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5-ALA is a precursor substance of tetrapyrrole biomolecule synthase, which is involved in the synthesis of porphyrins and is closely related to the synthesis of heme and the absorption and utilization of iron in animals, and has the effect of improving the antioxidant capacity of cells and the immune function of body. 5-ALA is involved in synthesis of chlorophyll in plants, which can affect the photosensitive biosynthesis system and enhance fruit coloring. 5-ALA is a precursor to the production of the photosensitizer protoporphyrin IX (PpIX), which can be produced by heme biosynthesis and is used clinically as a photosensitizer for light detection and cancer photodynamic therapy (PDT). In the paper, the biological function of 5-ALA and its application in animals are reviewed, in order to provide reference for the further application of 5-ALA in livestock and poultry breeding. [ABSTRACT FROM AUTHOR]

Mallory-Denk-bodies (MDBs) are hepatic protein aggregates associated with inflammation both clinically and in MDB-inducing models. Similar protein aggregation in neurodegenerative diseases also triggers inflammation and NF-κB activation. However, the precise mechanism that links protein aggregation to NF-κB-activation and inflammatory response remains unclear. Herein we find that treating primary hepatocytes with MDB-inducing agents (N-methylprotoporphyrin (NMPP), protoporphyrin IX (PPIX), or Zinc-protoporphyrin IX (ZnPP)) elicited an IκBα-loss with consequent NF-κB activation. Four known mechanisms of IκBα-loss i.e. the canonical ubiquitin-dependent proteasomal degradation (UPD), autophagic-lysosomal degradation, calpain degradation and translational inhibition, were all probed and excluded. Immunofluorescence analyses of ZnPP-treated cells coupled with 8 M urea/CHAPS-extraction revealed that this IκBα-loss was due to its sequestration along with IκBβ into insoluble aggregates, thereby releasing NF-κB. Through affinity pulldown, proximity biotinylation by antibody recognition, and other proteomic analyses, we verified that NF-κB subunit p65, which stably interacts with IκBα under normal conditions, no longer binds to it upon ZnPP-treatment. Additionally, we identified 10 proteins that interact with IκBα under baseline conditions, aggregate upon ZnPP-treatment, and maintain the interaction with IκBα after ZnPP-treatment, either by cosequestering into insoluble aggregates or through a different mechanism. Of these 10 proteins, the nucleoporins Nup153 and Nup358/RanBP2 were identified through RNA-interference, as mediators of IκBα-nuclear import. The concurrent aggregation of IκBα, NUP153, and RanBP2 upon ZnPP-treatment, synergistically precluded the nuclear entry of IκBα and its consequent binding and termination of NF-κB activation. This novel mechanism may account for the protein aggregate-induced inflammation observed in liver diseases, thus identifying novel targets for therapeutic intervention. Because of inherent commonalities this MDB cell model is a bona fide protoporphyric model, making these findings equally relevant to the liver inflammation associated with clinical protoporphyria.

Liquid biopsy is a minimally invasive alternative to surgical biopsy, encompassing different analytes including extracellular vesicles (EVs), circulating tumour cells (CTCs), circulating tumour DNA (ctDNA), proteins, and metabolites. EVs are released by virtually all cells, but at a higher rate by faster cycling, malignant cells. They encapsulate cargo native to the originating cell and can thus provide a window into the tumour landscape. EVs are often analysed in bulk which hinders the analysis of rare, tumour‐specific EV subpopulations from the large host EV background. Here, we fractionated EV subpopulations in vitro and in vivo and characterized their phenotype and generic cargo. We used 5‐aminolevulinic acid (5‐ALA) to induce release of endogenously fluorescent tumour‐specific EVs (EVPpIX). Analysis of five different subpopulations (EVPpIX, EVCD63, EVCD9, EVEGFR, EVCFDA) from glioblastoma (GBM) cell lines revealed unique transcriptome profiles, with the EVPpIX transcriptome demonstrating closer alignment to tumorigenic processes over the other subpopulations. Similarly, isolation of tumour‐specific EVs from GBM patient plasma showed enrichment in GBM‐associated genes, when compared to bulk EVs from plasma. We propose that fractionation of EV populations facilitates detection and isolation of tumour‐specific EVs for disease monitoring. [ABSTRACT FROM AUTHOR]

Photodynamic therapy (PDT) is a potential option for treatment of cancer since it can be performed non- invasive for superficial cancers or minimal-invasive with low traumatization. But PDT is intrinsically inefficient due to the complex accumulation of the photosensitizing drug inside the tumor and the processes of heme syntheses to create the needed cell killing components. To optimize the outcome of PDT and increase acceptance as viable option it is necessary to predict the optimal time for the start of the treatment based on measurable precursors. A former cell study proposed a new filter fluorometer in a complex and sensitive setup. In this work we now designed and tested a simplified system that can be used in combination with standard endoscopic imaging systems. This system will be used as base to prove viability of this approach for a future clinical study.

Changes in morphological and optical parameters of HeLa cells preincubated with 5-aminolevulinic acid (5-ALA), after photodynamic treatment (PDTr) with different intensity of excitation radiation are studied by digital holographic microscopy. Optimization of the algorithm of cell segmentation on their phase image is performed. The developed algorithm allows for automatic data acquisition on extended amounts of cells in a sample (over 600). The morphological and optical parameters of cells at different doses of PDTr were determined, and the lognormal distribution of average phase shift of the cells was demonstrated. Changes in the cell parameters at different radiation doses which initiate programmed cell death and necrosis were recorded. [ABSTRACT FROM AUTHOR]

Photobiomodulation (PBM) represents a promising and powerful approach for non-invasive therapeutic interventions. This emerging field of research has gained a considerable attention due to its potential for multiple disciplines, including medicine, neuroscience, and sports medicine. While PBM has shown the ability to stimulate various cellular processes in numerous medical applications, the fine-tuning of treatment parameters, such as wavelength, irradiance, treatment duration, and illumination geometry, remains an ongoing challenge. Furthermore, additional research is necessary to unveil the specific mechanisms of action and establish standardized protocols for diverse clinical applications. Given the widely accepted understanding that mitochondria play a pivotal role in the PBM mechanisms, our study delves into a multitude of PBM illumination parameters while assessing the PBM's effects on the basis of endpoints reflecting the mitochondrial metabolism of human cardiac myocytes (HCM), that are known for their high mitochondrial density. These endpoints include: i) the endogenous production of protoporphyrin IX (PpIX), ii) changes in mitochondrial potential monitored by Rhodamine 123 (Rhod 123), iii) changes in the HCM's oxygen consumption, iv) the fluorescence lifetime of Rhod 123 in mitochondria, and v) alterations of the mitochondrial morphology. The good correlation observed between these different methods to assess PBM effects underscores that monitoring the endogenous PpIX production offers interesting indirect insights into the mitochondrial metabolic activity. This conclusion is important since many approved therapeutics and cancer detection approaches are based on the use of PpIX. Finally, this correlation strongly suggests that the PBM effects mentioned above have a common "fundamental" mechanistic origin. • Photobiomodulation (PBM) increases the production of protoporphyrin IX (PpIX). • Measuring the PpIX production indirectly reflects PBM effects in the mitochondria. • An increase of metabolic activity is only observed with specific PBM conditions. • We found optimal PBM radiometric/spectral parameters maximizing metabolic activity. • Our study suggests that different PBM effects have a common mechanistic origin. [ABSTRACT FROM AUTHOR]

