Part 1: Encapsulation and responsive release of polymersomes from protocells
Components of the previously established protocell system were successfully synthesized, including modified amyloses and membrane forming terpolymer. The amphiphilic polymers for forming the polymersomes were synthesized following a ring opening polymerization of caprolactone and trimethylene carbonate monomers. To increase nucleic acid encapsulation, one solution is the incorporation of imidazole containing groups in the hydrophobic block. To achieve this an activated ester monomer was synthesized, para-pentafluorophenol-trimethylene carbonate (TMC(OPhF5)), which was then polymerized in the same conditions as previously, to give PEG-p(CL-TMC(OPhF5)). The activated ester was successfully modified with imidazolepropylamine. At a 50:50 ratio of PEG-P(CL-TMCimid) and NBenz-PEG-P(CL-TMCimid) in the polymersome formulation, stable polymersomes formed and the surface charge of the polymersomes increased from from negative to positive after UV irradiation. Encapsulation studies with cy5-siRNA in polymersomes showed between 15-20% encapsulation efficiency. Polymersomes were incorporated into the established coacervate system, and were released from coacervates after UV irradiation as followed by confocal microscopy. The supernatant from the release experiment was also analysed by DLS, which also indicated release of nanoparticles by an increasing count rate over time.
Exploitation and dissemination: This work will be published in a journal article, and was presented at CHAINS chemistry conference 2022 and PTN Dutch Polymer Days 2023. A review article was published as part of this work in Accounts of Materials Research: ‘Complex Coacervate Materials as Artificial Cells’ 10.1021/accountsmr.2c00239.
Part 2: exosome-like RNA transfer between protocells and living cells
Initial studies incubating coacervates containing polymersomes with HeLa cells have been performed . Cells were incubated in microscopy slides and coacervates added. The coacervates and cells were irradiated with UV light for 10 min, and then imaged. An increase in intracellular fluorescent signal was seen with the UV responsive polymersomes compared to the non-responsive polymersomes. These results indicate that the UV responsive polymersomes are able to be uptaken into living cells after UV charge switching is triggered. Further studies are being carried out to determine if the RNA remains functional, by repeating the experiments with eGFP-siRNA. These results will be a first example of RNA signalling between artificial cells and living cells. The research article resulting from these studies will be of high impact.
Exploitation and dissemination: This work will be published in a journal article, and was presented at Advanced Materials and Nanotechnology (Rotorua, NZ) and CHAINS chemistry conference 2023.
