Periodic Reporting for period 4 - EXACTYMER (ADVANCED NANOMEMBRANES FOR EXACT POLYMER PRODUCTION)
Período documentado: 2023-01-01 hasta 2023-12-31
Polymers are macromolecules comprising monomers linked in a chain. Nature makes extensive use of polymers with exquisitely defined monomer sequences for transmitting information (DNA, RNA) and for fabricating unique 3-D structures (peptides and proteins).
However, current processes for production of synthetic polymers with exactly defined sequences of monomers fall far short of Nature’s ideal. As yet, there are only limited successful reports describing synthesis of specific polymers, and so the potential benefits to society of defined monomer sequence polymers cannot be realised. Iterative synthesis, in which monomers are added one-at-a-time to the end of a growing polymer chain, affords exquisite control over the monomer sequence. However it requires separation of the growing polymer from the monomers with each and every cycle, which is the crucial, and hardest, step. To solve this problem, the EXACTYMER Advanced Grant works with attaching multiple growing polymer chains to a rigid hub, so that a macromolecular „nanostar“ is created which can be separated from reaction debris by membrane separation – we have named this „Nanostar Sieving“, and the defined monomer sequence exact polymers that are produced, „Exactymers“.
The objective of EXACTYMER is curiosity-driven, multidisciplinary research at the boundaries of membrane technology, polymer chemistry, and process engineering to create this new platform for exactymer synthesis using Nanostar Sieving. This technology can bring benefits to society through access to synthetic polymers of unprecedented accuracy. Potential applications for exactymers include medicine (where they could be used to conjugate drugs, and to link drugs to functional moieties such as affintiy ligands, cell penetrating agents, and imgaging agents), nanotechnology and information storage.
The overall objectives are:
Create ultra-selective, high permeance molecular separation membranes;
Expanding the chemistry of building blocks and iterative synthesis cycles for use in Nanostar Sieving
Integrate iterative chemistry and nanomembrane purification in the homostar nanofiltration platform for rapid, automated production of exactymers;
Exploring exactymer applications in healthcare, nanotechnology and information storage.
From the Exactymer AdG we conclude that (i) the speed and accuracy of membrane separations, that is the ability to separate different molecules present in solution, can be greatly increased through manipulation of the molecular architecture of the membrane separating layers; (ii) fouling at the surface of membranes can be greatly reduced by attachment of a flexible grafting layer to the membrane surface; (iii) membrane separation is massivley influenced by the concentration of molecules in the mixture to be separated; (iv) exact PEG polymers allow exact determination of the PEGylations sites in biopharmaceutical proteins; (v) single nanopores drilled into ultrathin polyamide films show great potential for reading the monomer sequence on synthetic polymers; (vi) novel electroactive polymers can be formed under mild conditions using organic electron acceptors.
However, current processes for production of synthetic polymers with exactly defined sequences of monomers fall far short of Nature’s ideal. As yet, there are only limited successful reports describing synthesis of specific polymers, and so the potential benefits to society of defined monomer sequence polymers cannot be realised. Iterative synthesis, in which monomers are added one-at-a-time to the end of a growing polymer chain, affords exquisite control over the monomer sequence. However it requires separation of the growing polymer from the monomers with each and every cycle, which is the crucial, and hardest, step. To solve this problem, the EXACTYMER Advanced Grant works with attaching multiple growing polymer chains to a rigid hub, so that a macromolecular „nanostar“ is created which can be separated from reaction debris by membrane separation – we have named this „Nanostar Sieving“, and the defined monomer sequence exact polymers that are produced, „Exactymers“.
The objective of EXACTYMER is curiosity-driven, multidisciplinary research at the boundaries of membrane technology, polymer chemistry, and process engineering to create this new platform for exactymer synthesis using Nanostar Sieving. This technology can bring benefits to society through access to synthetic polymers of unprecedented accuracy. Potential applications for exactymers include medicine (where they could be used to conjugate drugs, and to link drugs to functional moieties such as affintiy ligands, cell penetrating agents, and imgaging agents), nanotechnology and information storage.
The overall objectives are:
Create ultra-selective, high permeance molecular separation membranes;
Expanding the chemistry of building blocks and iterative synthesis cycles for use in Nanostar Sieving
Integrate iterative chemistry and nanomembrane purification in the homostar nanofiltration platform for rapid, automated production of exactymers;
Exploring exactymer applications in healthcare, nanotechnology and information storage.
From the Exactymer AdG we conclude that (i) the speed and accuracy of membrane separations, that is the ability to separate different molecules present in solution, can be greatly increased through manipulation of the molecular architecture of the membrane separating layers; (ii) fouling at the surface of membranes can be greatly reduced by attachment of a flexible grafting layer to the membrane surface; (iii) membrane separation is massivley influenced by the concentration of molecules in the mixture to be separated; (iv) exact PEG polymers allow exact determination of the PEGylations sites in biopharmaceutical proteins; (v) single nanopores drilled into ultrathin polyamide films show great potential for reading the monomer sequence on synthetic polymers; (vi) novel electroactive polymers can be formed under mild conditions using organic electron acceptors.
We have created new ranges of separation membranes by employing polymerisation at an interface between two separate liquid phases, to create very thin polymer separating layers. The key to these new membranes was the synthesis of novel types of monomers for use in the interfacial polymerisation reaction. Further we have found that when we use membranes in Nanostar Sieving, the concentrations of solutes in the liquids is high and this can lead to fouling. Hence we have grafted flexible layers of polymers on the surface of the membranes to avoid the „sticking“ of the solutes to the membrane surfaces.
