Therapy1 | Main Goals

In the period of 2015 to 2020, the group plans to further implement current and novel advanced molecular approaches aiming (i) to study biological samples from patients (peripheral blood, fibroblasts, cancer biopsies, post mortem brain tissue), both in culture and, with some of them, upon implantation in animals, to further validate/correlate with goal (ii) which is, to establish in vitro (induced pluripotent stem cells, neural stem cells, cancer stem cells) and in vivo (vector-mediated knock-down or overexpression in the brain and subcutaneous and orthotopic tumors) models of disease, (iii) . The knowledge gathered from these approaches will help define (iv) novel therapeutic targets, and (v) lead to the new molecular therapeutic candidates, which will be tested in previously validated cellular and animal models; the effects, at the molecular level, of these novel candidates on the identified deregulated pathways will be investigated, complemented by both immunohistochemistry and pathological analyses and the assessment of the resulting behavioural impact and/or disease-tissue dynamics, taking advantage of state of the art imaging technologies (small animal MRI, PET scanners and multiphoton microscopy), vi) aiming at translation into clinical trials. Overall, this is expected to enable effective translation of new biological discoveries to solutions for unmet medical needs in cancer and neurodegenerative disorders.

These objectives will be reached by developing the following tasks:

Task 1 – Experimental models of disease (neurodegeneration and cancer) will be established through (a) isolation of biological samples including peripheral blood, fibroblasts and tumor cells, including cancer stem cells from patients; (b) introduction of specific disease-associated gene sequences in the genome of cell lines, as well as in vivo transduction of rodent tissues using viral vector platforms; (c) reprogramming of patient fibroblasts into induced pluripotent stem cells, to be differentiated into neural cells. Collaboration with the biomarkers group will provide genomic support to help pinpoint key genetic features involved in pathogenesis.

Task 2 – These validated models and patient samples will allow the study of disease-modified pathways – by mRNA- and microRNA-microarray screening complemented by mechanistic studies of cell death and proliferation, protein aggregation, using whole genome shRNA lentiviral libraries coupled to transcriptional microarray, qPCR and proteomic analysis. The candidate disease pathways will then be further investigated by counteracting the dysfunctions, upon overexpression or silencing of the identified relevant genes in in vitro models, including isolated cancer stem cells, cancer cell lines and ips-derived neurons, and in vivo models such as transgenic and viral induced models as well as immunocompromised mice.

Task 3 – Translational molecular therapy approaches will be investigated for cancer and neurodegenerative diseases (e.g. Machado-Joseph) using viral and non-viral vectors to carry nucleic acids and chemical compounds, in collaboration with the groups of medicinal chemistry and pharmacometry. Imaging studies coupled with biomarker, animal behavior and pathology will allow extensive multilevel evaluation of disease alleviation. New targeted nanoparticles will be developed to promote specific and efficient delivery to neuronal and cancer cells in animal models, aiming at maximizing therapeutic activity in Machado Joseph disease and cancer (e.g. glioblastoma and breast cancer), while decreasing secondary effects. In situ brain delivery of lentiviral vectors, as well as systemic delivery of AAV9 vectors will be investigated towards aleviating neurodegeneration with a special focus on Machado-Joseph disease.

The accomplishment of these objectives implies strengthen ongoing collaborations with other groups within the consortium, as Neuroendocrinology and Aging, Molecular Biotechnology, Immunometabolic Pharmacology, Biomarkers Group.

It is expected that until 2020 the generated intellectual property will result in creation of novel spin-offs and that the creation of knowledge will translate into clinical trials leading to health benefits for the population.