Endobronchial photodynamic therapy under fluorescence control: Photodynamic theranostics

BackgroundPhotodynamic therapy (PDT) has several advantages. However, one of the disadvantages is its inability to be individualized according to biological characteristics of malignant tumors. The objective of this study was to investigate a strategy for individualized endobronchial PDT in the treatment of centrally located non-small cell lung cancer.MethodsNew approach suggests taking fluorescence-based measurements of chlorine E6 photosensitizer (PS) accumulation in the malignant tumor tissue, and assess PS consumption rate during PDT. Two randomized groups of 45 patients took part in the comparative study of standard PDT procedure, 662 nm, pulse-periodic mode, therapeutic light (reference group – RG) versus the investigated individualized approach under fluorescence control after irradiation with violet light, 408 nm, diagnostic light (study group – SG). The PDT-treatment parameters and results of follow-up bronchoscopy were compared between the groups.Results43 (96%) of 45 patients in SG demonstrated intense fluorescence in the area of the tracheal/bronchial tumor stenosis. 4 (9%) of 45 patients (SG) demonstrated fluorescence of mucosa areas distant from the main tumor lesion after violet light irradiation. Mean fluence during the whole PDT procedure was 95 ± 20 J/cm² (range 60–130 J/cm²), which was significantly lower than in RG (p = 0.01). Total exposure time was significantly lower in SG (365 ± 65 s), compared with RG (690 ± 65 s), P = 0.001. According to the follow-up bronchoscopy the difference in the PDT-treatment results between the groups is statistically insignificant.ConclusionsThe investigated strategy suggests using fluorescence control of the efficacy of PDT-treatment (photodynamic theranostics) to optimize and individualize the PDT procedure.     

Photodynamic therapy using a cytotoxic photosensitizer porphyrus envelope that targets the cell membrane

BackgroundSubcellular localization of a photosensitizer is known to determine the therapeutic efficacy of photodynamic therapy (PDT). Cell membrane is an optimal target that promises an effective treatment outcome.ObjectivesWe previously developed a novel photosensitizer named porphyrus envelope (PE) by combining hemagglutinating virus of Japan envelope (HVJ-E) with lipidated protoporphyrin IX (PpIX lipid). In the current study, the cellular localization of PE and its ability to induce multiple anti-tumor effect were characterized.Materials and MethodsThe localization and uptake of PpIX lipid in cells were evaluated with confocal laser scanning microscopy and a cell-based fluorescent assay, respectively. The ability of PE to suppress the migration and proliferation of cancer cells was assessed using a scratch-wound assay. The synergistic effect of PDT and HVJ-E treatment was evaluated using an in vitro experiment with PC-3 cells.ResultsPE localized along the cell membrane and PpIX lipid accumulated selectively in the prostate cancer cells within 10 min. Also, PE maintained the ability to undergo fusion and induce cancer cell death even after light irradiation at the dose for PDT. Incubation with PE resulted in delayed migratory and proliferative activity of PC-3 cells. PE-mediated PDT was twice as effective when cells were further incubated with PE following PDT.ConclusionsPE allows rapid drug delivery targeting the cell membrane. Because the cytotoxicity of HVJ-E was maintained, synergistic effect of HVJ-E and the photochemical reactions resulted in highly effective killing of prostate cancer cells in vitro and thus represents a promising treatment for prostate cancer.     

Fluorescence analysis of a tumor model in the chorioallantoic membrane used for the evaluation of different photosensitizers for photodynamic therapy

