Periodic Reporting for period 2 - I-GENE (In-vivo Gene Editing by NanotransducErs)
Okres sprawozdawczy: 2020-11-01 do 2022-04-30
Most of the efforts of the 1st reporting period were devoted to the synthesis of the NT, its chemical, physical and biological characterisation and to simulate its plasmonic behaviour in the water environment during radiation. Prochimia Surfaces synthesised the AuNPs via its seed formation and grow methodology accompanied by ligand exchange. These AuNPs have a very narrow distribution and high stability. They have been functionalised with two kinds of distal functional group ligands for subsequent covalent linkage to the enzyme. UNIPI confirmed the ability of AuNPs to efficiently conjugate the enzyme. Furthermore, the NT is not cytotoxic and possesses the ability to localize to the correct genomic location. The NT also possess ability of spontaneous cell internalization and nuclear localization, to some degree, but these features need further improvements. Upon preparation, a long term storage of the NT at 4°C (up to several weeks) is possible. The Consortium is now working on the production and optimisation of all the NT components to develop a proprietary formulation for pushing future commercial exploitation of this product.
Many project efforts have been focused on mathematical simulations. The simulation activity of IIT has produced a clear representation of the process and has well-defined the experimental conditions allowing to obtain the parameters (radiation wavelength, power density, time) required for NT activation. We also made advances in the development of our technological solutions: the development of the laser workstation and the lab-on-chip. M Squared worked to design the laser configuration and the optical schemes for inducing NT activation in the radiated samples (cells and living organisms), according to specifications coming from simulation studies. A lab-on-chip device was designed by Lionix which follows the requirements set in the project. The configuration was conceived for handling cells in suspension (previously electroporated with the NTs) that enter the fluidic channel, are exposed to the laser radiation for milliseconds for NT activation and, consequently, induction of I-GENE genome editing.
I-Gene project intend to achieve the expected impact through four different impact pathways:
● Conceptual development, by providing a new paradigm of genome editing, characterized by safety and low cost, relative to other methods
● Technology development, by providing new vectors, devices and software for genome editing available for marketing
● Economical development, removing barriers to widespread adoption of genome editing and creating new opportunities and applications for EU industry, especially in healthcare
● Policy influence, removing/revisit ethical concern, safety issues of genome editing, which will make policy makers, public authorities and international organisations more aware of the possibility of a safe genome editing, contributing to the development of a European genome editing strategy ethically acceptable, sustainable and societally desirable.