chlorophyll-a and pyropheophorbide-a

The literature on the synthesis and the biological properties of boron-containing chlorins and phthalocyanines is reviewed. A series of homologous derivatives of pyropheophorbide A is described. The compounds contain the B(12)H(11)SH(2-) cluster attached to the single carboxyl group and vary in the length of the alkyl chain (methyl, propyl, pentyl, heptyl and nonyl) attached via an ether linkage to the former vinyl group. Cellular uptake was found for all derivatives except the nonyl sidechain. The compounds were moderately cell-toxic. Localization in lysosomes could be excluded; the compounds localized probably in the mitochondria.

Topics: Animals; Antineoplastic Agents; Boron Compounds; Boron Neutron Capture Therapy; Chlorophyll; Indoles; Isoindoles; Porphyrins; Pyrazines

Pyropheophorbide

Topics: Chlorophyll; DNA; G-Quadruplexes; Nucleic Acids; Photochemotherapy; Photosensitizing Agents; Singlet Oxygen

The wild bitter gourd (WBG) is a commonly consumed vegetable in Asia that has antioxidant and hypoglycemic properties. The present study aimed to investigate the anti-adipogenic activities of isolated compounds from WBG on 8-day differentiated cultures of 3 T3-L1 adipocytes that were then stained with Oil Red O (ORO) or diamidino-2-phenylindole (DAPI).. ORO stains of the methanol extracts of de-seeded HM86 cultivar of WBG (WBG-M) and the ethyl acetate fractions (WBG-M-EA) showed anti-adipogenic activities against differentiated adipocytes. Two chlorophyll-degraded compounds, pheophorbide a (1) and pyropheophorbide a (2), were isolated from WBG-M-EA. Treatments with 1 (5, 10, and 20 μmol L. Both 1 and 2 showed anti-adipogenic activities in cell models. These chlorophyll-degraded compounds commonly exist in several vegetables during storage or edible seaweeds, which will provide resources for further investigations aiming to test anti-obesity in animal studies. © 2022 Society of Chemical Industry.

Topics: Animals; Antioxidants; Chlorophyll; Hypoglycemic Agents; Lactate Dehydrogenases; Lipids; Methanol; Momordica charantia; Plant Extracts

Inspired by natural saccharide-protein complexes, a stimuli-responsive biodegradable and branched glycopolymer-pyropheophorbide-a (Ppa) conjugate (BSP) with saccharide units for cancer therapy is constructed. A linear glycopolymeric conjugate (LSP), a branched glycopolymeric conjugate (BShP) from Ppa with long carbon chains, and a branched conjugate (BHSP) based on poly[N-(2-hydroxypropyl) methacrylamide] (polyHPMA) without saccharide units are prepared as controls. Through structure-activity relationship studies, BSP with a 3D network structure forms stable nanostructures via weak intermolecular interactions, regulating the stacking state of Ppa to improve the singlet oxygen quantum yield and the corresponding photodynamic therapy (PDT) effect. BSP shows high loading of olaparib, and are further coated with tumor cell membranes, resulting in a biomimetic nanomedicine (CM-BSPO). CM-BSPO shows highly efficient tumor targeting and cellular internalization properties. The engulfment of CM-BSPO accompanied with laser irradiation results in a prominent antitumor effect, evidenced by disruption of cell cycles in tumor cells, increased apoptosis and DNA damage, and subsequent inhibition of repair for damaged DNA. The mechanism for the synergistic effect from PDT and olaparib is unveiled at the genetic and protein level through transcriptome analysis. Overall, this biodegradable and branched glycopolymer-drug conjugate could be effectively optimized as a biomimetic nanomedicine for cancer therapy.

Topics: Animals; Antineoplastic Agents; Apoptosis; Biomimetic Materials; Cell Line, Tumor; Chlorophyll; DNA Damage; Drug Carriers; Genomic Instability; Humans; Light; Mice; Nanomedicine; Nanostructures; Neoplasms; Photochemotherapy; Phthalazines; Piperazines; Polymethacrylic Acids; Polysaccharides; Reactive Oxygen Species

