (mouse BBS1); AAV-hBBS1 (transduced human BBS1); β-actin (β-actin positive control); and nephrin (negative control). [Figure 4] Expression of human BBS1 by RT-PCR in the retina and CNS (central nervous system) at P180 (180 days post-injection) after P0 intracranial and systemic delivery in mice. A) Clear expression of human BBS1 is observed in the brain and eye of intravenously (systemically) injected animals (IV). B) Intracranial injection (IC) of AAV2 / 8-EFS-BBS1 shows clear expression of human BBS1 in the brain, indicating successful transduction and expression. Specific human primers were designed to distinguish between mouse and human BBS1. M390R = Bbs1M390R / M390R animals. + / + = wild-type animals. RT- = RT-PCR negative control. [Figure 5] Phenotypic expression in treated and untreated Bbs1M390R / M390R demonstrates weight loss in P0 intracranially treated animals. When intracranially treated with the AAV2 / 8-EFS-BBS1 construct, Bbs1M390R / M390R animals show statistically significant weight recovery to wild-type levels in both males and females (see p-values). IC = intracranial. Het = heterozygous animals Bbs1M390R / M390R. WT = wild-type animals. M390R = homozygous animals Bbs1M390R / M390R. Error bars indicate standard error of the mean (SEM). p-values were obtained after nonlinear curve fitting (Gompertz growth curve fitting) followed by analysis of variance and Tukey's test. [Figure 6]Phenotypic expression in treated and untreated Bbs1M390R / M390R demonstrates weight loss in P0 systemically treated animals. When treated intravenously with the AAV2 / 8-EFS-BBS1 construct, Bbs1M390R / M390R animals show attenuated weight gain to wild-type levels in males and females (see p-values). IV = intravenous. Het = heterozygous animals Bbs1M390R / M390R. WT = wild-type animals. M390R = homozygous animals Bbs1M390R / M390R. Error bars indicate standard error of the mean (SEM). p-values were obtained after nonlinear curve fitting (Gompertz growth curve fitting) followed by analysis of variance and Tukey's test. [Figure 7] Graphical representation of Gompertz growth nonlinear regression analysis. Gompertz growth best fitted to all nonlinear regression curve functions tested. A. Intracranial P0 treated male. B. Systemic P0 treated male. C. Intracranial P0 treated female. D. Systemic P0 treated female. IC = intracranial. IV = intravenous. [Figure 8] Serum leptin plasma concentration / body weight. AAV2 / 8-EFS-BBS1 IV and IC Bbs1M390R / M390R treated animals had reduced serum leptin concentrations 6 months after treatment. M390R = Bbs1M390R / M390R homozygous animals. WT = wild-type animals. IC = intracranial treatment. IV = intravenous treatment. [Figure 9] Eosin-hematoxylin retinal sections showing that AAV2 / 8-EFS-BBS1 IV-treated Bbs1M390R / M390R animals had attenuated photoreceptor loss 6 months after treatment. For quantification, see Figure 11. ONL = outer nuclear layer. IN = inner nuclear layer. All images were taken with a 40x objective. [Figure 10] Eosin-hematoxylin retinal sections showing that AAV2 / 8-EFS-BBS1 IC-treated Bbs1M390R / M390R animals had attenuated photoreceptor loss 6 months after treatment. For quantification, see Figure 11. ONL = outer nuclear layer. IN = inner nuclear layer. All images were taken with a 40x objective. [Figure 11]Quantification of retinal thickness in the outer nuclear layer section. The number of granules in the ONL recovered in Bbs1M390R / M390R treated animals is twice as high as in untreated animals. [Figure 12] A map of the vector, including EFS and human BBS1 (EFS-hBBS1), used in the following examples, shows key features. This vector was used to generate the AAV2 / 8-EFS-BBS1 virus. In certain embodiments of the invention, the promoter, ciliopathic gene, and restriction sites may vary depending on the final product. [Figure 13] Expression of human BBS1 by RT-PCR in the eyes and brain 52 weeks after P0 intracranial AAV2 / 8-EFS-BBS1 delivery. Clear expression of human BBS1 was observed in the eyes and brain of injected animals after intracranial injection (IC treatment) of AAV2 / 8-EFS-BBS1, indicating successful transduction and long-term persistent expression. Specific human primers were designed to distinguish between mouse and human BBS1, and no detection of mouse Bbs1 was observed. M390R = Bbs1M390R / M390R, WT: wild-type animals, UT: untreated, IC treatment: intracranial treatment. [Figure 14] Figure 1 Construct efficacy one year after injection. Twelve months after perinatal intracranial treatment with the construct AAV2 / 8-EFS-BBS1, Bbs1M390R / M390R-treated males continued to show weight loss compared to wild-type untreated littermates. WT = wild-type animals. M390R = Bbs1M390R / M390R homozygous animals. Error bars indicate standard error of the mean (SEM). [Figure 15]Circulating leptin levels after 52 weeks of treatment. Male and female Bbs1M390R / M390R animals show elevated leptin levels. Male AAV2 / 8-EFS-BBS1-treated animals have leptin levels similar to untreated wild-type animals. Treated Bbs1M390R / M390R vs. untreated wild-type is not significantly different (p-value 0.647). Untreated Bbs1M390R / M390R vs. treated Bbs1M390R / M390R has a significant p-value of 0.027. Female AAV2 / 8-EFS-BBS1-treated animals show significantly reduced leptin levels compared to untreated Bbs1M390R / M390R animals (p-value 0.041). M: male, F: female, WT: wild type; HOM: Bbs1M390R / M390R, UT: untreated, IC: intracranial. [Figure 16] Real-time PCR showing the expression of BBS1 after individual transfection with all constructs of SEQ ID NO: 15 to SEQ ID NO: 29. After transfection of the constructs, total RNA was extracted and the RNA levels were quantified and normalized to the construct with SEQ ID NO: 15 (EFS-WTBBS1). [Figure 17] Real-time PCR showing expression of BBS10 after individual transfection with all constructs of SEQ ID NO: 30 to SEQ ID NO: 44. After transfection of the constructs, total RNA was extracted and RNA levels were quantified and normalized to construct with untransfected HEK293T total mRNA. [Figure 18] Western blot showing BBS1 protein expression after individual transfection with constructs of SEQ ID NO: 15 to SEQ ID NO: 29. Blots were quantified using Image J, and BBS1 expression levels were normalized to SEQ ID NO: 15 (EFS-WTBBS1). [Figure 19] The blots in Figure 18 were quantified using Image J, and all BBS1 expression levels were normalized to SEQ ID NO: 15 (EFS WTBBS1). Nearly all constructs and the new BBS1 sequence showed increased BBS1 protein expression compared to the initial EFS-WTBBS1 construct. [Figure 20] RT-PCR of COSEQ1-BBS1 from samples of animals intracranially injected with P0 AAV2 / 9-CAG-COSEQ1-BBS1. Bbs1+M390R animals were sacrificed 8, 14, and 40 days after injection with P0 AAV2 / 9-CAG-COSEQ1-BBS1. Total RNA was extracted and cDNA was synthesized. PCR of COSEQ1-BBS1, with the predicted band of 188 nucleotides, was performed to detect gene expression in the brain and eye. P: days after injection; C: injected control vehicle; bp: base pairs. DETAILED DESCRIPTION OF THE INVENTION

