Peptide-based solutions for light-triggered delivery of macromolecular therapeutics and nanoparticles

Lead Research Organisation: University of Bath
Department Name: Pharmacy and Pharmacology

Abstract

The effectiveness of many important drugs is much reduced because they cannot be adequately delivered to the fluid inside cells where their biological targets are located. This means that higher doses are needed, leading to an increase in potential side effects and reduced patient quality of life. Such drugs are often poorly absorbed in the body because they are taken up into cells by endocytosis, where the cell wall or membrane envelops drug molecules leaving them trapped inside small compartments (endosomes, lysosomes) within the cell from which they must escape in order to reach the right part of the cell (e.g. the nucleus). If the drug cannot escape efficiently, it may instead be broken down by enzymes, or expelled from the cell. The technique of photochemical internalisation (PCI) is a novel way to get around this problem. Here, the drug of interest is administered along with a photosensitiser, a molecule that can facilitate the escape process when activated by light. Ideally, the photosensitiser is activated with a low dose of red light which causes minimal damage to healthy tissue and also allows light-activation to take place deep within the target tissue, as tissue absorption at red light wavelengths is weak. In contrast, UV light-activated drug release cannot be used effectively in tissue not only due its mutagenic effects, but also because tissue absorption of UV light is too strong and limits the effect to a depth of few cell layers from the surface. Drugs that can be delivered with PCI range from toxins for cancer treatment to molecular agents for gene therapy, and light can either be shone directly onto the target tissue or guided from a laser down optical fibres placed within the tissue to allow illumination of larger volumes. When cells are exposed to PCI light treatment, to activate the photosensitiser, these molecules absorb energy and generate short-lived reactive chemical compounds that break down the walls of the drug-containing compartments, releasing the drug to allow it to reach its target. However, in order for PCI to work effectively, the drug and photosensitiser employed must be incorporated into the same compartment inside the cell and must of course both efficiently enter the cell in the first place. The initial aim of our project is therefore to develop new photosensitiser molecules for PCI that are water-soluble, cross cell membranes effectively, and are also taken up into cells by endocytosis so that they may be localised in the right cell compartments with drugs that are administered at the same time. We have already shown that prototype molecules of this sort give a much more efficient PCI effect than that obtained with a simple photosensitiser. To further improve the PCI approach, we then want to develop systems where both a drug and a photosensitiser are associated with the same carrier molecule so that the uptake of both components is enhanced and their localisation in exactly the same cell compartment is guaranteed. To make this approach as general and as flexible as possible, we aim to develop systems where the carrier and photosensitiser can be easily interchanged, and a wide range of drug molecules can be incorporated in such a way that they can be released from the carrier inside the cell once the PCI light treatment has been carried out. As a final refinement, we will also look at the possibility of making delivery systems that can be targeted to a specific part of the body and switched on specifically in diseased tissue only, so that the PCI therapy can be performed with pin-point selectivity, exactly where it is required.
All the results of this project will be of direct benefit to the healthcare field by providing a new means to more effectively deliver diverse chemotherapeutic agents, improving efficacy, lowering dosage, and minimising side-effects.

