Higher Light Doses Research Articles

Periopathogens differ in terms of the susceptibility to toluidine blue O-mediated photodynamic inactivation

BackgroundThe main goal of periodontal therapy is to eliminate the infection spreading in periodontium. Antimicrobial photodynamic therapy may be applied in order to eradicate pathogens remaining in periodontal tissues after conventional mechanical debridement, to improve the treatment results. The aim of this in vitro study was to evaluate the susceptibility of selected key periopathogens to toluidine blue O-mediated photodynamic inactivation and the influence of photosensitizer’s concentration and light dose on the effectiveness of this process. MethodsFollowing bacterial strains were used in the experiments: Porphyromonas gingivalis ATCC 33277, Aggregatibacter actinomyctemecomitans ATCC 33384, Fusobacterium nucleatum ATCC 10953. Toluidine blue O (TBO) was used in concentration ranging from 0.004 to 0.5mg/mL. Irradiation was performed by a non-laser red light source. ResultsComplete eradication of P. gingivalis was obtained upon the application of TBO in the concentration of 0.1mg/mL and the highest light dose. A, actinomycetemcomitans was, in turn, not susceptible to photodynamic inactivation regardless of the dosimetric parameters applied. High viability reductions were also obtained for F. nucleatum, however no complete eradication. The effectiveness of photodynamic inactivation of susceptible periopathogens was dependent on the light dose and photosensitizer’s concentration. ConclusionsPeriopathogens differ in terms of their susceptibility to photodynamic inactivation. Antimicrobial PDT may be valuable in the treatment of those cases of periodontal disease, in which P. gingivalis is a dominating pathogen. Microbiological examination prior to clinical application of aPDT may be recommended.

Histologic changes associated with talaporfin sodium-mediated photodynamic therapy in rat skin.

Alternative treatments are needed to achieve consistent and more complete port wine stain (PWS) removal, especially in darker skin types; photodynamic therapy (PDT) is a promising alternative treatment. To this end, we previously reported on Talaporfin Sodium (TS)-mediated PDT. It is essential to understand treatment tissue effects to design a protocol that will achieve selective vascular injury without ulceration and scarring. The objective of this work is to assess skin changes associated with TS-mediated PDT with clinically relevant treatment parameters. We performed TS (0.75 mg/kg)-mediated PDT (664 nm) on Sprague Dawley rats. Radiant exposures were varied between 15 and 100 J/cm2 . We took skin biopsies from subjects at 9 hours following PDT. We assessed the degree and depth of vascular and surrounding tissue injury using histology and immunohistochemical staining. TS-mediated PDT at 0.75 mg/kg combined with 15 and 25 J/cm2 light doses resulted in vascular injury with minimal epidermal damage. At light dose of 50 J/cm2 , epidermal damage was noted with vascular injury. At light doses >50 J/cm2 , both vascular and surrounding tissue injury were observed in the forms of vasculitis, extravasated red blood cells, and coagulative necrosis. Extensive coagulative necrosis involving deeper adnexal structures was observed for 75 and 100 J/cm2 light doses. Observed depth of injury increased with increasing radiant exposure, although this relationship was not linear. TS-mediated PDT can cause selective vascular injury; however, at higher light doses, significant extra-vascular injury was observed. This information can be used to contribute to design of safe protocols to be used for treatment of cutaneous vascular lesions. Lasers Surg. Med. 49:767-772, 2017. © 2017 Wiley Periodicals, Inc.

Effects of ascorbic acid and light on reactions in fresh-cut apples by microcalorimetry

During the manufacturing of fresh-cut apples, a number of biochemical events, overall exothermic, contribute to increasing the reaction rate of the fruit and the browning of its wounded surface. This work applied isothermal microcalorimetry to compare the overall effect of such complex events before and after treatments with ascorbic acid solutions, pulsed lights or UV-C lights. Briefly, apple samples were cut into cylinders and dipped in solutions containing ascorbic acid (0–2.5%) or exposed to high energy doses of light (from 6 to 175kJ/m2). In general, the heat-flow signal recorded by microcalorimetry was inversely proportional to the intensity of the applied treatment. In case of treatments with ascorbic acid, the heat-flow signal was empirically deconvoluted in three distinctive signals, respectively, (I) an exponential decay, (II) a gaussian central curve and (III) a final logistical function. The first and the third functions were constant regardless of the concentration of ascorbic acid used. Only the second Gaussian function was correlated with the concentration of ascorbic acid and the area was used to evaluate the efficacy of the process. Overall, this work contributes to the understanding of the heat produced by fruit after wounding and, from a practical standpoint, can help compare the effects of different treatments on fresh cut fruits.