[0080] Detailed Description of the Invention The inventors have developed a single vector that is administered by a simple injection and targets multiple organs at once. This approach is simpler and certainly more cost-effective than alternative multi-vector approaches. Restoring the function of more than one organ (e.g., vision and weight loss) could be life-changing, improving the health and quality of life of patients with ciliopathies.

[0081] Given the widespread organ involvement in ciliopathies, multisystemic treatments that address both central nervous system and visceral manifestations are needed. Furthermore, the early onset of symptoms in young children necessitates that such treatments ideally be administered as early as possible. Delivery of a single therapeutic agent in the neonatal period effectively targets multiple organs, prevents irreversible pathology, and is cost-effective.

[0082] To achieve these goals, we utilized a gene therapy-based protocol that utilizes adeno-associated viruses (AAVs) to achieve multi-organ therapy. Until recently, viral vectors capable of targeting peripheral organs and crossing the blood-brain barrier were unavailable, making multi-organ gene delivery difficult. However, AAV8 and AAV9 have been successfully used in mice (Foust KD et al. Nat Biotechnol. 2009;27(1):59-65) and non-human primates (Foust KD et al. Nat Biotechnol. 2010;28(3):271-4; Bevan AK et al. Mol Ther. 2011;19(11):1971-80). The discovery that AAV8 or AAV9 vectors can cross the blood-brain barrier and mediate highly efficient gene delivery to the central nervous system has transformed the outlook for this field. It is now feasible to explore simplification of vector administration, allowing for treatment of multiple organs with fewer administration routes rather than utilizing a single administration per organ regimen. For example, it has been demonstrated that intravenous administration of AAV8 or AAV9 vectors carrying the green fluorescent protein (GFP) gene in newborn mice leads to widespread and comprehensive transduction of the brain and nervous system, including the eye. Furthermore, the inventors have data showing that this approach also leads to widespread systemic transduction, including visceral organs and the muscular system (FASEB J. 2015 Sep;29(9):3876-88).

[0083] Multisystem and progressive disorders that present in infants, such as Bardet-Biedl syndrome, are ideal candidates for neonatal therapeutic AAV delivery. Because the most common mutation that causes BBS is found in BBS1, we developed a murine model of BBS, Bbs1. M390R / M390RWe tested AAV8 and AAV9 vectors carrying the human gene BBS1 in mice. This model is a "knock-in" of a common mutation and is well-validated and characterized to recapitulate human BBS phenotypes (e.g., blindness and obesity). Transcription of the exogenous BBS1 gene is driven by the ubiquitous mammalian EFS promoter, which is widely expressed within target organs. This approach demonstrates the ability of AAV8 and AAV9 to cross the blood-brain barrier and subsequently target neuronal defects, such as the retina, hippocampus / dentate gyrus, or hypothalamic appetite center, thereby restoring retinal function and obesity, respectively. Effective systemic gene transfer in these disease models will provide the necessary proof-of-principle, optimal dosing information, efficacy of recovery, and toxicity and safety profile of the vectors prior to clinical trials in patients.

[0084] We cloned human BBS1 cDNA under the control of the short elongation factor promoter (EFS) into an AAV-2 vector pseudotyped with the capsid protein from adeno-associated virus-8 (AAV2 / 8). These data demonstrate that the EFS-BBS1 construct efficiently transfects and expresses human BBS1 in HEK293T cells. After virus generation and infection via intracranial or systemic (IV) delivery of P0 pups, successful transduction was demonstrated in the retina and brain. No toxic effects were observed in treated mice. We demonstrated that the mutant Bbs1 M30R / M390R It could be demonstrated that when animals were treated, the obesity and retinal phenotype could be highly rescued.