Supramolecular self-assemblies of dendritic peptides with well-organized nanostructures have great potential as multifunctional biomaterials, yet the complex self-assembly mechanism hampers their wide exploration. Herein, a self-stabilized supramolecular assembly (SSA) constructed from a PEGylated dendritic peptide conjugate (PEG-dendritic peptide-pyropheophorbide a, PDPP), for augmenting tumor retention and therapy, is reported. The supramolecular self-assembly process of PDPP is concentration-dependent with multiple morphologies. By tailoring the concentration of PDPP, the supramolecular self-assembly is driven by noncovalent interactions to form a variety of SSAs (unimolecular micelles, oligomeric aggregates, and multi-aggregates) with different sizes from nanometer to micrometer. SSAs at 100 nm with a spherical shape possess extremely high stability to prolong blood circulation about 4.8-fold higher than pyropheophorbide a (Ppa), and enhance tumor retention about eight-fold higher than Ppa on day 5 after injection, which leads to greatly boosting the in vivo photodynamic therapeutic efficiency. RNA-seq demonstrates that these effects of SSAs are related to the inhibition of MET-PI3K-Akt pathway. Overall, the supramolecular self-assembly mechanism for the synthetic PEGylated dendritic peptide conjugate sheds new light on the development of supramolecular assemblies for tumor therapy.

Topics: Animals; Breast Neoplasms; Chlorophyll; Disease Models, Animal; Mice; Nanoparticles; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols

Biomarker-activatable theranostic formulations offer the potential for removing specific tumors with a high diagnostic accuracy and a significant pharmacological effect. Herein, we developed a novel activatable theranostic nanoformulation UAS-PD [upconversion nanophosphor (UCNP)-aptamer/ssDNA-pyropheophorbide-a (PPA)-doxyrubicin (DOX)], which can recognize specific cancer cells with sensitivity and trigger the localized photodynamic destruction and enhanced chemotherapy. UAS-PD was constructed by the conjugation of UCNPs and aptamer probes containing the photosensitizer PPA and the chemotherapeutic drug DOX. When cancer cells are present, the UAS-PD specifically binds to PTK7, an overexpressed protein present on the surface of cancer cells, through conformational recombination of the aptamer structure and switches its upconversion luminescence from 655 to 540 nm. This long-lived ratiometric optical signal provides an ultrasensitive detection limit as low as 3.9 nM for PTK7. Changes in the conformation of UAS-PD can also induce PPA to approach UCNPs, which can produce cytotoxic singlet oxygens under near-infrared excitation to destroy the cell membrane and enhance its permeability for the simultaneously released DOX that targets cellular DNA degradation, which results in a highly effective tumor-killing effect by synergistic extra-intracellular sequential damage.

Topics: Animals; Antineoplastic Agents; Aptamers, Nucleotide; Cell Adhesion Molecules; Cell Line, Tumor; Chlorophyll; Doxorubicin; Female; Humans; Light; Mice, Nude; Nanoparticles; Neoplasms; Photochemotherapy; Photosensitizing Agents; Receptor Protein-Tyrosine Kinases; Singlet Oxygen; Theranostic Nanomedicine; Xenograft Model Antitumor Assays

To deliver photosensitizers with PEGylated heparin (HP) into tumor cells for photodynamic therapy, we prepared two polyethylene glycol (PEG)-functionalized HP-based polymers conjugated with pyropheophorbide-a (Ppa): a non-GSH-responsive nanoagent (HP-Ppa-mPEG) with the mPEG moiety chemically attached to HP directly; and a GSH-responsive nanoagent (HP-Ppa-SS-mPEG) with the mPEG moiety conjugated to HP via a disulfide linkage. The Ppa-functionalized HP without PEGylation (HP-Ppa) was designed as another control. These amphiphilic polymers could aggregate into nanoparticles. Cellular uptake of three nanoparticles by 4T1 cells led to abundant production of reactive oxygen species after irradiation by a 660 nm laser, inducing cell apoptosis. HP-Ppa-SS-mPEG was found to achieve the highest tumor accumulation, the longest retention time and the best penetration into tumor tissues, resulting in the highest in vivo anticancer efficacy with 94.3 % tumor growth inhibition rate, suggesting that tumor microenvironment-responsive PEGylated HP-based nanomedicines may act as efficient anticancer agents.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Survival; Chlorophyll; Female; Heparin; Lasers; Light; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Nanoconjugates; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols; Reactive Oxygen Species; Spheroids, Cellular; Treatment Outcome; Tumor Burden; Tumor Microenvironment

