Patricia Soo-Ping Thong

Photodynamic-therapy-activated immune response against distant untreated tumours in recurrent angiosarcoma

192 Ir) source for a total dose of 36 Gy over six fractions. The tumours showed regression, but recurred within 1 year. Further treatment options, including radiotherapy, surgery and photodynamic therapy, were discussed. The patient subsequently consented to photodynamic therapy, which was carried out in an outpatient clinic. We used a new generation photosensitiser, Fotolon (Haemato-science GmbH, Luckenwalde, Germany), which comprises chlorin e6 and polyvinylpyrrolidone (molecular weight 12 000) in the ratio of 1:1. Unlike fi rst generation photosensitisers, which rendered patients light sensitive for up to a few weeks, Fotolon clears rapidly from the body within 24 h. 1 Fotolon was prepared freshly by dissolving 0·2 g of Fotolon in 100 mL of 0·9% saline. The preparation was administered intravenously at a dose of 2·0–5·7 mg/kg body weight over 10–20 min. 3 h later, the tumours were irradiated with laser light of wavelength 665 nm (± 3 nm) for a total light dose of 65–200 J/cm², delivered at a fl uence rate of 80–150 mW/cm². The table shows the treatment parameters used during each administration. We did not note any side-eff ects or complications during or after treatment, other than transient pain at the

Laser confocal endomicroscopy as a novel technique for fluorescence diagnostic imaging of the oral cavity.

Malignancies of the oral cavity are conventionally diagnosed by white light endoscopy, biopsy, and histopathology. However, it is often difficult to distinguish between benign and premalignant or early lesions. A laser confocal endomicroscope (LCE) offers noninvasive, in vivo surface and subsurface fluorescence imaging of tissue. We investigate the use of an LCE with a rigid probe for diagnostic imaging of the oral cavity. Fluorescein and 5-aminolevulinic acid (ALA) were used to carry out fluorescence imaging in vivo and on resected tissue samples of the oral cavity. In human subjects, ALA-induced protoporphyrin IX (PpIX) fluorescence images from the normal tongue were compared to images obtained from patients with squamous cell carcinoma (SCC) of the tongue. Using rat models, images from normal rat tongues were compared to those from carcinogen-induced models of SCC. Good structural images of the oral cavity were obtained using ALA and fluorescein, and morphological differences between normal and lesion tissue can be distinguished. The use of a pharmaceutical-grade solvent Pharmasolve enhanced the subsurface depth from which images can be obtained. Our initial results show that laser confocal fluorescence endomicroscopy has potential as a noninvasive optical imaging method for the diagnosis of oral cavity malignancies.

Targeted Therapy of Cancer Using Photodynamic Therapy in Combination with Multi-faceted Anti-Tumor Modalities

Photodynamic therapy (PDT) has emerged as one of the important therapeutic options in the management of cancer and other diseases. PDT involves a tumor-localized photosensitizer (PS), which when appropriately illuminated by visible light converts oxygen into cytotoxic reactive oxygen species (ROS), that attack key structural entities within the targeted cells, ultimately resulting in necrosis or apoptosis. Though PDT is a selective modality, it can be further enhanced by combining other targeted therapeutic strategies that include the use of synthetic peptides and nanoparticles for selective delivery of photosensitizers. Another potentially promising strategy is the application of targeted therapeutics that exploit a myriad of critical pathways involved in tumorigenesis and metastasis. Vascular disrupting agents that eradicate tumor vasculature during PDT and anti-angiogenic agents that targets specific molecular pathways and prevent the formation of new blood vessels are novel therapeutic approaches that have been shown to improve treatment outcome. In addition to the well-documented mechanisms of direct cell killing and damage to the tumor vasculature, PDT can also activate the body’s immune response against tumors. Numerous pre-clinical studies and clinical observations have demonstrated the immuno-stimulatory capability of PDT. Herein, we aim to integrate the most important findings with regard to the combination of PDT and other novel targeted therapy approaches, detailing its potential in cancer photomedicine.

Diffuse correlation spectroscopy with a fast Fourier transform-based software autocorrelator

Abstract. Diffuse correlation spectroscopy (DCS) is an emerging noninvasive technique that probes the deep tissue blood flow, by using the time-averaged intensity autocorrelation function of the fluctuating diffuse reflectance signal. We present a fast Fourier transform (FFT)-based software autocorrelator that utilizes the graphical programming language LabVIEW (National Instruments) to complete data acquisition, recording, and processing tasks. The validation and evaluation experiments were conducted on an in-house flow phantom, human forearm, and photodynamic therapy (PDT) on mouse tumors under the acquisition rate of ∼400  kHz. The software autocorrelator in general has certain advantages, such as flexibility in raw photon count data preprocessing and low cost. In addition to that, our FFT-based software autocorrelator offers smoother starting and ending plateaus when compared to a hardware correlator, which could directly benefit the fitting results without too much sacrifice in speed. We show that the blood flow index (BFI) obtained by using a software autocorrelator exhibits better linear behavior in a phantom control experiment when compared to a hardware one. The results indicate that an FFT-based software autocorrelator can be an alternative solution to the conventional hardware ones in DCS systems with considerable benefits.