[0085] The BBS1 nucleotide and amino acid sequences are highly conserved between humans and mice (92.2%). A knock-in mouse containing the M390R mutation in the Bbs1 gene, the most common mutation in patients, was generated (Proc Natl Acad Sci USA. 2007 Dec 4; 104(49): 19422-19427). M390R / M390RSerial histological examination of mouse retinas showed gradual and complete progressive degeneration of the inner and outer segments (IS and OS) by 6 months of age. M390R / M390R Electroretinograms (ERGs) of the knock-in mice showed significant attenuation of the a- and b-waves, with a lesser attenuation of the c-wave, suggesting that the degeneration preferentially affected cone and columnar photoreceptor cells, but not the retinal pigment epithelium (RPE). M390R / M390R The mice also developed obesity, which is associated with high serum levels of the adipocyte-derived hormone leptin, suggesting leptin resistance, increased food intake, and decreased locomotor activity. Numerous neuroanatomical defects are detected, including reduced size of key areas, the striatum and hippocampus. These phenotypes make this an ideal mouse model to recapitulate the human disease and evaluate novel treatments. Both mouse and human Bbs10 genes are encoded by two exons. Their proteins are conserved with 67% identical amino acid sequences. Bbs10 null (Bbs10 - / - ) mice completely lack exon 2 of Bbs10. - / - Bbs10 mice display a typical BBS phenotype, being perinatally dwarfed and developing obesity from 8 onwards, becoming overweight by 3 months of age. - / - Mice also develop hyperphagia and high levels of circulating leptin. - / - Mice develop severe retinal degeneration, with clear loss of the inner IS and OS of photoreceptors and the ONL by 3 months of age (Cilia 2015 4:10).

[0086] Materials and Methods Human BBS1 cDNA (SEQ ID NO:1 - NM_024649.4) A construct was generated that was cloned under the control of the EFS promoter (human eukaryotic translation elongation factor 1 alpha short promoter) within the AAV2 / 8 viral plasmid. Because the goal of this experimental design was to initiate viral gene therapy for Bardet-Biedl syndrome 1 (BBS1), an adeno-associated virus (AAV) containing human wild-type BBS1 cDNA and driven by the elongation factor 1 alpha short (EFS) promoter was generated. Standard methods were used for virus generation. 4000 cm 2 HEK293T cell monolayers were transfected with the EFS-BBS1-AAV-ITR-containing plasmid, the AAV2 Rep-Cap plasmid, and helper plasmids. Upon cytopathic effects, cells were harvested and lysed to release the virus. The adeno-associated virus was purified by centrifugation through two successive cesium chloride gradients. The final product was desalted and titered spectrophotometrically for viral particles and by plaque formation assay for PFU / IFU.

[0087] We also cloned BBS10 wild-type cDNA (SEQ ID NO: 1) under the control of the EFS promoter. Completely new codon-optimized sequences for BBS1 (SEQ ID NO: 2), BBS1 (SEQ ID NO: 11 and 12), and BBS10 (SEQ ID NO: 13 and 14) were cloned to improve gene expression and efficacy levels. The new sequences were cloned under the control of the EFS, CAG, CMV, CBA, and UBC promoters. All possible combinations of the described promoters with the described BBS1 and BBS10 sequences were cloned into the pAV-AAV-ITR-containing plasmid. The promoters were cloned between the SpeI and EcoRI restriction sites, and then the BBS coding sequence 3' downstream of the promoter was inserted using EcoRI and SalI restriction enzymes. Clones were sequenced to check for unwanted mutations in the promoter and coding sequences. All sequences containing the promoter and gene sequences are listed in SEQ ID NO: 15 to SEQ ID NO: Presented as 44.

[0088] To test for improved gene expression, HEK293T cells were transfected with 1 μg / μl of Lipofectamine 2000. All different constructs using DNA: pAV-EFS-WTBBS1, pAV-EFS-COSEQ1-BBS1, pAV-EFS-COSEQ2-BBS1, pAV-UBC-WTBBS1, pAV-UBC-COSEQ1-BBS1, pAV-UBC-COSEQ2-BBS1, pAV-CMV-WTBBS1, pAV-C MV-COSEQ1-BBS1, pAV-CMV-COSEQ2-BBS1, pAV-CBA-WTBBS1, pAV-CBA-COSEQ1-BBS1, pAV-CBA-COSEQ2-BBS1, pAV-CAG-WTBBS1, pAV-CAG-COSEQ1-BBS1, pAV-CAG-COSEQ2-BBS1, pAV- EFS-WTBBS10, pAV-EFS-COSEQ1-BBS10, pAV-EFS-COSEQ2-BBS10, pAV-UBC-WTBBS10, pAV-UBC-COSEQ1-BBS10, pAV-UBC-COSEQ2-BBS10, pAV-CMV-WTBBS10, pAV-CMV-COSEQ1-BBS10 , pAV-CMV-COSEQ2-BBS10, pAV-CBA-WTBBS10, pAV-CBA-COSEQ1-BBS10, pAV-CBA-COSEQ2-BBS10, pAV-CAG-WTBBS10, pAV-CAG-COSEQ1-BBS10, pAV-CAG-COSEQ2-BBS10.

[0089] Cells were harvested with 0.5 ml Trizol for total mRNA. Total mRNA was quantified, and real-time PCR was performed using 1 μg of mRNA for each transfection. Sequence-specific primers were used for each construct to quantify the level of human BBS1 expression. Ct value expression levels were normalized to EFS-BBS1 for all BBS constructs and to the untransfected sample for the BBS10 construct.

[0090] In a separate experiment, cells were also transfected for BBS1 protein expression. Total protein was extracted using RIPA buffer, and the total protein was quantified for each transfection. 1 μg / μl of sample protein for each transfection was loaded onto a 4-20% acrylamide gel. Western blots were performed with a specific antibody against BBS1, and the gels were scanned and analyzed. As a loading control, a second Western blot was performed with a GAPDH antibody. Blots were quantified by first normalizing each lane to GAPDH, and then normalizing each gel to EFS-WTBBS1 expression.