Sae regulator factor impairs the response to photodynamic inactivation mediated by Toluidine blue in Staphylococcus aureus

Photodynamic inactivation (PDI) involves the combined use of light and a photosensitizer, which, in the presence of oxygen, originates cytotoxic species capable of inactivating bacteria. Since the emergence of multi-resistant bacterial strains is becoming an increasing public health concern, PDI becomes an attractive choice.The aim of this work was to study the differential susceptibility to Toluidine blue (TB) mediated PDI (TB-PDI) of S. aureus mutants (RN6390 and Newman backgrounds) for different key regulators of virulence factors related to some extent to oxidative stress.Complete bacteria eradication of planktonic cultures of RN6390 S. aureus photosensitized with 13μM TB was obtained upon illumination with a low light dose of 4.2J/cm2 from a non-coherent light source. Similarly, complete cell death was achieved applying 1.3μM TB and 19J/cm2 light dose, showing that higher light doses can lead to equal cell death employing low photosensitizer concentrations. Interestingly, RN6390 in planktonic culture responded significantly better to TB-PDI than the Newman strain.We showed that deficiencies in rsbU, mgrA (transcription factors related to stress response) or agr (quorum sensing system involved in copper resistance to oxidative stress) did not modify the response of planktonic S. aureus to PDI. On the other hand, the two component system sae impaired the response to TB-PDI through a mechanism not related to the Eap adhesin.More severe conditions were needed to inactivate S. aureus biofilms (0.5mM TB, 157J/cm2 laser light). In mutant sae biofilms, strain dependant differential susceptibilities are not noticed.

Flow velocity and light level drive non-linear response of seagrass Zostera noltei to ammonium enrichment

We investigated the interactive effects of light (low and high light doses) and flowvelocity (low, medium and high levels) under NH4+ enriched conditions on dynamic and morphologicalvariables of Zostera noltei plants in a 5 wk flume experiment. Our results showed a nonlinearresponse of Z. noltei in this factorial design, with the strongest negative effect of NH4+enrichment recorded at intermediate flow velocities for almost all the dynamic response variables(i.e. survival, net production, shoot appearance rate). This negative effect of NH4+ enrichment wasintensified under low light conditions, where net production was only positive in plants growingat high flow velocity. This positive effect of flow velocity was ascribed to the more horizontal positionof the leaves, which allows for higher levels of light capture than under lower flow velocities.However, enhancing current velocity may increase NH4+ uptake rates until they reach adverselevels, which can potentially trigger NH4+ toxicity. This negative effect of flow velocity seems to beneutralized by the higher light capture at high flow, resulting in intermediate current velocitiesbeing more harmful for plants. In summary, our results highlight the importance of studying thecomplexity of interactions among multiple stressors that frequently co-occur in nature to improveour ability to forecast the response of seagrass populations to possible interaction effects in futureglobal change scenarios.

CdSe/ZnS quantum dots induce photodynamic effects and cytotoxicity in pancreatic cancer cells.

To investigate the photodynamic effect of CdSe/ZnS quantum dots (QDs) on pancreatic cancer cells and elucidate the probable mechanisms. The pancreatic cancer cell line SW1990 was treated with different concentrations of CdSe/ZnS QDs (0, 0.5, 1.0, 1.5, 2.0, 2.5 μmol/L), with or without illumination. The viability of SW1990 cells was tested using the Cell Counting Kit-8 (CCK-8) assay. The ultrastructural changes of SW1990 cells were observed by transmission electron microscopy. Apoptosis was detected by nuclear staining and flow cytometry (FCM). Reactive oxygen species (ROS) were measured by dichlorofluorescein diacetate via fluorescence microscopy. Expression of Bax, Bcl-2 and caspase-3 was measured by real-time polymerase chain reaction (PCR) and protein immunoblotting 24 h after SW1990 cells were treated with CdSe/ZnS QDs and illuminated. The CCK-8 assay results showed that both CdSe/ZnS QDs with and without illumination suppressed SW1990 cell proliferation. Cell viability was significantly lower when illuminated or with a longer incubation time and a higher light dose. CdSe/ZnS QDs with illumination caused ultrastructural changes in SW1990 cells, such as organelle degeneration and chromatin condensation and aggregation at the periphery of the nucleus. Fluorescence microscopy and FCM showed that CdSe/ZnS QDs (1.5 μmol/L) with illumination increased SW1990 cell apoptosis (53.2%) and ROS generation compared with no illumination. Real-time PCR showed that expression of Bax and caspase-3 was upregulated and Bcl-2 was downregulated. Immunoblotting results were consistent with real-time PCR results. Inhibition of ROS and apoptosis both attenuated QD-photodynamic-therapy-induced cell death. CdSe/ZnS QDs can be used as a photosensitizer to inhibit SW1990 cell proliferation through ROS generation and apoptotic protein expression regulation.

