RESUMO
Triple-negative breast cancer (TNBC) overexpresses the Epidermal Growth Factor Receptor (EGFR), a characteristic of different types of tumors, linked to worse disease prognosis and risk of recurrence. Conventional treatments are also aggressive and can be morbid.. Therefore, t improvement and development of new methods are notorious. Photodynamic Therapy (PDT) is an effective method for treating different types of cancer by using light radiation to activate a photosensitizing agent (drug) in molecular oxygen presence, promoting cell death., Improving drug uptake in target cells could contribute to PDT efficiency. Accordingly, we developed a bifunctional nanoprobe (BN), used in PDT as a a treatment method in vivo against breast cancer. The BN uses gold nanoparticles with active targeting through the Epidermal Growth Factor (EGF) protein and Chlorine e6 (Ce6) carriers. We evaluated the therapeutic efficacy of in vivo xenograft in 4 groups: Saline, BN, Ce6+PDT, and BN+PDT. As a result, we observed that the BN+PDT group exhibited an excellent effect with greater selectivity to tumor tissue and tissue damage when compared to the Saline, BN, and Ce6+PDT groups. The results indicate a potential impact on breast cancer treatment in vivo.. In conclusion, our data propose that the BN developed heightened PDT efficacy through cellular DNA repair effects and tumor microenvironment.
Assuntos
Clorofilídeos , Nanopartículas Metálicas , Nanopartículas , Fotoquimioterapia , Porfirinas , Neoplasias de Mama Triplo Negativas , Linhagem Celular Tumoral , Ouro/farmacologia , Ouro/uso terapêutico , Xenoenxertos , Humanos , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Microambiente TumoralRESUMO
The identification of substances that prevent or minimize the detrimental effects of ionizing radiation is an essential undertaking. The aim of this paper was to evaluate and compare the radioprotective potential of chlorophyllin, protoporphyrin and bilirubin, with amifostine®, an US Food & Drug Administration approved radioprotector Using the somatic mutation and recombination assay in the Drosophila melanogaster wing, it was found that pretreatment (1-9â¯h) with any of the porphyrins or amifostine® alone, did not affect the larva-adult viability or the basal frequency of mutation. However, they were associated with significant reductions in frequency of somatic mutation and recombination compared with the gamma-irradiated (20â¯Gy) control as follows: bilirubin (69.3 %)> chlorophyllin (40.0 %)> protoporphyrin (39.0 %)> amifostine® (19.7 %). Bilirubin also caused a 16 % increase in larva-adult viability with 3â¯h of pretreatment respect to percentage induced in 20â¯Gy control group. Whilst amifostine® was associated with lower genetic damage after pre-treatment of 1 and 3â¯h, this did not attain significance. These findings suggest that the tested porphyrins may have some potential as radioprotectant agents.
Assuntos
Amifostina/farmacologia , Bilirrubina/farmacologia , Clorofilídeos/farmacologia , Drosophila melanogaster/efeitos dos fármacos , Drosophila melanogaster/efeitos da radiação , Protoporfirinas/farmacologia , Protetores contra Radiação/farmacologia , Animais , Drosophila melanogaster/genética , Feminino , Masculino , Testes de Mutagenicidade , Mutação/efeitos dos fármacos , Recombinação Genética/efeitos dos fármacos , Asas de Animais/efeitos dos fármacos , Asas de Animais/efeitos da radiaçãoRESUMO
The antimicrobial photodynamic therapy (aPDT) has stood out as an alternative and promising method of disinfection and has been exploited for the treatment of oral bacteria. In this study, we evaluate in vitro the action of aPDT, mediated by methylene blue, chlorin-e6, and curcumin against clinical subgingival plaques that were resistant to metronidazole. The sensitivity profile of the samples to metronidazole was analyzed by the agar dilution method. Cell viability in the planktonic and biofilm phase was assessed by CFU / mL. The composition of the biofilm was evaluated by the checkboard DNA-DNA Hibrydization technique. Photosensitizers internalization was qualitatively assessed by confocal fluorescence microscopy (CLSM). The aPDT mediated by the three photosensitizers tested was able to reduce the totality of the planktonic microbial load and partially reduce the biofilm samples. The analysis performed by CLSM showed that the photosensitizers used in the application of aPDT were able to permeate the interior of the biofilm. The aPDT has been shown to be useful in a supportive and effective approach to the treatment of periodontal disease.