INDUCTION OF VANADIUM ACCUMULATION AND NUCLEAR SEQUESTRATION CAUSING CELL SUICIDE IN HUMAN CHANG LIVER CELLS

Very little is known about the modulation of vanadium accumulation in cells, although this ultratrace element has long been seen as an essential nutrient in lower life forms, but not necessarily in humans where factors modulating cellular uptake of vanadium seem unclear. Using nuclear microscopy, which is capable of the direct evaluation of free and bound (total) elemental concentrations of single cells we show here that an NH4Cl acidification prepulse causes distinctive accumulation of vanadium (free and bound) in human Chang liver cells, concentrating particularly in the nucleus. Vanadium loaded with acidification but leaked away with realkalinization, suggests proton-dependent loading. Vanadyl(4), the oxidative state of intracellular vanadium ions, is known to be a potent source of hydroxyl free radicals (OH.). The high oxidative state of nuclei after induction of vanadyl(4) loading was shown by the redox indicator methylene blue, suggesting direct oxidative damage to nuclear DNA. Flow cytometric evaluation of cell cycle phase-specific DNA composition showed degradation of both 2N and 4N DNA phases in G1, S and G2/M cell cycle profiles to a solitary 1N DNA peak, in a dose-dependent manner, effective from micromolar vanadyl(4) levels. This trend was reproduced with microccocal nuclease digestion in a time response, supporting the notion of DNA fragmentation effects. Several other approaches confirmed fragmentation occurring in virtually all cells after 4 mM V(4) loading. Ultrastructural profiles showed various stages of autophagic autodigestion and well defined plasma membrane outlines, consistent with programmed cell death but not with necrotic cell death. Direct intranuclear oxidative damage seemed associated with the induction of mass suicide in these human Chang liver cells following vanadium loading and nuclear sequestration.

Review of Confocal Fluorescence Endomicroscopy for Cancer Detection

The cancer burden is increasing worldwide and there is a need to develop new technologies for cancer diagnosis. Confocal laser endomicroscopy (CLE) is a minimally invasive optical technique that enables in vivo confocal imaging of tissue structures. With the use of fluorescent dyes, the technique allows confocal fluorescence endomicroscopy of tissue from surface to subsurface layers. CLE has been applied to the surveillance and diagnosis of cancer in numerous clinical studies recently, and also holds potential for optical and guided biopsy procedures. The first part of this mini review is focused on the application of CLE for cancer detection and surveillance. The second part is focused on the application of CLE to imaging of the oral cavity. We have previously demonstrated the potential of CLE for diagnostic imaging of oral cavity lesions. To move toward real-time 3-D imaging, we interfaced an endomicroscope to an embedded computing system. The prototype system is capable of automated image acquisition and real-time volume rendering. Rendering results provide topographical and depth information. Our aim is to achieve a real-time 3-D fluorescence imaging system that can be used for diagnostic imaging and guided biopsy procedures of oral cavity lesions in a clinical setting.

The use of diffuse optical spectroscopy and diffuse correlation spectroscopy system for monitoring of tumor response to photodynamic therapy

Photodynamic therapy (PDT) of cancer works via direct cytotoxicity, causing damage to tumor vasculature and stimulating the body’s anti-tumor immune response. PDT outcome depends on the parameters used; therefore an in vivo tumor response monitoring system is useful for optimization of the treatment protocol. The combined use of diffuse optical spectroscopy and diffuse correlation spectroscopy allows us to measure the tissue oxygen saturation (StO2) and relative blood flow (rBF) in tumors. These parameters were measured before and after PDT in mouse tumor models and were calculated as ratios relative to the baseline in each tumor (rStO2 and rBF). Readings were also measured in drugonly control tumors. In responders (mice with tumor eradication), significant PDT-induced decreases in both rStO2 and rBF levels were observed at 3h post-PDT. The rStO2 and rBF readings in these mice remained low until 48h post-PDT, with recovery of these parameters to baseline values observed 2 weeks after PDT. In non-responders (mice with partial or no response), the rStO2 and rBF levels decreased less sharply at 3h post-PDT, and the rBF values returned toward baseline values at 48h post-PDT. By comparison, the rStO2 and rBF readings in drug-only control tumors showed only fluctuations about the baseline values. Thus tumor response can be predicted as early as 3h post-PDT. Recovery or sustained decreases in rStO2 and rBF up till 48h post-PDT were correlated to long-term tumor control. Diffuse optical measurements can thus facilitate early assessment of tumor response to PDT to aid in treatment planning.

MICROPIXE INVESTIGATIONS: TIME SEQUENCING STUDIES IN DEGENERATIVE DISEASES

The simultaneously applied techniques of Particle Induced X-ray Emission (PIXE), Rutherford Backscattering Spectrometry (RBS) and Scanning Transmission Ion Microscopy (STIM), have been successful in mapping and quantifying trace elements during the progression of several human diseases, in particular those degenerative diseases which have a corresponding animal model. In atherosclerosis, iron has been shown to be present in increased concentrations at the early stage of lesion formation, and when the animal model has been kept anaemic, the artery wall shows a reduced uptake of iron and a delay in lesion formation compared with controls. In Parkinsons disease, there is also an increased concentration of iron in the substantia nigra region of the brain. Although the increase in bulk iron appears to lag behind the dopaminergic cell death, we have detected an increase in localized deposits of iron at the onset of cell death. These two results infer that iron may play a role in both diseases, perhaps through the mediation of free radicals. The induction of epilepsy through the injection of kainic acid has shown that the cell death is accompanied by an increase in calcium levels as early as one day after injection. The increase in calcium is consistent with activation of phospholipase A2 and free radical damage.