NANOTECHNOLOGIES AND PHAGE DISPLAY: SELECTION OF PEPTIDES MIMICKING BDNF

Brain derived neurotrophic factor is a neurotrophin of vital importance for neuron survival. It may therefore be of great help to people affected by sensorineural hearing loss and cochlear implant users, as it has already been demonstrated that it maintains neuronal cells alive and this proved to be necessary for the device to work. The issue with its clinical application is related to its large size that makes it difficult to deliver. In this thesis I took on one of the tasks of a European project. The objective of the Nanoear project is the creation of multifunctional nanoparticles that could be used as drug carriers for a target delivery in the inner ear. The final aim is to find a substitute for BDNF protein, able to mimic its action but small enough to be attached to the nanoparticles and carry them to their target in the inner ear. The best method to obtain small peptides with these features is the phage display. This technique allows selection of a pool of specific peptides binding specifically to an immobilized target. In collaboration with an English company involved in the project as well, we worked to set all the parameters in order to make the phage display for this project feasible. As our goal was attaining BDNF mimetic peptides our target had to be its functional receptor, TrkB. Starting from a polyA+ RNA library from adult human brain we could get the genes coding for TrkB and BDNF and follow all the process up to the expression of the two proteins in bacteria and in eukaryotic cells, as well. In order to include into nanoparticles the most specific peptides many approaches for the phage display were taken into consideration and experimented on. This technique was in fact performed in vivo on guinea pigs and all the parameters were set for different trials in vitro either on eukaryotic cells or on the specific binding domain purified from bacteria. Therefore the results and the data obtained in this study can lead to a new generation of drug with great clinical potential.