[0091] Virus dose and titer Bbs1 M390R / + Male and Bbs1 M390R / + Timed matings were arranged between females. P0 pups were genotyped for sex and Bbs1 genotype. The adenovirus-associated vector was administered intracranially (3.5 × 10 13 vg / ml (vector genome / ml) 5 μl) and systemically (IV) (3.5 × 10 13 The PO animals were given 20 μl of 20 μg / ml of 1000 mg ...

[0092] We investigated three different animal groups: Bbs1 M390R / M390R Animals, wild-type and heterozygous, were injected. Non-injected controls served as controls for each group. A total of 6 animals per group were used. Treated animals did not show any physical or behavioral distress 6 months after injection.

[0093] Based on the results with the codon-optimized sequence and construct, we decided to test the ability of new constructs to deliver and express human BBS1 in different tissues. The authors tested a new viral capsid (AAV2 / 9), a new CAG promoter, and a new CAG-COSEQ1-BBS1 construct (SEQ ID NO: 28). An AAV2 / 9 vector containing the CAG-COSEQ1-BBS1 construct (SEQ ID NO: 28) was generated and tested for efficacy in restoring Bbs1 activity by administering it to P0 newborn dogs. Bbs1 M390R / M390R For newborn animals, 0.175 × 10 12 vg / animal was injected intracranially in a 5 μl injection. Control, Bbs1 + / M390 and WT animals were injected with AAV2 / 9-CAG-COSEQ1-BBS1 or vehicle and tested for human BBS1 expression at 14 and 40 days post-injection.

[0094] result We demonstrate for the first time the use of gene therapy to treat multiple tissues affected by the ciliopathy Bardet-Biedl syndrome. Human BBS1 cDNA under the control of the ubiquitous promoter EFS transduced BBS1 protein expression in an AAV2 / 8 vector. Figure 1 shows the correct size of the cloned pAV-EFS-BBS1 after digestion (lane 4), demonstrating accurate cloning. To generate this AAV, producer plasmids pHGTI, pLT-AAV2-8, and pAV-EFS-BBS1 were obtained. The helper plasmid is pHGTI, which contains sequences encoding herpes simplex virus proteins. These are essential for efficient AAV production. The pLT-AAV2-8 plasmid contains sequences encoding the AAV rep and cap genes from AAV2 and AAV8, respectively. The rep gene is essential for AAV replication, and the cap gene encodes the capsid protein and determines AAV tropism. Finally, the pLT-AAV2-8 plasmid contains the AAV inverted terminal sequences, controlled by the EFS promoter, along with the BBS1 cDNA. It is these sequences that are packaged within the AAV and delivered to cells.

[0095] To assess whether the resulting producer plasmids were as expected, restriction enzyme digests were performed by digesting pLT-AAV2-8, pHGTI, and pAV-EFS-BBS1 plasmid DNA (Figure 1). The band sizes for the pLT-AAV2-8 plasmid were 186 bp, 2109 bp, and 4844 bp, respectively (lane 3, Figure 1). For the transgene pAV-EFS-BBS1 construct, the band sizes were 236 bp, 643 bp, 1195 bp, and 5078 bp (lane 4, Figure 1). For the helper plasmid, the visible band sizes were 6318 bp and 11549 bp, respectively (lane 2, Figure 1), verifying the absence of additional unwanted DNA within the plasmids.

[0096] High expression of BBS1 was observed when HEK293T cells were transfected with the pAV-EFS-BBS1 plasmid. This data indicates that EFS can drive human BBS1 expression in vitro (Figure 2). We generated and purified AAV2 / 8-EFS-BBS1 and injected it into P0 wild-type embryos via intracranial and systemic delivery. The transduction ability of AAV2 / 8-EFS-BBS1 was demonstrated in both retinal and brain tissues. Specific expression of human BBS1 in the retina and brain was observed (Figures 3 and 4).

[0097] Bbs1 M390R / M390R Functional studies were performed to assess the efficacy of BBS1 expression in mouse models. Wild-type, heterozygous Bbs1 M390R / + and Bbs1 M390R / M390R Littermates were injected with AAV2 / 8-EFS-BBS1 at PO. In parallel, cohorts of untreated animals from all three genotypes were bred as controls. We followed this cohort for 26 weeks, measuring the body weight of each animal weekly. Both intracranial and systemic injections of Bbs1 M390R / M390RSignificant improvements were demonstrated in weight maintenance in animals (Figures 5-7). No difference in weight was observed between wild-type animals treated with AAV2 / 8-EFS-BBS1 and untreated animals. Human BBS1 was found to be expressed in the eye and retina 52 weeks after intracranial injection, indicating that human BBS1 expression levels persist for at least one year (Figure 13). Improvements in weight regulation were also maintained throughout the year, as shown in Figure 14. Nonlinear regression, one-way analysis of variance followed by Tukey's test statistical analysis, demonstrated that weight rescue was significantly greater in Bbs1 treated with AAV2 / 8-EFS-BBS1 compared to untreated animals. M390R / M390R The difference was statistically significant in the animals treated with AAV2 / 8-EFS-BBS1. Serum leptin levels were analyzed. M390R / M390R Normal leptin levels were restored in animals (Figure 8). Normal leptin levels were maintained for 52 weeks in both males and females treated with AAV2 / 8-EFS-BBS1 at birth (Figure 15).

[0098] Bbs1 treated with AAV2 / 8-EFS-BBS1 at P0 M390R / M390R The animals also showed an attenuated loss of external granule cell (ONL) numbers, which was greater than that observed in untreated Bbs1 mice. M3 90R / M390R This effect was demonstrated by quantifying the number of surviving photoreceptor granules present in the retinas of treated animals at 6 months compared to untreated animals. This effect was observed in both the intravenous and intracranial treatment groups (Figures 9, 10, and 11).