Nanoparticle loaded coatings and patches on live cells

Event Abstract Back to Event Nanoparticle loaded coatings and patches on live cells Brad Berron1, Pei-Jung Wu1, Jacob L. Lilly1 and Roberto Arreaza1 1 University ofKentucky, Chemical and Materials Engineering, United States Introduction: Cells possess biodistribution characteristics which are unique from all traditional modes of drug delivery. For example, a red blood cell can carry large payloads of oxygen and remain in circulation for weeks. Other cells actively defeat natural barriers to which are difficult to drug delivery (blood-brain barrier, blood-retinal barrier). The present work develops the fabrication techniques for loading cells with drug-eluting nanoparticles for cell-directed drug delivery using nanoscale coatings. For prolonged systemic delivery, we develop techniques for depositing a nanoparticle loaded coating on the surface of red blood cells. For cell-directed drug delivery, we develop techniques for restricting the drug-loaded coating to a small area on the cell. Methods: For uniform coatings, cell surfaces are first labeled with an eosin photoinitiator through immunolabeling with a biotinylated antibody and subsequent immersion in a streptavidin eosin conjugate. Cells are then immersed in a pH buffered solution of the desired nanoparticles, PEG diacrylate, triethanol amine, and vinyl pyrollidinone. Irradiation with 530 nm light (20 mW/cm2 for 20 minutes) results in the conformal coating of the cell. Cell patches are prepared similarly, with the additional use of a photomask to restrict polymer formation to irradiated regions. Results Discussion: Conformal cell coatings were deposited on human red cells, Jurkats, A549, and H358 cells. Diffusion of drugs through the coatings was limited to materials <10 kDa. Transport of low MW materials was rapid; equilibrium release rates are established within seconds. The release kinetics of doxorubicin from nanoparticles from the coatings is consistent with that of the doxorubicin from the nanoparticles, again supporting the minimal mass transfer resistance of the coatings for small molecules. >80% cell viability was confirmed by calcein, ethidium, and caspase assays. The coated cells are able to deform to traverse <10 micron openings. Patterned coatings are formed on cells with photomasks to limit the area of light exposure. The parameter space was evaluated to determine their influence on gelation kinetics and the associated quality of the pattern. Additionally, the influence of the parameter space on cell viability was evaluated. Key findings include: 1) Rapid gelation kinetics promote accuracy of patterning, 2) High light doses negatively impact viability, 3) The presence of oxygen during polymerization promotes redox damage of cells. Conclusion: Photopolymerization offers a convenient route to load nanoparticles into coatings on cells. The cells retain viability during the coating process. The coatings offer minimal resistance to mass transfer of small molecules, while large species are trapped in the coatings. Photopatterning of these coatings is simple. The polymerization design is focused on cell-compatible conditions while encouraging rapid gelation for accurate patterning on the micron scale. Keywords: Drug delivery, material design, Cell modulation, bioinerface Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: General Session Oral Topic: Biomaterials for therapeutic delivery Citation: Berron B, Wu P, Lilly JL and Arreaza R (2016). Nanoparticle loaded coatings and patches on live cells. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.02101 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2016; Published Online: 30 Mar 2016. Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Brad Berron Pei-Jung Wu Jacob L Lilly Roberto Arreaza Google Brad Berron Pei-Jung Wu Jacob L Lilly Roberto Arreaza Google Scholar Brad Berron Pei-Jung Wu Jacob L Lilly Roberto Arreaza PubMed Brad Berron Pei-Jung Wu Jacob L Lilly Roberto Arreaza Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Photodynamic proprieties of Hypericin and its aqueous soluble derivative Glucamine-Hypericin

s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 351 for low and high light dose, respectively for Vero and HEp-2. The results suggest that HY-G may be as potent photosensitizer as HY with the advantage of being more soluble. Acknowledgements: CEPOF, CAPES, FAPESP. http://dx.doi.org/10.1016/j.pdpdt.2015.07.102 Cellular distribution and PDT response of chlorin based photosensitizer in murine melanoma Bruno A. Ono, Sebastiao Pratavieira, Cristina Kurachi Sao Carlos Institute of Physics, University of Sao

