Retinitis pigmentosa is a collective term for a group of inherited retinal dystrophies characterized by loss of rod photoreceptors, followed by loss of cone photoreceptors. It is a disease with a... Show moreRetinitis pigmentosa is a collective term for a group of inherited retinal dystrophies characterized by loss of rod photoreceptors, followed by loss of cone photoreceptors. It is a disease with a variable clinical and genetic presentation, with regards to age at onset, severity, and disease progression. For many patients, no treatment is available at the moment, but promising advances are made in genomic medicine. In preparation for these novel therapies, this thesis focuses on the clinical characteristics and natural course of candidate genes, in order to define optimal clinical endpoints. Additionally, this thesis investigates current treatment modalities, as retinitis pigmentosa is associated with ocular comorbidities such as cataract and cystoid macular edema, which can significantly impact a patient's quality of life when left untreated. Show less
Biallelic CRB1 gene variations can cause retinitis pigmentosa (RP), Leber congenital amaurosis, or in some cases macular degeneration. This thesis describes the generation and analysis of RP-CRB1... Show moreBiallelic CRB1 gene variations can cause retinitis pigmentosa (RP), Leber congenital amaurosis, or in some cases macular degeneration. This thesis describes the generation and analysis of RP-CRB1 mouse and human retinas (mouse: Crb1KOCrb2LowMGCs; chapter 2. Human RP-CRB1-patient-derived organoids (chapter 4 and 5). The data indicates that the human RP-CRB1 disease can be studied in mice and human organoids. Then, we show that recombinant adeno-associated viral (rAAV)-CRB gene supplementation therapy to Müller glial cells (MGCs) of the Crb1KOCrb2LowMGCs mouse retina can protect it from stress-induced vision loss, and that human CRB2 cDNA was superior to human CRB1 cDNA (chapter 2). We then developed an improved rAAV tropism assay on human donor eyes (chapter 3). This assay shows that rAAV5 can efficiently infect Müller glial cells and photoreceptors, the target cells of a RP-CRB1 gene therapy. Also, rAAV5 infection studies outperformed rAAV9 on human retinal organoids and human donor retinas (chapter 4). Finally, we find much more early endosomes and an increase of the degradative cellular vesicles which is linked to decrease of RAB11A-postive recycling endosomes in RP-CRB1 patient organoids (chapter 5). Thus, this thesis on both human and mouse models provides new insight into retinal degeneration and rAAV gene supplementation therapies. Show less
Retinal dystrophies comprise relatively rare but devastating causes of progressive vision loss. They represent a spectrum of diseases with marked genetic and clinical heterogeneity. Mutations in... Show moreRetinal dystrophies comprise relatively rare but devastating causes of progressive vision loss. They represent a spectrum of diseases with marked genetic and clinical heterogeneity. Mutations in the same gene may lead to different diagnoses, e.g. retinitis pigmentosa or cone dystrophy. Conversely, mutations in different genes may lead to the same phenotype. The age at symptom onset, as well as the rate of vision decline, may vary widely per disease group and even within families. For most IRD cases, no effective treatment is currently available. However, preclinical studies and phase I/II/III gene therapy trials are ongoing for several IRD subtypes, and recently the first retinal gene therapy has been approved by the United States Food and Drug Administration for RPE65-associated IRDs: voretigene neparvovec-rzyl (Luxturna®). With these rapid advances in gene therapy studies, insight into the phenotypic spectrum and long-term disease course becomes crucial. The vast clinical heterogeneity presents an important challenge in the evaluation of potential efficacy in future treatment trials, and in establishing treatment candidacy criteria. This thesis responds to these challenges, providing detailed clinical descriptions of several forms of IRD that are caused by genes of interest for ongoing and future gene or cell-based therapy trials. Show less
By investigating the roles of CRB proteins in mouse, non-human-primate, human fetal retina, and iPSC-derived retinal organoids, this thesis describes important insights to pathobiology in CRB1... Show moreBy investigating the roles of CRB proteins in mouse, non-human-primate, human fetal retina, and iPSC-derived retinal organoids, this thesis describes important insights to pathobiology in CRB1-retinitis pigmentosa (RP) and CRB1-Leber congenital amaurosis (LCA) disease models. The thesis describes AAV gene and cell therapy-based tools as therapeutic strategies for alleviation of RP and LCA due to loss of CRB1. Show less
Inherited retinal diseases encompass a large group of clinically and genetically heterogeneous diseases estimated to affect two million people worldwide. Among these people, approximately 80,000... Show moreInherited retinal diseases encompass a large group of clinically and genetically heterogeneous diseases estimated to affect two million people worldwide. Among these people, approximately 80,000 are or will become blind in their first decades of life due to mutations in both alleles of the Crumbs homologue-1 (CRB1) gene. Microglia are the resident immune surveyor cells in the retina, and their roles have been heavily studied in several retinal diseases, including retinitis pigmentosa (RP), age-related macular degeneration, and diabetic retinopathy. However, very little is known about the role of microglia in CRB1-associated retinopathies. Thus, we here summarize the main findings described in the literature concerning inflammation and the role of microglia in CRB1-patients and CRB1-rodent models. Show less