The development of a tumor in the chicken chorioallantoic membrane (CAM) enables a more individualized understanding of the dynamics of the photosensitizer (PS) interaction with the tumor blood vessels and cells. Photogem^® and 5-aminolevulinic acid (ALA), a protoporphyrin IX (PpIX) precursor, were used as PS and their red fluorescence enabled the monitoring of PS dynamic distribution in the vessels and in the tumor. With a tumor model in CAM and fluorescence assessment, the aim of this study was to evaluate the PDT parameters comparing different photosensitezers. In this model, the topical application was chosen as the best way of drug delivery and widefield fluorescence images were at every 30 min. The images were processed in a MATLAB^® routine for a semi-quantitative analysis of the red fluorescence. PpIX formation in the blood vessels and in the tumor region was observed after 3 h and 1.5 h, respectively, whereas Photogem^® was accumulated in the tumor region after 2 h. The illumination was performed by a diode laser with emission centered at 635 nm and irradiance of 80 mW/cm² for 10 min. After PDT irradiation, the photobleaching for both compounds was observed. Photogem^® showed a reduced photobleaching, however, both PS induced a destruction of the tumor mass and vascular network in the treated area.     

A quantitative study of in vivo protoporphyrin IX fluorescence build up during occlusive treatment phases

BackgroundTopical photodynamic therapy (PDT) is a non-invasive light based therapy used to treat non-melanoma skin cancer (NMSC) and dysplasia. During PDT, the light sensitive molecule protoporphyrin IX (PpIX) is activated, resulting in the production of singlet oxygen, which subsequently leads to cell death. PpIX is metabolised from a topically applied pro-drug and the strong fluorescence signal associated with PpIX can be utilised as an indicator of the amount of PpIX present within the tumour tissue. In this work we measure the build up PpIX during the occlusive treatment phase and investigate how the PpIX production rate is affected by different lesion and patient characteristics.MethodsFluorescence measurements were used to investigate the build up of PpIX within the tumour tissue during the 3 h long occlusive treatment prior to irradiation. The study included in vivo measurements of 38 lesions from 38 individual patients. Actinic keratosis (AK) and basal cell carcinoma (BCC) were the lesion types included in this study. The resulting data from the study was analysed using generalised linear mixed effects models.ResultsIt was found that the surface fluorescence signal linearly increased with occlusive treatment time. The predictive models suggest that there is a significant difference in PpIX production between lesion location, however no significant difference is demonstrated between different lesion types, gender and skin type.ConclusionsThe study extends and supports previous knowledge of PpIX production during the occlusive treatment phase.     

Effects of photodynamic therapy with talaporfin sodium on squamous cell carcinoma and sarcoma cells

BackgroundPhotodynamic therapy (PDT) uses a photosensitizer and light to destroy abnormal cells. Talaporfin sodium (NPe6) is a second-generation photosensitizer.MethodsWe evaluated the toxic effects of different combinations of laser and NPe6 doses on squamous cell carcinoma (KLN205) and sarcoma (Meth A) cell lines. The cells were incubated with 0, 5, 10, or 30 μg/mL NPe6 for 24 h. The cells were then irradiated with 0, 5, 15, or 30 mW/cm² of laser power, and 0, 1, 5, 10, or 20 J/cm² of laser energy. Cell viability was evaluated after 24 h. We also evaluated the cytotoxic effects of continuous wave or square-wave modulated laser irradiations (2, 5, or 10 Hz, 50% duty) on Meth A cells.ResultsThe median lethal doses of NPe6 against the KLN205 and Meth A cells after treatment at a fluence rate of 15 mW/cm² and a light dose of 20 J/cm² were 18.6 and 5.0 μg/mL, respectively. Meth A cells were more sensitive to PDT than KLN205 cells. There was no significant difference between the effects of continuous wave and square-wave modulated lasers on Meth A cell viability.ConclusionsNPe6 PDT induced cell death in a dose-dependent manner in KLN205 and Meth A cells. More work is required to evaluate the cytotoxic effects of square-wave modulated laser therapy at low light doses.     

