Periodic Reporting for period 1 - ICE-METs (Interstitium-on-a-chip to explore its role in cancer metastasis) Reporting period: 2019-06-01 to 2021-05-31 Summary of the context and overall objectives of the project In the European Union (EU) alone, the annual number of deaths due to metastasis was projected to reach almost 3 million in 2018. In 2017, the cost of cancer treatment in the EU amounted to between €270 and 610 billion. One of the key factors in metastatic cancer is the ability of the cancer cells to migrate from a primary tumor to the blood vessels and lymphatics. A better understanding of how cancer cells interact with the microenvironment and migrate from blood vessels to the lymphatics to create distant metastases, is key for discovering efficient targeted therapy drugs that will cover the growing needs of the worldwide population. The possibility of developing biomimetic engineered devices to understand metastasis proliferation and testing of drugs is a crucial goal for the research community, both in science and medicine, translating to major benefits for future patients. The overall aim of ICE-METs was to create, for the first time, a microfluidic in vitro model to study whether the interstitium plays a role in cancer cell migration, by integrating biomimetic conducting scaffolds with optimal mechanical properties and electrical conductivity. The scaffolds integrate impedance sensing functionality to perform in-line measurements of the ongoing migration and proliferation process. The objectives of ICE-METs were:- Develop new electroactive 3D scaffolds mimicking the interstitium to study cell interaction, migration and proliferation. - Conducting polymer impedance sensor fabrication and parameter optimisation for monitoring cell migration and proliferation and sensing of cell and cell-specific biomarkers.- Integration of the 3D scaffold and sensors inside microfluidics to generate the Interstitium-on-a-Chip.No website has been developed for the project Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far WP1. Electroactive 3D porous scaffold engineering.The first step was the establishment of the 3D matrices. Two strategies were followed to prepared conducting polymers to be used as polymer matrixes. Firstly, dispersions of poly(3,4-ethylenedioxythiophene) PEDOT:hyaluronic acid and collagen type I were prepared (in collaboration with Prof. Mecerreyes, Polymat 1.5 month secondment) and secondly, a blend composed of the conducting polymer PEDOT:polystyrene sulfonate (PEDOT:PSS) and hyaluronic acid, collagen type I and laminin. The addition of these biopolymers determine the mechanical properties of the scaffold. HA and Collagen type I were selected because they represent the biological component of the interstitium. The 3D scaffolds mimicking the interstitium were prepared by lyophilisation and characterised for their mechanical and electrical properties.Moreover, the possibility of enhancing cell migration and proliferation was assessed by developing anisotropic scaffolds in collaboration with Prof. Ruth Cameron from Materials Science and Metallurgy Department in University of Cambridge.Sw480 adenocarcinoma cancer cells and neuroblastoma derived SHY-5Y cells were used to assess both scaffolds cytocompatibility and their ability to influence cell proliferation and in the case of the neuronal cells, differentiation. Results are summarised in three manuscripts in preparation.WP2. OECT fabrication, monitoring of cell migration and proliferation, and biomarker detection. Organic electronic electrodes (instead of organic electrochemical transistors) were developed, for the capture and release of cancer cells. The capture and release process was used as a way of pre-concentrating cells and monitor the process both electrically and optically. For this PEDOT:PSS was blended with N-isopropylacrylamide polymer (pNIPAAm) to create PEDOT:PSS/pNIPAAm copolymers. Thanks to the inherent capacity of pNIPAAm to shrink above its lower critical solution temperature (>32 C), the polymer can undergo a conformational change and release cells. For the capture of cells, the copolymer was functionalized with fibronectin protein. The results have been published in Biosensors and Bioelectronics. A PhD student at the group is currently applying this technology in clinical samples and two more publications are expected in future. WP3. Integration of the 3D scaffold and the OECTs inside microfluidics to generate the Interstitium-on-a-Chip. Electrodes were generated in cyclic olefin polymer (COP) as substrate and integrated into a microfluidic device to generate the interstitium-on-a-chip. The electrodes were fabricated by depositing gold onto the COP and integrating a slice of PEDOT:PSS scaffolds. The electrodes were closed by using a pressure sensitive adhesive (PSA). Sw480 cells and sw480 Tag RFP transfected cells were seeded dynamically and flow rates and time was optimised. Cell migration and proliferation both electrically and optically was assessed. Parts of this work was performed in collaboration with Dr. Fairen-Jimenez from University of Cambridge.The results will be published in a manuscript in preparation and a conference paper has been submitted. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) Three main breakthroughs were achieved at the end of the project that are linked to each objective and work package.First, different scaffolds mimicking the interstitium were developed with a variety of configurations. Pore alignment and composition were evaluated obtaining PEDOT:HA and collagen scaffolds as the composition of choice. Cancer cells on these scaffolds have proven to migrate and proliferate as the material has improved mechanical and biocompatible properties. These developed scaffolds have a potential impact in the development of biomimetic tissues to try to understand the interaction of cancer cells with their microenvironment.Secondly, electrodes with both, electrical sensing and thermoresponsive properties were developed for the capture and release of cancer cells made of PEDOT:PSS (conducting) and pNIPAAm (thermo-responsive). The electrodes showed a capture and release performance, if compared to bare PEDOT:PSS electrodes. PEDOT/pNIPAAm electrodes were able to sense the presence of cells and after a slightly thermos-treatment the release of cells. The application is important in the development of point-of-care devices for the cell sorting of cells in clinic, showing evidence of translation.Third, an interstitium-on-chip platform was developed for the study of the migration and proliferation of the cancer cells through the biomimetic interstitium. The device showed enhanced electrical and optical sensing capabilities for cancer cell monitoring. This platform enables the monitoring of cancer cell seeding under perfusion conditions and the electrical and optical monitoring of their interaction with the biomimetic interstitium. The platform demonstrates that bioelectronic electrodes integrated into microfluidics are important tools in the discovery of new mechanisms of interaction between cancer cells and the extracellular matrix. During the duration of the project, I have gained skills that I had not previously such as fabrication of electrodes, cell culture and a competent handling of different microscopy techniques. All this knowledge has been transferred to PhD students at the group through training and mentoring. The project’s activities have been benefited by the collaboration of Prof. David Mecerreyes and Dr. David Fairen-Jimenez, two stablished PIs in different research fields. Finally, the MSCA fellowship and the ICE-METs project have helped me to secure my current position as Ikerbasque Research Fellow. Figure 1: Schematic representation of the Interstitium-on-a-Chip.