5‐Aminolevulinic acid‐based photodynamic therapy (ALA‐PDT) comprises photochemical and biological reactions, finally generating tumor destruction. Tumor selectivity of ALA‐PDT relies on the disturbed heme synthesis pathway in neoplastic cells. Tumor pretreatment with ALA induces two reactions: the first, self‐activation of the key enzyme porphobilinogen synthase and then protoporphyrin (PpIX) synthesis at low Fe++ availability. We emphasize that the toxic mechanism of ALA‐PDT triggered by 1O2 targets intracellular organelle destruction death pathways. PpIX fluorescence of tumors, particularly in glioblastoma, is used clinically for fluorescence guidance during neurosurgery. Targeting residual and metastatic cancer stem cells is a crucial challenge of ALA‐PDT, plausibly combined with other therapeutic modalities to encounter metastatic recurrences. We overview the different aspects of ALA‐PDT mechanism and future research and prospects. [ABSTRACT FROM AUTHOR]

5-ALA photodynamic therapy (5-ALA-PDT) could be a novel approach to treat glioblastoma. However, we observed an in vitro resistance of human glioblastoma cells to 5-ALA-PDT. ABCG2, a transporter responsible of the efflux of the photosensitizer protoporphyrine IX (PpIX) could be implied in this resistance. We aimed here to correlate PpIX accumulation and ABCG2 transporter expression with the efficacy of in vitro PDT on glioblastoma. For each of 6 cell lines (4 glioblastomas, 2 controls), assessment of: cell death after PDT (Cell viability); PpIX metabolization and efflux (fluorometry); ABCG2 transporter transcriptomic expression (RTqPCR) and proteinic expression (Western-blot; flow cytometry); PpIX and ABCG2 transporter localization (Confocal microscopy). We have observed different time and dose dependent PDT efficacy and PpIX metabolization regarding the different cell lines studied. There is a correlation between intracellular PpIX, its efflux, and PDT efficacy. Our preliminary results need to be correlated with ABCG2 expression. [ABSTRACT FROM AUTHOR]

Low-power fluorescence microscopy of 5-ALA-induced PpIX has emerged as a valuable intraoperative imaging technology for improving the resection of malignant gliomas. However, current fluorescence imaging tools are not highly sensitive nor quantitative, which limits their effectiveness for optimizing operative decisions near the surgical margins of gliomas, in particular non-enhancing low-grade gliomas. Intraoperative high-resolution optical-sectioning microscopy can potentially serve as a valuable complement to low-power fluorescence microscopy by providing reproducible quantification of tumor parameters at the infiltrative margins of diffuse gliomas. In this forward-looking perspective article, we provide a brief discussion of recent technical advancements, pilot clinical studies, and our vision of the future adoption of handheld optical-sectioning microscopy at the final stages of glioma surgeries to enhance the extent of resection. We list a number of challenges for clinical acceptance, as well as potential strategies to overcome such obstacles for the surgical implementation of these in vivo microscopy techniques.

Objective: 5-Aminolevulinic acid (5-ALA)–guided resection of gliomas in adults enables better delineation between tumor and normal brain, allowing improved resection and improved patients' outcome. Recently, several reports were published regarding 5-ALA for resection of pediatric brain tumors. The aim of the study was to determine the intracellular fluorescence of protoporphyrin IX (PPIX) in pediatric brain tumors by hyperspectral imaging and to compare it with visually observed intraoperative fluorescence. Methods: 5-ALA was administered orally 4 h prior to surgery. During tumor resection, the surgeon assessed the fluorescence signal to be strong, weak, or absent. Subsequently, fluorescence intensity of tumor samples was measured via spectroscopy. In addition, clinical data, imaging, and laboratory data were analyzed. Results: Eleven children (1–16 years) were operated. Tumor entities included three (n = 3) medulloblastomas, two (n = 2) pilocytic astrocytomas (PA), two (n = 2) anaplastic ependymomas and one (n = 1) diffuse astrocytoma, anaplastic astrocytoma (n = 1), pilomyxoid astrocytoma (n = 1) and anaplastic pleomorphic xanthoastrocytoma (n = 1). Strong fluorescence was visible in all anaplastic tumors and one PA; one PA demonstrated weak fluorescence. Visible fluorescence was strongly associated with intracellular fluorescence intensity and PPIX concentration (P < 0.05). Within all tumors with visible fluorescence, the intracellular PPIX concentration was greater than 4 μg/ml. Except for moderate and transient elevation of liver enzymes, no 5-ALA related adverse events were reported. Conclusion: We demonstrate a strong association between intraoperative observations and spectrometric measurements of PPIX fluorescence in tumor tissue. As in former studies, fluorescence signal was more commonly observed in malignant glial tumors. Further prospective controlled trials should be conducted to investigate the feasibility of 5-ALA-guided resection of pediatric brain tumors. [ABSTRACT FROM AUTHOR]

Low-power fluorescence microscopy of 5-ALA-induced PpIX has emerged as a valuable intraoperative imaging technology for improving the resection of malignant gliomas. However, current fluorescence imaging tools are not highly sensitive nor quantitative, which limits their effectiveness for optimizing operative decisions near the surgical margins of gliomas, in particular non-enhancing low-grade gliomas. Intraoperative high-resolution optical-sectioning microscopy can potentially serve as a valuable complement to low-power fluorescence microscopy by providing reproducible quantification of tumor parameters at the infiltrative margins of diffuse gliomas. In this forward-looking perspective article, we provide a brief discussion of recent technical advancements, pilot clinical studies, and our vision of the future adoption of handheld optical-sectioning microscopy at the final stages of glioma surgeries to enhance the extent of resection. We list a number of challenges for clinical acceptance, as well as potential strategies to overcome such obstacles for the surgical implementation of these in vivo microscopy techniques. [ABSTRACT FROM AUTHOR]

Highlights • Photodynamic therapy (PDT) is a modality with promising results for the treatment of various cancers. • At different stages of delivery, PDT requires imaging to plan, evaluate and monitor treatment. • MRI is a good estimator of the PDT lesional effects. • Effect of fluence rate and fractionating schemes during PDT delivery can be assessed by MRI imaging. Abstract Background There is currently no therapy that prevents high-grade glioma recurrence. Thus, these primary brain tumors have unfavorable outcomes. Recently, 5-ALA photodynamic therapy (PDT) has been proposed to delay relapse and is highly expected to have potential synergistic effects with the current standard of care. However, PDT treatment delivery needs to be optimized by evaluating the impact of both the number of fractions and the light power used. Objectives Previous studies have reported MRI examination-based outcomes for PDT in glioblastoma. Our study aimed to compare MRI markers across different treatment schemes that use interstitial PDT in high-grade glioma in a preclinical model. Materials and methods Forty-eight "nude" rats were grafted with human U87 cells into the right putamen and subsequently submitted to interstitial PDT. The rats were randomized into six groups, including two different sham groups and four different treated groups (5 fractions at 5 mW or 30 mW and 2 fractions at 5 mW or 30 mW). After photosensitizer (PS) precursor (5-ALA) intake, an optical fiber was introduced into the tumor. Treatment effects were assessed with early high-field MRI to acquire T1 and T2 diffusion and perfusion images. Results There was no difference in the variation of the diffusion coefficient among the six groups (p = 0.0549, Kruskal-Wallis test). However, a significant difference was identified among the six groups in terms of variation in perfusion (p = 0.048, Kruskal-Wallis test), supporting a lesional effect in the treated groups. Additionally, the sham groups had significantly smaller edema volumes than were observed in the treated groups. Moreover, the 5-fraction group treated with 30 mW was associated with edema volumes that were significantly greater than those in the 5-fraction group treated with 5 mW (p = 0.019). Conclusion Based on observations of MRI data and considering treatment effects, the 5-fraction group treated at 5 mW was not significantly different from the other treated groups in terms of cell deaths, characterized by diffusion imaging, or necrosis level. However, the significantly lower level of edema observed in this group indicated that this treatment scheme had limited toxicity. [ABSTRACT FROM AUTHOR]

