Radon-222 is a naturally occurring radioactive gas that is responsible for approximately half of the human annual background radiation exposure globally. Chronic exposure to radon and its decay ...products is estimated to be the second leading cause of lung cancer behind smoking, and links to other forms of neoplasms have been postulated. Ionizing radiation emitted during the radioactive decay of radon and its progeny can induce a variety of cytogenetic effects that can be biologically damaging and result in an increased risk of carcinogenesis. Suggested effects produced as a result of alpha particle exposure from radon include mutations, chromosome aberrations, generation of reactive oxygen species, modification of the cell cycle, up or down regulation of cytokines and the increased production of proteins associated with cell-cycle regulation and carcinogenesis. A number of potential biomarkers of exposure, including translocations at codon 249 of TP53 in addition to HPRT mutations, have been suggested although, in conclusion, the evidence for such hotspots is insufficient. There is also substantial evidence of bystander effects, which may provide complications when calculating risk estimates as a result of exposure, particularly at low doses where cellular responses often appear to deviate from the linear, no-threshold hypothesis. At low doses, effects may also be dependent on cellular conditions as opposed to dose. The cellular and molecular carcinogenic effects of radon exposure have been observed to be both numerous and complex and the elevated chronic exposure of man may therefore pose a significant public health risk that may extend beyond the association with lung carcinogenesis.
(1) Background: The protoporphyrin IX (PpIX)-mediated fluorescence-guided resection and interoperative photodynamic therapy (PDT) of remaining cells may be effective adjuvants to the resection of ...glioma. Both processes may be enhanced by increasing intracellular PpIX concentrations, which can be achieved through iron chelation. AP2-18 is a novel combinational drug, which ester-links a PpIX precursor (aminolaevulinic acid; ALA) to an iron-chelating agent (CP94). (2) Methods: Human glioma U-87 MG cells were cultured in 96-well plates for 24 h and incubated for 3 or 6 h with various test compound combinations: ALA (±) CP94, methyl aminolevulinate (MAL) (±) CP94 and AP2-18. PpIX fluorescence was measured at 0, 3 or 6 h with a Bio-tek Synergy HT plate reader, as well as immediately after irradiation with a 635 nm red light (Aktilite CL16 LED array), representing the PDT procedure. Cell viability post-irradiation was assessed using the neutral red assay. (3) Results: AP2-18 significantly increased PpIX fluorescence compared to all other test compounds. All treatment protocols effectively achieved PDT-induced cytotoxicity, with no significant difference between test compound combinations. (4) Conclusions: AP2-18 has potential to improve the efficacy of fluorescence-guided resection either with or without the subsequent intraoperative PDT of glioma. Future work should feature a more complex in vitro model of the glioma microenvironment.
Photodynamic therapy (PDT) is a light activated drug therapy that can be used to treat a number of dermatological cancers and precancers. Improvement of efficacy is required to widen its application. ...Clinical protoporphyrin IX (PpIX) fluorescence data were obtained using a pre-validated, non-invasive imaging system during routine methyl aminolevulinate (MAL)-PDT treatment of 172 patients with licensed dermatological indications (37.2% actinic keratosis, 27.3% superficial basal cell carcinoma and 35.5% Bowen's disease). Linear and logistic regressions were employed to model any relationships between variables that may have affected PpIX accumulation and/or PpIX photobleaching during irradiation and thus clinical outcome at three months. Patient age was found to be associated with lower PpIX accumulation/photobleaching, however only a reduction in PpIX photobleaching appeared to consistently adversely affect treatment efficacy. Clinical clearance was reduced in lesions located on the limbs, hands and feet with lower PpIX accumulation and subsequent photobleaching adversely affecting the outcome achieved. If air cooling pain relief was employed during light irradiation, PpIX photobleaching was lower and this resulted in an approximate three-fold reduction in the likelihood of achieving clinical clearance. PpIX photobleaching during the first treatment was concluded to be an excellent predictor of clinical outcome across all lesion types.