Antimicrobial peptides (AMPs) are originally developed for anti-infective treatments. Because of their membrane-lytic property, AMPs have been considered as candidates of antitumor agents for a long time. However, their antitumor applications are mainly hampered by fast renal clearance and high systemic toxicities. This study proposes a strategy aiming at addressing these two issues by conjugating AMPs with porphyrins, which bind to albumin increasing AMPs' resistance against renal clearance and thus enhancing their antitumor efficacies. Porphyrins' photodynamic properties can further augment AMPs' antitumor effects. In addition, circulating with albumin ameliorates AMPs' systemic toxicities, i.e. hemolysis and organ dysfunctions. As an example, we conjugated an AMP, K6L9, with pyropheophorbide-a (PPA) leading to a conjugate of PPA-K6L9. PPA-K6L9 bound to albumin with a K

Topics: Albumins; Animals; Antineoplastic Agents; Binding Sites; Cell Proliferation; Cell Survival; Chlorophyll; Drug Screening Assays, Antitumor; Female; Humans; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Pore Forming Cytotoxic Proteins; Porphyrins; Tumor Cells, Cultured

Topics: Adult; Aged; Aged, 80 and over; Air Pollutants; Air Pollution; Animals; Anti-Bacterial Agents; Anti-Infective Agents; Anti-Inflammatory Agents; Antibodies, Monoclonal, Humanized; Antigens, Surface; Antineoplastic Agents; Antioxidants; Antiviral Agents; Aporphines; Atherosclerosis; Benzoyl Peroxide; beta Catenin; Biofilms; Biomarkers; Brain; Cannabis; Carcinoma, Squamous Cell; Case-Control Studies; CD4 Lymphocyte Count; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Line; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Child; China; Chlorides; Chlorophyll; Cholesterol, LDL; Coinfection; Corylus; Cross-Sectional Studies; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Developmental Disabilities; Disease Models, Animal; Drug Evaluation, Preclinical; Drug Screening Assays, Antitumor; Electroencephalography; Environmental Exposure; Enzyme Inhibitors; Epilepsy, Generalized; Ethnicity; Female; Fertilization in Vitro; Fluorescent Dyes; Follow-Up Studies; Forecasting; Glutamate Carboxypeptidase II; Glycine; Half-Life; Head and Neck Neoplasms; Health Communication; Heart Ventricles; Hepacivirus; Hepatitis C; Heterosexuality; HIV Infections; Humans; Hypercholesterolemia; Immunoassay; Inhalation Exposure; Isocitrate Dehydrogenase; Laryngeal Neoplasms; Ligands; Light; Lipopolysaccharide Receptors; Liver Cirrhosis; Lung; Lung Neoplasms; Magnetic Resonance Imaging, Cine; Male; Maternal Age; Mechanical Phenomena; Mice; Mice, Nude; Mice, SCID; Microglia; MicroRNAs; Microscopy, Fluorescence; Microsomes, Liver; Middle Aged; Minority Groups; Mitochondrial Membrane Transport Proteins; Models, Biological; Molecular Structure; Molecular Weight; Monte Carlo Method; Muscle Hypotonia; Mutagenesis, Site-Directed; Mutation, Missense; Natriuretic Peptide, Brain; Neoplasms; Nickel; Nitric Oxide; Optical Imaging; Oxides; Particle Size; Particulate Matter; PCSK9 Inhibitors; Peptide Fragments; Phenotype; Photochemotherapy; Photosensitizing Agents; Phytochemicals; Piper; Placenta Growth Factor; Plant Extracts; Plant Leaves; Plant Stems; Platinum; Point-of-Care Testing; Population Surveillance; Postpartum Period; Pregnancy; Pregnancy, Twin; Prevalence; Prospective Studies; Prostatic Neoplasms; Pseudomonas aeruginosa; Pyridines; Pyridones; Racial Groups; Rats; Respiratory Physiological Phenomena; Retrospective Studies; Risk Factors; RNA, Long Noncoding; Semiconductors; Sexual and Gender Minorities; Sexual Behavior; Social Media; Sodium; Solubility; Stereoisomerism; Stochastic Processes; Structure-Activity Relationship; Substance-Related Disorders; Sustained Virologic Response; Sweat; Temperature; Time Factors; Tissue Distribution; Titanium; Transplantation, Heterologous; Tumor Cells, Cultured; Tungsten; Tyramine; United States; Up-Regulation; Ventricular Dysfunction, Left; Ventricular Function, Left; Veterans; Xenograft Model Antitumor Assays; Young Adult