5-Aminolevulinic acid (5-ALA) is an amino acid derivative and a precursor of protoporphyrin IX (PpIX). The photophysical feature of PpIX is clinically used in photodynamic diagnosis (PDD) and photodynamic therapy (PDT). These clinical applications are potentially based on in vitro cell culture experiments. Thus, conducting a systematic review and meta-analysis of in vitro 5-ALA PDT experiments is meaningful and may provide opportunities to consider future perspectives in this field. We conducted a systematic literature search in PubMed to summarize the in vitro 5-ALA PDT experiments and calculated the effectiveness of 5-ALA PDT for several cancer cell types. In total, 412 articles were identified, and 77 were extracted based on our inclusion criteria. The calculated effectiveness of 5-ALA PDT was statistically analyzed, which revealed a tendency of cancer-classification-dependent sensitivity to 5-ALA PDT, and stomach cancer was significantly more sensitive to 5-ALA PDT compared with cancers of different origins. Based on our analysis, we suggest a standardized in vitro experimental protocol for 5-ALA PDT.

Introduction: 5-ALA-based fluorescence-guided surgery has been shown to be a safe and effective method to improve intraoperative visualization and resection of malignant gliomas. However, it remains ineffective in guiding the resection of lower-grade, non-enhancing, and deep-seated tumors, mainly because these tumors do not produce detectable fluorescence with conventional visualization technologies, namely, wide-field (WF) surgical microscopy.Methods: We describe some of the main factors that limit the sensitivity and accuracy of conventional WF surgical microscopy, and then provide a survey of commercial and research prototypes being developed to address these challenges, along with their principles, advantages and disadvantages, as well as the current status of clinical translation for each technology. We also provide a neurosurgical perspective on how these visualization technologies might best be implemented for guiding glioma surgeries in the future.Results: Detection of PpIX expression in low-grade gliomas and at the infiltrative margins of all gliomas has been achieved with high-sensitivity probe-based visualization techniques. Deep-tissue PpIX imaging of up to 5 mm has also been achieved using red-light illumination techniques. Spectroscopic approaches have enabled more accurate quantification of PpIX expression.Conclusion: Advancements in visualization technologies have extended the sensitivity and accuracy of conventional WF surgical microscopy. These technologies will continue to be refined to further improve the extent of resection in glioma patients using 5-ALA-induced fluorescence. [ABSTRACT FROM AUTHOR]

Background Pseudomonas aeruginosa (PA) frequently develops antibiotic-resistant characteristics, which is clinically problematic. The main reason behind the rise of antibiotic-resistant PA is the extensive use of antibiotics. Therefore, a novel technique is needed to treat PA infections. Photodynamic therapy (PDT) is thought to have the potential to be a non-antibiotic treatment for infections. 5-Aminolevulinic acid (ALA), which works as a photosensitizer after being metabolized into protoporphyrin IX (PpIX) in the heme synthetic pathway, is used for PDT. Thus far, the in vivo effectiveness of PDT using ALA against PA is unknown. Objective In this study, we investigated PDT using ALA both in vitro and in vivo . Methods and results Although PDT with ALA alone did not show a bactericidal effect on PA, PDT with both ALA and EDTA-2Na had a bactericidal effect in vitro . In in vivo experiments, wounds healed faster in PA-infected mice treated with PDT using both EDTA-2Na and ALA compared to non-PDT. Conclusion These results suggest that PDT with EDTA-2Na and ALA is a potential novel treatment option for PA-infected wounds. [ABSTRACT FROM AUTHOR]

Protoporphyrin IX (PpIX) is widely used in photodynamic diagnosis. To date, the details of molecular mechanisms underlying PpIX accumulation in malignant cells after 5-ALA administration remain unclear. The fluorescence of PpIX was studied in human glioma cells. Several cell cultures were established from glioma tumor tissue to study the differences between fluorescence-positive and fluorescence-negative human glioma tumors. The cell cultures demonstrated fluorescence profiles similar to those of source tumor tissues, which allows us to use these cultures in experimental research. Dynamics of the rates of synthesis and degradation of fluorescent protoporphyrin IX was studied in the cultures obtained. In addition, the expression of CPOX, an enzyme involved in PpIX synthesis, was evaluated. mRNA levels of heme biosynthesis enzymes were analyzed, and PpIX fluorescence proved to correlate with the CPOX protein level, whereas no such correlation was observed at the mRNA level. Fluorescence intensity decreased at low levels of the enzyme, which indicates its critical role in PpIX fluorescence. Finally, the fluorescence intensity proved to correlate with the proliferative activity. [ABSTRACT FROM AUTHOR]

Photodynamic therapy (PDT) is a modality with promising results for the treatment of various cancers. PDT is increasingly included in the standard of care for different pathologies. This therapy relies on the effects of light delivered to photosensitized cells. At different stages of delivery, PDT requires imaging to plan, evaluate and monitor treatment. The contribution of molecular imaging in this context is important and continues to increase. In this article, we review the contribution of nuclear medicine imaging in oncology to PDT for planning and therapeutic monitoring purposes. Several solutions have been proposed to plan PDT from nuclear medicine imaging. For instance, photosensitizer biodistribution has been evaluated with a radiolabeled photosensitizer or with conventional radiopharmaceuticals on positron emission tomography. The effects of PDT delivery have also been explored with specific SPECT or PET radiopharmaceuticals to evaluate the effects on cells (apoptosis, necrosis, proliferation, metabolism) or vascular damage. Finally, the synergy between photosensitizers and radiopharmaceuticals has been studied considering the Cerenkov effect to activate photosensitized cells. [ABSTRACT FROM AUTHOR]

The present study describes the development and evaluation of a novel biocompatible dendrimer-based nano drug delivery system which is readily soluble in water prepared by condensing Phloroglucinol and Succinic Acid (PGSA) and could efficiently encapsulate a well known hydrophobic photodynamic therapy (PDT) agent, protoporphyrin Ⅸ (PpⅨ). The (dark and photo) cytotoxicity of the PGSA-PpⅨ (dendrimer-drug) formulation towards Dalton Lymphoma Ascites (DLA) cancer cell lines upon visible light treatment is reported and evaluated the cytotoxic Reactive Oxygen Species (ROS) generation efficiency of the Protoporphyrin Ⅸ in free and dendrimer encapsulated forms. The in vitro toxicity demonstrated by PpⅨ loaded PGSA dendrimer nanoformulation, on DLA cells reveals that this novel PGSA nanocarrier reduces the toxic nature of PpⅨ when compared to free PpⅨ without light treatment which satisfy the aim of our study. On the other hand, treatment of DLA cells with PGSA dendrimer formulation in combination with light resulted in significant enhancement in therapeutic efficacy of the PDT agent, PpⅨ.