Technical Summary

The use of several important biopharmaceuticals is severely hampered by their limited ability to reach intracellular targets. This is either due to poor diffusion across the cell membrane or endosomal/lysosomal sequestration upon uptake . Photochemical internalisation (PCI) is a promising solution which exploits the photodynamic action of sub-toxic doses of photosensitisers to promote rupture of lysosomal vesicles, so that entrapped drugs can reach their targets. PCI is designed to use red light where tissue absorption is relatively weak, unlike blue or UV light, so light-triggered, site-specific drug release can be effected at therapeutically useful depths in tissue.
We aim to develop innovative solutions to 3 key challenges in the PCI approach, using peptide-based delivery systems.
1) Photosensitisers for PCI must be lysosomotropic, to localise in the same vesicles as administered drugs. This makes many photosensitisers clinically used for photodynamic therapy unsuitable for PCI. We aim to overcome this by conjugating photosensitisers to cell-penetrating peptides (CPPs) to both improve their uptake and control sub-cellular localisation.
2) PCI requires the photosensitiser and drug to localise in the SAME intracellular vesicle. We can achieve this by covalently attaching a drug cargo to a peptide carrier, from which it may dissociate and reach its target post-PCI. Alternatively, a peptide carrier, with the photosensitiser attached, may be used to generate a drug-encapsulating nanoparticle, such as a liposome.
3) For maximum efficacy, the combination of photosensitiser and drug should be delivered only to specific tissues. A CPP construct that is activated for uptake by disease-dependent levels of a protease activity is an ideal way to achieve this.
We will characterise our constructs in detail with respect to their photophysical properties and efficiency at delivering diverse drug molecules in a variety of in vitro cell models and validate them in vivo.

Planned Impact

Our research has the potential to deliver impact for a wide range of beneficiaries, including clinicians and their patients, biomedical scientists and the pharmaceutical industry, as well as the direct academic participants at Bath and UCL. For the general public, novel methodologies that can facilitate the delivery of poorly absorbed chemotherapeutic agents will lead to the development of safer and more effective treatments, allowing the use of lower doses and reduction in chemotherapy side effects/morbidity. This outcome will have an impact upon quality of life in the UK and increase the effectiveness of public healthcare. In the pharmaceutical sector, biotherapeutics now comprise a significant proportion of all drugs on the market. Achieving the efficient delivery of such large often hydrophilic molecules to targeted tissues is a major challenge that can limit the therapeutic potential of otherwise promising clinical candidates, and may result in their abandonment, despite huge investments in their discovery and development. The development of tools that expand the scope of the PCI technique are likely to be of great benefit to the pharmaceutical industry in the next 5-10 years (half the new drugs in late-stage clinical trials will soon be antibodies, peptides, nucleic acids, and other macromolecules). For companies in the UK, this could impact positively on economic performance and competitiveness in the global market place, and thus the wealth of the UK.

As well as being of direct benefit to academic researchers in the field of drug delivery, our research will also be of benefit to the academic community at large, who will gain from the knowledge and reagents obtained in these studies that can be applied to other research endeavours (e.g. in gene therapy). The research will also have a key impact for the academic institutions involved, in that our work should produce intellectual property that would be considered very valuable by pharmaceutical companies, thus leading to the possibility to benefit from revenue streams obtained from licensing opportunities. Finally, the researchers who will perform the proposed studies will benefit from the opportunities to participate in a multidisciplinary research project, with valuable training in a range of chemical and biological techniques. This will greatly enhance their value for ultimate employment in either industrial or academic settings. As such, the project will constitute a long-term training investment in the researchers and the creative output of the UK.

We are already in discussion with potential beneficiaries and end users of our proposed research in both the clinical and commercial arena. For example, the Norwegian pharmaceutical company PCI Biotech AS (specialists in the development of PCI technology), and their research director, Dr Anders Hogset; and Mr Colin Hopper, Academic Head of the Unit of Oral and Maxillofacial Surgery, at UCL Hospital. We have similarly taken taken steps to engage with other leading academic figures in light-based drug delivery research (Prof. Kristian Berg, Institute for Cancer Research, University of Oslo, and Prof. Alex Lou, National Taiwan University). Furthermore, the National Medical Laser Centre, where the UCL group are based, is a translational research institute with extensive experience of converting fundamental bioscience into clinical applications and also exploiting its commercial potential. We are therefore very well placed to both identify results of potential interest to beneficiaries in the biomedical and pharmaceutical arenas and also maximise their economic and societal impact.
 
