Engenharia avançada do nicho inervado pré-metastático humano: o impacto das vesículas extracelulares de cancro da próstata na neurotização patológica da metástase óssea

This proposal aims to develop advanced – fully human – in vitro model to unravel the functional impact of the EVs, released from the primary PCa, on the sensory innervation pattern of bone microenvironment to support PCa metastases. To achieve the main goal, the proposal will be focused on the following specific objectives (SO): SO1: Set a fully humanized in vitro bone preMN comprising human induced sensory neurons (iSN), vasculature and bone cells; SO2: Set a portfolio of high resolution and high content analytical tools adapted to the microfluidic device scale; SO3: Address the effect of patients’ PCa-derived EVs on sensory innervation pattern, at the engineered bone preMN, upon characterization of its cargo: SO4: Evaluate the engraftment yield of perfused PCa into the engineered preMN exposed to the patient PCa-derived EVs; SO5: Identify the signalling pathways and molecular players of human sensory neurons modulation and preMN conditioning, triggered by patient PC-derived EVs; SO6: Proof of Concept – Pharmacologically manipulate the signalling pathways unravelled in SO5 and evaluate the engraftment and growth of PCa cells in the advanced innervated in vitro preMN. The specific objectives outlined demonstrate a high level of ambition and innovation, surpassing the current state-of-the-art in several respects. Firstly, the establishment of a fully humanized in vitro bone preMN comprising human iSN, vasculature, and bone cells (SO1) represents a novel and sophisticated approach to modeling PCa metastasis in vitro. This integration of multiple cell types and tissue engineering techniques, and high resolution analytical tools to monitor the NeurONiche microsystem (SO2) pushes the boundaries of current research methodologies. Furthermore, characterizing the neurotrophic profile of patient-derived EVs and elucidating their role on sensory innervation pattern of bone preMN (SO3) constitute novel concepts that expand our understanding of PCa communication, within the multicellular bone microenvironment (SO4). These objectives leverage cutting-edge molecular biology techniques to uncover new insights into the mechanisms driving PCa metastasis. Identifying signaling pathways and molecular players involved in the modulation of human sensory neurons (SO5) and pharmacologically manipulating these pathways to disrupt PCa metastasis (SO6) represent ambitious objectives that go beyond current knowledge. To achieve the objectives the interdisciplinary collaboration, secured by this team, is required and the integration of advanced techniques from various fields, illustrating the innovative nature of the proposed research. Overall, the proposed research addresses important challenges in cancer biology and metastasis while demonstrating ambition and innovation through its novel concepts, interdisciplinary approaches, and development beyond the current state-of-the-art. NeurONiche will set the foundation to understand the preMN establishment and the impact of sensory innervation at bone metastases progression. Ultimately, this work will be INSTRUMENTAL IN IDENTIFYING THE CHRONOLOGY OF PCa PROGRESSION and the optimal timing and mode of action to DISRUPT THE DISEASE ADVANCE.