Improving the delivery of 5-aminolaevulinic acid in photodynamic therapy (PDT): synthesis and biological studies of novel peptide prodrugs
Lead Research Organisation:
University of Bath
Department Name: Pharmacy and Pharmacology
Abstract
The main aim of this project is to design new drugs for photodynamic therapy (PDT) which is a treatment that can be used for destroying cancers and killing infectious bacteria. The key feature feature of PDT is that it uses a light-activated drug known as a 'photosensitiser'. When light activates the drug this results in a toxic effect to tumour cells or bacteria leading to destruction of the tumour or healing of the bacterial infection. In effect, the light 'switches on' the drug but only in areas exposed to light. We propose to look at a new type of drug based on 5-aminolaevulinic acid (ALA). This is a natural compound present in many cells but normally at very low levels. However if cells or bacteria are exposed to high concentrations of ALA then a photosensitiser is produced (a porphyrin) which absorbs light over a wide range from red to blue and can therefore be used for PDT treatment. For example, a skin tumour can be treated by first covering the tumour with a lotion containing the ALA which is then left for a few hours. After ALA has seeped into the tumour, light is shone onto it for a few minutes to kill the tumour cells. Alternatively, a solution of ALA can be injected or given orally for the treatment of tumours inside the body where an endoscope can be used to guide the light onto the tumour. One main problem with using ALA however is that it is not quickly taken up by cells. We plan to change its chemical structure so that cells or bacteria can absorb it more easily, specifically by attaching amino acids to the ALA to help its uptake by cells. After entering the cell the drug would break down to release the ALA. If the modified ALA drug can enter cells more easily then in principle it should be more effective for tumour treatment, and since amino acids are natural compounds which make up proteins these new drugs should be safe to use. As well as treating cancers, there are several types of bacterial or even fungal infection which can be treated using PDT. One advantage of PDT for treating bacteria is that bacteria resistant to antibiotics can be treated. For example, burn wounds often become infected and in this case the PDT drug can be applied to the wounds and then illuminated after the drug has penetrated the infected area. If the new types of ALA drug can be taken up more efficiently by the bacteria then again the PDT treatment should be more effective. We also hope to design modified ALA drugs that are broken down specifically by bacteria and not by healthy cells by attaching amino acids whose structures are the mirror image of those which are normally found naturally.
Technical Summary
Photodynamic therapy (PDT)is an emerging therapy for the treatment for cancer and various other human disorders. In PDT, destruction of tumours or pathogenic organisms is achieved with light following the administration of a light-activated photosensitising drug which is ideally selectively retained in, or targeted to diseased tissue relative to normal adjacent tissue. The exogenous administration of 5-aminolaevulinic acid (ALA) has attracted considerable interest for PDT since it is a naturally occurring compound present in prokaryotic and eukaryotic cells which can be metabolised to a porphyrin photosensitiser, protoporphyrin IX (PpIX)via the haem biosynthetic pathway. ALA-PDT is a powerful approach for both the detection and treatment of cancers, in particular the treatment of basal cell carcinomas using topical ALA administration, and the visualisation of early tumours in hollow organs. Moreover, it has considerable potential for use in antimicrobial applications since since various Gram-positive and Gram-negative bacteria, yeasts and fungi are able to assimilate exogenous ALA for porphyrin synthesis, thus rendering them susceptible to photosensitisation. Currently, a key limitation of ALA-PDT is the zwitterionic nature of ALA at physiological pH which limits its passage through cellular membranes and other biological barriers. Furthermore, the selective delivery and release of ALA in specific tissues is difficult to achieve. To address these issues, this proposal aims to develop novel ALA peptide prodrugs with improved cellular uptake and targeting properties and to demonstrate the effectiveness of these compounds in a variety of cell lines and microorganisms. The project will build on preliminary synthetic and biological studies that indicate that a variety of ALA peptide prodrugs are more effectively internalised than ALA itself in cell and tissue explant models and are effectively converted to ALA by intracellular esterases and peptidases, leading to PpIX production and photocytotoxicity upon irradiation.
People |
ORCID iD |
Ian Eggleston (Principal Investigator) |
Publications
Bourré L
(2009)
Protoporphyrin IX enhancement by 5-aminolaevulinic acid peptide derivatives and the effect of RNA silencing on intracellular metabolism.
in British journal of cancer
Bourré L
(2008)
5-Aminolaevulinic acid peptide prodrugs enhance photosensitization for photodynamic therapy.
in Molecular cancer therapeutics
Bourré L
(2010)
Effective photoinactivation of Gram-positive and Gram-negative bacterial strains using an HIV-1 Tat peptide-porphyrin conjugate.
in Photochemical & photobiological sciences : Official journal of the European Photochemistry Association and the European Society for Photobiology
Di Venosa G
(2015)
The use of dipeptide derivatives of 5-aminolaevulinic acid promotes their entry to tumor cells and improves tumor selectivity of photodynamic therapy.
in Molecular cancer therapeutics
Dixon MJ
(2007)
Novel prodrug approach to photodynamic therapy: Fmoc solid-phase synthesis of a cell permeable peptide incorporating 5-aminolaevulinic acid.
in Bioorganic & medicinal chemistry letters
Dondi R
(2016)
Flexible synthesis of cationic peptide-porphyrin derivatives for light-triggered drug delivery and photodynamic therapy.
in Organic & biomolecular chemistry
Giuntini F
(2009)
Improved peptide prodrugs of 5-ALA for PDT: rationalization of cellular accumulation and protoporphyrin IX production by direct determination of cellular prodrug uptake and prodrug metabolization.
in Journal of medicinal chemistry
Giuntini F
(2008)
Quantitative determination of 5-aminolaevulinic acid and its esters in cell lysates by HPLC-fluorescence.
in Journal of chromatography. B, Analytical technologies in the biomedical and life sciences
Giuntini, F.