Photodynamic therapy (PDT) with the pro‐drugs 5‐aminolevulinic acid (ALA) or methyl aminolevulinate (MAL) utilizes the combined interaction of a photosensitizer, light and molecular oxygen to ablate ...tumor tissue. To potentially increase accumulation of the photosensitizer, protoporphyrin IX (PpIX), within tumor cells an iron chelator can be employed. This study analyzed the effects of ALA/MAL‐induced PDT combined with the iron chelator 1, 2‐diethyl‐3‐hydroxypyridin‐4‐one hydrochloride (CP94) on the accumulation of PpIX in human glioma cells in vitro. Cells were incubated for 0, 3 and 6 h with various concentrations of ALA/MAL with or without CP94 and the resulting accumulations of PpIX, which naturally fluoresces, were quantified prior to and following light irradiation. In addition, counts of viable cells were recorded. The use of CP94 in combination with ALA/MAL produced significant enhancements of PpIX fluorescence in human glioma cells. At the highest concentrations of each prodrug, CP94 enhanced PpIX fluorescence significantly at 3 h for ALA and by more than 50% at 6 h for MAL. Cells subsequently treated with ALA/MAL‐induced PDT in combination with CP94 produced the greatest cytotoxicity. It is therefore concluded that with further study CP94 may be a useful adjuvant to photodiagnosis and/or PpIX‐induced PDT treatment of glioma.
Abstract
Background:
Protoporphyrin IX (PpIX)-induced photodynamic therapy (PDT) is being utilised as a topical method of localised ablation of certain non-melanoma skin cancers and precancers. ...Standardised protocols have been implemented to good effect when the disease remains superficial but improvement is required to treat thicker or acrally located conditions. Concurrent administration of an iron chelator during PpIX-PDT has been demonstrated to increase cellular accumulation of PpIX by reducing its bioconversion to haem (an iron dependent process) thus increasing cell kill on subsequent irradiation. Iron however, can also play a role in reactive oxygen species (ROS) generation and limiting its availability via chemical chelation could theoretically reduce the efficacy of PpIX-PDT, so that a response less than that maximally feasible is produced.
Materials and methods:
The effects of iron availability and chelation on PpIX-PDT have therefore been investigated via fluorescence quantification of PpIX accumulation, single-cell gel electrophoresis (comet assay) measurement of ROS-induced DNA damage and trypan blue exclusion assessment of cell viability. Cultured human cells were utilised and incubated in standardised iron conditions with the PpIX precursor’s aminolaevulinic acid (ALA) or its methyl ester (MAL) in the presence or absence of either of the iron chelating agents desferrioxamine (DFO) or hydroxypyridinone (CP94), or alternatively iron sulphate as a source of iron.
Results:
ALA or MAL incubation was found to significantly increase cellular PpIX accumulation pre-irradiation as anticipated and this observation correlated with both significantly increased DNA damage and reduced cellular viability following irradiation. Co-incubation with either of the iron chelators investigated (DFO or CP94) significantly increased pre-irradiation PpIX accumulation as well as DNA damage and cell death on irradiation indicating the positive effect of iron chelation on the effectiveness of PpIX-induced PDT. The opposite effects were observed however, when the cells were co-incubated with iron sulphate, with significant reductions in pre-irradiation PpIX accumulation (ALA only) and DNA damage (ALA and MAL) being recorded indicating the negative effects excessive iron can have on PpIX-PDT effectiveness. Some dark toxicity produced by iron sulphate administration in non-irradiated control groups was also observed.
Conclusion:
Iron chelation and availability have therefore been observed to positively and adversely affect the PpIX-PDT process respectively and it is concluded that the effects of increased PpIX accumulation pre-irradiation produced via iron chelation outweigh any limitations reduced iron availability may have on the ability of iron to catalyse ROS generation/cascades following PpIX-induced PDT. Further investigation of iron chelation within dermatological applications where enhanced PpIX-PDT treatment effects would be beneficial is therefore warranted.
Summary Background Methyl-aminolevulinate photodynamic therapy (MAL-PDT) is a successful treatment for non-melanoma skin cancers in the UK. Monitoring the photobleaching of the photosensitiser, ...protoporphyrin IX (PpIX) during treatment has been demonstrated to indicate the efficacy of the treatment. This study investigated photobleaching during light irradiation. Methods A validated non-invasive fluorescence imaging system was utilised to monitor changes in PpIX fluorescence during light irradiation. Fifty patients were recruited to this study, with patients monitored before, during (forty patients at the half way stage and ten at regular intervals in the initial phase of treatment) and after light irradiation. Results Phased PpIX photobleaching was observed during light irradiation with a significantly greater change ( P < 0.001) in PpIX photobleaching during the first half of light treatment. Within the ten patients monitored periodically the phased photobleaching observed fitted a double exponential decay curve ( r2 = 0.99, P < 0.005) suggesting a rapid initial phase of reaction when the light treatment was commenced. Conclusions Photobleaching was observed to be maximal in the initial phases of treatment, however photobleaching of PpIX continued until the completion of light treatment indicating that current clinical protocols for MAL-PDT do not over-treat the lesion with light.