[0099] Using the novel codon-optimized sequences, the researchers demonstrated that the relative expression of BBS1 mRNA improved after transfection of the BBS1 construct (see Figure 16). The CMV promoter showed the highest level of expression, followed by the CAG promoter. The BBS1 codon-optimized sequences, COSEQ1-BBS1 and COSEQ2-BBS1, had significantly better expression than wild-type human BBS1 cDNA. The constructs with better expression were CMV-COSEQ1-BBS1 (19-fold increase), CMV-WTBBS1 (10-fold increase), and CAG-COSEQ1-BBS1 (7-fold increase). All expression was normalized to EFS-WTBBS1 expression. Transfection with the novel BBS10 construct also demonstrated increased human BBS10 expression, and in most promoter-BBS10 sequence combinations, the new codon-optimized BBS10 sequence delivered better yields of BBS10 RNA (Figure 17).

[0100] Western blots of protein extracts from transfections of all BBS1 constructs showed increased protein expression for all BBS1 constructs. Specific bands for BBS1 (65 kDa) and GAPDH (38 kDa) were detected (see Figure 18). Protein abundance analysis and normalization were performed as shown in Figure 19.

[0101] Analysis of the gel shows how the new codon-optimized sequences, COSEQ1-BBS1 and COSEQ1-BBS2, are able to express BBS1 better than wild-type BBS1, regardless of the promoter used to drive expression. The highest expression was found with the sequence COSEQ1-BBS1, which achieved a 33-fold increase with the CMV promoter and a 24-fold increase with the CAG promoter.

[0102] We demonstrated expression of the new codon-optimized sequence COSEQ1-BBS1 in the AAV2 / 9-CAG-COSEQ1-BBS1 construct in the brain and eye 40 days after vector transduction (see Figure 20). To check the specificity of the band, the band was excised, washed, and Sanger sequenced to observe the correct sequence of COSEQ1-BBS1.

[0103] Consideration These results demonstrate that we were able to deliver a functional human BBS1 gene expressing wild-type BBS1 protein to multiple affected organs with a single administration of a gene therapy vector. Increased expression of WT Bbs1 in the CNS was followed by restoration of hypothalamic leptin-regulated function, as indicated by weight loss and reduced circulating leptin levels. Similarly, expression of human BBS1 in the eye was followed by attenuation of retinal degeneration 6 months after systemic delivery (Figures 3-15).

[0104] The multi-systemic nature of most ciliopathies makes it difficult to treat several or all different affected organs with a single procedure. Even in situations where gene therapies are currently being developed to treat specific organs, the gene therapy will only be useful for that specific phenotype and will not treat the disease more generally.

[0105] All ciliopathies have the same organs affected with different severity (see review in N Engl J Med 2011; 364:1533-1543 April 21, 2011). Of all of them, BBS is more BBS is a type of ciliopathies in which many organs are directly affected by mutations in the BBS gene. The present inventors have demonstrated that gene therapy can target multiple affected tissues with the administration of a single vector. Therefore, the present invention provides a method for targeting specific ciliopathies genes in affected organs with a single administration. It may be possible to target and restore function.

[0106] Even when a ciliopathic disease primarily affects a single organ, as is the case for some ciliary dysfunctional retinal disorders, intravenous, intracranial, and / or intravenous and intracranial administration may be more effective and less risky than current techniques for subretinal treatment.

[0107] Treatment of other ciliary diseases The previously described experiments demonstrate that systemic expression of a protein that replaces the function of a mutant gene responsible for a ciliopathic disease, in this case Bardet-Biedl syndrome, can be an effective method for treating some or all organs affected by the disease. This therefore provides a more effective method for treating ciliopathies than previous attempts. All ciliopathies are part of a similar spectrum of disorders that affect cilia function or structure in some way. The phenotypic relevance of these associations means that the same gene has been found to be causally involved in more than one ciliopathic disease. Shared genes can be found in different ciliopathies; for example, MKKS / BBS6 is associated with Bardet-Biedl syndrome and McKusick-Kaufman syndrome. The shared phenotypic expression, meaning the same organs are affected, and the genetic homogeneity, meaning the same gene is involved in more than one ciliopathic disease, make this gene delivery and expression method of the present invention a unique approach for treating many ciliopathies. Therefore, this approach is not limited to Bardet-Biedl syndrome but is applicable to many ciliopathies. Furthermore, all ciliopathies are caused by mutations within a single gene, and therefore systemic expression of the appropriate non-mutated gene will systemically ameliorate ciliopathic pathology.

[0108] As previously demonstrated, Bardet-Biedl syndrome can be treated using this gene therapy approach. The table below lists a number of genes that can become mutated and cause the phenotypic pathology associated with Bardet-Biedl syndrome. Therefore, Bardet-Biedl syndrome can be treated using the previously described gene therapy vectors containing the appropriate genes that express wild-type, non-mutated proteins.

[0109] In addition, some of the genes associated with Bardet-Biedl syndrome have also been implicated in other related ciliopathies. Consequently, the above approach using the appropriate genes can also be used to treat other ciliopathies, such as Joubert syndrome, Meckel-Gruber syndrome, medullary cystic kidney disease, Senior-Loken syndrome, McKusick-Kaufman syndrome, and Leber congenital amaurosis. For example, McKusick-Kaufman syndrome is caused by a mutation in the MKKS / BBS6 gene. Therefore, McKusick-Kaufman syndrome and Bardet-Biedl syndrome can be treated or ameliorated using a vector that provides expression of the MKKS / BBS6 gene so that wild-type MKKS / BBS6 protein is expressed. This also applies to various other ciliopathies referenced in the table below.