Cellular distribution and PDT response of chlorin based photosensitizer in murine melanoma

s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 351 for low and high light dose, respectively for Vero and HEp-2. The results suggest that HY-G may be as potent photosensitizer as HY with the advantage of being more soluble. Acknowledgements: CEPOF, CAPES, FAPESP. http://dx.doi.org/10.1016/j.pdpdt.2015.07.102 Cellular distribution and PDT response of chlorin based photosensitizer in murine melanoma Bruno A. Ono, Sebastiao Pratavieira, Cristina Kurachi Sao Carlos Institute of Physics, University of Sao

Evaluation of photodynamic therapy in three-dimensional cell culture by magnetic levitation

s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 351 for low and high light dose, respectively for Vero and HEp-2. The results suggest that HY-G may be as potent photosensitizer as HY with the advantage of being more soluble. Acknowledgements: CEPOF, CAPES, FAPESP. http://dx.doi.org/10.1016/j.pdpdt.2015.07.102 Cellular distribution and PDT response of chlorin based photosensitizer in murine melanoma Bruno A. Ono, Sebastiao Pratavieira, Cristina Kurachi Sao Carlos Institute of Physics, University of Sao

Acute phase injury of vascular endothelial cells under extra-cellular talaporfin sodium existence: In vitro study

s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 351 for low and high light dose, respectively for Vero and HEp-2. The results suggest that HY-G may be as potent photosensitizer as HY with the advantage of being more soluble. Acknowledgements: CEPOF, CAPES, FAPESP. http://dx.doi.org/10.1016/j.pdpdt.2015.07.102 Cellular distribution and PDT response of chlorin based photosensitizer in murine melanoma Bruno A. Ono, Sebastiao Pratavieira, Cristina Kurachi Sao Carlos Institute of Physics, University of Sao

Stability enhanced polyelectrolyte-coated gold nanorod-photosensitizer complexes for high/low power density photodynamic therapy

Photodynamic therapy (PDT) is a promising treatment modality for cancer and other malignant diseases, however safety and efficacy improvements are required before it reaches its full potential and wider clinical use. Herein, we investigated a highly efficient and safe photodynamic therapy procedure by developing a high/low power density photodynamic therapy mode (high/low PDT mode) using methoxypoly(ethylene glycol) thiol (mPEG-SH) modified gold nanorod (GNR)-AlPcS4 photosensitizer complexes. mPEG-SH conjugated to the surface of simple polyelectrolyte-coated GNRs was verified using Fourier transform infrared spectroscopy; this improved stability, reduced cytotoxicity, and increased the encapsulation and loading efficiency of the nanoparticle dispersions. The GNR-photosensitizer complexes were exposed to the high/low PDT mode (high light dose = 80 mW/cm2 for 0.5 min; low light dose = 25 mW/cm2 for 1.5 min), and a high PDT efficacy leads to approximately 90% tumor cell killing. Due to synergistic plasmonic photothermal properties of the complexes, the high/low PDT mode demonstrated improved efficacy over using single wavelength continuous laser irradiation. Additionally, no significant loss in viability was observed in cells exposed to free AlPcS4 photosensitizer under the same irradiation conditions. Consequently, free AlPcS4 released from GNRs prior to cellular entry did not contribute to cytotoxicity of normal cells or impose limitations on the use of the high power density laser. This high/low PDT mode may effectively lead to a safer and more efficient photodynamic therapy for superficial tumors.