Methyl-aminolevulinate-based photodynamic therapy (MAL-PDT) is utilised clinically for the treatment of non-melanoma skin cancers and pre-cancers and the hydroxypyridinone iron chelator, CP94, has ...successfully been demonstrated to increase MAL-PDT efficacy in an initial clinical pilot study. However, the biochemical and photochemical processes leading to CP94-enhanced photodynamic cell death, beyond the well-documented increases in accumulation of the photosensitiser protoporphyrin IX (PpIX), have not yet been fully elucidated. This investigation demonstrated that MAL-based photodynamic cell killing of cultured human squamous carcinoma cells (A431) occurred in a predominantly necrotic manner following the generation of singlet oxygen and ROS. Augmenting MAL-based photodynamic cell killing with CP94 co-treatment resulted in increased PpIX accumulation, MitoSOX-detectable ROS generation (probably of mitochondrial origin) and necrotic cell death, but did not affect singlet oxygen generation. We also report (to our knowledge, for the first time) the detection of intracellular PpIX-generated singlet oxygen in whole cells via electron paramagnetic resonance spectroscopy in conjunction with a spin trap.
Summary Background Methyl-aminolevulinate (MAL) photodynamic therapy (PDT) involves selective accumulation of a photosensitiser, protoporphyrin IX (PpIX), primarily in tumour tissue, which in ...combination with visible light and tissue oxygen results in reactive oxygen species (ROS) production and thus cellular destruction. Methods A non-invasive fluorescence imaging system (Dyaderm, Biocam, Germany) has been employed to acquire colour (morphological) and fluorescent (physiological) images simultaneously during dermatological PDT. This system had been previously utilised for fluorescence diagnosis, however, here changes in PpIX concentration within the skin lesions and normal tissue were followed after MAL application. Measurements were also recorded from a synthetic PpIX standard. Results Results indicated that imaging distance, imaging angle, position of the region of interest and light conditions all altered the PpIX levels acquired from the synthetic PpIX standard. The imaging system was therefore adapted and a standard operating procedure developed allowing reproducible images of dermatological lesions to be acquired. Different concentrations of synthetic PpIX were analysed with the system and a linear relationship was observed between the PpIX concentration and the mean greyscale value calculated for the images acquired up to 10 μM. Conclusions The Dyaderm imaging system can now be used reproducibly with confidence to semi-quantify PpIX (within the range of 0–10 μM) within dermatological lesions using the standard operating procedure derived from this work.
Summary Background Dermatological methyl-aminolevulinate photodynamic therapy (MAL-PDT) is utilized to successfully treat dermatological conditions. This study monitored fluorescence changes ...attributed to the accumulation and destruction of the photosensitizer, protoporphyrin IX (PpIX), at several different stages during the first and second treatments of clinical dermatological MAL-PDT. Methods A commercially available, non-invasive, fluorescence imaging system (Dyaderm, Biocam, Germany) was utilized to monitor fluorescence changes during the first and second MAL-PDT treatments in seventy-five lesions. Results The clinical data indicated statistically significant increases in fluorescence within lesions following the application of MAL for both treatments ( P < 0.001 and P < 0.01 respectively) and subsequent statistically significant decreases in fluorescence within the lesions following light irradiation for both treatments ( P < 0.001 and P < 0.01 respectively) whilst normal skin fluorescence remained unaltered. Lesions receiving a second treatment accumulated and dissipated significantly less PpIX ( P < 0.05) than during the first treatment. No significant differences were noted in PpIX accumulation or dissipation during MAL-PDT when gender, age, lesion type and lesion surface area were considered. Conclusions It can therefore be concluded that PpIX fluorescence imaging can be used in real-time to assess PpIX levels during dermatological PDT. Similar observations were recorded from the three currently licensed indications indicating that the standard ‘one size fits all’ protocol currently employed appears to allow adequate PpIX accumulation, which is subsequently fully utilized during light irradiation regardless of patient age, gender or lesion surface area.
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