[0110] [Table 1]

[0111] array SEQ ID NO. 1 - Human Bardet-Biedl syndrome 1 (BBS1) nucleotide sequence (WT), cDNA (NM_024649.4) SEQ ID NO. 2 - Human Bardet-Biedl syndrome 10 (BBS10) nucleotide sequence (WT), cDNA (NM_024685.3) SEQ ID NO. 3 - Short elongation factor (EFS) promoter sequence SEQ ID NO. 4 - CAG promoter sequence SEQ ID NO. 5 - Ubiquitin C (UBC) promoter sequence SEQ ID NO. 6 - cytomegalovirus (CMV) immediate early promoter sequence; SEQ ID NO. 7 - phosphoglycerate kinase (PGK) promoter sequence; SEQ ID NO. 8 - chicken beta actin (CBA) promoter sequence. SEQ ID NO. 9 - Human BBS1 full-length protein sequence (Q8NFJ9) SEQ ID NO. 10 - Human BBS10 full-length protein sequence (Q8TAM1) SEQ ID NO. 11 - Codon-optimized nucleotide sequence encoding human BBS1 protein (referred to as COSEQ1-BBS1) SEQ ID NO. 12 - Codon-optimized nucleotide sequence encoding human BBS1 protein (designated COSEQ2-BBS1) SEQ ID NO. 13—Codon-optimized nucleotide sequence encoding human BBS10 protein (designated COSEQ1-BBS10) SEQ ID NO. 14 - Codon-optimized nucleotide sequence encoding human BBS10 protein (designated COSEQ2-BBS10) SEQ ID NO 15 - EFS promoter (nt 41-272) and wild-type B A construct containing the BS1 nucleotide sequence (nt 1238 to 3019) (designated EFS-WTBBS1) SEQ ID NO 16 - EFS promoter (nt 41-272) and COSE A construct containing the Q1-BBS1 nucleotide sequence (nt 1243 to 3024) (EFS-C (referred to as OSEQ1-BBS1) SEQ ID NO 17 - Construct containing the EFS promoter (nt 41 to 272) and the COSEQ2-BBS1 nucleotide sequence (nt 1243 to 3024) (designated EFS-COSEQ2-BBS1) SEQ ID NO 18 - UBC promoter (nt 29-1198) and wild type A construct containing the BBS1 nucleotide sequence (nt 1281 to 3062) (designated UBC-WTBBS1) SEQ ID NO 19 - UBC promoter (nt 29-1198) and COS A construct containing the EQ1-BBS1 nucleotide sequence (nt 1285 to 3066) (designated UBC-COSEQ11BBS1) SEQ ID NO 20 - UBC promoter (nt 29-1198) and COS A construct containing the EQ2-BBS1 nucleotide sequence (nt 1285 to 3066) (designated UBC-COSEQ2-BBS1) SEQ ID NO 21 - CMV promoter (nt 367-570) and wild type A construct containing the BBS1 nucleotide sequence (nt 626 to 2407) (designated CMV-WTBBS1) SEQ ID NO 22 - CMV promoter (nt 367-570) and COS A construct containing the EQ1-BBS1 nucleotide sequence (nt 630 to 2411) (designated CMV-COSEQ1-BBS1) SEQ ID NO 23 - CMV promoter (nt 367-570) and COS A construct containing the EQ2-BBS1 nucleotide sequence (nt 630 to 2411) (designated CMV-COSEQ2-BBS1) SEQ ID NO 24 - CBA promoter (nt 42-319) and wild-type B A construct containing the BS1 nucleotide sequence (nt 469 to 2250) (designated CBA-WTBBS1) SEQ ID NO 25 - CBA promoter (nt 42-319) and COSE A construct containing the Q1-BBS1 nucleotide sequence (nt 473 to 2254) (designated CBA-COSEQ1-BBS1) SEQ ID NO 26 - CBA promoter (nt 42-319) and COSE A construct containing the Q2-BBS1 nucleotide sequence (nt 473 to 2254) (designated CBA-COSEQ2-BBS1) SEQ ID NO 27 - CAG promoter (nt 35-562) and wild-type B A construct containing the BS1 nucleotide sequence (nt 712 to 2493) (designated CAG-WTBBS1) SEQ ID NO 28 - CAG promoter (nt 35-562) and COSE A construct containing the Q1-BBS1 nucleotide sequence (nt 716 to 2497) (designated CAG-COSEQ1-BBS1) SEQ ID NO 29 - Construct containing the CAG promoter (nt 35 to 562) and the COSEQ2-BBS1 nucleotide sequence (nt 716 to 2497) (designated CAG-COSEQ2-BBS1) SEQ ID NO 30 - EFS promoter (nt 41-272) and wild-type B A construct containing the BS10 nucleotide sequence (nt 1243 to 3414) (designated EFS-WTBBS10) SEQ ID NO 31 - EFS promoter (nt 41-272) and COSE A construct containing the Q1-BBS10 nucleotide sequence (nt 1243 to 3414) (designated EFS-COSEQ1-BBS10) SEQ ID NO 32 - EFS promoter (nt 41-272) and COSE A construct containing the Q2-BBS10 nucleotide sequence (nt 1243 to 3414) (designated EFS-COSEQ2-BBS10) SEQ ID NO 33 - UBC promoter (nt 29-1198) and wild type A construct containing the BBS10 nucleotide sequence (nt 1285 to 3456) (designated UBC-WTBBS10) SEQ ID NO 34 - UBC promoter (nt 29-1198) and COS A construct containing the EQ1-BBS10 nucleotide sequence (nt 1285 to 3456) (designated UBC-COSEQ1BBS10) SEQ ID NO 35 - UBC promoter (nt 29-1198) and COS A construct containing the EQ2-BBS10 nucleotide sequence (nt 1285 to 3456) (designated UBC-COSEQ2-BBS10) SEQ ID NO 36 - CMV promoter (nt 367-570) and wild type A construct containing the BBS10 nucleotide sequence (nt 630 to 2801) (designated CMV-WTBBS10) SEQ ID NO 37 - CMV promoter (nt 367-570) and COS A construct containing the EQ1-BBS10 nucleotide sequence (nt 630 to 2801) (designated CMV-COSEQ1-BBS10) SEQ ID NO 38 - CMV promoter (nt 367-570) and COS A construct containing the EQ2-BBS10 nucleotide sequence (nt 630 to 2801) (designated CMV-COSEQ2-BBS10) SEQ ID NO 39 - CBA promoter (nt 42-319) and wild-type B A construct containing the BS10 nucleotide sequence (nt 473 to 2644) (designated CBA-WTBBS10) SEQ ID NO 40 - CBA promoter (nt 42-319) and COSE A construct containing the Q1-BBS10 nucleotide sequence (nt 473 to 2644) (designated CBA-COSEQ1-BBS10) SEQ ID NO 41 - CBA promoter (nt 42-319) and COSE A construct containing the Q2-BBS10 nucleotide sequence (nt 473 to 2644) (designated CBA-COSEQ2-BBS10) SEQ ID NO 42 - CAG promoter (nt 35-562) and wild-type B A construct containing the BS10 nucleotide sequence (nt 716 to 2887) (designated CAG-WTBBS10) SEQ ID NO 43 - CAG promoter (nt 35-562) and COSE A construct containing the Q1-BBS10 nucleotide sequence (nt 716 to 2887) (designated CAG-COSEQ1-BBS10) SEQ ID NO 44 - CAG promoter (nt 35-562) and COSE A construct containing the Q2-BBS10 nucleotide sequence (nt 716 to 2887) (designated CAG-COSEQ2-BBS10) SEQ ID NO. 45 - Alternative CMV promoter sequence SEQ ID NO. 46 - Alternative short elongation factor (EFS) promoter sequence SEQ ID NO. 47 - Alternative CAG promoter sequence SEQ ID NO. 48 - Alternative Ubiquitin C (UBC) promoter sequence SEQ ID NO. 49 - Alternative chicken beta actin (CBA) promoter sequence