Assuntos
Antibacterianos/farmacologia , Placa Dentária/microbiologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Metronidazol/farmacologia , Fotoquimioterapia/métodos , Biofilmes/efeitos dos fármacos , Clorofilídeos , Curcumina/farmacologia , Humanos , Azul de Metileno/farmacologia , Testes de Sensibilidade Microbiana , Doenças Periodontais/tratamento farmacológico , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Porfirinas/farmacologiaRESUMO
The self-produced extracellular polymeric matrix of biofilms renders them difficult to eliminate once they are established. This makes the inhibition of biofilm formation key to successful treatment of biofilm infection. Antimicrobial photodynamic therapy (aPDT) and antimicrobial peptides offer a new approach as antibiofilm strategies. In this study sub-lethal doses of aPDT (with chlorin-e6 (Ce6-PDT) or methylene blue (MB-PDT)) and the peptides AU (aurein 1.2 monomer) or (AU)2K (aurein 1.2 C-terminal dimer) were combined to evaluate their ability to prevent biofilm development by Enterococcus faecalis. Biofilm formation was assessed by resazurin reduction, confocal microscopy, and infrared spectroscopy. All treatments successfully prevented biofilm development. The (AU)2K dimer had a stronger effect, both alone and combined with aPDT, while the monomer AU had significant activity when combined with Ce6-PDT. Additionally, it is shown that the peptides bind to the lipoteichoic acid of the E. faecalis cell wall, pointing to a possible key mechanism of biofilm inhibition.
Assuntos
Antibacterianos/química , Biofilmes , Peptídeos/química , Fármacos Fotossensibilizantes/química , Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Clorofilídeos , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/fisiologia , Peptídeos/farmacologia , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Porfirinas/químicaRESUMO
Bacterial resistance to available antibiotics nowadays is a global threat leading researchers around the world to study new treatment modalities for infections. Antimicrobial photodynamic therapy (aPDT) has been considered an effective and promising therapeutic alternative in this scenario. Briefly, this therapy is based on the activation of a non-toxic photosensitizing agent, known as photosensitizer (PS), by light at a specific wavelength generating cytotoxic singlet oxygen and free radicals. Virtually all studies related to aPDT involve a huge screening to identify ideal PS concentration and light dose combinations, a laborious and time-consuming process that is hardly disclosed in the literature. Herein, we describe an antimicrobial Photodynamic Therapy (aPDT) study against Enterococcus faecalis and Propionibacterium acnes employing methylene blue, chlorin-e6 or curcumin as PS. Similarities and discrepancies between the two bacterial species were pointed out in an attempt to speed up and facilitate futures studies against those clinical relevant strains. Susceptibility tests were performed by the broth microdilution method. Our results demonstrate that aPDT mediated by the three above-mentioned PS was effective in eliminating both gram-positive bacteria, although P. acnes showed remarkably higher susceptibility to aPDT when compared to E. faecalis. PS uptake assays revealed that P. acnes is 80 times more efficient than E. faecalis in internalizing all three PS molecules. Our results evidence that the cell wall structure is not a limiting feature when predicting bacterial susceptibility to aPDT treatment.
