RESUMO
Phthalocyanine (Pc) 4, like many photosensitizers for photodynamic therapy (PDT), localizes to intracellular membranes, especially mitochondria. Pc 4-PDT photodamages Bcl-2 and Bcl-xL, antiapoptotic proteins interacting with the permeability transition pore complex that forms at contact sites between the inner and outer mitochondrial membranes. These complexes and the inner membrane are unique in containing the phospholipid cardiolipin. Nonyl-acridine orange (NAO) is a specific probe of cardiolipin. Here we show evidence for fluorescence resonance energy transfer from NAO to Pc 4, defining a binding site for the photosensitizer. This observation establishes an innovative tool for exploring the localization of other photosensitizers and additional fluorescent, mitochondrion-localizing drugs having appropriate spectral properties.
Assuntos
Laranja de Acridina/análogos & derivados , Cardiolipinas/metabolismo , Transferência Ressonante de Energia de Fluorescência/métodos , Indóis/metabolismo , Fármacos Fotossensibilizantes/metabolismo , Neoplasias da Próstata/metabolismo , Laranja de Acridina/química , Sítios de Ligação , Cardiolipinas/química , Corantes/química , Humanos , Indóis/química , Masculino , Microscopia Confocal , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/química , Neoplasias da Próstata/tratamento farmacológico , Espectrometria de Fluorescência , Frações Subcelulares/metabolismo , Células Tumorais CultivadasRESUMO
OBJECTIVE: The purpose of this prospective, randomized, double-blind study was to compare the anesthetic efficacy of the intraligamentary injection of 4% articaine with 1:100,000 epinephrine and of 2% lidocaine with 1:100,000 epinephrine, administered with computer-controlled local anesthetic delivery system, in mandibular posterior teeth. STUDY DESIGN: Using a crossover design, intraligamentary injections of 1.4 mL of 4% articaine with 1:100,000 epinephrine and of 1.4 mL of 2% lidocaine with 1:100,000 epinephrine were randomly administered with a computer-controlled local anesthetic delivery system, in a double-blind manner on the mesial and distal aspects of a mandibular first molar, at 2 separate appointments to 51 subjects. A pulp tester was used to test for anesthesia, in 2-minute cycles for 60 minutes, of the mandibular first and second molars and second premolar. Anesthesia was considered successful when 2 consecutive 80 readings (highest output) were obtained within 20 minutes. RESULTS: Successful pulpal anesthesia was obtained 86% of the time for the first molar using the articaine solution and 74% of the time using the lidocaine solution. There were no significant differences (P > .05) between the articaine and lidocaine solutions. The mean onset times of pulpal anesthesia for the first molar were 1.3 minutes with articaine solution and 2.2 minutes with lidocaine solution. Duration of pulpal anesthesia for the first molar was 34 minutes for the articaine solution and 31 minutes for the lidocaine solution. CONCLUSION: The efficacy of 4% articaine with 1:100,000 epinephrine was similar to the efficacy of 2% lidocaine with 1:100,000 epinephrine for intraligamentary injections.
