3D modeling and RNA-based therapeutics for Dutch-type cerebral amyloid angiopathy

Dutch-type cerebral amyloid angiopathy (D-CAA) is a rare neurodegenerative/neurovascular disease caused by a point mutation in the amyloid precursor protein (APP) gene that leads to aggregation of the amyloid beta peptide in the brain vasculature. It is characterized by intracerebral hemorrhages, infarcts, cognitive decline and vascular dementia. To date there are no therapies that prevent or delay D-CAA onset or progression. In this thesis we aimed to model the disease using patient-derived cell models, develop and test an RNA-targeting therapy and look into the mutated protein trafficking in the cell. Using D-CAA patient-derived induced pluripotent stem cells as template, we generated state-of-the-art 3D brain organoids and successfully modeled the amyloid beta aggregation and other D-CAA disease signatures, also found in the D-CAA human brain. We developed and tested an RNA-targeting therapy that showed efficient and significant reduction of the amyloid beta peptide in D-CAA patient-derived cells and control mice. Finally, when looking into the mutated APP protein trafficking we have found that the Dutch mutation affects the processing and trafficking of APP protein prior to the generation of the amyloid beta fragment, a phenotype that was reversed when the RNA-targeting therapy was applied. With the data generated in this thesis we hope to further advance the knowledge on D-CAA disease mechanisms as well as the possibility for therapeutics that will benefit D-CAA patients.LUMC / Geneeskund

Students´ Knowledge about Nanotechnology and the Importance to Introduce Nanotechnology into Chemistry Lessons

The primary objective of this study was to find out what students already know about nanotechnology in order to design a context based module with a nanotechnology background. Therefore a questionnaire was distributed to 116 German students in grade 11. Questions referred to the first thought of the students´ mind when they hear the word nanotechnology, to the size of a nanometer, to if something can be seen at the nanometer scale with the unaided eye, to their self-assessment concerning nanotechnology and to the surface-to-volume ratio of nanoparticles. The findings of the students´ answers were informative and allowed us to design a module in the future that can lead students at school level to understanding nanotechnology

Antisense oligonucleotide-induced amyloid precursor protein splicing modulation as a therapeutic approach for Dutch-type cerebral amyloid angiopathy

Dutch-type cerebral amyloid angiopathy (D-CAA) is a monogenic form of cerebral amyloid angiopathy and is inherited in an autosomal dominant manner. The disease is caused by a point mutation in exon 17 of the amyloid precursor protein (APP) gene that leads to an amino acid substitution at codon 693. The mutation is located within the amyloid beta (A beta) domain of APP, and leads to accumulation of toxic A beta peptide in and around the cerebral vasculature. We have designed an antisense oligonucleotide (AON) approach that results in skipping of exon 17, generating a shorter APP isoform that lacks part of the A beta domain and the D-CAA mutation. We demonstrate efficient AON-induced skipping of exon 17 at RNA level and the occurrence of a shorter APP protein isoform in three different cell types. This resulted in a reduction of A beta 40 in neuronally differentiated, patient-derived induced pluripotent stem cells. AON-treated wild-type mice showed successful exon skipping on RNA and protein levels throughout the brain. These results illustrate APP splice modulation as a promising therapeutic approach for D-CAA.Functional Genomics of Muscle, Nerve and Brain Disorder

Generation of 3 human induced pluripotent stem cell lines LUMCi005-A, B and C from a Hereditary Cerebral Hemorrhage with Amyloidosis-Dutch type patient

Hereditary Cerebral Hemorrhage with Amyloidosis-Dutch type (HCHWA-D) is an autosomal dominant hereditary disease caused by a point mutation in exon 17 of the APP gene. We generated human induced pluripotent stem cells (hiPSCs) from a symptomatic HCHWA-D patient by using non-integrating Sendai virus (SeV). The newly generated hiPSCs express all pluripotency markers, have a normal karyotype, carry the Dutch mutation, can differentiate in the three germ layers in vitro and are SeV free