Sub-additive effects of photodynamic therapy combined with erlotinib for the treatment of epidermoid carcinoma: An in vitro study

BackgroundPhotodynamic therapy (PDT) is an antitumour treatment that employs the combination of a photosensitive compound, oxygen and visible light. To improve the antitumour activity of PDT, the present study used the strategy of combining PDT with erlotinib (ERL), a drug frequently used in the treatment of epidermoid carcinoma.MethodsAn MTT cell viability assay was used to evaluate the cytotoxicity of PDT combined with ERL on A431 epidermoid carcinoma cells in vitro. This study evaluated the cytotoxicity of the following treatments: red laser irradiation (660 nm) at different power densities (1.25–180 J/cm²), the photosensitizer methylene blue (MB) at concentrations of 0.39–100 μM, PDT (12.5 μM MB and laser power densities from 1.25 to 180 J/cm²), and PDT (12.5 μM MB and a laser density of 120 J/cm²) plus ERL (1 μM).ResultsThe laser power densities that were tested showed no cytotoxicity in A431 cells. MB showed a dose-dependent cytotoxicity. In PDT, an increase in the dose of light resulted in an increase in the cytotoxicity of MB. In addition, there was a sub-additive effect between PDT and ERL compared to the effect of each therapy alone.ConclusionsThe sub-additive effect between PDT and ERL suggests that their combination may be an important strategy in the treatment of epidermoid carcinoma.     

Phototoxicity in a laryngeal cancer cell line enhanced by a targeting amphiphilic chlorin photosensitizer

BackgroundPhotodynamic therapy (PDT) has been established in several countries as an alternative therapy for the treatment of various malignancies. This therapy involves the incorporation of a photosensitizer (PS) that is activated by visible light and form reactive oxygen species leading to target cell death by apoptosis or necrosis. Previously, our group has demonstrated that CHL-T (semi-synthesized from chlorophyll a and containing a linked solubilizing group TRISMA^®) presented a pronounced potential to induce death in HeLa cell line after PDT. In the present study, besides confirm the high cytotoxicity in another cell line, we have further investigated the cell death mechanisms caused by CHL-T as a photosensitizer in laryngeal carcinoma cells.MethodsCells were exposed to different concentrations of three photosensitizers, namely, hypericin (HY), unmodified chlorin (CHL) and a synthesized amphiphilic chlorin derivative (CHL-T). PSs accumulation and localization were accessed by fluorescence assays. Photosensitization was induced at 6 J cm⁻² using red LEDs (630 ± 10 nm). Viability was assessed by mitochondrial function (MTT); whereas apoptosis/necrosis was evaluated by fluorescence microscopy and flow cytometry. Expression of pro-apoptotic p53 protein was studied by Western blot.Results and conclusionsAll PS showed similar localization profile in the HEp-2 cells. The use of CHL-T increased the percentage of apoptotic cells and also p53 expression in comparison with the use of HY and CHL as photosensitizers. This study shows a significant effect of CHLT associated with red light (630 ± 10 nm and 18 mW cm⁻²) irradiation on a cancer cell line, indicating the potential of this amphiphilic chlorin in enhancing the therapeutic effectiveness of Photodynamic Therapy (PDT).     

Molecular analysis of apoptosis pathway after photodynamic therapy in breast cancer: Animal model study

BackgroundMolecular investigation of breast tumors has permitted better understanding about interaction of genes and pathways involved in tumor progression.ObjectiveThe aim of this study was to evaluate the association between genes belonging to the pathway of apoptosis with tumor response to photodynamic therapy.Study Design/Materials and methodsThe mammary tumors were induced in twenty-four Spraguey-Dawley female rats by oral gavage of 7,12-dimethylbenz(a)anthracene (8 mg/Kg body weight). Animals were divided into three groups: G1 (normal tissue), G2 (tumors without treatment), G3 (animals euthanized 48 h after treatment). The photosensitizer used was a chlorin, 5,15-bis-(2-bromo-5-hydroxyphenyl) chlorin in the dose of 8 mg/kg for each animal. Light source of diode laser at a wavelength of 660 nm, fluence rate of 100 mW/cm, and light dose of 100 J/cm was delivery to lesions for treatment. A sample from each animal was investigated by quantitative real time PCR using Rat Apoptosis RT² Profiler™ PCR Array platform.ResultsPro-apoptotic BAK1, CARD6, CASP8, CIDEA, CIDEB, DAPK1, TNF, TNFRSF10B, FASLG, LOC687813, and TP73 genes showed increased expression, and CD40 anti-apoptotic gene showed decreased expression in the group who underwent PDT (G3) in relation to G2.ConclusionThe results indicated that these genes are involved more directly with cellular apoptosis induced by PDT using the Chlorin photosensitizer.     