(2008)
A novel approach to improve cellular delivery of 5-aminolaevulinic acid: new ALA-containing peptide prodrugs for photodynamic therapy
in Journal of Peptide Science
Wang JT
(2012)
Photochemical internalisation of a macromolecular protein toxin using a cell penetrating peptide-photosensitiser conjugate.
in Journal of controlled release : official journal of the Controlled Release Society
Yaghini E
(2017)
Endolysosomal targeting of a clinical chlorin photosensitiser for light-triggered delivery of nano-sized medicines.
in Scientific reports
Yaghini E
(2014)
Fluorescence lifetime imaging and FRET-induced intracellular redistribution of Tat-conjugated quantum dot nanoparticles through interaction with a phthalocyanine photosensitiser.
in Small (Weinheim an der Bergstrasse, Germany)
Description | In this project, we have synthesised and tested a range of new drugs for photodynamic therapy (PDT) - a treatment for destroying cancers and harmful microorganisms using light. In particular, we have prepared and evaluated a series of 5-aminolaevulinic acid (ALA)-containing peptide compounds with the aim of improving the cell uptake of ALA, which can then be converted to a light-activatable drug for PDT (a porphyrin photosensitiser) once inside a cell. By selecting the right peptide to link with the ALA, the resulting compound can be designed to pass through cell membranes more easily and be far more stable in water compared to ALA itself. The results we obtained using four different types of cancer cell demonstrated that the incorporation of ALA into a short peptide derivative is an effective general approach for increasing cellular delivery of ALA. Several compounds show significantly elevated cellular accumulation in cells compared to ALA, and are indeed able to induce porphyrin production at concentrations where ALA is not effective. Since the release of ALA from the compound depends upon the action of specific peptidase enzymes it should be possible to fine-tune the structures of our ALA prodrugs to target and effect selective ALA release in normal or malignant tissue for diagnostic or therapeutic purposes. The synthetic route that we have developed is straightforward, simple to scale up, and allows for a wide range of structural variations to be easily introduced. We have also investigated the use of other types of peptide known as cell-penetrating peptides (CPP) as a novel way of improving the delivery of ALA or other "preformed" photosensitisers to cells and bacteria. We were able to demonstrate the concept of using a CPP to deliver multiple units of ALA into cells, and develop the chemistry that will allow the therapeutic and diagnostic applications of this approach to be explored in detail. We have also prepared and tested a new porphyrin-containing CPP compound which was very effective for light-induced killing of bacteria. Again the chemistry we have developed will allow the antibacterial potential of this type of compound to be investigated further, using other peptides and photosensitisers. Such CPP-photosensitisers also have a range of other interesting applications for light-activated drug delivery which we are now in a position to exploit. |
Exploitation Route | The ALA peptide prodrugs that we have developed offer a new way to effectively deliver ALA for the targeted treatment of both cancer and infectious diseases. This is being exploited in our laboratory, and various clinicians have expressed an interest in working with these compounds. The CPP-photosensitisers that we designed are of great interest for the light-activated delivery of drugs and biological probes. We have started a new BBSRC-funded project that takes this idea forward, with the support of clinicians and industrial researchers who would like to make use of these techniques for the delivery of therapeutic agents against cancer. |
Sectors | Chemicals Healthcare Pharmaceuticals and Medical Biotechnology |
Description | The UCL group have presented work on Photodynamic Therapy at the Royal Society Science Exhibition, 2009, which was attended by the media, the general public and school groups. Details of our work on ALA-PDT on the University of Bath website, and presentation at open fora at Bath have prompted direct enquiries from a member of the public regarding the availability of this treatment in the UK. The research findings have been used as the basis of successful applications for both BBSRC and non-RCUK funding. |
First Year Of Impact | 2009 |
Sector | Chemicals,Healthcare,Pharmaceuticals and Medical Biotechnology |
Impact Types | Societal |
Description | Evaluation of ALA peptide prodrugs in an oncogenic cell line |
Organisation | University of Buenos Aires |
Country | Argentina |
Sector | Academic/University |
PI Contribution | Compounds provided from BBSRC-funded project |
Collaborator Contribution | Studies of ALA and derivatives were undertaken in the Hb4a immortalised breast epithelial line and its tumourigenic rastransfected counterpart, which was part of the original proposal. The research was carried out jointly between UCL and Buenos Aires, and a research assistant (Gabriella Di Venosa) also spent 2 months in the UK at UCL and Bath. It was shown that the ras-transfected cell line took up more ALA although the photocytotoxicity threshold for this cell line was also higher. |
Impact | Manuscript in submission |
Start Year | 2008 |
Description | Peptide targeted agents for photodynamic therapy and photobiology |
Form Of Engagement Activity | Scientific meeting (conference/symposium etc.) |
Part Of Official Scheme? | Yes |
Type Of Presentation | keynote/invited speaker |
Geographic Reach | Regional |
Primary Audience | Participants in your research and patient groups |
Results and Impact | Invited lecture 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). 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 |