Photodynamic therapy (PDT) has gained much attention in tumor therapy because of its special advantages. PDT heavily depends on the oxygen, yet the tumor microenvironment (TME) is a hypoxic and acid milieu, which weakens the PDT effect. Based on the consideration that the TME deteriorated by the PDT oxygen consumption could activate the hypoxic-sensitive small-molecule drug, we designed and prepared an integrated nanocomposite including zirconium ion metal organic framework (carrier), pyropheophorbide-a (PPa, photosensitizer), and 6-amino flavone (AF, hypoxic-sensitive drug), aiming to exert a cascaded PDT-chemotherapy (CT) antitumor effect and to solve the hypoxic challenge. The prepared nanocomposite showed great stability under the physiological (pH 7.4) condition and could continuously release PPa and AF under slightly acidic pH condition (pH 6.4), suggesting a tumor microenvironment responsive feature. Systematical in vitro and in vivo researches under various conditions (light, dark, hypoxic and normoxic) have showed that the obtained Zr-MOF@PPa/AF@PEG nanoparticles (NPs) had good biocompatibility and could achieve efficient antitumor effects based on PDT- chemotherapy (CT) cascade process. Finally, bright red fluorescence was observed in the tumor cells after internalization implying an application potential in tumor imaging.

Topics: Animals; Biocompatible Materials; Cell Line, Tumor; Cell Survival; Chlorophyll; Drug Liberation; Female; Flavonoids; Humans; Metal-Organic Frameworks; Mice; Mice, Inbred BALB C; Nanocomposites; Neoplasms; Photochemotherapy; Polyethylene Glycols; Singlet Oxygen; Theranostic Nanomedicine; Tumor Microenvironment; Zirconium

Nanocarriers are employed to deliver photosensitizers for photodynamic therapy (PDT) through the enhanced penetration and retention effect, but disadvantages including the premature leakage and non-selective release of photosensitizers still exist. Herein, we report a

Topics: Animals; Cell Line, Tumor; Cell Survival; Chlorophyll; Infrared Rays; Melanoma, Experimental; Mice; Microscopy, Confocal; Nanoparticles; Particle Size; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols; Polymers; Porphyrins; Positron-Emission Tomography; Singlet Oxygen

Metabolic demand of cancer is quite unique compared to normal tissues and this is an emerging hallmark of cancer, which brings a potential opportunity to discover drugs that target cancer cell metabolism. Herein, the development of a dendronized pyropheophorbide a (Ppa)-conjugated polymer (DPP) is reported, and a linear Ppa-conjugated polymer (LPP) is reported as a control. DPP is found to disturb cellular metabolism including increased energy depletion, dysfunctional H

Topics: Animals; Cell Line, Tumor; Cell Survival; Chlorophyll; Humans; Lasers; Mice; Nanostructures; Neoplasms; Photochemotherapy; Photosensitizing Agents; Polymers; Tissue Distribution; Xenograft Model Antitumor Assays

Photodynamic therapy (PDT) is a method for killing cancer cells by employing reactive singlet oxygen (

Topics: Animals; Cell Hypoxia; Cell Line, Tumor; Chlorophyll; Choroid Neoplasms; Fluorocarbons; Humans; Hyaluronic Acid; Melanoma; Mice, Inbred BALB C; Mice, Nude; Nanoparticles; Photochemotherapy; Photosensitizing Agents; Singlet Oxygen

Marine environments are known to be a new source of structurally diverse bioactive molecules. In this paper, we identified a porphyrin derivative of Pyropheophorbide a (PPa) from the mussel Musculus senhousei (M. senhousei) that showed broad anti-influenza A virus activity in vitro against a panel of influenza A viral strains. The analysis of the mechanism of action indicated that PPa functions in the early stage of virus infection by interacting with the lipid bilayer of the virion, resulting in an alteration of membrane-associated functions, thereby blocking the entry of enveloped viruses into host cells. In addition, the anti-influenza A virus activity of PPa was further assessed in mice infected with the influenza A virus. The survival rate and mean survival time of mice were apparently prolonged compared with the control group which was not treated with the drug. Therefore, PPa and its derivatives may represent lead compounds for controlling influenza A virus infection.