Assuntos
Anestesia Dentária/métodos , Anestesia Local/métodos , Anestésicos Locais/administração & dosagem , Carticaína/administração & dosagem , Sistemas de Liberação de Medicamentos/instrumentação , Quimioterapia Assistida por Computador , Lidocaína/administração & dosagem , Ligamento Periodontal , Adulto , Dente Pré-Molar , Computadores , Teste da Polpa Dentária , Método Duplo-Cego , Feminino , Humanos , Injeções , Masculino , Pessoa de Meia-Idade , Dente Molar , Estudos ProspectivosRESUMO
The purpose of this prospective, randomized, double-blind study was to compare the pain of injection, heart rate increase, and postinjection pain of the intraligamentary injection of 4% articaine with 1:100,000 epinephrine and 2% lidocaine with 1:100,000 epinephrine administered with a computer-controlled local anesthetic delivery system. Using a crossover design, intraligamentary injections of 1.4 mL of 4% articaine with 1:100,000 epinephrine and 1.4 mL of 2% lidocaine with 1:100,000 epinephrine were randomly administered on the mesial and distal aspects of the mandibular first molar with a computer-controlled local anesthetic delivery system in a double-blind manner at 2 separate appointments to 51 subjects. The results demonstrated the incidence of moderate pain was 14%-27% with needle insertion, with 0%-4% reporting severe pain. For solution deposition, moderate pain was reported 8%-18% of the time, with no reports of severe pain. There were no significant differences between the articaine and lidocaine solutions. Regarding heart rate changes, neither anesthetic solution resulted in a significant increase in heart rate over baseline readings. On day 1 postinjection, there was a 31% incidence of moderate/severe pain with the articaine solution and 20% incidence of moderate/severe pain with the lidocaine solution. The moderate/severe pain ratings decreased over the next 2 days. There were no significant differences between the articaine and lidocaine solutions. We concluded that the intraligamentary injection of 4% articaine with 1:100,000 epinephrine was similar to 2% lidocaine with 1:100,000 epinephrine for injection pain and postinjection pain in the mandibular first molar when administered with a computer-controlled local anesthetic delivery system. For both anesthetic solutions, heart rate did not significantly increase with the intraligamentary injection using the computer-controlled local anesthetic system.
Assuntos
Anestesia Dentária/métodos , Anestesia Local/instrumentação , Anestésicos Locais/administração & dosagem , Carticaína/administração & dosagem , Dor Facial/etiologia , Injeções/efeitos adversos , Lidocaína/administração & dosagem , Ligamento Periodontal , Terapia Assistida por Computador/instrumentação , Adulto , Estudos Cross-Over , Método Duplo-Cego , Feminino , Frequência Cardíaca , Humanos , Masculino , Mandíbula , Pessoa de Meia-Idade , Dente Molar , Estudos ProspectivosRESUMO
Cardiolipin is a unique phospholipid of the mitochondrial inner membrane. Its peroxidation correlates with release of cytochrome c and induction of apoptosis. The phthalocyanine photosensitizer Pc 4 binds preferentially to the mitochondria and endoplasmic reticulum. Earlier Förster resonance energy transfer studies showed colocalization of Pc 4 and cardiolipin, which suggests cardiolipin as a target of photodynamic therapy (PDT) with Pc 4. Using liposomes as membrane models, we find that Pc 4 binds to cardiolipin-containing liposomes similarly to those that do not contain cardiolipin. Pc 4 binding is also studied in MCF-7c3 cells and those whose cardiolipin content was reduced by treatment with palmitate. Decreased levels of cardiolipin are quantified by thin-layer chromatography. The similar level of binding of Pc 4 to cells, irrespective of palmitate treatment, supports the lack of specificity of Pc 4 binding. Thus, factors other than cardiolipin are likely responsible for the preferential localization of Pc 4 in mitochondria. Nonetheless, cardiolipin within liposomes is readily oxidized by Pc 4 and light, yielding apparently mono- and dihydroperoxidized cardiolipin. If similar products result from exposure of cells to Pc 4-PDT, they could be part of the early events leading to apoptosis following Pc 4-PDT.