In this paper, we investigate the possibility of using accurate prediction models for the prediction of protoporphyrin bleaching dynamics to achieve photobleaching-insensitive methods to improve the evaluation of data in an existing clinical fluorescence-guided resection technique. To simulate the scenario, measurements were carried out in vivo on skin of healthy volunteers using a compact fiber-based fluorescence spectroscopy system. We have developed an effective method for the parameterization of sequences of bleaching spectra. We analyze convergence and decay rates with respect to initial conditions and excitation irradiance. We also discuss the consequences and the potential for bleaching-insensitive measurements and their applicability in a few examples from in vivo open brain surgery.

Surgical guidance with fluorescent tracers has seen significant adoption in the last decade1, including the ability to image tissue perfusion2,3. More recently, tissue metabolism tracers have been approved for human use, such as protoporphyrin IX (PpIX)4,5, which is induced by oral or topical administration of aminolevulinic acid (ALA)6. One of the most striking things about the ALA-PpIX system is that it is produced throughout most tissues of the body and demarcates glioma tumors in neurosurgery, skin lesions and bladder cancers. However, a less studied PpIX phenomenon in surgical guidance is its delayed fluorescence (DF) signal, which is directly related to the ambient oxygen level around it7,8. The lack of oxygen, or hypoxia, induces a strong DF component which has significant relevance to oncology, where the vast majority of tumors have microregional hypoxia present9. Hypoxia imaging for surgical guidance has never been possible, but here it is shown that PpIX emits unique signal in hypoxic tumor tissue regions, as a delayed fluorescence (DF) signal in the red to near-infrared spectrum. PpIX is endogenously synthesized by mitochondria in most tissues and the particular property of DF emission is directly related to low microenvironmental oxygen concentration10,11. Most tumors have microregional chronic and/or cycling hypoxia present as well as chaotic blood flow. In this report, we show the first demonstration of imaging hypoxia signals from PpIX in a pancreatic cancer model, exhibiting >5X contrast relative to surrounding normal oxygenated tissues. Additionally, tissue palpation amplifies the signal and provides intuitive temporal contrast based upon neoangiogenic blood flow differences. This new mechanism for contrast could easily be translated to human use as an intrinsic contrast mechanism for oncologic surgical guidance. [ABSTRACT FROM AUTHOR]

• Photosensitizers and skin properties alter the depth penetration of light. • Commonly used metrics of illuminance provide inadequate comparisons of delivered light. • Methods for estimating the effective light dose in skin photo therapy are introduced. • Considering the effective light dose at varying depths is necessary to provide adequate comparisons of photo therapy regimens. Skin-based photodynamic therapy (PDT) is used for the clinical treatment of actinic keratosis (AKs) and other skin lesions with continued expansion into the standard of care. Due to the spectral dependency of photosensitizer activation and skin optical fluence, there is a need for more accurate methods to estimate the delivered dose at depth from different PDT light sources and treatment regimens. Develop radiometric methods for calculating photosensitizer-effective fluence and dose at depth and determine differences between red-lamp, blue-lamp, and daylight-based PDT treatments. Radiometric measurements of FDA-approved PDT lamp sources, outdoor daylight, and indoor daylight were performed for clinically relevant AK treatments. The protoporphyrin IX (PpIX) equivalent irradiance, fluence, and dose for each light source were calculated from the PpIX absorption spectrum and a 7-layer skin fluence model. The effective fluence and dose at depth was estimated by combining the spectral attenuation predicted at each wavelength and depth with the source fluence at each wavelength. The red-lamp source had the highest illuminance (112,000 lumen/m2), but lowest PpIX-effective irradiance (9.6 W/m2), and highest effective fluence at depth (10.8 W/m2 at 500 µm). In contrast, the blue light source had the lowest illuminance (2300 lumen/m2), but highest PpIX effective irradiance (37.0 W/m2), and ultimately the lowest effective fluence at depth (0.18 W/cm2 at 500 µm). The daylight source had values of (outdoor | indoor) illuminance of (49,200 | 37,800 lumen/m2), effective irradiance of (19.2 | 10.7 W/m2), and effective fluence of (1.50 | 1.08 W/m2 at 500 µm). The effective fluence and dose at depth facilitated the comparison of treatment regimens, for example, calculating an equivalent dose for a 2 hr indoor daylight treatment and a 10 min red-light treatment for the 300–1000 μm depth range. The consideration of PpIX-effective fluence at varying depths is necessary to provide adequate comparisons of the delivered dose from PDT light sources. Methods for calculating radiometric fluence and delivered dose at depth were introduced, with open source MATLAB code, to help overcome the limitations of commonly used photometric and irradiance-based reporting. [ABSTRACT FROM AUTHOR]

This study evaluated the potential of coated microneedles for improved dermal delivery of 5-aminolevulinic acid (5-ALA), which naturally gets converted by cells of the tissue in to a photosensitizer called protoporphyrin IX (PPIX). Microneedle patches containing 57 microneedles were coated with 5-ALA using an in-house developed micro-precision dip coater. The coating process was optimized to achieve higher 5-ALA loading on microneedles and a high delivery efficiency into porcine cadaver skin. Using 5 dips with 25% w / v 5-ALA solution, a mass of about 350 μg of 5-ALA was coated per patch, which gave a delivery efficiency of about 90% in porcine cadaver skin. Bright-field and scanning electron microscopy established that coatings of 5-ALA on microneedles of the patch were uniform. In vivo dermal pharmacokinetics showed that delivery of just 350 μg of 5-ALA using coated microneedles led to about 3.2-fold higher PPIX formation after 4 h, as compared to topical application of 20% w /w 5-ALA in a conventional cream formulation (25 mg cream). Furthermore, with use of coated microneedles, PPIX was observed in deeper regions of the skin (~ 480 μm) as compared to topical 5-ALA cream formulation (~ 150 μm). The potential of PPIX for photodynamic therapy was tested in vivo . After light exposure (633 nm; 118 J/cm 2 ), PPIX got photosensitized, and due to higher initial amount of PPIX in the coated microneedle group, about twice the amount of PPIX was photobleached compared to topical cream application. Finally, even with a lower dose of just 1.75 mg 5-ALA, coated microneedles suppressed the growth of subcutaneous tumors by ~ 57%, while a topical cream containing 5 mg of 5-ALA did not suppress the tumor volume and led to tumor growth comparable to the untreated control group. Overall, the strategy of delivering 5-ALA using coated microneedles could be a promising approach for photodynamic therapy of skin tumors. [ABSTRACT FROM AUTHOR]