Treatment of peritoneal carcinomatosis with photodynamic therapy: Systematic review of current evidence

BackgroundPeritoneal carcinomatosis results when tumour cells implant and grow within the peritoneal cavity. Treatment and prognosis vary based on the primary cancer. Although therapy with intention-to-cure is offered to selective patients using cytoreductive surgery with chemotherapy, the prognosis remains poor for most of the patients. Photodynamic therapy (PDT) is a cancer-therapeutic modality where a photosensitiser is administered to patients and exerts a cytotoxic effect on cancer cells when excited by light of a specific wavelength. It has potential application in the treatment of peritoneal carcinomatosis.MethodsWe systematically reviewed the evidence of using PDT to treat peritoneal carcinomatosis in both animals and humans (Medline/EMBASE searched in June 2017).ResultsThree human and 25 animal studies were included. Phase I and II human trials using first-generation photosensitisers showed that applying PDT after surgical debulking in patients with peritoneal carcinomatosis is feasible with some clinical benefits. The low tumour-selectivity of the photosensitisers led to significant toxicities mainly capillary leak syndrome and bowel perforation. In animal studies, PDT improved survival by 15–300%, compared to control groups. PDT led to higher tumour necrosis values (categorical values 0–4 [4 = highest]: PDT 3.4 ± 1.0 vs. control 0.4 ± 0.6, p < 0.05) and reduced tumour size (residual tumour size is 10% of untreated controls, p < 0.001).ConclusionPDT has potential in treating peritoneal carcinomatosis, but is limited by its narrow therapeutic window and possible serious side effects. Recent improvement in tumour-selectivity and light delivery systems is promising, but further development is needed before PDT can be routinely applied for peritoneal carcinomatosis.     

In vitro assessment of anti-tumorigenic mechanisms and efficacy of NanoALA,a nanoformulation of aminolevulic acid designed for photodynamic therapy of cancer

BackgroundThe development of nanocarriers is an important approach to increase the bioavailability of hydrophilic drugs in target cells. In this work, we evaluated the anti-tumorigenic mechanisms and efficacy of NanoALA, a novel nanoformulation of aminolevulic acid (ALA) based on poly(lactide-co-glycolide) (PLGA) nanocapsules designed for anticancer photodynamic therapy (PDT).MethodsFor this purpose, physicochemical characterization, prodrug incorporation kinetics, biocompatibility and photocytotoxicity tests, analysis of the cell death type and mitochondrial function, measurement of the intracellular reactive oxygen species production and DNA fragmentation were performed in murine mammary carcinoma (4T1) cells.ResultsNanoALA formulation, stable over a period of 90 days following synthesis, presented hydrodynamic diameter of 220 ± 8.7 nm, zeta potential of −30.6 mV and low value of polydispersity index (0.28). The biological assays indicated that the nanostructured product promotes greater ALA uptake by 4T1 cells and consequently more cytotoxicity in the PDT process. For the first time in the scientific literature, there is a therapeutic efficacy report of approximately 80%, after only 1 h of incubation with 100 μg mL⁻¹ prodrug (0.6 mM ALA equivalent). The mitochondria are probably the initial target of treatment, culminating in energy metabolism disorders and cell death by apoptosis.ConclusionsNanoALA emerges as a promising strategy for anticancer PDT. Besides being effective against a highly aggressive tumor cell line, the treatment may be economically advantageous because it allows a reduction in the dose and frequency of application compared to free ALA.     