Assuntos
Cardiolipinas/química , Cardiolipinas/metabolismo , Indóis/metabolismo , Fármacos Fotossensibilizantes/metabolismo , Sítios de Ligação , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Feminino , Transferência Ressonante de Energia de Fluorescência , Humanos , Técnicas In Vitro , Lipossomos , Potencial da Membrana Mitocondrial , Membranas Mitocondriais/metabolismo , Oxirredução , Palmitatos/farmacologia , Processos Fotoquímicos , FotoquimioterapiaRESUMO
Photodynamic therapy (PDT) is emerging as a promising non-invasive treatment for cancers. PDT involves either local or systemic administration of a photosensitizing drug, which preferentially localizes within the tumor, followed by illumination of the involved organ with light, usually from a laser source. Here, we provide a selective overview of our experience with PDT at Case Western Reserve University, specifically with the silicon phthalocyanine photosensitizer Pc 4. We first review our in vitro studies evaluating the mechanism of cell killing by Pc 4-PDT. Then we briefly describe our clinical experience in a Phase I trial of Pc 4-PDT and our preliminary translational studies evaluating the mechanisms behind tumor responses. Preclinical work identified (a) cardiolipin and the anti-apoptotic proteins Bcl-2 and Bcl-xL as targets of Pc 4-PDT, (b) the intrinsic pathway of apoptosis, with the key participation of caspase-3, as a central response of many human cancer cells to Pc 4-PDT, (c) signaling pathways that could modify apoptosis, and (d) a formulation by which Pc 4 could be applied topically to human skin and penetrate at least through the basal layer of the epidermis. Clinical-translational studies enabled us to develop an immunohistochemical assay for caspase-3 activation, using biopsies from patients treated with topical Pc 4 in a Phase I PDT trial for cutaneous T-cell lymphoma. Results suggest that this assay may be used as an early biomarker of clinical response.
Assuntos
Indóis/farmacologia , Fotoquimioterapia/métodos , Animais , Apoptose/efeitos dos fármacos , Ensaios Clínicos Fase I como Assunto , Avaliação Pré-Clínica de Medicamentos , Humanos , Indóis/química , Indóis/uso terapêutico , Modelos Biológicos , Estrutura Molecular , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêuticoRESUMO
The effect of linker chain length on the energy transfer from CdSe quantum dots (QDs) to silicon phthalocyanine (Pc) photodynamic therapy agents was investigated by steady-state and femtosecond time-resolved spectroscopy with 500 nm light for the specific excitation of the QD energy donor. The conjugation between the QD and the Pc was achieved with linker chains varying from 4 to 9 bond lengths by incorporating 1-6 methylene groups into the axial ligand of the Pc. With increasing chain length, the energy-transfer efficiency increased, which appears to be opposed to a purely Förster-type resonance energy-transfer behavior that is commonly discussed for the energy transfer in QD conjugates. The obtained results provide strong evidence for a capping-layer-mediated energy transfer in the QD-based donor-acceptor conjugates.
Assuntos
Indóis/química , Pontos Quânticos , Transferência de Energia , IsoindóisRESUMO
Eleven silicon phthalocyanines which can be grouped into two homologous series [SiPc[OSi(CH3)2(CH2)(n)N(CH3)2]2, n = 1-6 (series 1), and SiPc[OSi(CH3)2(CH2)3N((CH2)(n)H)2]2, n = 1-6 (series 2)] as well as an analogous phthalocyanine, SiPc[OSi(CH3)2(CH2)3NH2]2, were synthesized. The ground state absorption spectra, the triplet state dynamics, and singlet oxygen quantum yields of 10 of these phthalocyanines were measured. All compounds displayed similar ground state absorption spectral properties in dimethylformamide solution with single Q band maxima at 668 +/- 2 nm and B band maxima at 352 +/- 1 nm. Photoexcitation of all compounds in the B bands generated the optical absorptions of the triplet states which decayed with lifetimes in the hundreds of microseconds region. Oxygen quenching bimolecular rate constants near 2 x 10(9) M(-1) s(-1) were measured, indicating that energy transfer to oxygen was exergonic. Singlet oxygen quantum yields, phi(delta), were measured, and those phthalocyanines in which the axial ligands are terminated by dimethylamine residues at the end of alkyl chains having four or more methylene links exhibited yields near > or = 0.35. Others gave singlet oxygen quantum yields near 0.2, and still others showed singlet oxygen yields of <0.1. The reduced singlet oxygen yields are probably caused by a charge transfer quenching of the 1pi,pi* state of the phthalocyanine by interaction with the lone pair electrons on the nitrogen atoms of the amine termini. In some cases, these can approach and interact with the electronically excited pi-framework, owing to diffusive motions of the flexible oligo-methylene tether.
