CEO collects high scientific and technical competences. Associated with this Centre are:

1. CEO – Centre of Excellence in Optronicsno profit Consortium set up in Florence in 1989 promotes and develops scientific research and technological innovation in the field of optoelectronics CEO collects high scientific and technical competences. Associated with this Centre are: • The University of Florence through its: • Department of Physics • Department of Electronics and Telecommunications • Department of Clinical Physiopathology • The National Research Council (CNR) trough its: • Institute of Quantum Electronics (IEQ) • Institute of Research on Electromagnetic Waves (IROE) • The National Institute of Applied Optics (INOA) • The El.En. S.p.A.

3. ACTIVITY

4. CLAM -Centre for Laser Application in Medicine Development of new optoelectronic techniques for biomedical and environmental applications. MAIN ACTIVITIES: • Multispectral Imaging Autofluorescence Microscopy • Laser Systems in Photochemotherapy and Photosurgery • Optical Tomography Systems • Consultant for the Regional Reference Centre for Genetics of Jaundice and Hereditary Colestatic Diseases

5. CLAM -Centre for Laser Application in MedicineAutofluorescence imaging: diagnostic applications C E L L T I S S U E

6. THEMATICAREA: 1.1.2 Antiblastic Drug Resistance (ANDREA)

7. Antiblastic Drug Resistance (ANDREA) • Treatment failure in cancer is one of the greater problems in the management of human diseases. • The limited efficacy of cancer chemotherapy is due to two main factors: • The extremely low therapeutic index of the drugs presently available; • The almost invariable occurrence of "innate" or treatment-induced resistance of tumor cells to anticancer drugs.

8. Antiblastic Drug Resistance (ANDREA) • The aim of the present programme is to attack the problem of anticancer drug resistance through a wide and multidisciplinary research action. • The ultimate goal is to achieve a marked improvement of cancer chemotherapy, thus contributing to realise the objectives of the priority thematic area (1.1.2 – 1.1.2.2).

9. Antiblastic Drug Resistance (ANDREA) The strategy of this proposal consists of establishing a network of excellence in which the biomedical, biotechnological and biophysical approaches are integrated in a way that allows a better exploitation of the potentialities of different disciplines such as molecular biology, pharmacology, chemistry optoelectronics, etc.…..

10. Antiblastic Drug Resistance (ANDREA) • Research will be performed to identify the molecular determinants of chemotherapy with the aim to sensitize tumor cells to the cytotoxic agents of clinical use. • The molecular targets of the biomodulators of chemoresistance studied will be as follows: • proteins that are responsible for drug efflux; • proteins involved in drug internalization; • DNA repair systems; • systems required for modulating the expression of enzymes involved in the metabolism of cytotoxic drugs; • networks involved in cell cycle, apoptosis, as well as survival in the hypoxic niches occurring in solid tumors.

11. Antiblastic Drug Resistance (ANDREA) • Pharmacodynamic studies will be performed by Autofluorescence Microspectroscopy (AMS) and Multispectral Imaging Autofluorescence Microscopy (MIAM) techniques, that allow the monitoring of drug intracellular distribution on single living cell. • Ultrasensitive fluorescence techniques for single molecule detection will be applied for studies on molecule diffusion in cell membrane and within the cytoplasm. • Optical tweezers, an optical technique for selective capture and translation of dielectric microparticles, will be applied to study surface interactions.

12. Antiblastic Drug Resistance (ANDREA) Fluorescence imaging of HL60 cell treated with doxorubicin

13. Antiblastic Drug Resistance (ANDREA) Fluorescence imaging of HL60 cell treated with doxorubicin

14. Antiblastic Drug Resistance (ANDREA) Emission spectra recorded from HL60 cells treated and untreated with doxorubicin

15. Antiblastic Drug Resistance (ANDREA) RESEARCH OBJECTIVES: • To highlight the pharmacodynamics of different antiblastic drug classes at single cell level. • To study cell-drug interaction in terms of surface phenomena, drug transport, cell response. • To identify drug resistance mechanisms. • To implement analytical methods to individuate resistant cells. • To individuate possible strategies opposing resistance mechanisms in cells. • To individuate more efficacious drugs.

16. Antiblastic Drug Resistance (ANDREA) EXPLOITATION OF RESULTS, BENEFITS AND PERSPECTIVES: • Development of new diagnostic and prognostic techniques. • Identification of new therapeutic strategies. • Identification of new modalities for drug administration. • Development of strategies to reverse resistance mechanisms.

17. Antiblastic Drug Resistance (ANDREA) SECTORS INVOLVED: • Pharmaceutical Industries; • Centres for Cancer Diagnostic and Therapy; • Committees and Institutions of Public Safety; • Research Institutions; • Other Industries; • Other Institutions.