Topics: Animals; Antiviral Agents; Betacoronavirus; Bivalvia; Chlorophyll; Dogs; Host-Pathogen Interactions; Influenza A Virus, H1N1 Subtype; Lipid Bilayers; Madin Darby Canine Kidney Cells; Male; Mice; Orthomyxoviridae Infections; Respiratory Syncytial Viruses; SARS-CoV-2; Seafood; Survival Analysis; Virion; Virus Internalization

Pyropheophorbide a (Pyro) is a widely used photosensitizer for photodynamic therapy (PDT). However, poor water solubility, aggregation-induced fluorescence quenching, and lack of selectivity to targeted cells seriously limit its application. In this work, we prepared aptamer-Pyro conjugates (APCs) by linking Pyro to hydrophilic nucleic acid aptamer to enhance its water solubility and endow it with protein tyrosine kinase 7 (PTK7) overexpressed tumor spheroid specific targeting and penetration abilities for photodynamic therapy. The molecular conjugate was successfully synthesized and dissolved well in an aqueous solution. The APCs showed strong near-infrared fluorescence in the aqueous solution and produced singlet oxygen both in the solution and cells under laser irradiation, indicating its generation of singlet oxygen during PDT was guaranteed. Owing to the cancer cell targeting ability of the aptamer, the APCs specifically bound with PTK7 overexpressed cancerous cells and showed fluorescence signal for tumor cell imaging and diagnosis. The APCs exhibited favorable enhanced phototoxicity to target tumor cells compared with control cells. More importantly, due to the small size of the molecular conjugate, the APCs efficiently penetrated into the interior of multicellular tumor spheroids (MCTS) and caused cell damage. All these results indicated that the robust aptamer-Pyro conjugate is a promising selective tumor-targeting and penetrable molecule for cancer photodynamic therapy.

Topics: Aptamers, Peptide; Cell Adhesion Molecules; Cell Line, Tumor; Chlorophyll; Fluorescence; HeLa Cells; Hep G2 Cells; Humans; Hydrophobic and Hydrophilic Interactions; MCF-7 Cells; Photochemotherapy; Photosensitizing Agents; Singlet Oxygen; Solubility; Spheroids, Cellular

Conjugate Sn(iv)(pyropheophorbide a)dichloride-(peptide nucleic acid) catalyzes reduction of azobenzene derivatives in the presence of complementary nucleic acid (NA) upon irridiation with red light (660 nm). This is the first red light-induced NA-templated photoreduction. It is highly sensitive to single mismatches in the NA-template and can detect down to 5 nM NAs.

Topics: Azo Compounds; Base Sequence; Catalysis; Chlorophyll; Coordination Complexes; Light; Limit of Detection; Nucleic Acids; Oxidation-Reduction; Peptide Nucleic Acids; Tin

To enhance the drug delivery efficiency of hyaluronic acid (HA), we designed and prepared glycodendron and pyropheophorbide-a (Ppa)-functionalized HA (HA-Ppa-Dendron) as a nanosystem for cancer photodynamic therapy. Linear Ppa-modified HA (HA-Ppa) was also prepared as a control. Cellular uptake of both polymers by MDA-MB-231 cells led to mitochondrial dysfunction and generation of reactive oxygen species under the irradiation of a laser. Compared to the linear polymer, HA-Ppa-Dendron had higher molecular weight, a more compact nanoscale particle size, and a dendritic structure, resulting in a much longer blood circulation time and higher tumor accumulation. HA-Ppa-Dendron outperformed HA-Ppa in inhibiting cell growth, with 60 % of tumors was eradicated under laser irradiation. Tumor growth inhibition (TGI) up to 99.2 % was achieved from HA-Ppa-Dendron, which was much higher than that of HA-Ppa (50.6 %). Therefore, glycodendron-functionalized HAs by integration of HA and dendritic polymers may act as efficient anti-cancer nanomedicine.

