Protoporphyrin IX fluorescence photobleaching is a useful tool to predict the response of rat ovarian cancer following hexaminolevulinate photodynamic therapy.
ABSTRACT Accurate dosimetry was shown to be critical to achieve effective photodynamic therapy (PDT). This study aimed to assess the reliability of in vivo protoporphyrin IX (PpIX) fluorescence photobleaching as a predictive tool of the hexaminolevulinate PDT (HAL-PDT) response in a rat model of advanced ovarian cancer.
Intraperitoneal 10(6) NuTu 19 cells were injected in 26 female rats Fisher 344. Peritoneal carcinomatosis was obtained 26 days post-tumor induction. Four hours post-intraperitoneal HAL (Photocure ASA, Oslo, Norway) injection, a laparoscopic procedure (D-light AutoFluorescence system, Karl Storz endoscope, Tuttlingen, Germany) and a fluorescence examination were made for 22 rats. The first group (LASER group, n=26) was illuminated with laser light using a 532 nm KTP laser (Laser Quantum, Stockport, UK) on 1 cm(2) surface at 45 J/cm(2). The second group (NO LASER group, n=26) served as controls. Biopsies were taken 24 hours after PDT. Semi-quantitative histology was performed and necrosis value was determined: 0--no necrosis to 4--full necrosis. Fluorescence was monitored before and after illumination on complete responders (NV=3-4; n=20) and non-responders (NV=0-2; n=6).
High PpIX photobleaching corresponded with complete responders whereas low photobleaching corresponded with non-responders (P<0.05). A direct linear correlation was shown between photobleaching and necrosis (R(2)=0.89).
In vivo PpIX fluorescence photobleaching is useful to predict the tissue response to HAL-PDT.
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ABSTRACT: Photodynamic therapy requires the combined interaction of a photosensitiser, light and oxygen to ablate target tissue. In this study we examined the effect of iron chelation and oxygen environment manipulation on the accumulation of the clinically useful photosensitiser protoporphyrin IX (PpIX) within human squamous epithelial carcinoma cells and the subsequent ablation of these cells on irradiation. Cells were incubated at concentrations of 5%, 20% or 40% oxygen for 24h prior to and for 3h following the administration of the PpIX precursors aminolevulinic acid (ALA), methyl aminolevulinate (MAL) or hexylaminolevulinate (HAL) with or without the iron chelator 1,2-diethyl-3-hydroxypyridin-4-one hydrochloride (CP94). PpIX accumulation was monitored using a fluorescence plate reader, cells were irradiated with 37J/cm(2) red light and cell viability measured using the neutral red uptake assay. Manipulation of the oxygen environment and/or co-administration of CP94 with PpIX precursors resulted in significant changes in both PpIX accumulation and photobleaching. Incubation with 5% or 40% oxygen produced the greatest levels of PpIX and photobleaching in cells incubated with ALA/MAL. Incorporation of CP94 also resulted in significant decreases in cell viability following administration of ALA/MAL/HAL, with oxygen concentration predominantly having a significant effect in cells incubated with HAL. Experimentation with human squamous epithelial carcinoma cells has indicated that the iron chelator CP94 significantly increased PpIX accumulation induced by each PpIX congener investigated (ALA/MAL/HAL) at all oxygen concentrations employed (5%/20%/40%) resulting in increased levels of photobleaching and reduced cell viability on irradiation. Further detailed investigation of the complex relationship of PDT cytotoxicity at various oxygen concentrations is required. It is therefore concluded that iron chelation with CP94 is a simple protocol modification with which it may be much easier to enhance clinical PDT efficacy than the complex and less well understood process of oxygen manipulation.Photodiagnosis and photodynamic therapy 12/2013; 10(4):575-582.
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ABSTRACT: Common methods to characterize treatment efficacy based on morphological imaging may misrepresent outcomes and exclude effective therapies. Using a three-dimensional model of ovarian cancer, two functional treatment response metrics are used to evaluate photodynamic therapy (PDT) efficacy: total volume, calculated from viable and nonviable cells, and live volume, calculated from viable cells. The utility of these volume-based metrics is corroborated using independent reporters of photodynamic activity: viability, a common fluorescence-based ratiometric analysis, and photosensitizer photobleaching, which is characterized by a loss of fluorescence due in part to the production of reactive species during PDT. Live volume correlated with both photobleaching and viability, suggesting that it was a better reporter of PDT efficacy than total volume, which did not correlate with either metric. Based on these findings, live volume and viability are used to probe the susceptibilities of tumor populations to a range of PDT dose parameters administered using 0.25, 1, and 10 μM benzoporphyrin derivative (BPD). PDT with 0.25 μM BPD produces the most significant reduction in live volume and viability and mediates a substantial shift toward small nodules. Increasingly sophisticated bioengineered models may complement current treatment planning approaches and provide unique opportunities to critically evaluate key parameters including metrics of therapeutic response.Journal of Biomedical Optics 09/2013; 18(9):98004. · 2.88 Impact Factor
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ABSTRACT: While Photodynamic therapy (PDT) is a promising treatment for peritoneal carcinomatosis, its use is often limited because of the toxicity of photosensitizers. In this study, safety of PDT with hexaminoevulinate (HAL), a second generation photosensitizer, is assessed. PDT of the peritoneal cavity was performed in a rat model of peritoneal carcinomatosis. Rats were treated according to different protocols: with full or half HAL dose, after intraperitoneal or oral administration of HAL, 4 or 8hours after its injection, using red or green light, after protection of the liver or cooling of the abdominal wall. Toxicity was assessed by blood tests quantifying hematocrit, liver and muscular enzymes and by pathological examination of abdominal and intrathoracic organs after treatment. The results were analyzed in the light of quantification of fluorescence and protoporphyrin IX (PPIX) content of the same organs. PDT with HAL induced rhabdomyolysis, intestinal necrosis and liver function test anomalies, leading to death in 2 out of 34 rats. The liver and the intestine contained high levels of PPIX (3 to 5 times more than tumor nodules). HAL PDT lacked specificity. However, the strategy associating diagnosis, treatment and evaluation of the results in one single procedure was effective and should be tested with other photosensitizers.Photodiagnosis and photodynamic therapy 04/2014;