18. Antiblastic Drug Resistance (ANDREA) SOME OF THE PARTNERS: • Consorzio CEO, Centre of Excellence in Optronics, CLAM Unit, Centre for Laser Application in Medicine, Florence • Unit of Haematology, Careggi Hospital, Florence • Dept. of Clinic Physiopathology, University of Florence • Dept. of Pharmacology, University of Florence • Dept. of Pharmacology, University of Siena • Single Molecule Spectroscopy Group, Forschungszentrum Julich, Germany • The Department of Physics, Imperial College, London, UK • Dep. of oncology, Lund University Hospital, Sweden • Dep. of Physics, Lund Institute of Technology, Sweden We are in touch with other possible partners, besides those mentioned.

19. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests. (DIET) THEMATIC AREA: 1.1.5

20. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) • Life in industrial countries is characterized by high-pressure rhythms and often not enough care is dedicated to the importance of diet. • Nutritionists indicate that incorrect alimentation can cause or increase the risk ofmany “modern” diseases • Thus, the demand for products improving performance and appearance is increasing exponentially.

21. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests. (DIET) • The market offers a wide variety of products to integratedietary deficiencies and advertising campaigns encourage consumers to use dietary supplements, often without an adequate, expert filter (doctors, nutritionists, etc.). • Moreover, the wide use of diet integrators in zootechnics has great relevance for at least two reasons: • the transmission of the substances to the human organisms through the food chain; • the development of products for human use on the basis of substances previously utilized for zootechnics.

22. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) • Products suggested primarily for athletes or subjects performing an intense physical activity are now widely used also by people “not physically stressed”, in order to improve their own fitness and appearance. • At present, we do not have enough information about the risks of a chronic exposure to dietary supplements, assumed both directly and indirectly (through the food chain). • The performed studies have been preferentially directed at a population of sportsmen and mostly concerned acute effects. • A further weak point in the prevention of risks for human health is the scarce quality control performed on these products.

23. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) • The aim of this programme is to elucidate the real necessity of supplements for “normal” population (if compared to athletes with intense physical activity), to evaluate the efficacy of the treatments, to investigate and monitor acute and long-term effects of integrator intake. • Besides, part of the program will be devoted to develop new detection methods as well as new toxicity tests in vitro (alternative to animals), utilizing suitable experimental models as cell and tissue cultures.

24. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) RESEARCH OBJECTIVES: • To clear up the therapeutical need. • To study acute and long-term systemic effects. • To study the mechanisms of action at the cellular level. • To study metabolic and catabolic pathways. • To characterize new classes of molecules. • To develop new analytical tests.

25. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) The ultimate goal is to achieve a marked improvement in the knowledge of the complex interaction between integrators intake and metabolism, genetic background, environmental and behavioural factors to identify key risk factors as well as beneficial effects.

26. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) RESEARCH ACTIVITIES Studies will be performed at molecular, cellular and systemic level and will concern: • Identification and monitoring of pharmacologically active compounds; • Characterization of new classes of molecules; • Study of the metabolic and catabolic pathways; • Studies on drug distribution at cellular, tissue and systemic level; • Studies on the effect at cellular level: genotypic, fenotypic and metabolic aspects; • Monitoring of parameters relevant for studying acute and long-term systemic effects; • Statistical studies; • Development of new analytical tests.

27. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests. (DIET) METHODS and TECHNIQUES: Standardized as well as new and advanced techniques will be utilized: • Chromatographic techniques; • Techniques and methods of clinical chemistry; • Cytochemical and immunocytochemical methods; • Molecular biology techniques; • Optoelectronic techniques.

28. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests. (DIET) Fluorescence imaging of blood cells after treatment with fluorocarbons

29. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) Autofluorescence Imaging Control Treated Cell autofluorescence (emission at 450 nm) monitored during treatment increasing the reduced form of NAD(P)

30. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) EXPLOITATION OF RESULTS, BENEFITS, PERSPECTIVES: • Fit use; • Improvement of salutary effects; • Decrease of risks for acute or long-term toxicity; • Decrease of undesired side effects; • Availability of new analytical methods and toxicity tests.

31. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) SECTORS INVOLVED: • Pharmaceutical Industries; • Committees and Institutions of Public Safety; • Sport Committees, Sport Unions, Sport Leagues; • Research Institutions; • Sport preparer, sport physicians, nutritionists; • Fitness centres; • Other Industries; • Other Institutions.

32. Diet integrators: therapeutic need, acute and chronic effects, development of suitable analytical tests (DIET) SOME OF THE PARTNERS: • Consorzio CEO, Centre of Excellence in Optronics, CLAM Unit, Centre for Laser Application in Medicine, Florence • CISM - Centre for Mass Spectrometry, University of Florence • Dept. of Clinic Physiopathology, University of Florence • Ist. Superiore di Sanità, Divisione Alimenti per Zootecnia, Rome • Dept. of Anatomy, Histology and Forensic Medicine, University of Florence • Single Molecule Spectroscopy Group, University of Regensburg, Germany • Laboratoire Anti-Dopage, Ins. de Medecine Legale, Losanna, Switzerland • The Department of Physics, Imperial College, London, UK • Dept. of Anatomy, University of Vienna, Austria We are in touch with other possible partners, besides those mentioned.