Topics: Animals; Anthracenes; Apoptosis; Breast Neoplasms; Cell Proliferation; Chlorophyll; Female; Humans; Hyaluronic Acid; Mice; Mice, Inbred BALB C; Nanoparticles; Photochemotherapy; Photosensitizing Agents; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

A nanoemulsion with a porphyrin shell (NewPS) was created by the self-assembly of porphyrin salt around an oil core. The NewPS system has excellent colloidal stability, is amenable to different porphyrin salts and oils, and is capable of co-loading with chemotherapeutics. The porphyrin salt shell enables porphyrin-dependent optical tunability. The NewPS consisting of pyropheophorbide a mono-salt has a porphyrin shell of ordered J-aggregates, which produced a narrow, red-shifted Q-band with increased absorbance. Upon nanostructure dissociation, the fluorescence and photodynamic reactivity of the porphyrin monomers are restored. The spectrally distinct photoacoustic imaging (at 715 nm by intact NewPS) and fluorescence increase (at 671 nm by disrupted NewPS) allow the monitoring of NewPS accumulation and disruption in mice bearing KB tumors to guide effective photodynamic therapy. Substituting the oil core with Lipiodol affords additional CT contrast, whereas loading paclitaxel into NewPS facilitates drug delivery.

Topics: Animals; Chlorophyll; Drug Carriers; Emulsions; Ethiodized Oil; Humans; KB Cells; Mice, Nude; Nanoparticles; Neoplasms; Paclitaxel; Particle Size; Photoacoustic Techniques; Porphyrins; Theranostic Nanomedicine; Xenograft Model Antitumor Assays

The utilization of the light-harvesting and electron-transferring function of chlorophylls (Chls) has received attention for visible-light driven hydrogen production. In this work, a series of Chl derivatives based on pyropheophorbide-a (Pyro-a) conjugated with a viologen moiety, including a Pyro-a methyl ester directly bonded with the viologen at the 3-position 1, its 31-methylene analog 2 and Pyro-a connected with the viologen in the 17-substituent 3, were synthesized from chemical modification of naturally occurring Chl-a and characterized in terms of their photochemical and photophysical properties. As the photoexcited singlet state of the Pyro-a moiety was strongly quenched by the viologen moiety in a molecule, the effective photoinduced intramolecular electron transfer from Pyro-a to the bonded viologen moiety occurred. Moreover, these molecules were applied as a photosensitizer in the system for visible-light driven hydrogen production with platinum nanoparticles via intramolecular reduction of the bonded viologen moiety. Efficient photoreduction of external methyl viologen and successive hydrogen production on platinum nanoparticles were achieved using the synthetic conjugate of Pyro-a with the viologen moiety as a photosensitizer. In particular, effective visible-light driven hydrogen production was accomplished using 3 and platinum nanoparticles via the reduction of external methyl viologen.

Topics: Chlorophyll; Electron Transport; Hydrogen; Light; Metal Nanoparticles; Photosensitizing Agents; Platinum; Quantum Theory; Viologens

Novel nanoscale drug delivery biomaterials are of great importance for the diagnosis and treatment of different cancers. We have developed a new pegylated squalene (SQ-PEG) derivative with self-assembly properties. Supramolecular assembly with a lipophilic photosensitizer pyropheophorbide-a (Ppa) by nanoprecipitation gave nanoconstructs SQ-PEG:Ppa with an average size of 200 nm in diameter and a drug loading of 18% (w/w). The composite material demonstrates nanoscale optical properties by tight packing of Ppa within Sq-PEG:Ppa resulting in 99.99% fluorescence self-quenching. The biocompatibility of the nanomaterial and cell phototoxicity under light irradiation were investigated on PC3 prostate tumor cells in vitro. SQ-PEG:Ppa showed excellent phototoxic effect at low light dose of 5.0 J/cm

Topics: Animals; Apoptosis; Cell Proliferation; Chick Embryo; Chlorophyll; Chorioallantoic Membrane; Drug Delivery Systems; Female; Glioblastoma; Humans; Light; Male; Mice; Nanoparticles; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols; Prostatic Neoplasms; Squalene; Theranostic Nanomedicine; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Pyropheophorbide a (Pyro) is a promising photosensitizer; however, it has no tumor selectivity and enrichment capability. In our former work, the prepared folic acid (FA)-Pyro conjugates showed considerably improved tumor accumulation and photodynamic therapy (PDT) activity in cell- and animal-based studies. However, the targeting capability, selectivity and water solubility of the conjugate remain problematic. Here, we evaluated the installation of hydrophilic polyethylene glycol chains as the linker between Pyro and FA, by avoiding direct conjugation of Pyro with FA, aiming to improve tumor selectivity and accumulation. However, PEGylation may have negative effects on the PDT activity and cutaneous phototoxicity. Therefore, we chose various lengths of PEGs as linkers to optimize the molecular weight, hydrophilicity, and PDT activity and, thus, to balance the tumor selectivity and biological function of the conjugate. One optimized conjugate, Pyro-PEG1K-FA, exhibited excellent tumor enrichment and was able to eradicate subcutaneous tumors at a considerably reduced dose. We report the synthesis and characterization of these conjugates as well as the evaluation of their tumor accumulation ability and the corresponding PDT efficiency through in vitro and in vivo experiments.