Evaluation of DNA damage reversal during medium-pressure UV disinfection

Ultraviolet (UV) disinfection relies on the principal that DNA exposure to UV irradiation leads to the formation of cytotoxic lesions resulting in the inactivation of microorganisms. Cyclobutane pyrimdine dimers (CPDs) account for the majority of DNA lesions upon UV exposure. Past research has demonstrated reversal of CPDs in extracted DNA formed at high UV-C wavelength irradiation (280 nm) upon subsequent irradiation at lower UVC wavelengths (230–240 nm). Medium-pressure (MP) UV lamps produce a polychromatic emission giving rise to the possibility that cellular DNA in a target pathogen may undergo simultaneous damage and repair when exposed to multiple wavelengths during the disinfection process, decreasing the efficiency of MP UV lamp disinfection. Culture techniques and a quantitative polymerase chain reaction (qPCR) assay were used to examine cell viability and DNA damage reversal. qPCR results indicated direct photoreversal of UV-induced DNA damage through sequential irradiations of 280 nm followed by 228 nm in Escherichia coli DNA. However, significant photoreversal was only observed after high initial doses and secondary doses of UV light. The doses where significant photoreversal took place were more than 10 times higher than those typically used in UV disinfection. Despite evidence of CPD photoreversal, bacterial growth assays showed no indication that sequential-wavelength irradiations result in higher survival rates than single-wavelength irradiations.

Safety Study of Photodynamic Therapy Using Talaporfin Sodium in the Pancreas and Surrounding Tissues in the Syrian Golden Hamster

Aim. To assess the safety of photodynamic therapy (PDT) using talaporfin sodium on the pancreas and surrounding organs in normal hamsters.Methods. Fluorescence microscopy documented talaporfin levels in liver, duodenum, and pancreas up to 24 hours after photosensitisation. Lesion size in liver 3 days after PDT (50 J, 5 mg/kg, variable drug-light interval (DLI)) was documented to optimise the DLI. Using optimum DLI, pancreas and surrounding organs were treated with laser fibre touching the surface and animals were killed at 3 or 21 days.Results. Peak fluorescence was seen in duodenum and pancreas at 15 mins (second lower peak at 2 hours). Liver fluorescence was consistently high (peak 1 hour) until after 4 hours. Optimum DLI was seen at 15 minutes. The pancreas was relatively resistant to direct PDT injury (small lesions at high doses) but surrounding stomach, duodenum, and liver were more susceptible with evidence of adhesions and full thickness damage (localised peritonitis and duodenal perforation at highest doses).Conclusion. The safety profile is similar to PDT with longer acting photosensitisers. The pancreas appears safe to treat, but care is required to avoid high light doses to the intestinal tract, particularly the duodenum.

Lethal photosensitization of Porphyromonas gingivalis by their endogenous porphyrins under anaerobic conditions: An in vitro study

Lethal photosensitization has been previously demonstrated in Porphyromonas gingivalis, but oxygen is considered to be essential to this process. However, since P. gingivalis is a periodontal pathogen which grows in the low oxygen conditions found in the subgingival crevice, it was considered prudent to study its photosensitivity in anaerobic conditions. A series of experiments were undertaken to attempt to induce lethal photosensitization in P. gingivalis (ATCC 33277) under strict anaerobic conditions using two different 405 nm light sources. Samples of P. gingivalis were grown on a blood-containing, solid growth medium before being suspended in saline and then exposed to 405 nm light delivered by either a hand-held light source (Toothcare™) (11.4 mW/cm(2)) or a laser pointer (328.5 mW/cm(2)). With the exception of the adjustment of the P. gingivalis suspensions to a fixed optical density, the experiments were carried out in their entirety within an anaerobic chamber. The lowest Toothcare light dose tested (0.34 J/cm(2); 30s) yielded a statically significant kill of 63.4% which increased to 94.1% kill at higher light doses (3.42 J/cm(2); 300 s). The laser pointer similarly achieved kills of 90.2% at the lower light dose tested (9.86 J/cm(2); 30s) and 94.5% kill at the highest light dose (98.55 J/cm(2); 300 s). Lethal photosensitization can be instigated in planktonic suspensions of P. gingivalis at 405 nm delivered by hand-held devices under anaerobic conditions. This suggests the possibility that lethal photosensitization occurred by the oxygen-independent type I pathway as oppose to the oxygen-dependent type II pathway.