We have worked on making new and improved polymer building blocks, or monomers. The type of reactive group at each end of the polymer has been manipulated, so that the polymer can be attached to other molecules, for example in the PEGylation of biopharmaceuticals such as proteins.
Next, we have worked on engineering of a synthesizer for carrying out Nanostar Sieving. We have worked to reduce the volume of liquid and improve the mixing in the synthesizer, so that all our reactions progress equally and there are no „dead zones“ of liquid. We have also automated the synthesizers so that there is less operator intervention required.
We have use the membranes, monomers and synthesize to produce the most accurate PEGs reported to date. We have used these to attach to a protein, and shown that by using these accurate PEGs the exact site of attachment of the PEG to the protein is easily revealed. We have also proved the concept of drilling single nanopores in a thin polymder film so that we can „read“ the monomer sequence of the defined polymers we synthesise. Finally, by chance, we have discovered how to produce electroactive polymers from the same polymers we used to produce membranes.
To exploit the results of this project, a company - Exactmer Limited - has formed and has focussed on the commercialisation of the Nanostar Sieving technology for production of pharmaceutically relevant defined monomer sequence polymers. The major achievements to date have been (i) the development of a process based on liquid phase synthesis for producing oligonucleotide drugs with pharmaceutical partners Novartis, AstraZeneca, and Alnylam and (ii) the opening in September 2023 of an Advanced Membrane Manufacturing Suite in which the PBI membranes developed in the EXACTYMER AdG will be manufactured at commercial scale. Exactmer employs 40 staff as at end 2023, of whom 30 have PhDs, and is based in the east of London, UK.
We have worked on making new and improved polymer building blocks, or monomers. The type of reactive group at each end of the polymer has been manipulated, so that the polymer can be attached to other molecules, for example in the PEGylation of biopharmaceuticals such as proteins.
Next, we have worked on engineering of a synthesizer for carrying out Nanostar Sieving. We have worked to reduce the volume of liquid and improve the mixing in the synthesizer, so that all our reactions progress equally and there are no „dead zones“ of liquid. We have also automated the synthesizers so that there is less operator intervention required.
We have use the membranes, monomers and synthesize to produce the most accurate PEGs reported to date. We have used these to attach to a protein, and shown that by using these accurate PEGs the exact site of attachment of the PEG to the protein is easily revealed. We have also proved the concept of drilling single nanopores in a thin polymder film so that we can „read“ the monomer sequence of the defined polymers we synthesise. Finally, by chance, we have discovered how to produce electroactive polymers from the same polymers we used to produce membranes.
To exploit the results of this project, a company - Exactmer Limited - has formed and has focussed on the commercialisation of the Nanostar Sieving technology for production of pharmaceutically relevant defined monomer sequence polymers. The major achievements to date have been (i) the development of a process based on liquid phase synthesis for producing oligonucleotide drugs with pharmaceutical partners Novartis, AstraZeneca, and Alnylam and (ii) the opening in September 2023 of an Advanced Membrane Manufacturing Suite in which the PBI membranes developed in the EXACTYMER AdG will be manufactured at commercial scale. Exactmer employs 40 staff as at end 2023, of whom 30 have PhDs, and is based in the east of London, UK.
Polymers labelled as narrow dispersity are in fact a mixture of many different molecular weights, giving a bell shaped curve as in Figure 1A obtained from a „narrow dispersity“ PEG of 5kDa molecular weight. We have been able to create PEG molecules with outstanding purity – Figure 1B shows the single peak resulting from synthesis of PEG 5kDa using nanostar sieving.
We have created membranes with high flux rates and high selectivity between organic solutes, using interfacial polymerisation to control the exact molecular structures in the separating layer of the membrane. We have also created a series of „non-stick“ membranes that can operate under very harsh solvent conditions with high substrate concentrations, without suffering from fouling on their surfaces.
A selection of exact 5 kDa PEGylation agents was synthesised and used in a comparative study on the random PEGylation and subsequent characterisation of the protein bovine serum albumin (BSA). It was demonstrated that the defined Mw PEGylation agents eanble the straightforward identification of the PEGylated fragments within a PEGylated protein through a simple peptide mapping approach using UPLC/MS.
We have made an exciting discovery on reacting Polybenzimidazole polymers with organic electron acceptor, generating an electroactive polymer with built-in charge within in the backbone and counterion side groups. Extensive physical analysis conclusively indicates a new polymer has been synthesised. Further work revealed the same approach also works on further polymers including Polyetherimide, Polyacrylontirile, PEG, PEEK and Polyethylene.
We have created membranes with high flux rates and high selectivity between organic solutes, using interfacial polymerisation to control the exact molecular structures in the separating layer of the membrane. We have also created a series of „non-stick“ membranes that can operate under very harsh solvent conditions with high substrate concentrations, without suffering from fouling on their surfaces.
A selection of exact 5 kDa PEGylation agents was synthesised and used in a comparative study on the random PEGylation and subsequent characterisation of the protein bovine serum albumin (BSA). It was demonstrated that the defined Mw PEGylation agents eanble the straightforward identification of the PEGylated fragments within a PEGylated protein through a simple peptide mapping approach using UPLC/MS.
We have made an exciting discovery on reacting Polybenzimidazole polymers with organic electron acceptor, generating an electroactive polymer with built-in charge within in the backbone and counterion side groups. Extensive physical analysis conclusively indicates a new polymer has been synthesised. Further work revealed the same approach also works on further polymers including Polyetherimide, Polyacrylontirile, PEG, PEEK and Polyethylene.