Topics: Animals; Cell Line, Tumor; Cell Survival; Chlorophyll; Drug Carriers; Female; Folic Acid; Humans; Hydrophobic and Hydrophilic Interactions; Injections, Intravenous; Mice; Neoplasms; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols; Solubility; Tissue Distribution; Toxicity Tests, Acute; Xenograft Model Antitumor Assays

Photodynamic therapy (PDT) is an established therapeutic modality for the management of cancers. Conjugation with tumor-specific small molecule ligands (e.g., short peptides or peptidomimetics) could increase the tumor targeting of PDT agents, which is very important for improving the outcome of PDT. However, compared with antibody molecules, small molecule ligands have a much weaker affinity to their receptors, which means that their tumor enrichment is not always ideal. In this work, we synthesized multimeric RGD ligand-coupled conjugates of pyropheophorbide-a (Pyro) to increase the affinity through multivalent and cluster effects to improve the tumor enrichment of the conjugates. Thus, the dimeric and trimeric RGD peptide-coupled Pyro conjugates and the monomeric one for comparison were efficiently synthesized via a convergent strategy. A short polyethylene glycol spacer was introduced between two RGD motifs to increase the distance required for multivalence. A subsequent binding affinity assay verified the improvement of the binding towards integrin α

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Chlorophyll; Drug Design; Female; Humans; Integrin alphaVbeta3; Mice; Mice, Inbred BALB C; Molecular Structure; Oligopeptides; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols; Protein Binding; Protein Multimerization; Structure-Activity Relationship; Tissue Distribution

Photosensitizer-based photodynamic therapy (PDT) has attracted great attention in cancer treatment. However, achieving efficient delivery of photosensitizers is still a great challenge for their clinical applications. The photosensitizer-encapsulating delivery nanosystem usually suffers from poor stability, complex preparation process and low drug loading. Herein, we utilize a surfactant-like chemotherapeutic agent, mitoxantrone (MTX), as a nanocarrier to deliver a photosensitizer pyropheophorbide a (PPa) for antitumor therapy. MTX consists of aromatic rings (hydrophobic part) and two amino-groups and two hydroxyl-groups (hydrophilic part) with planar structure, which could interact with PPa via π-π stacking, hydrophobic interactions, intermolecular hydrogen bonding and electrostatic interactions. This system (PPa@MTX) spontaneously forms near-spherical nanostructures (∼150 nm), has a high loading capacity for PPa (56.5%) and exhibits a pH-responsive drug release manner in vitro. In vivo antitumor efficacy evaluations show that the pegylated PPa@MTX nanosystem has increased accumulation in tumor tissues and enhanced antitumor efficacy in female BALB/c mice bearing murine mammary carcinoma (4T1) tumor cells, compared to free PPa. Employing the surfactant-like drug as nanocarriers, our results show that the "drug-delivering-drug" strategy is a good foundation for the development of novel PDT-based drug delivery system against cancer.

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Chlorophyll; Drug Carriers; Drug Delivery Systems; Drug Liberation; Female; Mice, Inbred BALB C; Mitoxantrone; Nanostructures; Neoplasms; Photosensitizing Agents; Surface-Active Agents

There is an urgent need to develop efficient combination drug delivery approaches to address the low efficiency of clinical cancer monotherapy. However, how to achieve high-efficient synchronous co-delivery and synergistic therapy remains a big challenge. Herein, we report a self-facilitated nanoassembly of a heterotypic chemo-photodynamic dimer for multimodal cancer therapy. A reactive oxygen species (ROS)-responsive dimer of paclitaxel (PTX) and pyropheophorbide a (PPa) is rationally designed and synthesized. The "Two-in-One" dimer serves as both carrier material and cargo, could self-assemble into nanoparticles in water with ultrahigh co-loading capacity and self-facilitated ROS-responsive drug release. The endogenous ROS overproduced in tumor cells together with PPa-generated ROS under laser irradiation synergistically facilitate on-demand drug release from the nano-assembly. The disintegration of nanoassembly triggered by ROS effectively addresses the dilemma of aggregation-caused quenching (ACQ) effect of photosensitizer (PPa). Both in vitro and in vivo results suggest that PTX-initiated chemotherapy in combination with PPa-mediated PDT exhibits synergistic antitumor activity. Our findings provide a strategy for the rational design of nanocarrier for high-efficient synergetic cancer therapy.