[0112] Other aspects of the present disclosure include, for example, the following. [1] A vector for treating a ciliopathic disease, the vector comprising a promoter operably linked to a ciliopathic gene, the vector capable of providing transduction of the ciliopathic gene into multiple organs, the promoter being a ubiquitous promoter capable of providing expression of the ciliopathic gene in the transduced organs, and the ciliopathic gene encoding the protein corresponding to a functional human protein mutated in a ciliopathic disease. [2] Adeno-associated virus (AAV) vector or lentivirus vector, [1 ] A vector described in [3] The vector according to [1] or [2], which is an AAV vector. [4] The vector according to any one of [1] to [3], which is selected from AAV8, AAV9, AAV vectors pseudotyped with capsid proteins from AAV8 or AAV9, AAV-PHP.A, AAV-PHP.B, AAV9.47, AAV-B1, AAV8(Y733F), or AAV2-TT. [5] The vector according to any one of [1] to [4], which is an AAV8 vector, an AAV9 vector, or an AAV vector pseudotyped with a capsid protein from AAV8 or AAV9. [6] The vector according to any one of [1] to [5], wherein the promoter is selected from the group consisting of a short elongation factor promoter (EFS), a CAG promoter, a cytomegalovirus immediate early promoter (CMV), a ubiquitin C promoter (UBC), a phosphoglycerate kinase promoter (PGK), and a beta-actin promoter. [7] The promoter is selected from SEQ ID NO. 3, SEQ ID NO. 4, SEQ The vector according to any one of [1] to [6], having a sequence selected from SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 and SEQ ID NO.45. [8] The vector according to any one of [1] to [7], wherein the promoter is a CAG promoter. [9] The vector according to any one of [1] to [8], wherein the promoter is a CAG promoter which may have the nucleotide sequence of SEQ ID NO. 4 or SEQ ID NO. 47.

[10] The vector according to any one of [1] to [9], wherein the ciliopathy gene encodes a functional human protein selected from BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10, BBS11 / TRIM32, BBS12, BBS13 / MKS1, BBS14 / CEP290, BBS15 / C2ORF86, BBS16 / SDCCAG8, BBS17 / LZTFL1, BBS18 / BBIP1, BBS19 / IFT27, BBS20 / IFT74, and BBS21 / C8ORF37 proteins.

[11] The vector described in any one of [1] to

[10] , wherein the ciliopathy gene encodes a functional human protein selected from BBS1, BBS2, BBS3 / ARL6, BBS4, BBS5, BBS6 / MKKS, BBS7, BBS8, BBS9, BBS10 and BBS12 proteins.

[12] The vector according to any one of [1] to

[11] , wherein the ciliopathy gene encodes a functional human protein selected from BBS1 and BBS10 proteins.

[13] A vector described in any of [1] to

[12] , wherein the ciliopathic gene has the nucleotide sequence of SEQ ID NO. 1 or has at least 70% sequence identity thereto and encodes a functional human BBS1 protein.

[14] The ciliopathic gene has the nucleotide sequence of SEQ ID NO. 11 or 12. The vector according to any one of [1] to

[13] ,

[15] A vector described in any of [1] to

[12] , wherein the ciliopathic gene has the nucleotide sequence of SEQ ID NO. 2 or has at least 70% sequence identity thereto and encodes a functional human BBS10 protein.

[16] The ciliopathic gene has the nucleotide sequence of SEQ ID NO. 13 or 14. The vector according to any one of [1] to

[12] and

[15] ,

[17] A vector described in any of [1] to

[14] , wherein the ciliopathic gene encodes a functional human BBS1 protein having the protein sequence of SEQ ID NO 9 or has 80% sequence identity thereto.