Efficient fluorescence detection of protoporphyrin IX in metastatic lymph nodes of murine colorectal cancer stained with indigo carmine

Protoporphyrin IX (PpIX), a biochemical converted from 5-aminolevulinc acid (5-ALA) in living cells, is useful for intraoperative fluorescent detection of cancer metastasis in lymph nodes (LNs). However, unknown is whether the fluorescence of PpIX can be detected in the LNs when they coexist with indigo carmine, a blue dye commonly used for identification of sentinel LNs during surgery. To address this issue, we sought to evaluate the diagnostic usefulness of PpIX fluorescence in the presence of indigo carmine in a mouse LN metastasis model of rectal cancer after administration of 5-ALA. Spectral analysis of pure chemicals revealed that the absorption spectrum of indigo carmine widely overlapped with the fluorescence spectrum of PpIX specifically at the peak of 632 nm, a common emission wavelength for detecting PpIX, but not at the other peak of 700 nm. Due to such spectral overlap, the PpIX fluorescence intensity was significantly attenuated by mixture with indigo carmine at 632 nm, but not at 700 nm. Accordingly, fluorescent measurements of the mouse metastatic LN revealed more intense presentation of PpIX at 700 nm than at 632 nm, indicating that the diagnostic usefulness is greater at 700 nm than at 632 nm for the indigo carmine-dyed LNs after administration of 5-ALA. From these observations, we propose that the fluorescence measurement is more efficient at 700 nm than at 632 nm for detection of PpIX in metastatic LNs stained with indigo carmine.     

Skin fluorescence following photodynamic therapy with NPe6 photosensitizer

BackgroundThe second-generation photosensitizer NPe6 has strong anti-tumor effects with a much shorter photosensitive period than the first-generation photosensitizer Photofrin. Although photosensitive period has been reduced, skin photosensitivity is still a major side effect of photodynamic therapy (PDT). Therefore, we conducted a prospective study to investigate whether the NPe6 fluorescence intensity in skin after PDT could be measured effectively in human patients to improve the management of a patient's photosensitive period.MethodsThe NPe6 fluorescence measurements using a constructed fluorescence sensing system at the inside of the arm were acquired prior to and 5 and 10 min after NPe6 administration as well as at the time of PDT (4–5 h after administration), at discharge (2 or 3 days after PDT), and at 1 or 2 weeks after PDT. Participants were interviewed as to whether they had any complications at 2 weeks after PDT.ResultsNine male patients and one female patient entered this study. Nine patients were inpatients and one patient was an outpatient. All of the measurements of NPe6 fluorescence in the skin could be obtained without any complications. The spectral peak was detected at the time of discharge (2–3 days after administration) in most cases and it decreased at 1 or 2 weeks after PDT.ConclusionsThe fluorescence of NPe6 in the skin could be detected feasibly using the fluorescence sensing system in human patients. Measuring the relative concentration of NPe6 in the skin indirectly by measuring fluorescence intensity might be useful to predict the period of skin photosensitivity after PDT.     

In vitro effects of photodynamic therapy induced by chloroaluminum phthalocyanine nanoemulsion

BackgroundThe photodynamic therapy (PDT) has been used to treat cancer mainly by inducing oxidative stress. Our aim was to evaluate the effect of PDT and its combination with methoxyamine (MX), a blocker of base excision repair (BER), in cells expressing high levels of the APE1 protein, which is involved in cell oxidative damage response.MethodsThe HeLa and A549 cells were treated for 3 h with chloroaluminum phthalocyanine incorporated into a well-designed nanoemulsion (ClAlPc/NE); and then irradiated by visible light (@670 nm) with doses of 0.1, 0.5 and 1.0 J/cm². A simultaneous combination of MX + ClAlPc/NE was performed and then irradiated with the selected dose of 0.5 J/cm². The treatments were evaluated in terms of viability, clonogenicity, DNA fragmentation, and cell death mechanism by apoptosis and/or necrosis.ResultsThe APE1 protein expression observed was higher in HeLa than in A549. Both cell lines exhibited substantial differences in cell cytotoxicity. The PDT decreased the clonogenicity of HeLa by inducing apoptosis (sub-G1 and annexin detection). Additionaly, the MX potentiates the PDT-effects in HeLa. Otherwise, low cytotoxicity was observed in A549 cells.ConclusionThe PDT induced apoptosis in high APE1 expressive HeLa cells, and the blockage of BER by MX increased its effects.     