Malignant gliomas are locally invasive tumors that offer a poor prognosis. Evidence shows that complete resection of the tumor at the time of surgery confers a significant improvement in overall survival. In recent years, 5- aminolevulinic acid (ALA)-induced fluorescence has been used by neurosurgeons to good effect in increasing the rate of complete resection. Despite the considerable interest in the use of 5-ALA in fluorescence-guided neurosurgery, the mechanisms behind the accumulation of Protoporphyrin IX (PpIX) in neoplastic tissue are unclear. In this review, we summarize the evidence in the literature on the mechanisms underlying the selective production of PpIX with a specific focus on gliomas. [ABSTRACT FROM AUTHOR]

BACKGROUND Coaxial electrospray is an emerging technology to produce multilayer micro- and nano-particles ( NPs) with well controlled shape and size. ALA-PDT is a fascinating technology used in the treatment for skin and other cancers. In this work, ALA-loaded poly(lactide-co-glycolide) ( PLGA) ( PLGA-ALA) NPs with high drug loading efficiency were produced via coaxial electrospray and used for the PDT treatment in HSC-3 cells. RESULTS Results showed that 200-1000 nm PLGA-ALA NPs with 60-75% ALA loading efficiency were successfully produced. ALA and PLGA were well integrated and no extra chemical reaction occurred during the electrospray. PLGA-ALA NPs displayed a delayed release of ALA in PBS solution up to 7 days. A massive accumulation of PpIX in HSC-3 cells happened after the incubation of PLGA-ALA NPs. With light irradiation, these PLGA-ALA NPs showed a remarkable cytotoxicity against HSC-3 cells, leading to 40-70% cell death depending on the incubation dose. CONCLUSIONS Coaxial electrospraying is a facile method to prepare PLGA-ALA NPs with high drug loading content and efficiency, and PLGA-ALA NPs turned out to be an effective vehicle for delivering ALA to tumor cells. © 2015 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

In this work, we developed an experimental apparatus to directly measure transmittance and fluorescence in the stratum corneum (SC) ex vivo. The SC transmittance varied from ~6 to ~52 % in the wavelength range of 280-850 nm. For 260-300 nm excitation, the SC autofluorescence showed a strong emission band between 290 and 425 nm, which is associated with tryptophan, and another in the 600-670 nm range, which we attributed to a process involving resonance energy transfer to very hydrophobic keratin filaments. Weaker emission associated with less hydrophobic keratin filaments was also observed in the wavelength range of 350-480 nm. Protoporphyrin IX (PpIX) was incorporated into SC membranes, and its penetration was further increased by the addition of nerolidol to the treatment suspension. Both PpIX and the endogenous porphyrins showed fluorescence anisotropy consistent with their localization in SC membranes, and their molecular dynamics increased significantly in the presence of 1 % nerolidol. The emission and excitation spectra of PpIX and the endogenous SC porphyrins showed similar alterations during the photobleaching induced by 405-nm irradiation. This work also highlights the SC contribution to skin autofluorescence, which could be useful for fluorescence spectroscopy applications in the early diagnosis of skin diseases. [ABSTRACT FROM AUTHOR]

Background and objective 5-Aminolevulinic acid (ALA)-mediated photodynamic therapy (ALA-PDT) is effective for the treatment of condyloma and intraepithelial neoplasia associated with human papillomaviruses (HPV). However, the mechanism of PDT on HPV-infected host cells is not fully understood. The aim of this in vitro study was to examine ALA-PDT induced apoptosis in a high-risk HPV E7 gene transfected cell line. Materials and methods HPV16 E7 gene obtained from HPV16 positive human cervical carcinoma was transfected into a human keratinocyte cell line. PpIX generation and ALA-PDT-induced apoptosis and mitochondrial damage were investigated in vitro . Results Fluorescence examination showed that HPV16E7 transfected cells could produce PpIX fairly quickly in the presence of 1 mM ALA. Apoptosis occurred after ALA-PDT at dose levels of 4, 8, and 12 J/cm 2 . Mitochondrial potential collapse occurred as early as 2 h post PDT. ALA-PDT induced a relatively weak and shorter caspase-9 activation but a strong and long lasting caspase-3 activation. However, mRNA levels of caspase-9 and -3 genes in cells treated with ALA-PDT were unchanged. Conclusion This in vitro study demonstrated that HPV16 E7 transfected keratinocytes could generate PpIX in the presence of ALA. Under light irradiation, ALA-PDT could induce mitochondrial-dependent apoptosis in HPV16 E7 transfected cells. [ABSTRACT FROM AUTHOR]

Objective Photodynamic therapy (PDT) has become a well-established modality for the treatment of many cancers. Photodynamic eradication of tumor cells depends on the presence of a photosensitizer, oxygen and light. However, oxygen depletion during PDT is a well known problem. Modulation of light delivery could address this issue by counteracting tumor hypoxia, thereby improving tumor cell killing. This preclinical study was designed to validate an animal model incorporating 5-aminolaevulinic acid (5-ALA)-PDT using U87 glioblastoma cells. We aimed to evaluate the effects of light modulation for inducing specific tumoral lesions in this model (i.e., necrosis or apoptosis). Materials and methods U87 glioblastoma cells were stereotactically engrafted into the brains of male fox1 rnu/rnu rats. Light delivery was studied after 5-ALA injection (100 mg/kg i.p.). 26 J of 635 nm light was interstitially delivered to U87 tumor-bearing rats at a radiant power of either 30 mW (high fluence rate) or 4.8 mW (low fluence rate). In each group, half of the population received illumination in 2 fractions with a refractory interval of 120 s, whereas the other half received continuous illumination. Results Twenty-two animals received 5-ALA-PDT, and the level of necrosis was scored. In the high-fluence-rate group, we observed a greater degree of tumor necrosis in rats receiving fractionated delivery than in rats receiving continuous illumination. Similar differences were not observed in the low-fluence-rate group, which exhibited only sparse necrosis. Higher morbidity and mortality rates were observed in the high-fluence-rate group. Conclusion We have developed a reproducible and reliable rodent model for interstitial 5-ALA PDT. We found that the effects of 5-ALA-PDT are dependent on light delivery conditions. Although the low-fluence-rate treatment was better tolerated, 5-ALA-PDT induced more necrosis using fractionated delivery at a high fluence rate. These results require confirmation with further studies involving larger populations and additional fractionation schemes. [ABSTRACT FROM AUTHOR]

Renal cell carcinoma (RCC) remains one of the greatest challenges of urological oncology and is the third leading cause of death in genitourinary cancers. Surgery may be curative when patients present with localized disease. Our previous results demonstrated the autofluorescence of blood PpIX in primary RCC mouse model and an increase in fluorescence intensity as a function of growth of the subcutaneous tumor mass. In another work, a nice correlation between the growth of the tumor mass and tissue fluorescence intensity was found. The aim of this study was to evaluate the expression profile of porphyrin biosynthesis pathway-related genes of human kidney cells. We used two kidney cell lines, one normal (HK2) and another malignant (Caki-1). Endogenous and 5-aminolevolinic acid (ALA) induced protoporphyrin IX (PpIX) HK2 and Caki-1 cells were analyzed by fluorescence spectroscopy. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to measure mRNA of those genes. Emission spectra were obtained by exciting the samples at 405 nm. For ALA untreated cells the maximum fluorescence intensity was detected at 635 nm. The mean peak area of emission spectra in both cells types increased linearly in function of cell number. Besides, basal levels of PpIX autofluorescence of each cell concentration of HK2 samples were significantly lower than those of Caki-1 samples. For ALA-treated cells the mean PpIX spectra shows PpIX emission peak at 635 nm with a shoulder at 700 nm. Analysis of PpIX fluorescence intensity ratio between tumor cells and HK2 cells showed that fluorescence intensity was, on average, 26 times greater in tumor cells than in healthy cells. qRT-PCR revealed that in Caki-1 ALA-treated cells, PEPT gene was significantly up-regulated and FECH and HO-1 genes were significantly down regulated in comparison with HK2 ALA-treated cells. In conclusion, our results demonstrate the preferential accumulation of ALA-induced PpIX in human RCC and also indicate that PEPT1, FECH and HO-1 genes are major contributors to this accumulation. [ABSTRACT FROM AUTHOR]