Description The effectiveness of many important drugs is much reduced because they cannot be adequately delivered to the fluid inside cells where their biological targets are located. This means that higher doses are needed, leading to an increase in potential side effects and reduced patient quality of life. Such drugs are often poorly absorbed in the body because they are taken up into cells by endocytosis, where the cell wall or membrane envelops drug molecules leaving them trapped inside small compartments (endosomes, lysosomes) within the cell from which they must escape in order to reach the right part of the cell (e.g. the nucleus). If the drug cannot escape efficiently, it may instead be broken down by enzymes, or expelled from the cell. The technique of photochemical internalisation (PCI) is a novel way to get around this problem. Here, the drug of interest is administered along with a photosensitiser, a molecule that can facilitate the escape process when activated by light. Ideally, the photosensitiser is activated with a low dose of red light which causes minimal damage to healthy tissue and also allows light-activation to take place deep within the target tissue, as tissue absorption at red light wavelengths is weak.

When cells are exposed to PCI light treatment, to activate the photosensitiser, these molecules absorb energy and generate short-lived reactive chemical compounds that break down the walls of the drug-containing compartments, releasing the drug to allow it to reach its target. However, in order for PCI to work effectively, the drug and photosensitiser employed must be incorporated into the same compartment inside the cell and must of course both efficiently enter the cell in the first place. The initial aim of our project has been to develop new photosensitiser molecules for PCI that are water-soluble, cross cell membranes effectively, and are also taken up into cells by endocytosis so that they may be localised in the right cell compartments with drugs that are administered at the same time. We have devised efficient syntheses of a range of photosensitiser molecules that can be selectively and efficiently attached to cell-penetrating peptides. As desired, this makes the photosensitisers which are largely water-insoluble, soluble in water and also able to efficiently cross cell membranes. Importantly, they also localise in the right part of the cell, i.e. endosomes, where drug molecules of interest may be trapped. We have shown that our new molecules give a much more efficient PCI effect than that obtained with a simple photosensitiser alone. We have studied our novel photosensitisers both for the delivery of macromolecular drugs and also nanoparticles (quantum dots) that have potential in molecular imaging applications.

To make this approach as general and as flexible as possible, we have developed an approach that allows the cell-penetrating peptide carrier and photosensitiser to be easily interchanged. This means that we can make use of photosensitisers that absorb red light most efficiently, which is what is required to translate molecules of this type into in vivo applications.

To further improve the PCI approach, we have now developed liposomal systems where both a drug and a photosensitiser are associated with the same carrier peptide molecule so that the uptake of both components is enhanced and their localisation in exactly the same cell compartment is guaranteed. Our approach allows us as before to interchange the cell-penetrating peptide carrier and photosensitiser, and particularly to employ peptides that will target delivery to specific cell types.
Exploitation Route The compounds that we have developed so far have been extensively investigated for light-activated drug delivery in vitro. They are also of great interest for the specific intracellular delivery of biological probes, and we have begun to collaborate with other groups who have seen the potential of the research that we are developing. We have also been in contact with one of the major industrial players in the development of biological therapeutics (Medimmune), who have expressed an interest in our PCI approach.

As part of the exploitation of our findings, we now intend to explore the potential of our drug delivery platform for the targeted non-invasive delivery of anti-cancer agents.
Sectors Chemicals,Healthcare,Pharmaceuticals and Medical Biotechnology

 
Description To date, the findings of our research have been mainly exploited within the research programme itself and to stimulate further related investigations. Recent presentations and publications relating to results from our first completed objective (the development of a range of novel CPP-photosensitiser derivatives for PCI) have allowed us to initiate/discuss new collaborations and applications for these research tools. This includes researchers at other academic institutions, as well as clinicians and industrial researchers who have expressed initial interest in our research programme.
First Year Of Impact 2012
Sector Chemicals,Healthcare,Pharmaceuticals and Medical Biotechnology
Impact Types Societal

 
Description Cell-penetrating peptide-targeted photosensitiser for light-triggered intracellular delivery of macromolecules 
Organisation MedImmune
Country United States 
Sector Private 
PI Contribution MTA-based discussion re material supply, namely a cell-penetrating peptide-porphyrin conjugate for use in light-triggered intracellular delivery of macromolecules, synthesised in Bath.
Collaborator Contribution N/A at this time
Impact N/A
Start Year 2015
 