Photodynamic therapy: An option in mycosis fungoides

BackgroundPhotodynamic therapy (PDT) is a well-known and effective treatment for non-melanoma skin-cancer. Numerous studies have also shown its effectiveness in mycosis fungoides. The aim of the study was to analyse MF patients treated with PDT at the Dermatology Unit of Bologna University.MethodsWe retrospectively analysed MF patients treated with PDT over the last ten years. Each PDT protocol consisted of the appliance for 3 h under an occlusive film dressing on each lesion of a one-mm-thick layer of 16% methyl aminolaevulinate (MAL) 160 mg/g cream (Metvix^®, Galderma, Paris, France). The cream was then removed and the skin was exposed to 630 nm red light from a diode lamp (Aktilite^®, Galderma Benelux, Rotterdam, the Netherlands), with a total radiation dose of 37 J/cm2 for 9 mins. A protocol of one session every month was scheduled. The treated lesions were clinically examined, before each treatment.ResultsFour cases, three male and one female, had been treated with PDT. Two patch lesions on the plantar area, one leg and the pubic area were treated. The number of PDT sessions ranged from 4 to 9. Two complete remissions and two partial remissions were observed. A low-to-mild burning sensation was reported during the treatment, and persisted over the next day; no further side effects were observed.ConclusionsOur series shows that PDT can be considered an effective second-line treatment in patients characterised by a disease located in difficult-to-treat anatomical areas such as the feet and the pubic area.     

Photodynamic therapeutic efficacy of symmetrical diiodinated squaraine in in vivo skin cancer models

BackgroundPhotodynamic therapy (PDT) has clinical approval for use as a minimally invasive therapeutic procedure that is able to exert selective cytotoxic activity toward malignant cells. The dye selected for our study, symmetrical diiodinated squaraine, is one of the newly developed photosensitizers. The study is designed to determine the efficacy of PDT mediated by symmetrical diiodinated squaraine in skin tumor induced Swiss albino mice.MethodsSkin tumor was induced in mice with dimethyl benzanthracene (DMBA) and croton oil. After squaraine administration to the tumor mice, photodynamic treatment of tumors was performed using a 1000 W halogen lamp corresponding to the light dose of 100 J/cm². The mice were euthanized and skin flaps and tumor tissues from the back of mice were excised for biochemical studies. The biochemical parameters analyzed include some relevant tumor markers for epithelial tissues, inflammatory markers and markers of apoptosis. The gene expression studies were done by RT-PCR.ResultsAfter two weeks of the treatment, there was significant inflammation. However at 90 days after PDT, the parameters reverted to near-normal values. All marker parameters of tumor progression brought back to normal levels by PDT. Increased caspase-3 activity in PDT treated group shows that cell death might have occurred by apoptosis. The gene expression profile confirms the results.ConclusionsThe results of the whole study show the therapeutic efficacy and apoptosis mediated tumor destruction by squaraine PDT.     

A clinical evaluation of the efficacy of photodynamic therapy in the treatment of erosive oral lichen planus: A case series