Introduction: Photodynamic therapy for psoriasis showed promise in the early 1990s with reports of plaque clearance following topical aminolevulinic acid - photodynamic therapy (ALA-PDT). Methods: In December 2013, we conducted a systematic search of the PubMed Medline database using the keywords 'psoriasis' and 'photodynamic therapy'. Results: Numerous clinical studies have failed to demonstrate a consistent, efficacious response to topical ALA-PDT. Furthermore, severe pain and burning sensations were repeatedly reported, many cases being intolerable for patients. Discussion: The variability in clinical response and the painful side effects have made topical ALA-PDT an unsuitable treatment option for chronic plaque psoriasis. Nonetheless, early clinical studies of other modalities such as topical hypericin and methylene blue, as well as systemic ALA and verteporfin, have shown that these photosensitizers are efficacious and much better tolerated than topical ALA. Conclusion: With the current landscape of phototherapy dominated by psoralen combined with ultraviolet A (PUVA) and narrow-band ultraviolet B (NB-UVB), an alternative light therapy utilizing the visible spectrum is certainly promising and a worthwhile endeavor to pursue. [ABSTRACT FROM AUTHOR]

Sonodynamic therapy (SDT) is a relatively new approach in the treatment of various cancers including leukemia cells. The aim of this study is to investigate the occurrence of apoptosis and autophagy after treated by protoporphyrin IX (PpIX)-mediated SDT (PpIX-SDT) on human leukemia K562 cells as well as the relationship between them. Firstly, mitochondrial-dependent apoptosis was observed through morphological observation and biochemical analysis. Meanwhile, SDT was shown to induce autophagy in K562 cells, which caused an increase in EGFP-LC3 puncta cells, a conversion of LC3 II/I, formation of acidic vesicular organelles (AVOs) and co-localization between LC3 and LAMP2 (a lysosome marker). Besides, pretreatment with autophagy inhibitor 3-MA or bafilomycin A1 was shown to provide protection against autophagy and to enhance SDT-induced apoptosis and necrosis, while the apoptosis suppressor z-VAD-fmk failed to affect formation of autophagic vacuoles or partially prevented SDT-induced cytotoxicity, which suggested that SDT-induced autophagy functioned as a survival mechanism. Additionally, this study reported apparent apoptosis and autophagy with dependence on intracellular reactive oxygen species (ROS) production. Preliminary data showed that ROS scavenger N-acetylcysteine (NAC) effectively blocked the SDT induced accumulation of ROS, reversed sono-damage, cell apoptosis and autophagy. Taken together, these data indicate that autophagy may be cytoprotective in our experimental system, and the ROS caused by PpIX-SDT treatment may play an important role in initiating apoptosis and autophagy. [ABSTRACT FROM AUTHOR]

High-grade dysplasia (HGD) in Barrett's esophagus (BE) poses increased risk for developing esophageal adenocarcinoma. To date, early detection and treatment of HGD regions are still challenging due to the sampling error from tissue biopsy and relocation error during the treatment after histopathological analysis. In this study, CP-A (metaplasia) and CP-B (HGD) cell lines were used to investigate the "seek-and-treat" potential using 5-aminolevulinic acid-induced protoporphyrin IX (PpIX). The photodynamic therapy photosensitizer then provides both a phototoxic effect and additional image contrast for automatic detection and real-time laser treatment. Complementary to our studies on automatic classification, this work focused on characterizing subcellular irradiation and the potential phototoxicity on both metaplasia and HGD. The treatment results showed that the HGD cells are less viable than metaplastic cells due to more PpIX production at earlier times. Also, due to mitochondrial localization of PpIX, a better killing effect was achieved by involving mitochondria or whole cells compared with just nucleus irradiation in the detected region. With the additional toxicity given by PpIX and potential morphological/textural differences for pattern recognition, this cellular platform serves as a platform to further investigate real-time "seek-and-treat" strategies in three-dimensional models for improving early detection and treatment of BE. [ABSTRACT FROM AUTHOR]

β-Lapachone is a phytochemotherapeutic originally isolated from Lapacho tree whose extract has been used medicinally for centuries. It is well known that NAD(P)H:quinone oxidoreductase (NQO1) activity is the principal determinant of β-Lapachone cytotoxicity. As NQO1 is overexpressed in most common carcinomas, recent investigations suggest its potential application against cancer. Photodynamic therapy (PDT) is a clinically approved and rapidly developing cancer treatment. PDT involves the administration of photosensitizer (PS) followed by local illumination with visible light of specific wavelength. In the presence of oxygen molecules, the light illumination of PS can lead to a series of photochemical reactions and consequently the generation of cytotoxic reactive oxygen species (ROS). It has been reported that β-Lapachone synergistically interacts with ionizing radiation, hyperthermia and cisplatin and that the sensitivity of cells to β-Lapachone is closely related to the activity of NQO1. So, the present study aimed to investigate the feasibility of PDT to increase the anticancer effect of β-Lapachone by up-regulating NQO1 expression on breast cancer MCF-7c3 cells. NQO1 expression was evaluated by Western blot analysis at different times after PDT using ME-ALA as PS. The cytotoxicity of the photodynamic treatment and β-Lapachone alone or in combination was determined by MTT assay and the combination index (CI)-isobologram method and the dose reduction index (DRI) analysis were used to assess the effect of drug combinations. Our studies for the first time demonstrated that the expression of NQO1 is induced 24h after photodynamic treatment. The sensitivity of cancer cells to β-Lapachone treatment increased 24h after PDT and a synergistic inhibitory effect on MCF-7c3 cells was showed. Taken together, these results lead us to conclude that the synergistic interaction between β-Lapachone and PDT in killing cells was consistent with the up-regulation of NQO1. The combination of β-Lapachone and PDT is a potentially promising modality for the treatment of cancer. [ABSTRACT FROM AUTHOR]

Highlights: [•] This is the first study to show the structural requirements for the binding of a porphyrin compound to HSP90. [•] An in silico molecular docking model and experimental approaches revealed the anti-angiogenic activities of PPIX. [•] A tetrapyrrole macrocycle and propionate chains of porphyrins coordinately interact with the ATP-binding pocket of HSP90. [•] Our results may offer key structural information on the potential development of a new class of HSP90 inhibitors. [Copyright &y& Elsevier]