Description Delivery of therapeutic peptides by PCI 
Organisation University of Nottingham
Country United Kingdom 
Sector Academic/University 
PI Contribution Synthesis of reagents (ligatable photosensitisers) and preparation of peptide conjugates
Collaborator Contribution Evaluation of efficiency of labelled peptide in promoting light-activated intracellular relocalisation from endosomal compartments.
Impact None yet
Start Year 2014
 
Description CAGED IRON CHELATORS AND PEPTIDE TARGETING: NOVEL STRATEGIES FOR LIGHT-ACTIVATED THERAPIES AND PHOTOBIOLOGY 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact The invited presentation by Dr Ian Eggleston in January 2014 took place at the Faculty of Pharmacy, Charles Univerity of Prague, in Hradac Kralove, Czech Republic. The talk stimulated significant discussion with scientists, clinicians, and early career researchers at this institute.

We have previously hosted a visiting researcher from this Department. This talk has opened up the possibility for further collaborative visits, including undergraduate and postgraduate exchanges.
Year(s) Of Engagement Activity 2014
 
Description Clickable photosensitisers and peptides - efficient routes to targeted agents for PDT and photobiology 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact Invited lecture by Dr Ian Eggleston at Biennial meeting of European Society for Photobiology, Aveiro, Portugal, August-September 2015. The talk sparked discussions with a variety of researchers who expressed interest in collaborations, as well as requests for information from early career researchers who were interested in our research.
Year(s) Of Engagement Activity 2015
 
Description Light-Triggered Delivery of Macromolecular Therapeutics by Photochemical Internalisation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Industry/Business
Results and Impact Invited talk at Medimmune, Cambridge (November 2015). This invitation arose from a presentation given by the PDRA engaged on the project at another meeting. The presentation was given jointly by Dr Eggleston and the UCL project lead, Professor MacRobert. Following the meeting, discussions took place with staff in the Antibody Discovery and Protein Engineering Group at Medimmune regarding potential future collaborations.
Year(s) Of Engagement Activity 2015
 
Description Peptide targeted agents for photodynamic therapy and photobiology 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation keynote/invited speaker
Geographic Reach Regional
Primary Audience Other audiences
Results and Impact Invited lecture by Dr Ian Eggleston at 7th Cancer Research at Bath (CR@B) Symposium - a showcase of presentations and posters highlighting cancer research activities taking place in the South West (17 April, 2013 at University of Bath).
The talk stimulated a significant amount of discussion, and requests for information from early career researchers who were interested in our research. There were also enquiries from members of the public who were interested in access to the therapeutic techniques and drug delivery methods described.


After my talk, I was approached by a member of the public who wanted to know about the availability of photodynamic therapy and related therapies via the NHS. I was able to direct them to appropriate sources of information and advice.
Year(s) Of Engagement Activity 2013
 
Description Peptide targeting of photosensitisers for photodynamic therapy and drug delivery applications 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Professional Practitioners
Results and Impact Invited lecture at the School of Medicine and Photobiology Unit, University of Dundee (January 2017). The talk provided the focus for discussions regarding future collaborations with staff from the Photobiology Unit.
Year(s) Of Engagement Activity 2017
 
Description Peptide targeting of photosensitisers for photodynamic therapy and drug delivery applications 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Invited presentation at the School of Pharmacy, University of Cardiff (May 2017). The talk sparked questions and discussions from early career researchers who were present, and also discussions with members of academic staff regarding potential collaborations.
Year(s) Of Engagement Activity 2017
 
Description Peptide targeting of photosensitisers for photodynamic therapy and drug delivery applications 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Invited plenary lecture at Closing Meeting of Singlet Oxygen Strategy Marie Curie Network, Ghent, Belgium (June 2016). This sparked questions and discussion from early career researchers who were attending the meeting from several countries and also other speakers from the photobiology field.
Year(s) Of Engagement Activity 2016
 
