Laser-Guided nanoparticles and Cell Scalextrics

Lead Research Organisation: Cardiff University
Department Name: Welsh School of Pharmacy

Abstract

Steering nanoparticle transport in human cells - why is this important?
Viruses seem to travel effortlessly into tissues and cells, being transported selectively in the body to reach the sites where they can cause most harm. They do this by breaching cellular barriers such as the outer or plasma membrane of cells and use human cellular machinery to make copies of themselves. Drug molecules on the other hand spread non-selectively throughout the body thus reducing effects where they are most needed, and causing adverse side-effects where they are not wanted. One reason for this is that current synthetic carriers for drugs and diagnostic agents, unlike viruses, are unable to effectively cross biological barriers and then reach specific sites inside cells. An artificial particle that could transport through tissue, in a manner analogous to a virus and then into defined cell locations but without causing disease, would therefore revolutionise healthcare applications. This would be particularly important for the early stage diagnostics and therapeutics needed in developing nations and for ageing populations.

What do novel polymer-coated gold nanoparticles have to offer?
These materials are an optimal test platform for proof-of-concept studies described in this application. Firstly, gold particles can be tracked in cells using a number of microscopic approaches including the newly developed highly sensitive four-wave mixing imaging system available to this team. They can be coated with a variety of polymeric materials that will help to guide them into cells and into specific cell locations. We have previously shown that polymers which are capped with functional 'keys' to enter natural cell portals can have their entry switched on and off by small increases in temperature which cause them to change their conformations. We also have shown that we can generate these temperature increases at gold nanoparticles inside cells through laser pulses but without damaging the cells. By attaching the temperature-responsive polymers to gold, it should be possible to use laser pulses to switch the functional keys on and off, and in this way guide particles to reach defined cellular locations. This will help to unravel the mechanisms by which materials travel in cells, thus enabling us to guide diagnostics and therapeutics to where they are required.

Impact.
A major hurdle to effective therapy against major disease burdens such as cancer, coronary heart disease and neurodegeneration is our inability to direct therapeutic molecules such as genes and proteins to specific tissue and defined compartments inside cells. This is a major objective of this application and progress here could have widespread implications for academia, industry and the society that they serve. One could envisage a commercial application in which a combined imaging and guiding instrument (e,g, ultrasonic probes and imaging) is used with a set of nanoparticles with functionality for specific disease markers, with a potential for truly selective personalised therapies.
Better diagnostics are also needed that allow earlier detection of disease and thus better healthcare outcomes. Successful completion of this work could allow development of an imaging/detection platform where specific markers of disease could be detected through their interaction with selective receptors on gold nanoparticles guided to intracellular sites by the local laser-heating method. When it is considered that 1 in 3 individuals in the EU will be affected directly or indirectly by cancer by 2010, it is clear that earlier detection and intervention will bring marked benefits to patients, carers and society as a whole.
Longer-term development could generate impact through a new biomedical technology i.e. laser-guided therapeutics wherein local heating by focused ultrasound guides biodegradable responsive nanoparticles in humans.

Publications

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Sayers EJ (2014) Distal phenylalanine modification for enhancing cellular delivery of fluorophores, proteins and quantum dots by cell penetrating peptides. in Journal of controlled release : official journal of the Controlled Release Society

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Sayers EJ (2019) Endocytic Profiling of Cancer Cell Models Reveals Critical Factors Influencing LNP-Mediated mRNA Delivery and Protein Expression. in Molecular therapy : the journal of the American Society of Gene Therapy

 
Description Key Findings 1
RECEPTOR CROSSLINKING FOR DRUG DELIVERY
Through this award we have discovered a universal strategy for delivering therapeutic entities into cells. It is based on binding receptors on the surface of cells using a targeting entity and then driving a therapeutic into the interior of hte cell to reach its target.
The results were published in Molecular Therapy and as recognition of the potential impact of these findings the Journal wrote a two page commentary on this work in the same issue and also included one of our images on the cover.
Moody et al Molecular Therapy (2015); 23 12, 1888-1898. doi:10.1038/mt.2015.178 Key KEY FINDINGS 2 MAKING SMART NANOPOLYMERS TO DELIVER THERAPEUTICS Work in collaboration with Nottingham University and the University of Paduva in Italy demonstrated that we could design and synthesis nanoparticles that were in a controlled manner (1 temperature and 2. acidicy) able to bind selectively to cancer cells showing high levels of a target protein on their surfaces. This is work was published via two papers 1) Brazzale et al (2017) Nanoscale DOI: 10.1039/C7NR02595E 2) Sayers et al (2018) Bioconjugate Chemistry DOI: 10.1021/acs.bioconjchem.7b00704

