Patrizia Riccio

Hypericin photosensitization of tumor and metastatic cell lines of human prostate

We have investigated the photoactivating effect of hypericin on two cancer cell lines: PC-3, a prostatic adenocarcinoma non-responsive to androgen therapy and LNCaP, a lymphonodal metastasis of prostate carcinoma responsive to androgen therapy. The two cell lines are incubated for 24 h with hypericin at concentrations ranging from 0.001 to 0.3 microg/ml in cell culture medium. The cells are irradiated at 599 nm (fluence = 11 J/cm2) using a dye laser pumped by an argon laser. Hypericin exerts phototoxic effects on both cell lines, while it does not produce toxic effects in the absence of irradiation. These results suggest that photodynamic therapy (PDT) with hypericin could be an alternative approach to the treatment of prostatic tumors, and could be beneficial in tumors that are non-responsive to androgen therapy.

Elevated expression of the tyrosine phosphatase SHP-1 defines a subset of high-grade breast tumors

Objectives: Protein tyrosine phosphatases are key regulators of intracellular signaling that contribute to determining cancer cell growth, which thus makes them attractive targets for therapeutic and diagnostic agents. SHP-1 phosphotyrosine phosphatase is rarely expressed in epithelial tumor cells, but expression has been found in several breast cancer cell lines and tumors. To determine the potential significance of SHP-1 as a prognostic marker in the clinical setting, we examined SHP-1 protein expression in breast tumors. Methods: We analyzed SHP-1 expression by immunohistochemistry in a breast tissue microarray composed of 2,081 cores, either alone or in combination with known prognostic markers. Results: Our data showed that SHP-1 expression was confined to a well-defined subset of high-grade tumors characterized by unique biological parameters. SHP-1 expression correlated directly with expression of the tyrosine kinase receptor HER-2 and inversely with expression of the estrogen receptor, while it was weakly associated with Bcl-2 expression. Conclusions: Levels of SHP-1 were correlated with conventional pathologic parameters of tumor aggressiveness and were associated with reduced patient survival, suggesting that elevated expression of SHP-1 is a common molecular abnormality in a defined subset of breast tumors and might be used in routine diagnosis to identify patients with high-risk tumors.

Preliminary tests of a prototype system for optical and radionuclide imaging in small animals

We have assembled a prototype system for multimodal (radionuclide and optical) in vivo planar imaging of small animals (mice) using single photon emission radiotracers (Tc-99m) and a fluorescent marker (hematoporphyrin). Preliminary tests of the separate (optical and radionuclide) prototype imaging systems are presented, aimed at assessing their features and at determining the experimental protocol for in vivo imaging. Tests were performed on anesthetized healthy or tumor bearing mice. The gamma radiation detector is a small area (11 /spl times/ 11 mm/sup 2/) hybrid pixel detector based on the Medipix1 ASIC readout technology (64 /spl times/ 64 square pixels of 170 /spl mu/m by side), bump-bonded to a 300 /spl mu/m thick silicon detector. High spatial resolution in radioimaging (in the order of 1 mm) is achieved in vivo with a pinhole tungsten collimator (0.35 mm diameter, 90/spl deg/ acceptance angle, field of view of over 20 mm at 10 mm source distance). A future setup will use the Medipix2 hybrid detector (256 /spl times/ 256 square pixels, 55 /spl mu/m by side) bump-bonded to a 1 mm thick CdTe pixel detector. The laser-induced in vivo fluorescence imaging system comprises a pulsed light source (Nd:YAG laser, /spl lambda/=532 nm, energy/pulse = 30 mJ, pulse width = 50 ps, repetition rate = 10 Hz) used to excite the fluorescence emission (600-760 nm) of injected hematoporphyrin compound, a low sensitivity CCD camera and a commercial image analysis system. Images of normal and tumor regions are acquired by using a cut-on filter (/spl lambda/>600 nm). Digital image subtraction then enhances the tumor contrast with respect to the background. The final experimental protocol, only partly implemented here, includes independent and then combined optical/radio imaging of control mice and of a solid tumoral area (human thyroid derived anaplastic carcinoma) after injection of the radiotracer and/or of the fluorophore. In this work, the accumulation of the radionuclide in selected organs and of the fluorophore in the tumor provides the signal contrast in the two imaging modalities. Fluorescence spectroscopy of excised tissue samples is also performed to help the interpretation of fluorescence images. Results of in vivo combined imaging on tumor in mice will be shown in a next paper.

Multiple processor version of a Monte Carlo code for photon transport in turbid media

Abstract Although Monte Carlo (MC) simulations represent an accurate and flexible tool to study the photon transport in strongly scattering media with complex geometrical topologies, they are very often infeasible because of their very high computation times. Parallel computing, in principle very suitable for MC approach because it consists in the repeated application of the same calculations to unrelated and superposing events, offers a possible approach to overcome this problem. It was developed an MC multiple processor code for optical and IR photon transport which was run on the parallel processor computer CRAY-T3E (128 DEC Alpha EV5 nodes, 600 Mflops) at CINECA (Bologna, Italy). The comparison between single processor and multiple processor runs for the same tissue models shows that the parallelization reduces the computation time by a factor of about N , where N is the number of used processors. This means a computation time reduction by a factor ranging from about 10 2 (as in our case where 128 processors are available) up to about 10 3 (with the most powerful parallel computers with 1024 processors). This reduction could make feasible MC simulations till now impracticable. The scaling of the execution time of the parallel code, as a function of the values of the main input parameters, is also evaluated.