Assuntos
Aminas/química , Indóis/síntese química , Compostos de Organossilício/síntese química , Siloxanas/química , Fluorescência , Indóis/química , Ligantes , Estrutura Molecular , Compostos de Organossilício/química , FotoquímicaRESUMO
BACKGROUND AND OBJECTIVES: As a potential therapy for malignant glioma, we tested the phthalocyanine photosensitizer Pc 4 for: (1) rapid clearance from the vasculature, (2) specificity for glioma, and (3) tumoricidal photosensitizing capability. STUDY DESIGN/MATERIALS AND METHODS: Parenchymal injection of U87 cells into athymic rat brains (N = 100) was followed after 12 days by tail vein injection of 0.5 mg/kg Pc 4. After 1 day, the tumor was illuminated with either 5 (N = 11) or 30 (N = 16) J/cm(2) red light at 672 nm. Sacrifice was 1 day later. The brains from these 27 animals underwent H&E (necrosis) and TUNEL assay (apoptosis) histology. Pc 4 concentration of explanted brains and tumors (N = 16), and all blood samples (N = 52) were determined by HPLC-MS 1 day post Pc 4 administration. RESULTS: Tumor-specific apoptosis was almost uniformly seen; however, necrosis was found mostly in the high-light-dose group. Pc 4 concentration in bulk tumor averaged 3.8 times greater than in normal brain. CONCLUSIONS: These results warrant expanding this pre-clinical study to seek effective baseline Pc 4 drug- and light-doses and infusion-to-photoirradiation timing that would be necessary for a Pc 4-mediated PDT clinical trial for glioma patients.
Assuntos
Neoplasias Encefálicas/tratamento farmacológico , Glioma/tratamento farmacológico , Indóis/uso terapêutico , Fotoquimioterapia , Fármacos Fotossensibilizantes/uso terapêutico , Animais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Glioma/metabolismo , Glioma/patologia , Humanos , Indóis/administração & dosagem , Indóis/farmacocinética , Injeções Intravenosas , Fármacos Fotossensibilizantes/administração & dosagem , Fármacos Fotossensibilizantes/farmacocinética , Ratos , Ratos NusRESUMO
STATEMENT OF PROBLEM: The introduction of packable composite has expanded the choices of materials for the restoration of posterior teeth. Few independent studies are available on the fracture toughness (K(IC)) of the presently available packable composites compared with more conventional composite alternatives. PURPOSE: This investigation evaluated the relative fracture toughness of 3 packable composites, 2 conventional composites, and 1 laboratory-processed composite. MATERIALS AND METHODS: Six composite materials were tested in this study. These included: 3 packable composites (Alert, SureFil and Solitaire), 2 conventional composites (Herculite and Heliomolar), and 1 laboratory-processed composite (Belleglass). K(IC) was determined by preparing 8 mini-compact test specimens (8.2 mm diameter x 2 mm thickness) for each composite in a polytetrafluoroethylene split-mold with introduced precracks created with a razor blade. Specimens were stored in distilled water at 37 degrees +/- 2 degrees C for 7 days. Testing was performed on a universal testing machine at a displacement rate of 0.5 mm/min until fracture. Analysis of variance (P<.0001) and Ryan-Einot-Gabriel-Welsch multiple range tests (P<.05) were performed on all data. RESULTS: The mean fracture toughness of Alert (1.57 Mpa x m(1/2)) was significantly greater than any of the other composites tested. Solitaire, a packable composite, exhibited a mean fracture toughness (0.67 MPa x m(1/2)) that was significantly lower than any of the other materials tested. No significant difference was noted between Belleglass (1.27 MPa x m(1/2)), SureFil (1.25 MPa x m(1/2)) and Herculite (1.16 MPa x m(1/2)). CONCLUSION: Within the limitations of this study, the glass fiber-reinforced packable composite exhibited improved fracture toughness when compared with the other composite materials tested.