Key Findings 3
POLYMER SCIENCE CAN BE COOL AND EXCITING
Through this funding a large science exhibition was organised under the title of "Working Polymers" This was held over a eight day period at the Science and Technology Pavilion of the National Eisteddfod of Wales - Meifod Mid-Wales- August 1 - August 8 2015. The aim of this exhibit was to highlight the science underlying polymers and the fact that we are now totally dependent on them but to also bring introduce EPSRC funded research in the Jones laboratory on the use of polymers for designing nanomedicines to target diseases such as cancer. Approximately 130,000 visited the Eisteddfod over this eight day period and >20,000 were counted into the Science Pavilion. The Exhibit was staffed by Cardiff University academics and undergraduate students.
Feedback was extremely positive and there was much discussion surrounding the various polymer activities in display e.g. polymer surface of the world cup rugby ball, nappies and cardiac stents with controlled release of drugs. This also attracted the media and interviews were conducted with BBC Radio and TV and also S4C Wales. A major outcome of these is that the public now clearly view science as a fundamental part of Welsh culture and this is why the Science and Technology Pavilion sits proudly and comfortably at the heart of this very large cultural festival.

Key Findings 4
Work initiated during the award have resulted in an important publication in the Journal Nanoscale. This related to using fluorescence to track nanoparticles moving in cells. These particles are of significant interests as vectors for therapeutics and diagnostic agents. Using our newly discovered microscopy technique we have identifies (at the nanoscale) that the fluorescence read out may not perfectly align with the location of the nanoparticle. This has significant implications for the academic and industrial sectors who are studying nanoparticle dynamics in cells. Giannakopoulou et al (202) Nanoscale doi.org/10.1039/C9NR08512B


Key Findings 5 During the end of the award the Jones group teamed up with Astra Zeneca who were interested in a paper that was published in 2015 from EPSRC funding (Moody el al Molecular Therapy 2015). This led to a new collaboration and funding and a publication in 2019 where nanoparticles cell interactions have been studied in great detail suing a new technology platform called Endocytic profiling that also involved manufacturing a new reagent for analysing the pH (acidity) in cells. An image from the paper was placed as a cover image
Sayers et al 2019: Molecular Therapy https://doi.org/10.1016/j.ymthe.2019.07.018
Cover Image: https://www.sciencedirect.com/journal/molecular-therapy/vol/27/issue/11
Exploitation Route A. TAKEN FORWARD
The results form this paper and thus award have had immediate impact
1 New Grant Funding from Cancer Research Wales . Project Grant October 2015- April 2017 £101,000.
2 New contract and collaboration with Astra Zeneca to develop more efficient drug delivery systems. £135,000
We will continue to apply for funding to study this further with a view to translating the in vitro data into realistic therapeutic strategies.


B. USE BY OTHERS
1. We have discovered an approach for receptor targetting, crosslinking and internalization that we believe will be of significant benefit to therapeutic approaches to address a number of diseases with high societal burden such as cancer. We have developed methods for manufacturing new SMART polymer nanoplatforms for delivering therapeutics to cancer cells. In March 10 202 this article has 35 citations

2. We hope that our Endocytic Profile analysis methodological platform and new research tool will be of significant interest to academia and industry. This will allow them to perform more rigorous pre cinical testing in cell models of drug delivery, using nanoparticles and other vectors as delivery platforms
Sectors Education,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

URL http://www.nature.com/mt/journal/v23/n12/abs/mt2015178a.html(2)http://pubs.rsc.org/en/content/articlelanding/2017/nr/c7nr02595e#!divAbstract(3)https://pubs.acs.org/doi/10.1021/acs.bioconjchem.7b00704(4)https://www.sciencedirect.com/journal/molecular-therapy/vol/27/issue/11)
 
Description To provide funding for future Public Engagement activities. To collaborate with industry (Astrazeneca) on delivery of mRNA lipid nanoparticles- 12 month project grant extended to current date in collaborative research towards submission of a manuscript for publication - ongoing Our new Endocytic Platform methodology and pH probe (Sayers et al Molecular Therapy 2019) will be of significant interest to industry. This universal analytcal platform will allow them to perfom more rigourous pre clinical testing of drug delivery performance in cell models for cancer and other diseases.
Sector Education,Healthcare,Pharmaceuticals and Medical Biotechnology
Impact Types Cultural,Societal

 
Description Cancer Research Wales Project Grant
Amount £101,000 (GBP)
Organisation Cancer Research Wales 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2015 
End 04/2017
 
Description Industrial Research Project
Amount £135,000 (GBP)
Organisation AstraZeneca 
Sector Private
Country United Kingdom
Start 03/2016 
End 04/2017
 
Description Institutional Strategic Support Fund/Wellcome Trust for Public Engagement
Amount £6,300 (GBP)
Organisation Wellcome Trust 
Department Wellcome Trust Institutional Strategic Support Fund
Sector Charity/Non Profit
Country United Kingdom
Start 01/2016 
End 12/2016
 
Description Wellcome Trust Institutional Strategic Support Fund: - Collaboration Panel: Cross-Disciplinary Award
Amount £29,552 (GBP)
Organisation Wellcome Trust 
Department Wellcome Trust Institutional Strategic Support Fund
Sector Charity/Non Profit
Country United Kingdom
Start 08/2017 
End 02/2018
 