Abstract: An effective G-quadruplex-based probe has been constructed for rapid and sensitive detection of Cu2+. In this probe, an anionic porphyrin, protoporphyrin IX (PPIX) served as a reference signal, which binds to G-quadruplex specifically and the fluorescence intensity increases sharply. While, in the presence of Cu2+, the G-quadruplex can catalyze the related Cu2+ insertion into the protoporphyrin, the fluorescent intensity is decreased. The fluorescence of the response ligand could be selectively quenched in the presence of Cu2+ and not interfered by other metal ions. The probe provided an effective platform for reliable detection of Cu2+ with a detection limit as low as 3.0nM, the high sensitivity was attributed to the strong metalation of PPIX with Cu2+ catalyzed by G-quadruplex (PS5.M). Linear correlations were obtained over the logarithm of copper ion concentration in the range from 8×10−9 M to 2×10−6 M (R=0.998). The G-quadruplex-based probe also could be used to detect Cu2+ in real water samples. Additionally, these striking properties endow the G-quadruplex-ligand with a great promise for analytical applications. [Copyright &y& Elsevier]

The present study describes the development and evaluation of a novel biocompatible dendrimer-based nano drug delivery system which is readily soluble in water prepared by condensing Phloroglucinol and Succinic Acid (PGSA) and could efficiently encapsulate a well known hydrophobic photodynamic therapy (PDT) agent, protoporphyrin IX (PpIX). The (dark and photo) cytotoxicity of the PGSA-PpIX (dendrimer-drug) formulation towards Dalton Lymphoma Ascites (DLA) cancer cell lines upon visible light treatment is reported and evaluated the cytotoxic Reactive Oxygen Species (ROS) generation efficiency of the Protoporphyrin IX in free and dendrimer encapsulated forms. The in vitro toxicity demonstrated by PpIX loaded PGSA dendrimer nanoformulation, on DLA cells reveals that this novel PGSA nanocarrier reduces the toxic nature of PpIX when compared to free PpIX without light treatment which satisfy the aim of our study. On the other hand, treatment of DLA cells with PGSA dendrimer formulation in combination with light resulted in significant enhancement in therapeutic efficacy of the PDT agent, PpIX. [ABSTRACT FROM AUTHOR]

Abstract: In pursuit of a single molecule with potential applications for both in vitro fluorescence microscopy and in vivo PET imaging, novel targeted 69/71Ga/68Ga-protoporphyrin IX (PPIX) probes were developed. Optical analysis, fluorescence microscopy and radiolabeling with gallium-68 were performed to confirm potential applications. The use of a targeted-PPIX ring, in conjunction with 69/71Ga or 68Ga, eliminates the need for the attachment of multiple imaging tags, allowing for either in vitro fluorescent-based evaluation or in vivo nuclear-based imaging. [Copyright &y& Elsevier]

This collection of summaries on endoscopic diagnosis of Barrett's esophagus (BE) includes the best endoscopic markers of the extent of BE; the interpretation of the diagnosis of ultra-short BE; the criteria for endoscopic grading; the sensitivity and specificity of endoscopic diagnosis; capsule and magnifying endoscopy; narrow band imaging; balloon cytology; the distinction between focal and diffuse dysplasia; the techniques for endoscopic detection of dysplasia and the grading systems; and the difficulty of interpretation of inflammatory or regenerative changes. [ABSTRACT FROM AUTHOR]

Summary: Background: Patients with localised scleroderma receiving aminolevulinic acid (ALA)/methyl aminolevulinic acid (MAL)-photodynamic therapy (PDT) were noted to show a reduction in skin tightness, suggesting that this therapy reduces skin sclerosis . Karrer and colleagues treated patients with 5-ALA-PDT once or twice weekly for 3–6 months and in all patients the therapy was reported to be highly effective for sclerotic plaques. In view of the potential benefit of PDT in reducing skin sclerosis, the following study looks at the possible clinical and histological effects of topical PDT on the mechanism of scarring, looking particularly at hypertrophic scars. Methods: Patients with long standing hypertrophic scars were treated with MAL-PDT on two occasions at week apart, and repeated for 3 sessions at 6-weekly intervals. PDT effect was studied by means of fluorescence imaging throughout the treatment and biopsies were taken prior to and 6 weeks post-treatment to observe histological changes. Results and conclusions: Six weeks following the treatment the scarred areas had significantly softened and become more flexible clinically and histologically there had been a significant increase in elastin fibres. This suggests that ALA/MAL-PDT may be a useful treatment or adjuvant therapy in the treatment of scarring. [ABSTRACT FROM AUTHOR]

Light fractionation in aminolaevulinic acid photodynamic therapy (PDT) of superficial basal cell carcinoma has been shown to enhance the therapeutic efficacy significantly. We have shown previously that this increase in efficacy is not simply due to an increase in the amount of protoporphyrin utilized during therapy. The present study investigated the spatial distribution of protoporphyrin in 32 superficial basal cell carcinomas undergoing light-fractionated PDT. Superficial fluorescence imaging performed during therapy was compared with the microscopic analysis of protoporphyrin fluorescence in biopsies acquired immediately before the second of two light fractions. The microscopic distribution of fluorescence was also compared with tumour sections immunohistochemically stained for Ki-67. Large variations in superficial and microscopic protoporphyrin fluorescence were found in both control and treated lesions. Despite these variations there was a reasonable correlation between the two techniques (R2=0.86). The mean fluorescence intensity in control biopsies was greater than in illuminated lesions before the second light fraction, but there was no significant difference in the variation within and between regions of interest in each of these sets of lesions. There was no clear trend in depth of protoporphyrin reappearance during the dark interval between light fractions. The general distribution of cells stained positive for Ki-67 followed the protoporphyrin fluorescence that was associated with islands of basal cell carcinoma. In conclusion, this study confirms that the mean relative re-synthesis of protoporphyrin after PDT is consistent with that found previously in pre-clinical models. There are large variations in fluorescence within superficial basal cell carcinoma that include regions of tumour cells that do not synthesize protoporphyrin. [ABSTRACT FROM AUTHOR]

Abstract: Titanium oxide (Ti(O)) xerogel films functionalized by protoporphyrin IX (PPIX) and ferrocene carboxylic acid (FCA) were deposited on indium tin oxide (ITO) electrodes following a sol–gel synthesis. PPIX and FCA were first complexed to titanium oxide precursors, which were then subjected to hydrolysis to obtain a homogenous Ti(O) polymeric network gel doped with PPIX and FCA. The Ti(O) film cast on the ITO electrode has been characterized by UV–Vis absorption, X-ray photoelectron spectroscopy (XPS), scanning electron microscopy and cyclic voltammetry. Illumination of the PPIX doped Ti(O) films on the ITO electrode immersed in aqueous electrolytes onsets photoinduced electron transfer reactions, and a cathodic photocurrent was observed in most cases. This photocurrent response was investigated in detail using a kinetic model. Preliminary investigations of oxygen reduction, lithium and proton insertion into the Ti(O) film have also been carried out. [Copyright &y& Elsevier]