Assuntos
Anti-Infecciosos/farmacologia , Enterococcus faecalis/efeitos dos fármacos , Fármacos Fotossensibilizantes/farmacologia , Propionibacterium acnes/efeitos dos fármacos , Anti-Infecciosos/química , Clorofilídeos , Curcumina/química , Curcumina/farmacologia , Enterococcus faecalis/efeitos da radiação , Luz , Azul de Metileno/química , Azul de Metileno/farmacologia , Fármacos Fotossensibilizantes/química , Porfirinas/química , Porfirinas/farmacologia , Propionibacterium acnes/efeitos da radiação , Oxigênio Singlete/química , Oxigênio Singlete/metabolismoRESUMO
Candida albicans is an opportunistic fungal producing both superficial and systemic infections in immunocompromised patients. Furthermore, it has been described an increase in the frequency of infections which have become refractory to standard antifungal therapy. Photodynamic antimicrobial chemotherapy (PACT) is a potential antimicrobial therapy that combines visible light and a nontoxic dye, known as a photosensitizer, producing reactive oxygen species (ROS) that can kill the treated cells. The objective of this study was to investigate the effects of PACT, using chlorin e6, as a photosensitizer on C. albicans. In this work, we studied the effect of PACT on both cell growth and biofilm formation by C. albicans. In addition, both ROS production and cell permeability were determined after PACT. PACT inhibited both growth and biofilm formation by C. albicans. We have also observed that PACT increased both ROS production (six times) and cell membrane permeability (five times) in C. albicans. PACT decreased both cell growth and biofilm development. The effect of PACT using chlorin e6 on C. albicans could be associated with an increase in ROS production, which could increase cell permeability, producing permanent damage to the cell membranes, leading to the cell death.
Assuntos
Anti-Infecciosos/farmacologia , Biofilmes/crescimento & desenvolvimento , Candida albicans/fisiologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Permeabilidade da Membrana Celular/efeitos da radiação , Fármacos Fotossensibilizantes/farmacologia , Porfirinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Biofilmes/efeitos dos fármacos , Biofilmes/efeitos da radiação , Candida albicans/efeitos dos fármacos , Clorofilídeos , Testes de Sensibilidade MicrobianaRESUMO
Conformation of protein is vital to its function, but may get affected when processing to manufacture products. It is therefore important to understand structural changes during each step of production. In this study, we investigate secondary structure changes in the targeting protein Epidermal Growth Factor (EGF) during synthesis of theranostic bifunctional nanoparticle, devised for Photodynamic therapy of breast cancer. We acquired FTIR spectra of EGF; unconjugated, post treatment with α-lipoic acid, attached to gold nanoparticle, and bound to the bifunctional nanoprobe. We observed decreasing disordered structures and turns, and increasing loops, as the synthesis process progressed. There was an overall increase in ß-sheets in final product compared to pure EGF, but this increase was not linear and fluctuated. Previous crystal structure studies on EGF-EGFR complex have shown loops and ß-sheets to be important in the binding interaction. Since our study found increase in these structures in the final product, no adverse effect on binding function of EGF was expected. This was confirmed by functional assays. Such studies may help modify synthesis procedures, and thus secondary structures of proteins, enabling increased functionality and optimum results.
Assuntos
Fator de Crescimento Epidérmico/química , Nanopartículas Metálicas/química , Espectroscopia de Infravermelho com Transformada de Fourier , Neoplasias da Mama , Linhagem Celular Tumoral , Clorofilídeos , Fator de Crescimento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Feminino , Ouro , Humanos , Proteínas de Neoplasias/metabolismo , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Porfirinas/farmacologia , Ligação Proteica , Estrutura Secundária de Proteína , Ácido Tióctico/farmacologiaRESUMO
The use of eosin methylene blue according to Giemsa as photosensitizer is presented for the first time in this paper. The present study evaluated the potential application of chlorophyllin sodium copper salt (CuChlNa) and eosin methylene blue according to Giemsa (EMB) as antimicrobial photosensitizers (aPS) for photodynamic inactivation (PDI) of Staphylococcus aureus (gram-positive) and Escherichia coli (gram-negative) bacteria. The experiments were performed using S. aureus stain ATCC 25923 and E. coli ATCC 25922 in which five aPS concentrations (0.0, 1.0, 2.5, 5.0, 10.0, and 20.0 µM for S. aureus and 0.0, 5.0, 10.0, 20.0, 40.0, and 50.0 µM for E. coli) were prepared and added in 2 mL of a saline solution containing the bacterial inoculum. After aPS incubation, the samples were divided into two groups, one kept in the dark and another submitted to the illumination. Then, the bacterial inactivation was determined 18 h after the incubation at 37 °C by counting the colony-forming units (CFU). The results revealed that both EMB and CuChlNa can be used as aPS for the photoinactivation of S. aureus, while only EMB was able to photoinactivate E. coli. Nevertheless, a more complex experimental setup was needed for photoinactivation of E. coli. The data showed that EMB and CuChlNa presented similar photoinactivation effects on S. aureus, in which bacterial growth was completely inhibited at photosensitizer (PS) concentrations over 5 µM, when samples were previously incubated for 30 min and irradiated by a light dose of 30 J cm-2 as a result of an illumination of 1 h at 8.3 mW cm-2 by using a red light at 625 nm with a 1 cm beam diameter and output power of 6.5 mW. In the case of E. coli, bacterial growth was completely inhibited only when combining a PS incubation period of 120 min with concentrations over 20 µM.