[18] The ciliopathic gene encodes a functional human BBS10 protein having the protein sequence of SEQ ID NO: 10 or having 80% sequence identity thereto. The vector according to any one of [1] to

[12] and

[15] .

[19] The vector according to any one of [1] to

[10] , wherein the ciliopathy is selected from Bardet-Biedl syndrome, McKusick-Kaufmann syndrome, Joubert syndrome, Meckel-Gruber syndrome, medullary polycystic kidney disease, Senior-Loken syndrome, and Leber congenital amaurosis.

[20] The vector according to any one of [1] to

[19] , wherein the ciliopathy is Bardet-Biedl syndrome.

[21] The vector according to any one of [1] to

[20] , which is an AAV8 vector, an AAV9 vector, or an AAV vector pseudotyped with a capsid protein from AAV8 or AAV9, the promoter is a CAG promoter, A vector wherein the ciliopathic gene encodes a functional human protein selected from BBS1 and BBS10 proteins.

[22] The ciliopathic gene is selected from the group consisting of SEQ ID NOs. 11, 12, 13, and 14. A vector according to

[21] , having one nucleotide sequence.

[23] A pharmaceutical composition comprising the vector according to any one of [1] to

[22] and one or more pharmaceutically acceptable excipients.

[24] A method for treating a ciliopathic disease, comprising administering a therapeutically effective amount of the vector according to any one of [1] to

[22] to a patient suffering from the ciliopathic disease.

[25] The method according to

[24] , wherein the vector is administered intravenously.

[26] The method according to

[24] , wherein the vector is administered intracranially.

[27] The method of

[24] , wherein the vector is administered intravenously and intracranially.

[28] The method of

[27] , wherein the vector is administered intravenously and intracranially on the same day.

[29] The method of

[27] , wherein the vector is administered intravenously and intracranially simultaneously.

[30] The method according to any one of

[24] to

[29] , wherein the vector is additionally administered intrathecally.

[31] The method according to any one of

[24] to

[30] , wherein the vector is administered at a single time point and not repeatedly administered.

[32] The vector according to any one of [1] to

[22] for use in therapy.

[33] The vector according to any one of [1] to

[22] for use in treating ciliary diseases.

[34] Use of the vector according to any one of [1] to

[22] in the manufacture of a drug for treating a ciliopathic disease.

[35] The use according to

[33] or

[34] , wherein the vector is for intravenous administration.

[36] The use according to

[33] or

[34] , wherein the vector is for intracranial administration.

[37] The use according to

[33] or

[34] , wherein the vector is for intravenous and intracranial administration.

[38] The use according to

[37] , wherein the vector is for intravenous and intracranial administration on the same day.

[39] The use according to

[37] , wherein the vector is for simultaneous intravenous and intracranial administration.

[40] The use according to any one of

[33] to

[39] , wherein the vector is for additional intrathecal administration.

[41] The use according to any one of

[33] to

[40] , wherein the vector is for administration at a single time point and is not administered repeatedly.

Claims

1. A pharmaceutical composition for treating a ciliary disease, comprising a vector, the vector comprises a promoter operably linked to a ciliopathic gene; the promoter is a ubiquitous promoter capable of providing expression of the ciliopathic gene in the transduced organ; The ciliopathies gene has the nucleotide sequence of SEQ ID NO. 11 or 12 or has at least 90% sequence identity thereto and encodes a functional human BBS1 protein, and the ciliopathies are ciliopathies caused by mutations in the BBS1 gene, or The ciliopathies gene has the nucleotide sequence of SEQ ID NO. 13 or 14 or has at least 90% sequence identity thereto and encodes a functional human BBS10 protein, and the ciliopathies are ciliopathies caused by mutations in the BBS10 gene.

2. The pharmaceutical composition described in claim 1, wherein the vector is an adeno-associated virus (AAV) vector or a lentivirus vector.

3. A pharmaceutical composition described in claim 1 or 2, wherein the vector is an AAV vector.

4. A pharmaceutical composition described in any one of claims 1 to 3, wherein the vector is selected from AAV8, AAV9, an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9, AAV-PHP.A, AAV-PHP.B, AAV9.47, AAV-B1, AAV8(Y733F), or AAV2-TT.

5. A pharmaceutical composition described in any one of claims 1 to 4, wherein the vector is an AAV8 vector, an AAV9 vector, or an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9.

6. The promoter is selected from the group consisting of a short elongation factor promoter (EFS), a CAG promoter, a cytomegalovirus immediate early promoter (CMV), a ubiquitin C promoter (UBC), a phosphoglycerate kinase promoter (PGK), and a beta-actin promoter. The pharmaceutical composition according to any one of claims 1 to 5, wherein the pharmaceutical composition is selected from the group consisting of:

7. The pharmaceutical composition of any one of claims 1 to 6, wherein the promoter has a sequence selected from SEQ ID NO. 3, SEQ ID NO. 4, SEQ ID NO. 5, SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8, and SEQ ID NO.

45.

8. A pharmaceutical composition described in any one of claims 1 to 7, wherein the promoter is a CAG promoter.

9. The pharmaceutical composition according to any one of claims 1 to 8, wherein the promoter is a CAG promoter having the nucleotide sequence of SEQ ID NO. 4 or SEQ ID NO.

47.

10. The method of claim 1, wherein the vector is selected from AAV8, AAV9, or an AAV vector pseudotyped with capsid proteins from AAV8 or AAV9; The pharmaceutical composition according to any one of claims 1 to 9, wherein the promoter is a CAG promoter.

11. A pharmaceutical composition described in any one of claims 1 to 10, characterized in that it is administered intravenously and / or intracranially and provides transduction of the ciliopathic disease gene in multiple organs.

12. A pharmaceutical composition according to any one of claims 1 to 11, comprising one or more pharmaceutically acceptable excipients.

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