33. Owing to the complexity of the subjects and the wide-ranging research proposed in the programmes, an European-wide network is needed to reach the critical mass and the multidisciplinary skills required. Moreover, an European co-operation would ensure a faster spread of the expected scientific results and their exploitation by the sectors involved.

34. CLAM-CEO participates in a network of 58 partners that, encouraged by ELGRA (European Low-Gravity Research Association) in synergy with ESA (European Space Agency) and National Space Agencies, submitted the EoI: MIcrogravity and LIfe Sciences Network (MILIS) The network asked to the EU to consider MICROGRAVITY in relation to “Physical or Life sciences in Space research and applications” as a specific thematic since: a large scientific community exists in the domain of "Physical and Life Sciences and Applications in Space”. a new, huge, infrastructure, the International Space Station (ISS), will be available and it is important that such an investment is used effectively.

35. The Single Molecule Spectroscopy Group • Fluorescence detection and spectroscopy of single molecules in solution and on surfaces under native conditions; • Development and application of optical tweezers for single molecule and single cell studies. Measurement Systems: • Multichannel confocal microscope for spectrally and time-resolved fluorescence and fluorescence fluctuation spectroscopy measurements; • Wide-field fluorescence microscope for the direct imaging of single molecules on surfaces and in/on cells.

36. The Single Molecule Spectroscopy Group Some of the topics presently studied are: • Emission properties of single molecules at interfaces; • Absolute measurements of diffusion and concentration by fluorescence fluctuation spectroscopy; • Study of fast conformational dynamics of proteins and DNA; • Dynamics of molecular binding processes; • Mechano-dynamics of flagella and cilia; • Cellular chemotaxis; • Direct determination of the stoichiometry of molecular complexes.

37. CISM – Centre for Mass Spectrometry, University of FlorenceInterdepartmental Centre with the scope to assist public and private institutions in analytical researches, providing analytical services that require sophisticated mass spectrometry equipment. Scientific Committee: Prof. S. Turillazzi, Prof. G. Moneti, Dr. G. Pieraccini Associated with CISM are: Depts. of Animal Biology and Genetics, Agricultural Biotechnologies, Chemistry, Pharmacology, Clinical Physiopathology,Public Health and Analytical Chemistry, Earth Science and Plant Nutrition, Biochemistry, Pharmaceutical Sciences. Research activities: Neuropharmacology, entomology, environmental toxicology, clinical chemistry and diagnostics, toxicological and antidoping analyses, food and dietary analysis.

38. CISM Internet website: www.cism.unifi.it The Centre manages and utilizes the following equipments: • API 365 Sciex triple quadrupole( with APCI, ESI and mESI interfaces); • GC-MS ion traps with EI and positive CI ionisation and MSn; • GC-MS quadrupole with EI ionisation; • GC-MS quadrupole with EI and CI(positive and negative)ionisation; • ESI-TOF mass spectrometer; • MALDI-TOF/TOF instrument; • LC-MSn ion trap equipped with APCI and ESI interfaces.

39. Sergio Capaccioli Department of Experimental Pathol. and Oncol., Florence Main research topics: molecular oncology apoptosis antisense strategies Giuseppe Toffoli CRO - National Cancer Institute, Aviano Main research topics: farmacogenetics antifolates tumor drug resistance Lucio Tentori Department of Neuroscience, Rome Main research topics: mechanisms of resistance to alkylating agents preclinical models to evaluate pharmacological strategies to increase the antitumor activity of methylating agents new targets for the pharmacological therapy of drug-resistant tumors

40. Annarosa Arcangeli Department of Experimental Pathology and Oncology, Florence Main research topics: biophysical aspects of neoplastic transformation role of K+ channels in cancer cells biophysical mechanisms of cancer cell survival during hypoxia Elena Monti Dept. of Functional and Structural Biology, Busto Arsizio Main Research Topics: Molecular determinants of tumor cell response to anticancer drugs New drug combinations for the therapy of malignant gliomas Development of new photosensitizing agents for tumor photodynamic therapy

41. Angelo Nicolin School of Medicine, University of Milan Main research topics: • Bcl-2 modulation and drug resistance • Bcl-2 phosphorylation by the kinase mTOR • kinases modulation and bcl-2 level in human cells • Bcl-2 level and drug resistance Enrico Mini Dept. of Preclinical and Clinical Pharmacology, Univ. of Florence Main research topics: Molecular mechanisms of cancer drug resistance Antimetabolite agents and biochemical modulation of their cytotoxic effects Pharmacogenomic and pharmacogenetic determinants of therapeutic outcome gastrointestinal cancer