Actinic keratosis (AK) can be treated by photodynamic therapy (PDT), which is becoming a well-established tool in dermatology. Normally a precursor of the photosensitiser is applied topically and converted into protoporphyrin IX (PPIX) in the cells. By activating PPIX with light, the dysplastic cells will be destroyed. We report the results of two clinical studies investigating the properties of a novel self-adhesive 5-ALA-patch (PD P 506 A) intended for PDT of mild to moderate AK on the face and head. The studies investigated the influence of patch application duration on PPIX-specific fluorescence and the pharmacokinetic properties of the 5-ALA patch. The PPIX fluorescence in AK lesions and normal skin after patch application (intraindividual comparison; application for 2, 3, 4, 5 h) was investigated in 13 patients using DYADERM Professional (Biocam). In the subsequent pharmacokinetic study 12 patients were treated with 8 patches each (4 h application). 5-ALA and PPIX were analysed in plasma (over 24 h) and urine (over 12 h). PPIX-specific fluorescence measured immediately after patch removal increased with increasing application duration to a maximum at 4-h application. The fluorescence in AK lesions was more intense than in normal skin. A small increase of 5-ALA plasma concentrations was observed in 10 of 12 patients after applying 8 patches for 4 h, which rapidly declined to normal values after patch removal. The maximum increase was 3.7-fold of the pre-dose 5-ALA plasma concentration. No PPIX-concentrations above the lower limit of quantification were observed. PPIX-specific fluorescence in AK lesions can be steered by application duration of this novel 5-ALA patch. Application is safe and well tolerable. The observed small rise in 5-ALA plasma concentrations is regarded clinically irrelevant. Clinical efficacy of the patch in PDT will be investigated in further clinical trials. [ABSTRACT FROM AUTHOR]

Summary: Background: PDT is a treatment modality, which uses a combination of a photosensitising drug, oxygen and light to selectively destroy pre-malignant and malignant tissue. The most commonly used PDT photosensitiser precursor is 5-aminolevulinic acid (ALA). However recently the introduction of methyl aminolevulinate (MAL) another topical photosensitiser has posed the question of which may be of greater clinical advantage when used in PDT. Aims: The characteristics of PpIX fluorescence following the application of different quantities of both ALA and MAL were investigated in normal skin. Methods: PpIX fluorescence was detected using a non-invasive spectroscopy system. A total of 17 healthy volunteers were recruited, and varying quantities of both ALA and MAL were applied to the inner forearm. Results and conclusions: Topical application (within the range of 0.01–0.09ml) of both ALA and MAL to normal human skin-induced PpIX fluorescence. This induced fluorescence was not dose dependent within the range investigated. However, the time course and intensity of fluorescence was significantly different for ALA and MAL. PpIX-induced fluorescence peaked at 7h following the application of MAL, compared to 24h following the application of ALA. At all time points, ALA-induced fluorescence was significantly greater than that induced by MAL. These results help define some of the characteristics of ALA- and MAL-induced PpIX fluorescence in normal skin. [Copyright &y& Elsevier]

Objectives: To examine the efficacy and mechanism of δ- or 5-aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT) on a human hepatocellular carcinoma cell line. Materials and methods: The optimal uptake of photosensitizer ALA in HepG2 (p53 wild) cells was investigated by means of spectrometric measurement. Cell viability was determined by trypan blue exclusion assay. Morphological apoptotic changes in HepG2 cells before and after ALA-mediated PDT were determined by microscopic examination. Detection of apoptotic bodies was examined by DAPI staining. The changes in p53 expression were revealed by the immunostaining method. Results: ALA/protoporphyrin IX (PpIX) was mainly located in the cytoplasm of HepG2 cells. The maximal cellular uptake occurred after 18 h in vitro incubation. The photocytotoxic assay showed that ALA PDT induced 80% killing at 2 mM drug dose and 2 J/cm2 light intensity. Up to 70% of cells showed membrane blebbing and positive DAPI staining, indicating that ALA–PDT-mediated cell death was predominantly via apoptosis. In addition, p53 was upregulated after treatment, implying that p53 might evoke apoptotic cell death. Conclusions: HepG2 cell line is sensitive to ALA-mediated PDT. ALA–PDT induces apoptosis in the HepG2 cell line that may be mediated by a p53-dependent pathway. [ABSTRACT FROM AUTHOR]

Background: Photodynamic therapy with topical 5-aminolevulinic acid (ALA), followed by irradiation with red light (ALA-PDT), is used for non-melanoma skin cancer and other dermatological diseases. Pain during and after light exposure is a well-known adverse advent that may be a limiting factor for treatment, particularly, in viral warts. Methods: To assess the pain induced by ALA-PDT, we asked 45 patients enrolled in a randomized, placebo-controlled trial with six consecutive ALA- and placebo-PDT treatments for recalcitrant foot and hand warts to fill in questionnaires about pain immediately and 24 h after each treatment. Results: Immediately and 24 h after each of the six treatments, pain intensity was significantly higher in warts treated with ALA-PDT than in warts treated with placebo-PDT ( P<0.028). Severe or unbearable pain was reported from a median of 17% (6–31%) of the ALA -treated warts and from a median of 2% (0–15%) from the placebo-treated warts immediately after the treatments. With increasing treatments, no significant change in pain intensity was observed and no significant relation was found between the pain intensity and the relative change in wart area. The pain was primarily characterized as burning and shooting. The pain lasted about 30 h (range: 1–96 h). Conclusion: We conclude that pain induced by ALA-PDT is of such intensity in about one-fifth of the warts that pain relief is indicated. [ABSTRACT FROM AUTHOR]

Abstract: Photodynamic therapy (PDT) is a combination of light with a lesion-localizing photosensitizer or its precursor to destroy the lesion tissue. PDT has recently become an established modality for several malignant and non-malignant conditions, but it can be further improved through a better understanding of the determinants affecting its therapeutic efficiency. In the present investigation, protoporphyrin IX (PpIX), an efficient photosensitizer either endogenously induced by 5-aminolevulinic acid (ALA) or exogenously administered, was used to correlate its subcellular localization pattern with photodynamic efficiency of human oesophageal carcinoma (KYSE-450, KYSE-70) and normal (Het-1A) cell lines. By means of fluorescence microscopy ALA-induced PpIX was initially localized in the mitochondria, whereas exogenous PpIX was mainly distributed in cell membranes. At a similar amount of cellular PpIX PDT with ALA was significantly more efficient than photodynamic treatment with exogenous PpIX at killing all the 3 cell lines. Measurements of mitochondrial membrane potential and intracellular ATP content, and electron microscopy showed that the mitochondria were initially targeted by ALA-PDT, consistent with intracellular localization pattern of ALA-induced endogenous PpIX. This indicates that subcellular localization pattern of PpIX is an important determinant for its PDT efficiency in the 3 cell lines. Our finding suggests that future new photosensitizers with mitochondrially localizing properties may be designed for effective PDT. [Copyright &y& Elsevier]

The relationship between tumor cell differentiation and photosensitizer accumulation used in PDT is poorly defined. In the present work, specific cell differentiation of colon carcinoma CT26 cells induced by sodium butyrate was manifested by morphological changes, proliferation and protein expression and was correlated with the accumulation of endogenous and exogenous photosensitizes. Reduced accumulation of the endogenous protoporphyrin IX and the exogenous hypericin and MC540 was detected in differentiated cells. In contrast, a differentiation-dependent increase was measured with TPPS4, TMPyP, the pheophorbides (C5, C6, C12), HypS4 and helianthrone. In conclusion, PpIX, Hypericin and MC540 show specific binding and accumulation in poorly differentiated tumors, giving these tumors tissue-specific advantage in photo-diagnostic PDT applications. [Copyright &y& Elsevier]