Assuntos
Clorofilídeos/farmacologia , Amarelo de Eosina-(YS)/farmacologia , Escherichia coli/efeitos da radiação , Luz , Azul de Metileno/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Staphylococcus aureus/efeitos da radiação , Animais , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Camundongos , Células NIH 3T3 , Fármacos Fotossensibilizantes/farmacologia , Análise Espectral , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/crescimento & desenvolvimentoRESUMO
Photodynamic therapy is an alternative treatment for cancer based on cellular uptake of a photosensitizer, illuminated with an appropriate wavelength in the presence of oxygen. A cascade of reactions generates reactive oxygen species leading to cell death. Using carbodiimide chemistry, chlorin e6 (Ce6) was covalently bonded to thiourea, and (via the sulphur end group) to gold nanoparticles (AuNPs), forming the Ce6-AuNP complex. Ce6 absorbs in the range 650-680nm, where the coefficient of biological tissue absorption is low (part of the therapeutic window), which is ideal for biological application. Transmission Electron Microscopy, UV-vis spectroscopy, Fourier transform Infrared Spectroscopy and Zeta potential measurements were completed to characterize the Ce6-AuNP complex. The bare AuNPs have an average diameter of 18±4nm. A line of human breast carcinoma cells (MDA-MB-468) was used to determine whether Ce6 functionalization to AuNPs potentiate its activity. Trypan blue assays were used to assess cell viability. In the absence of light, Ce6 either alone or bounded to AuNPs was not cytotoxic. When irradiated at 660nm, the cytotoxicity of Ce6-AuNP was higher than Ce6 alone for MDA-MB-468 cells using 4h incubation. AuNPs without Ce6 showed no cytotoxic.
Assuntos
Ouro/química , Ouro/uso terapêutico , Nanopartículas Metálicas/uso terapêutico , Neoplasias Experimentais/tratamento farmacológico , Fotoquimioterapia/métodos , Porfirinas/administração & dosagem , Porfirinas/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Clorofilídeos , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Humanos , Nanopartículas Metálicas/química , Neoplasias Experimentais/patologia , Fármacos Fotossensibilizantes/administração & dosagem , Fármacos Fotossensibilizantes/síntese química , Resultado do TratamentoRESUMO
Fasciolosis is a food borne zoonosis, caused by the digenetic trematode Fasciola. Freshwater lymnaeid snails are the intermediate host of the trematodes. Chlorophyllin, a semi-synthetic derivative of chlorophyll and its formulations obtained from freeze dried cow urine (FCU) had their toxicity tested against redia and cercaria larvae of F. gigantica. The larvicidal activity of chlorophyllin and its formulations were found to depend on both, time and concentration used against the larvae. Toxicity of chlorophyllin + FCU (1:1 ratio) in sunlight against redia larva (8 h LC50: 0.03 mg/mL) was more pronounced than using just chlorophyllin (8 h LC50: 0.06 mg/mL). Toxicity of chlorophyllin + FCU in sunlight against redia (8 h LC50: 0.03 mg/mL) was higher than against cercaria (8 h LC50: 0.06 mg/mL). The larvicidal activity of chlorophyllin in sunlight (redia/cercaria larvae: 8 h LC50: 0.06 mg/mL) was more pronounced than under laboratory conditions (redia: 8 h LC50: 22.21 mg/mL/, cercaria 8 h LC50: 96.21 mg/mL). Toxicity of FCU against both larvae was lower than that of chlorophyllin and chlorophyllin + FCU. Chlorophyllin and its formulations + FCU were 357.4 to 1603.5 times more effective against redia/cercaria larvae in sunlight than under laboratory conditions. The present study has shown that chlorophyllin formulations may be used as potent larvicides against fasciolosis.