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Immunophotodynamic therapy of cancer: concepts and applications

Final Report Summary - IMMUNO PDT (Immunophotodynamic therapy of cancer: concepts and applications)

Photodynamic therapy of cancer, i.e. the generation of reactive oxygen species in the tumour environment which follows the irradiation of suitable photosensitizing molecules, is an attractive modality for the selective ablation of inoperable superficial neoplastic lesions, such as certain head and neck, gastrointestinal, urogenital and gynaecological tumours. It is likely that the scope and efficacy of photodynamic therapy could be enhanced by:
- the availability of novel efficient photosensitisers which absorb at in the red / near infrared region of the spectrum, where light penetration of tissues is maximal;
- the use of antibody-photosensitiser conjugates or conceptually related innovative delivery systems, which concentrate the photosensitizing molecules at suitable neoplastic sites.
The tumour neo-vasculature appears to be an attractive target, since the selective delivery of photosensitisers may occlude the tumour blood vessels, thus causing an avalanche of tumour cell deaths.

In this project, we have put together a network of academic research groups and companies, for the development of antibody-based targeted photodynamic therapy modalities. The planned research activity started with the synthesis of novel photosensitising molecules suitable for conjugation to antibodies, and with the identification of novel human monoclonal antibodies, capable of a selective targeting of the tumour neovasculature for immuno-PDT applications.

Following an extensive in vitro characterisation of the most promising antibodyphotosensitiser conjugates (including an innovative delivery concept in which photosensitisers are non-covalently bound to the antibody), the therapeutic potential of the best antibodyphotosensitiser conjugates have been tested in rodent models of cancer and have displayed an impressive tumour ablation performance. The L19, F16 and F8 antibodies have been moved to GMP manufacture, thus paving the way for future clinical applications.

We made substantial progress in terms of:
i. the synthesis of novel infrared photosensitisers with sufficiently water solubility (i.e. not sticky to unwanted cells and tissues), which absorb in the near-infrared and redshifted light spectrum and which efficiently generate singlet oxygen and / or other reactive oxygen species;
ii. isolation and validation (in vitro and in vivo) of novel human antibodies to accessible tumour-associated antigens;
iii. investigation of a novel method for the conjugation of the antibody and photosensitiser molecules;
iv. evaluation of the conjugates in vitro e in vivo. Our novel PDT agents have been extensively tested in vitro, in order to ascertain whether bound photosensitisers can retain singlet oxygen production activity upon irradiation;
v. The agents have been tested in rodent models of cancer and now will open novel therapeutic opportunities for the selective treatment of superficial tumours in accessible body cavities.

Most importantly, there is a reasonable expectation of a medical benefit for cancer patients stemming both directly and indirectly from this project:
- directly, since immuno-PDT procedures promise to be invaluable for the selective ablation of inoperable superficial neoplastic lesions, such as certain skin, head and neck, gastrointestinal, urogenital and gynecological tumours. Indeed, the discovery that squamous cell carcinoma of the skin (the second most common skin cancer) strongly reacts with the most advanced antibodies studied in the IMMUNOPDT project (F8 F16, L19) paves the way for Immuno-PDT applications in this cancer type, where multiple lesions often recur in certain patients (e.g. transplant patients) and where repeated surgical excision remains an undesirable and disfigurating medical procedure.
- indirectly, since the knowledge generated by the validation of novel antibodies for vascular targeting applications is likely to have an impact in other forms of immunotherapy, including the use of full IgGs and antibody-cytokine fusions for cancer therapy. We believe that our project and our network have contributed to the scientific and the technical objectives of the combating cancer area.

Furthermore, in addition to our regular meetings, we have fulfilled our dissemination activities duties by organising an experimental course on antibody phage technology. The course has been advertised in Nature, has been over-subscribed, and has brought together participants from over 20 countries.

The project also has resulted in:
- 40 publications in peer-reviewed scientific lournals;
- 10 manuscripts in press;
- 87 invited lectures at international congresses;
- 8 patent applications.

Project objectives and execution

The present project had as objectives the synthesis and conjugation of novel infrared and redshifted photosensitisers to the most promising antibodies against vascular tumour antigens obtained by human antibody technology, the immunohistochemical characterisation, the biodistribution and imaging targeting in vivo, in order to select the best antibodyphotosensitiser conjugates to be taken forward into clinical trials as a final objective.

The structure of the project was clearly defined:

- In the first year, synthesis of novel photosensitisers and identification of the most suitable antibodies for Immuno-PDT applications (fall back position: we already had a palette of photosensitisers and of antibodies which were used as a benchmark for the new molecules, and which were coupled together).
- In the second year, we continued to synthesise novel photosensitisers, investigated innovative conjugation strategies and performed an in vitro characterisation of antibodyphotosensitiser conjugates.
- In the third year, the therapeutic activity of the most promising antibodyphotosensitiser conjugates were tested in rodent models of cancer, in comparison to antibodies of irrelevant specificity and unconjugated photosensitisers.