ENHANCEMENT OF ANTITUMOR DRUG CYTOTOXICITY via LASER PHOTOACTIVATION

Abstract— —We investigate the efficacy of daunomycin, some imino‐ and amino‐substituted daunomycin analogues and the disubstituted aminoanthracenedione, mitoxantrone, in photosensitizing short‐term cell kill upon irradiation in the long wavelength visible range, during incubation of Fisher rat thyroid cells with the drugs. While all compounds exhibit similar cytocidal effects on our cell line, in the absence of irradiation, administering 86 J/cm2 at wavelengths either coincident or close to drug absorption peaks causes greater enhancement in cell mortality for the 4‐demethoxydaunomycin analogues than either the parent drug or its 5‐imino‐derivative. A lower enhancement is observed with mitoxantrone. In particular, C50 doses (i.e. concentrations that would kill 50% cells) as low as ∼10−9 M are found for both 6‐ and 11‐amino 4‐demethoxydaunomycin, compared with the values obtained in the absence of light, which are 2.59 × 10−4 and 0.43 × 10−4 M, respectively. Our previous studies of the photophysical and photochemical properties of the excited states of these drugs, and ESR and spin trapping studies of photosensitized generation of singlet oxygen, which were extended in this work to include mitoxantrone, indicate that the cytocidal effects proceed via type I rather than type II mechanisms.

Experimental study on in vivo optical and radionuclide imaging in small animals

We report on tests of a combined fluorescence and radionuclide planar imaging system for in vivo investigation on small animals. Combined images of anaesthetized mice bearing a surface solid tumor are presented. The fluorescent marker is a hematoporphyrin compound laser-excited with green light and imaged in the red fluorescence emission with a standard monochrome charge coupled device (CCD) camera. The gamma-ray (/sup 99m/Tc) pinhole imaging system uses a semiconductor pixel detector obtained by hybridizing a Silicon (300-/spl mu/m thick) or a CdTe (1-mm thick) pixel detector to the Medipix2 (55-/spl mu/m pitch) readout integrated circuit for single photon counting. The acquisition of combined images of the tumor area (fluorescence: animal top view; radionuclide: bottom view) shows that the tumor area can be imaged in a few minutes, with a few millimeter resolution (1-mm pinhole diameter), radioactively (/sup 99m/Tc MIBI, 74 MBq), and with the optical system. Combined imaging revealed also a different uptake of the two types of tumors studied (one grown from anaplastic human thyroid carcinoma-derived cells, the other from human papillary carcinoma-derived cells). Future progress will be toward a more compact optical setup and the use of a thicker CdTe detector.

Toward a Medipix2 coded aperture gamma microscope

We report on tests of a radionuclide imaging system for in vivo investigations in small animals with low energy gamma-rays as from /sup 125/I (27-35 keV). The system imaging optics features a high resolution coded aperture mask and a fine pitch silicon hybrid pixel detector of the Medipix2 series (55 /spl mu/m pitch). The coded aperture (no-two-holes-touching MURA 62/spl times/62) had 70 /spl mu/m holes in 75 /spl mu/m tungsten, and was used in a 2:1 magnification for a field of view of about 7 mm. Laboratory tests with a /sup 109/Cd 22-keV source and in vivo mouse thyroid imaging tests with /sup 125/I show a system resolution of about 110 /spl mu/m. This low energy, semiconductor-based, compact gamma camera is the basic imaging unit of a small animal single photon emission computed tomography system with deep sub-millimeter resolution.

Photon time-of-flight distributions through turbid media directly measured with single-photon avalanche diodes

We present measurements of photon time-of-flight distributions for a 9-ps, 532-nm laser pulse traveling through Intralipid suspensions and compare the measurements with the results of Monte Carlo simulations that yield the corresponding temporal point-spread function. We show that to obtain satisfactory agreement of experiments and simulation results, one must assume a quadratic dependence of the scattering coefficient on the Intralipid concentration.

MS‐2 fibrosarcoma characterization by laser induced autofluorescence

MS‐2 fibrosarcoma implanted in BALB‐CDF1 mice was investigated by frequency and time domain measurements of the autofluorescence (AF) radiation emitted upon excitation by a N2 laser beam (337.1 nm).

Multimodal system for in vivo tumor imaging in mice.

We devised a multimodal planar imaging system for in vivo mouse imaging, employing four modalities: optical imaging, green and red fluorescence reflectance imaging, radionuclide imaging and X-ray radiography. We are testing separately, and then in a combined way, each detection mode, via in vivo mouse imaging, with the final purpose of identifying small implanted tumor masses, of providing early tumor detection and following metastatic dissemination. We describe the multimodal system and summarize its main performance, as assessed in our research work in the various stages of the development, in fluorescence and radionuclide tests on healthy or tumor bearing mice. For gamma-ray detection we used a semiconductor pixel detector (Medipix1 or Medipix2) that works in single photon counting. Laser-induced fluorescence reflectance imaging was performed in vivo using a pulsed light source to excite the fluorescence emission of injected hematoporphyrin (HP) compound, a CCD camera, a low pass filter and a commercial image analysis system. The bimodal system was used for the acquisition of combined images of the tumor area (fluorescence: animal top view; radionuclide: bottom view). It was shown that the tumor area can be imaged in a few minutes, with a few millimeter resolution (1 mm pinhole diameter), radioactively (99mTc radiotracer), and with the fluorescence system and that, in one case, only one of the two modalities is able to recognize the tumor. A phantom study for thyroid imaging with 125I source embedded in a simulated tissue indicated a spatial resolution of 1.25 mm FWHM with a 1 mm pinhole.