BackgroundErosive oral lichen planus (EOLP) poses a substantial risk of malignant transformation into squamous cell cancer. The absence of established treatment gives way to alternative therapeutic strategies, including photodynamic therapy. The aim of the study was to evaluate the efficacy of PDT in the treatment of EOLP.MethodsTwelve female patients aged 63–80 with 22 OLP lesions (16 on the buccal mucosa, 6 on gingiva and tongue), underwent authors’ own PDT scheme with the use of 5% solution of 5-aminolevulinic acid (ALA) as photosensitizer. An ALA-saturated occlusive dressing was applied directly onto a lesion and surrounding mucosa 2 h prior to illumination with a custom-made diode lamp (light of 630 nm, dose of 300 mW). After a series of 10 weekly illumination sessions the patients were monitored for 12 months.ResultsThe mean size of lesions before treatment was 1.46 cm² ± 1.44. The lesions on the buccal mucosa were smaller (1.06 cm² ± 0.98) than those on the gingiva and tongue (2.63 cm² ± 1.93). Post-treatment improvement encompassed 16 lesions, 5 of which were in remission. The mean reduction in size after 10-session therapy was 8,05%. The healing continued and further reduction in size (by 69.13%) took place during the 12-month observation: 39.62% of lesions within the buccal mucosa and full remission of all lesions on the gingiva and tongue.ConclusionsThe results suggest that PDT offers non-invasive treatment of lesions in oral mucosa and may become an alternative and complementary method to those currently in use. Further studies involving larger groups of patients should be undertaken before it becomes routine practice.     

Photodynamic process induced by chloro-aluminum phthalocyanine nanoemulsion in glioblastoma

BackgroundGlioblastoma multiforme (GBM) is a tumor characterized by rapid cell proliferation and migration. GBM constitutes the most aggressive and deadly type of brain tumor and is classified into several subtypes that show high resistance to conventional therapies. There are currently no curative treatments for malignant glioma despite the numerous advances in surgical techniques, radiotherapy, and chemotherapy. Therefore, alternative approaches are required to improve GBM treatment.MethodsOur study proposes the use of photodynamic therapy (PDT) for GBM treatment, which uses chloro-aluminum phthalocyanine (AlClPc) encapsulated in a new drug delivery system (DDS) and designed as a nanoemulsion (AlClPc/NE). The optimal dark non-cytotoxic AlClPc/NE concentration for the U87 MG glioma cell model and the most suitable laser light intensity for irradiation were determined. Experimental U87 MG cancer cells were analyzed via MTT cell viability assay. Cellular localization of AlClPc, morphological changes, and cell death via the necrotic and apoptotic pathways were also evaluated.ResultsAlClPc remained in the cytoplasmic region at 24 h after administration. Additionally, treatment with 1.0 μmol/L AlClPc under light irradiation at doses lower than 140 mJ/cm resulted in morphological changes with 50 ± 6% cell death (p < 0.05). Moreover, 20 ± 2% of U87 MG cells underwent cell death via the necrotic pathway. Measurement of Caspase-9 and -3 activities also suggested that cells underwent apoptosis. Taken together, these results indicate that AlClPc/NE-PDT can be used in the treatment of glioblastoma by inducing necrotic and apoptotic cell death.ConclusionsOur findings suggest that AlClPc/NE-PDT induces cell death in U87 MG cells in a dose-dependent manner and could thus serve as an effective adjuvant treatment for malignant glioma. AlClPc/NE-PDT utilizes a low dose of visible light and can be used in combination with other classic GBM treatment approaches, such as a combination of chemotherapy and surgery.     

Methylene blue,curcumin and ion pairing nanoparticles effects on photodynamic therapy of MDA-MB-231 breast cancer cell

PurposeThe aim of current study was to use methylene blue-curcumin ion pair nanoparticles and single dyes as photosensitizer for comparison of photodynamic therapy (PDT) efficacy on MDA-MB-231 cancer cells, also various light sources effect on activation of photosensitizer (PS) was considered.MethodIon pair nanoparticles were synthesized using opposite charge ions precipitation and lyophilized. The PDT experiments were designed and the effect of PSs and light sources (Red LED (630 nm; power density: 30 mW cm⁻²) and blue LED (465 nm; power density: 34 mW cm⁻²)) on the human breast cancer cell line were examined. The effect of PS concentration (0–75 μg. mL⁻¹), incubation time, irradiation time and light sources, and priority in irradiation of blue or red lights were determined.ResultsThe results show that the ion pairing of methylene blue and curcumin enhance the photodynamic activity of both dyes and the cytotoxicity of ion pair nanoparticles on the MDA-231 breast cancer cell line. Blue and red LED light sources were used for photo activation of photosensitizers. The results demonstrated that both dyes can activate using red light LED better than blue light LED for singlet oxygen producing.ConclusionNano scale ion pair precipitating of methylene blue-curcumin enhanced the cell penetrating and subsequently cytotoxicity of both dyes together.     