Staphylococcus aureus small colony variants are susceptible to light activated antimicrobial agents

BackgroundAntibiotic therapy can select for small colony variants of Staphylococcus aureus that are more resistant to antibiotics and can result in persistent infections, necessitating the development of more effective antimicrobial strategies to combat small colony variant infections. Photodynamic therapy is an alternative treatment approach which utilises light in combination with a light-activated antimicrobial agent to kill bacteria via a non-specific mechanism of action. In this study, we investigated whether the combination of 665 nm laser light and the light-activated antimicrobial agent methylene blue was able to successfully kill S. aureus small colony variants. S. aureus and isogenic stable small colony variant were exposed to varying doses (1.93 to 9.65 J/cm2) of 665 nm laser light in the presence of varying concentrations (1 to 20 μM) of methylene blue.ResultsThe combination of 665 nm laser light and methylene blue was found to be an effective strategy for the killing of small colony variants. At the highest light dose (9.65 J/cm2) and methylene blue concentration (20 μM) tested, the number of viable bacteria decreased by approximately 6.9 log10 for the wild type and approximately 5 log10 for the small colony variant.ConclusionsThese results suggest that photodynamic therapy has potential for use in the treatment of superficial infections caused by small colony variants of S. aureus and supports further research in this field.

Acute Cytotoxic Effects of Photoimmunotherapy Assessed by 18F-FDG PET

We have recently developed a cancer-specific therapy, photoimmunotherapy, which uses an antibody-IR700 (phototoxic phthalocyanine dye) conjugate to bind to the cell membrane and near-infrared light to induce immediate and highly specific tumor killing in vivo. For monitoring the acute cytotoxic effects of photoimmunotherapy before the tumor begins to shrink, we used (18)F-FDG PET before and after this intervention in mice. Photoimmunotherapy was performed by binding panitumumab (anti-HER1)-IR700 to HER1-positive tumor cells (A431), followed by near-infrared light irradiation in vitro and in vivo. The uptake of (18)F-FDG in the tumor after photoimmunotherapy was evaluated in cellular uptake studies and PET imaging studies. Serial histologic analyses were conducted after photoimmunotherapy. The in vitro cellular uptake of (18)F-FDG was reduced as the dose of light increased, and at high light dose (2 J/cm(2)) the uptake was reduced by more than 99% within 1 h after photoimmunotherapy. In vivo (18)F-FDG PET imaging showed that the accumulation of radioactivity in the treated tumors decreased 76% at 75 min after photoimmunotherapy and did not change for 24 h. In contrast, no significant changes were demonstrated in nontreated tumors. None of tumors changed size within 24 h after photoimmunotherapy, although diffuse necrosis was observed in photoimmunotherapy-treated tumors. Immediate cytotoxic effects induced by photoimmunotherapy were clearly detected by decreased glucose uptake using (18)F-FDG PET even before changes in tumor size became evident. (18)F-FDG allows the clinical assessment of the therapeutic effects of photoimmunotherapy earlier than anatomic methods that rely on tumor size.

Development, characterization, and photocytotoxicity assessment on human melanoma of chloroaluminum phthalocyanine nanocapsules

In this work we have developed nanocapsules containing chloroaluminum phthalocyanine (ClAlPc) and assessed their phototoxic action on WM1552C, WM278, and WM1617 human melanoma cell lines. The ClAlPc-loaded nanocapsules were prepared by the nanoprecipitation method and optimized by means of a 23 full factorial design. The ClAlPc nanocapsules were characterized by particle size and distribution, zeta potential, morphology, encapsulation efficiency, singlet oxygen production, stability, and phototoxic action on melanoma cells. Both the development and optimization studies revealed that stable colloidal formulations could be obtained by using 1.75% (w/v) soybean lecithin, 1.25% (w/v) Poloxamer 188, 2.5% (v/v) soybean oil, and 0.75% (w/v) poly(D,L-lactide-co-glycolide). The nanocapsules had a mean diameter of 230nm, homogeneous size distribution (polydispersity index<0.3), and negative zeta potential (about −30mV). Their morphology was spherical, with evident polymer membrane coating droplet. The encapsulation efficiency was 70%, as expected for hydrophobic drugs, and the nanoencapsulated ClAlPc was able to produce high singlet oxygen quantum yield. ClAlPc nanocapsules exhibited good physical stability over a 12-month period. WM1552C primary melanoma cells were more sensitive (p<0.05) to the phototoxic effect elicited by ClAlPc nanocapsules (0.3μgml−1) under light irradiation at 20mJcm−2. On the other hand, the cell survival percentage for all the melanoma cell lines treated with the highest light dose (150mJcm−2) was lower than 10%. In summary, ClAlPc nanoencapsulation could enable application of this hydrophobic photosensitizer in the treatment of malignant melanoma with the use of both low sensitizer drug concentration and light dose.