Description Peptide-targeted agents for photodynamic therapy and photobiology 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation keynote/invited speaker
Geographic Reach International
Primary Audience Other audiences
Results and Impact Talk presented by Dr Ian Eggleston at Royal Society of Chemistry Peptide and Protein Group, 7th Chemistry and Biology of Peptides Meeting, MRC Laboratory, Cambridge, UK, Jul 2012. The talk stimulated a significant amount of discussion, and requests for information from early career researchers who were interested in our research.

no actual impacts realised to date
Year(s) Of Engagement Activity 2012
 
Description Peptide-targeted photosensitisers as tools for light-triggered delivery of macromolecular therapeutics 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact Poster by Dr Ruggero Dondi at Nanopeptide 2015 Meeting: International Meeting on Peptide Materials and Technologies, Strathclyde, March 2015. Poster sparked discussions with a variety of academic researchers and industrial scientists who expressed interest in collaborations.
Year(s) Of Engagement Activity 2015
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact Poster presentation by Dr Ian Eggleston at Photodynamic Therapy and Photodiagnosis in Clinical Practice Meeting, Brixen, Italy, October 2014).
Poster presentation sparked discussions with a variety of researchers who expressed interest in collaborations.
Year(s) Of Engagement Activity 2014
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation paper presentation
Geographic Reach International
Primary Audience Other audiences
Results and Impact Poster presentation by Dr Ruggero Dondi at Biennial meeting of European Society for Photobiology, Liege, Belgium, August-September 2013. Poster presentation sparked discussions with a variety of researchers who expressed interest in collaborations.

no actual impacts realised to date
Year(s) Of Engagement Activity 2013
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Other audiences
Results and Impact Poster presentation by Dr Ruggero Dondi at at Cancer Research at Bath (CR@B) Symposium - a showcase of presentations and posters highlighting cancer research activities taking place in the South West (November 2013 at University of Bath). Poster presentation sparked discussions with a variety of researchers who expressed interest in collaborations.
Year(s) Of Engagement Activity 2013
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact Poster presentation by by Dr Ruggero Dondi at Royal Society of Chemistry Peptide and Protein Group, 9th Chemistry and Biology of Peptides Meeting, Univ. Nottingham, UK, Jul 2014. Poster presentation sparked discussions with a variety of researchers who expressed interest in collaborations.
Year(s) Of Engagement Activity 2014
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Poster presentation by by Dr Ruggero Dondi at 8th Chemistry and Biology of Peptides Meeting, Univ. Nottingham, UK, Jul 2013.
Poster presentation sparked discussions with a variety of researchers who expressed interest in collaborations.
Year(s) Of Engagement Activity 2013
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Talk by Dr Ruggero Dondi at RSC Protein and Peptide Science Group Early Stage Researcher Meeting, Burlington House, London, UK, Nov-2014. Talk sparked discussions with a variety of academic researchers and industrial scientists who expressed interest in collaborations.
Year(s) Of Engagement Activity 2014
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Poster presentation by Dr Ruggero Dondi at RSC Chemical Biology and Bio-Organic Chemistry Forum, University of Southampton, May 2014.
Poster presentation sparked discussions with a variety of researchers who expressed interest in collaborations.
Year(s) Of Engagement Activity 2014
 
Description Peptide-targeted photosensitisers via efficient 'click' conjugations in solution 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact The presentation sparked discussion with a number of early career researchers and also potential industrial collaborators.

None so far.
Year(s) Of Engagement Activity 2014
 
Description Targeted agents for photodynamic therapy 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact Invited presentation at Skin@Bath Symposium, Bath (December 2017). The talk sparked questions and discussion from a variety of delegates at the meeting. This has led to discussions regarding future collaborations and funding applications with one business represented at the meeting.
Year(s) Of Engagement Activity 2017