Periodic Reporting for period 1 - NEXUS (automated in-line separatioN and dEtection of eXtracellular vesicles for liqUid biopsy applicationS)
Okres sprawozdawczy: 2022-06-01 do 2023-05-31
New modalities for non-invasive diagnostics, namely Liquid Biopsy, are considered a breakthrough innovation poised to have a relevant impact on timely and efficient cancer prevention and treatment. Extracellular Vesicles (EV) have the potential to outperform other liquid biopsy biomarkers such as circulating tumour cells (CTC), circulating tumour DNA (ctDNA), and traditional circulating protein biomarkers. EV is a collective term for a heterogeneous group of cell-released membranous vesicles abundantly present in the circulation and can be secured from blood, and from other body fluids in a non-invasive manner. They are natural multiplex biomarkers, enclosing different molecular species such as proteins, nucleic acids, glycans, and lipids that can be simultaneously investigated to provide real-time information on all tissue homeostasis alterations. However, despite their apparent abundancy, the heterogeneity and complexity of both EVs and the biofluids that contain them remain a huge challenge that is poorly addressed by state-of-art technologies for EV purification and characterization, which are still at research grade and don’t have clinical and commercial maturity.
The project NEXUS aims at industrializing a platform that integrates affinity isolation and in situ enrichment of target (cancer) sEVs from plasma to obtain pure, integral, and concentrated vesicles and Multiparameter analysis giving the simultaneous information on vesicle size, number, and phenotype. The aim of the project is to raise the TRL of the individual components and integrate them into a full-fledged analytical tool, characterized by an appealing “sample in – result out” design and featuring a real-time in-liquid measurement of scalable sample volumes as well as automatized and software-controlled flow.
The project NEXUS aims at industrializing a platform that integrates affinity isolation and in situ enrichment of target (cancer) sEVs from plasma to obtain pure, integral, and concentrated vesicles and Multiparameter analysis giving the simultaneous information on vesicle size, number, and phenotype. The aim of the project is to raise the TRL of the individual components and integrate them into a full-fledged analytical tool, characterized by an appealing “sample in – result out” design and featuring a real-time in-liquid measurement of scalable sample volumes as well as automatized and software-controlled flow.
The project is organized around three technical work packages, WP2, WP3, and W4 devoted to developing the separation module, the assay, and the detection module and its integration into the platform, respectively. The activity of WP5 end WP6 focuses on the analytical and clinical validation of the integrated instrument.
Partner CNR in WP 3 has introduced a method for the affinity separation of EVs from plasma providing, pure, intact, and concentrated vesicles. The method is essential for designing and developing separation and the detection modules addressed in WP 2 and WP 4. It is based on oligonucleotide-antibody conjugates that target specific EV subpopulations.
The activity of partner INOREVIA in WP2 focused on developing a microfluidic device for efficient magnetic isolation of nanoparticles. The microfluidic module for extraction and pre-concentration of exosomes from plasma (Task 2.1) uses magnetic beads functionalized with the DNA tagged antibodies (WP3) and the Magnetic Tweezers (MT) technology to mix magnetic beads with the sample, wash them with buffers and release component.
Iris Kinetics, in WP4, has provided a detailed overview of the design of the modular integrated instrument, including operative requirements and specifications for integration of isolation and detection modules, refinement of optical reader module, integrated instrument design with on-board computer control, and prototype development and manufacturability assessment.
CNR, in collaboration with partner UKLFR, has selected within WP5 reference extracellular vesicles and controls for the analytical validation of the two separate modules of the integrated instrument.
Partner UKLFR in WP6 has selected and reported EVs from cell cultures and antibodies for testing, standardization, and quantifications of EV isolation. The reference EVs exhibit on their surface antigen biomarkers identical to those on EVs of plasma samples. They will be used in repeated runs to determine the Reproducibility, Repeatability, and Precision of the integrated instrument, thus serving as Quality Control metrical standards.
Partner CNR in WP 3 has introduced a method for the affinity separation of EVs from plasma providing, pure, intact, and concentrated vesicles. The method is essential for designing and developing separation and the detection modules addressed in WP 2 and WP 4. It is based on oligonucleotide-antibody conjugates that target specific EV subpopulations.
The activity of partner INOREVIA in WP2 focused on developing a microfluidic device for efficient magnetic isolation of nanoparticles. The microfluidic module for extraction and pre-concentration of exosomes from plasma (Task 2.1) uses magnetic beads functionalized with the DNA tagged antibodies (WP3) and the Magnetic Tweezers (MT) technology to mix magnetic beads with the sample, wash them with buffers and release component.
Iris Kinetics, in WP4, has provided a detailed overview of the design of the modular integrated instrument, including operative requirements and specifications for integration of isolation and detection modules, refinement of optical reader module, integrated instrument design with on-board computer control, and prototype development and manufacturability assessment.
CNR, in collaboration with partner UKLFR, has selected within WP5 reference extracellular vesicles and controls for the analytical validation of the two separate modules of the integrated instrument.
Partner UKLFR in WP6 has selected and reported EVs from cell cultures and antibodies for testing, standardization, and quantifications of EV isolation. The reference EVs exhibit on their surface antigen biomarkers identical to those on EVs of plasma samples. They will be used in repeated runs to determine the Reproducibility, Repeatability, and Precision of the integrated instrument, thus serving as Quality Control metrical standards.
The impact of NEXUS is critically dependent on the assessment and advancement of TRL of proposed solutions and stabilization of specifications that are the premise for the successful product (a fully functional prototype featuring a Minimal Viable Product ) development and ultimately unlocking the investment readiness of NEXUS proposition. The first and early step towards delivering impact was a definition of product requirements and value proposition (D7.1). This gave input for the design and fabrication of components and modules to enter technical validation and integration, currently ongoing according to the work plan. The specifications will be revised and stabilized by the end of analytical validation when the functional prototype will be transferred to clinical validation partner premises. Partners have mapped the key stakeholders to engage, first of which reside in partners' own networks and will be approached by first dissemination actions, according to the schedule and plan submitted in D7.5. Stakeholders’ engagement started with bottom-up User Research performed by month 3, and is currently fostered by further user mapping within participation of D1 into the EIC Business Validation Program. CNR, IK and D1 actively work on the formulation and validation of the Business Plan as a ground for launching a new Spin-off company as a key NEXUS exploitation vehicle and beneficiary, foreseen by Month 24. Preliminary negotiations are therefore done in order to formulate provisional agreements with partners for resources and assets commitment and transfer (D7.2 D7.7). The exploitability of NEXUS technology is ensured by the ownership and efficient transfer of key IP including background, side-ground, and foreground IP to a Startup. The census and ranking of IPR assets are done to ensure the optimization of resources and inform the freedom-To-operate analysis prior to the MVP launch for beta testing. The development of NEXUS products takes into consideration the constraints regarding the standardization and regulatory clearance that are planned both for the instruments (CE Marking) and for the diagnostic Assays running on the integrated system (IVD marking).