Combination of hand-held probe and microscopy for fluorescence guided surgery in the brain tumor marginal zone

BackgroundVisualization of the tumor is crucial for differentiating malignant tissue from healthy brain during surgery, especially in the tumor marginal zone. The aim of the study was to introduce a fluorescence spectroscopy-based hand-held probe (HHF-probe) for tumor identification in combination with the fluorescence guided resection surgical microscope (FGR-microscope), and evaluate them in terms of diagnostic performance and practical aspects of fluorescence detection.Material and MethodsEighteen operations were performed on 16 patients with suspected high-grade glioma. The HHF-probe and the FGR-microscope were used for detection of protoporphyrin (PpIX) fluorescence induced by 5-aminolevulinic acid (5-ALA) and evaluated against histopathological analysis and visual grading done through the FGR-microscope by the surgeon. A ratio of PpIX fluorescence intensity to the autofluorescence intensity (fluorescence ratio) was used to quantify the spectra detected by the probe.ResultsFluorescence ratio medians (range 0 – 40) measured by the probe were related to the intensity of the fluorescence in the FGR-microscope, categorized as “none” (0.3, n = 131), “weak” (1.6, n = 34) and “strong” (5.4, n = 28). Of 131 “none” points in the FGR-microscope, 88 (67%) exhibited fluorescence with the HHF-probe. For the tumor marginal zone, the area under the receiver operator characteristics (ROC) curve was 0.49 for the FGR-microscope and 0.65 for the HHF-probe.ConclusionsThe probe was integrated in the established routine of tumor resection using the FGR-microscope. The HHF-probe was superior to the FGR-microscope in sensitivity; it detected tumor remnants after debulking under the FGR-microscope. The combination of the HHF-probe and the FGR-microscope was beneficial especially in the tumor marginal zone.     

Photodynamic therapy using methylene blue in lung adenocarcinoma xenograft and hamster cheek pouch induced squamous cell carcinoma

BackgroundPhotodynamic therapy (PDT) is used to treat early proximal bronchial cancer during a flexible bronchoscopy. The technique relies on the excitation of a photosensitizer by an appropriate wavelength, which is delivered into the bronchus in close contact with the tumor.ObjectiveTo assess methylene blue (MB) as a PDT agent for the treatment of respiratory tract cancer in animal models.MethodsMB-induced PDT was performed on 7 subcutaneous NCI-H460 lung adenocarcinoma xenografts in nude mice and 9 induced squamous cell cancer in the hamster cheek pouch model. In mice, PDT was carried out on right-sided tumors after intratumoral injection of methylene blue 1% (w/v) and illumination at 630 nm at 200 J/cm (Diomed PDT 630), with the left tumor used as control (illumination alone or MB alone). The tumoral volume was assessed before and 15 days after PDT.ResultsFourteen xenografts were treated in mice, including seven treated with MB-PDT, producing a 52% mean tumor volume regression (1568 mm³ vs. 544 mm³) compared to seven control cases in which tumor volume increased (p = 0.007; Mann-Whitney test). Nine cheek pouch induced carcinomas were treated in the hamster group, with a mean volume decrease of 85.8% (from 44.8% to 100%) (initial mean volume = 210 mm³ vs. post PDT mean volume = 97 mm³). Histology analysis showed 4/9 complete responses.ConclusionIntratumoral MB appears efficient as PDT agent for cancer treatment in animal models. Further studies are needed to assess the safety and efficacy of MB-associated PDT for the treatment of lung cancer in humans.