Periodic Reporting for period 2 - ICE_PACK (Polymer Materials for Cryogenic Storage and Distribution of Biologics)
Período documentado: 2022-03-01 hasta 2023-09-30
20 years ago many drugs were small molecules – for example paracetamol. These could be formulated into tablets and stored and distributed easily. However, since then there has been a revolution in the drug industry with the emergence of biologics – therapies typically based on proteins or living cells. These have made huge impact, especially in cancer treatment, but are very expensive and harder to store. To stop them degrading, it is essential to store them cold or even frozen. This means adding a cryoprotectant (‘antifreeze’) to protect them and ensure that they remain active after thawing. The current cryoprotection methods involve adding organic solvents, but these do not give 100 % recovery, their dose needs to be controlled to minimise cytotoxicity and there are logistical challenges.
This project aims to re-think how we freeze biologics by using macromolecular cryoprotectants – large soluble macromolecules which address the mechanisms of damage, which current cryoprotectants do not. These protective effects can be achieved through a combination of controlling ice growth and/or ice formation and stabilising cell membranes. This work was initially inspired by how extremophiles in nature survive in extreme cold environments, unlike humans who mostly prefer to be nice and warm!
Progress in this area is very important for society – ensuring therapies are optimal (i.e. if you freeze 100 therapeutic cells, ideally you want to recover 100 healthy cells after) and mitigating the cost of these exciting new biologic therapeutics by addressing the challenges of the cold chain and storage.
This project aims to re-think how we freeze biologics by using macromolecular cryoprotectants – large soluble macromolecules which address the mechanisms of damage, which current cryoprotectants do not. These protective effects can be achieved through a combination of controlling ice growth and/or ice formation and stabilising cell membranes. This work was initially inspired by how extremophiles in nature survive in extreme cold environments, unlike humans who mostly prefer to be nice and warm!
Progress in this area is very important for society – ensuring therapies are optimal (i.e. if you freeze 100 therapeutic cells, ideally you want to recover 100 healthy cells after) and mitigating the cost of these exciting new biologic therapeutics by addressing the challenges of the cold chain and storage.
This project is divided into 3 Workpackages. A) The synthesis and ice-binding interactions of macromolecular cryoprotectants; B) Cellular cryopreservation; C) Protein storage. The progress of each are summarised here:
A) This Workpackage focussed on the development of the macromolecular cryoprotectants to be deployed to stabilise biologics. We have made significant progress with our work on polyampholytes (a class of polymers made up of positively and negatively charged monomer subunits), which have demonstrated to have excellent cell stabilisation, and therefore cryoprotective, properties. We propose that this occurs through helping dehydration and stabilisation the membrane. We have shown how these can be used to store a range of important cell types and begun to obtain structure-function relationships for ‘how’ do these materials work. We have also made significant progress in developing materials which can nucleate ice – this is exceptionally challenging but controlling the temperature when ice actually forms is very important for cryopreservation. We have managed to develop the first synthetic polymers which can do this.
B) This Workpackage contains the cell biology to show how we can protect cells. We have managed to develop tools for protecting a range of cells and have even shown a version of our macromolecular cryoprotectants which is degradable yet retains its function in protecting cells. We have now made progress on the storage of t-cells: cells used in cancer therapy. In particular, e have identified key damage mechanisms which occur before and after freezing, allowing us to target our solutions to the exact mechanism, rather than random formulation.
C) For this Workpackage a key component is macromolecular engineering of our cryoprotective polymers to enable them to be conjugated (attached) to proteins. We have made progress in understanding the reaction parameters which enable us to install specific groups to attach these to proteins. We have also obtained preliminary data that shows our polymers are able to protect model proteins during freeze/thaw, without the need for other cryoprotectants and work is continuing on this.
A) This Workpackage focussed on the development of the macromolecular cryoprotectants to be deployed to stabilise biologics. We have made significant progress with our work on polyampholytes (a class of polymers made up of positively and negatively charged monomer subunits), which have demonstrated to have excellent cell stabilisation, and therefore cryoprotective, properties. We propose that this occurs through helping dehydration and stabilisation the membrane. We have shown how these can be used to store a range of important cell types and begun to obtain structure-function relationships for ‘how’ do these materials work. We have also made significant progress in developing materials which can nucleate ice – this is exceptionally challenging but controlling the temperature when ice actually forms is very important for cryopreservation. We have managed to develop the first synthetic polymers which can do this.
B) This Workpackage contains the cell biology to show how we can protect cells. We have managed to develop tools for protecting a range of cells and have even shown a version of our macromolecular cryoprotectants which is degradable yet retains its function in protecting cells. We have now made progress on the storage of t-cells: cells used in cancer therapy. In particular, e have identified key damage mechanisms which occur before and after freezing, allowing us to target our solutions to the exact mechanism, rather than random formulation.
C) For this Workpackage a key component is macromolecular engineering of our cryoprotective polymers to enable them to be conjugated (attached) to proteins. We have made progress in understanding the reaction parameters which enable us to install specific groups to attach these to proteins. We have also obtained preliminary data that shows our polymers are able to protect model proteins during freeze/thaw, without the need for other cryoprotectants and work is continuing on this.
When we began this project there were limited examples of macromolecular cryoprotectants, mainly focussed on polymers to control the rate of ice growth. We have exemplified how polyampholytes can protect cells and begun to understand ‘how’ they work, which is helping us use these in a range of applications. Perhaps the biggest advance so far has been the development of materials to control nucleation. Currently the only way to nucleate ice in cryopreservation is via physical stimulus (which is challenging) or the addition of bacterial extracts, or insoluble inorganic minerals. We have shown for the first time that soluble ice nucleators can protect cells. This is very important as it confirms that without changing the cryoprotectant, one can improve post-thaw outcomes simply by ensuring ice forms at the right temperature: counter intuitively, this means making the ice form ‘sooner’ and at warmer temperatures. We anticipate significant outcomes in this particular aspect as the project progresses.