In vitro evaluation of combined hyperthermia and photodynamic effects using magnetoliposomes loaded with cucurbit uril zinc phthalocyanine complex on melanoma

The aims of this study were two fold; to develop magnetoliposomes (MLs) loaded with zinc phthalocyanine (ZnPc) complexed with cucurbituril (CB) (CB:ZnPc-MLs) and to evaluate their in vitro photodynamic (PD) and/or hyperthermia (HT) effects while using melanoma cells (B16-F10) as model. The liposomal formulations were characterized by both average diameter and zeta potential. The vesicle average size ranged from 150 to 200nm and the polydispersity index (PdI) from 0.093 to 0.230. The zeta potential was significantly positive with values between 48 and 57mV. The cell viability (CV) after PD and HT treatments was assessed by colorimetric MTT method. Melanoma cells were initially treated with the liposome formulation without light and magnetic field application, revealing cell viability not different from the control cells (p>0.05). Photodynamic and hyperthermia assays were also applied separately, demonstrating that PD is more effective than HT in reducing the CV of the neoplastic cells. Combined application of both PD and HT treatments was even more effective in reducing the CV of B16-F10 cells. At the highest light dose (2J/cm2) and under magnetic field activation the CV was about half than PD applied alone. Therefore, the use of the photosensitizer-loaded magnetoliposome for combined photodynamic therapy (PDT) and magnetohyperthermia (MHT) application can be considered as a potential tool to treat malignant melanoma.

Abstract 4830: Target-molecular specific near infrared cancer photoimmunotherapy: Detection, treatment, and monitoring of tumors with a theranostic fluorescent probe

Abstract Targeted molecular therapies of cancer offer the possibility of effective tumor control with a minimum of side effects. However, to date this strategy has had modest results for most tumors. Antibody-based therapies have been among the more successful molecular approaches. The concept of using the antibody as a vector for additional therapy has been attempted with toxins and radio-isotopes with mixed results. Herein we describe an antibody conjugate with a photosensitive near infrared (NIR) phthalocyanine dye, IR700, which can be used as an optical imaging agent at low doses of light but becomes a photoimmunotherapeutic (PIT) at higher doses of light. In this study, we tested the efficacy of PIT with optimized regimen using an EGFR antbody-IR700 conjugate in a xenograft tumor model. For in vitro PIT, EGFR (HER1) expressing A431 cells were incubated with panitumumab-IR700 conjugates (Pan-IR700) which were then irradiated with NIR light. For in vivo PIT, A431 xenograft tumor bearing mice injected with 100 μg of Pan-IR700 every week. Mice were then treated with NIR light on day one (50 J cm−2) and day 2 (100 J cm−2) after Pan-IR700 injection which was administered every week for up to 4 weeks. NIR fluorescence images were obtained in each mouse and the effect of the PIT was analyzed. Pan-IR700 demonstrated specific binding to target receptors on the cell membrane. Target-specific cell death could be induced only in Pan-IR700 binding cancer cells immediately after exposure to NIR light. In vitro microscopy demonstrated that NIR exposure resulted in cellular swelling, bleb formation, and rupture of vesicles indicating necrotic cell death. No phototoxicity was observed in co-cultured HER1 negative Balb3T3 cells, even when MAb-IR700 was not removed from the medium. HER1 target-specific accumulation of MAb-IR700 was observed in mouse xenograft tumors by fluorescence imaging. Repeated administration of Pan-IR700 followed by repeated irradiation with NIR light resulted in effective tumor eradication and prolonged survival compared to control mice with no significant side effects. IR700 fluorescence from the conjugate allowed for image guidance of light delivery as well as monitoring of PIT. In conclusion, Pan-IR700 PIT was effective, only when the antibody conjugates were bound to the cell membrane, but showed no phototoxicity when they were not bound or irradiated with NIR light. Target selective PIT enables selective treatment of cancer with no apparent side effects even with multiple administrations of Pan-IR700 and NIR light irradiation. PIT using MAb-IR700 is a promising theranostics for highly selective treatment of cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4830. doi:1538-7445.AM2012-4830