Title Endocytic Profiling 
Description A methodological platform called Endocytic Profiling was developed to characterise the endocytic pathways of pre clinical cell models for drug delivery. A lipid nanoparticle vector enclosing mRNA as cargo was assessed using this platform that identified critical features regulation LNP transfection. We also designed an manufactured a new probe for analysing the pH on the endolysosomal pathway. 
Type Of Material Cell line 
Year Produced 2019 
Provided To Others? Yes  
Impact This will be of value to academic and industrial researchers with an interest in preclinical drug delivery analysis of drug delivery vector performance in one of a hundereds of cell models now available. In this respect, the method has significant universality. A manuscript was published on this work in Molecular Therapy that awared the study a cover image. The work was a collaboration between Cardiff University and Astra Zeneca 
URL https://www.sciencedirect.com/science/article/abs/pii/S1525001619303570
 
Title Receptor Crosslinking 
Description The method description is essentially in the title of the manuscript from this EPSRC funded project : Moody et al 2015, Receptor Crosslinking: A General Method to Trigger Internalization and Lysosomal Targeting of Therapeutic Receptor:Ligand Complexes It describes an in vitro crosslinking system that we find is applicable for internalisation of numerous plasma membrane receptors. Crosslinking could be exploited for a wide range of target receptors, for navigating therapeutics through the endolysosomal pathway, for significant therapeutic benefit. 
Type Of Material Model of mechanisms or symptoms - in vitro 
Year Produced 2015 
Provided To Others? Yes  
Impact Still very recent but already two new awards have been obatained based on this 2016-2017 Astra Zeneca Research Contract. Cellular Uptake of Drug Delivery Systems Targeting Cancer. A.T. Jones £135,486 2015-2017 Cancer Research Wales Project Grant. Inducing HER2 endocytosis and downregulation via enhanced antibody-receptor clustering as a therapeutic strategy in breast cancer. Jones, A.T., Hiscox, S., Westwell, A. £101,124 
URL http://www.nature.com/mt/journal/v23/n12/full/mt2015178a.html
 
Title Database for Moody et al 2015 
Description Allt the data supporting the research results reported in EPSRC funded Moody et al (2015); Molercular Therapy 23 12, 1888-1898. doi:10.1038/mt.2015.178 can be accessed via Cardiff University's data catalogue. This includes all the raw data for each indiviual figure via URL http://dx.doi.org/10.17035/d.2015.100121. This URL is displayed on the harcopy and web version of the article. 
Type Of Material Database/Collection of data 
Provided To Others? No  
Impact None yet 
URL http://dx.doi.org/10.17035/d.2015.100121.
 
Description Astra Zeneca Project 
Organisation AstraZeneca
Country United Kingdom 
Sector Private 
PI Contribution New collaboration with funding from AstraZeneca starting starting in April 2016 running to June 2017. This stems from EPSRC data presented at Nanotechnologies in Drug Delivery Congress, London, UK 2015
Collaborator Contribution AZ are synthesise targetted drug delivery vectors in the form of lipid nanaoparticles carrying mRNA for Cardiff University to test in in vitro models Submitted joint Cardiff University - AstraZeneca BBSRC-IPA application. Unsucessful
Impact Presentation of slides at international conferences since 2017. Abstract at International Conference: Puri, S., Ashford, M., Sayers E.J., Jones, A.T. (2018) The Subcellular Profile of Tumour Cells Influences the Intracellular Fate of mRNA Delivery Systems nanoDDS 16th International Nanomedicine & Drug Delivery Symposium, Portland USA. Poster 74
Start Year 2015
 
Description National Eisteddfod of Wales Meifod 2015 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Through EPSRC funding (EP/J021334/1) a large science exhibition was organised under the title of Working Polymers; This was held over a eight day period at the Science and Technology Pavilion of the National Eisteddfod of Wales - Meifod Mid-Wales- August 1 - August 8 and officially opened by the First Minister for Wales Carwyn Jones.
The aim of this exhibit was to highlight the science of polymers and the fact that we are now totally dependant on them but to also bring introduce EPSRC funded research in the ATJ laboratory on the use of polymers for designing nanomedicines to target diseases such as cancer. Approximately 130,000 visited the Eisteddfod over this eight day period and >20,000 were counted into the Science Pavilion. The Exhibit was staffed by Cardiff University academics and undergraduate students. Public engagement training was provided for all activity staff by Professor Jones.
Feedback was extremely positive and there was much discussion surrounding the various polymer activities in display e.g. surface of rugby balls, nappies, cardiac stents with controlled release of drugs. This also attracted the media and numerous interviews were conducted with BBC Radio and TV and also S4C. A major outcome of these is that the public now clearly view science as a fundamental part of culture and this is why the Science and Technology Pavilion sits proudly and comfortably at the heart of this very large cultural festival.
Year(s) Of Engagement Activity 2015
URL https://blogs.cardiff.ac.uk/pharmacy-engagement/2015/10/22/national-eisteddfod-2015/