Topics: Animals; Antineoplastic Agents; Carcinoma, Squamous Cell; Cell Line, Tumor; Chlorophyll; Combined Modality Therapy; Dimerization; Drug Delivery Systems; Drug Liberation; Female; Fluorescent Dyes; Humans; Mice, Inbred BALB C; Nanocapsules; Neoplasms, Experimental; Optical Imaging; Paclitaxel; Photochemotherapy; Photosensitizing Agents; Rats, Sprague-Dawley; Reactive Oxygen Species

The combination of chemotherapy with photodynamic therapy (PDT) holds promising applications in cancer therapy. However, co-encapsulation of chemotherapeutic agents and photosensitizers (PS) into the conventional nanocarriers suffers from inefficient co-loading and aggregation-caused quenching (ACQ) effect of PS trapped in dense carrier materials. Herein, we report a light-activatable photodynamic PEG-coated prodrug nanoplatform for core-shell synergistic chemo-photodynamic therapy. A novel photodynamic polymer is rationally designed and synthesized by conjugating pyropheophorbide a (PPa) to polyethylene glycol 2000 (PEG

Topics: Animals; Cell Death; Cell Line, Tumor; Chlorophyll; Drug Liberation; Endocytosis; Humans; Light; Mice, Nude; Nanoparticles; Oleic Acid; Paclitaxel; Photochemotherapy; Polyethylene Glycols; Prodrugs; Reactive Oxygen Species; Tissue Distribution; Xenograft Model Antitumor Assays

Hydrophile-lipophile balance (HLB) has a great influence on the self-assembly and physicochemical properties of amphiphiles, thus affecting their biological effects. It is shown that amphiphilic nanoparticles (NPs) with a moderate HLB value display enhanced stability and highly efficient tumor retention. 2,2-Bis(hydroxymethyl)propionic acid hyperbranched poly(ethylene glycol) (PEG)-pyropheophorbide-a (Ppa) amphiphiles (G320P, G310P, G220P, and G210P) are synthesized with a tunable HLB value from 6.1 to 9.9 by manipulating the number of generation of dendrons (G2 or G3) and the molecular weight of PEG chains (10 or 20 kDa). Molecular dynamics simulations reveal that G320P and G210P with a moderate HLB value (8.0 and 7.8) self-assemble into very stable NPs with a small solvent accessible surface area and high nonbonding interactions. G320P with a moderate HLB value (8.0) and a long PEG chain excels against other NPs in prolonging the blood circulation time of Ppa (up to 13-fold), penetrating deeply into multicellular tumor spheroids and accumulating in tumors, and enhancing the PDT efficacy with a tumor growth inhibition of 96.0%. Rational design of NPs with a moderate HLB value may be implemented in these NP-derived nanomedicines to achieve high levels of retention in tumors.

Topics: Animals; Cell Line, Tumor; Chlorophyll; Dendrimers; Hydrophobic and Hydrophilic Interactions; Mice; Molecular Dynamics Simulation; Nanoparticles; Polyethylene Glycols; Propionates; Protein Conformation

A pH and thermo dual-responsive supramolecular diblock copolymer is constructed by host-guest recognition of pillar[5]arene and viologen salt. The host polymer, poly(N,N-dimethylaminoethyl methacrylate) bearing pillar[5]arene as the terminal group (P[5]A-PDMAEMA) is synthesized by atom transfer radical polymerization (ATRP). Guest polymer, ethyl viologen-ended poly(N-isopropylacrylamide) (EV-PNIPAM) is prepared by reversible addition-fragmentation chain transfer polymerization. The supramolecular diblock copolymer can be self-assembled into stable supramolecular nanoparticles in aqueous solution at 40 °C, which show excellent pH and thermo responsiveness. The nanoparticles are further applied in the encapsulation of photosensitizers (pyropheophorbide-a, PhA) for photodynamic therapy (PDT). The dual-responsive nanoparticles can efficiently release PhA in acidic environment at 25 °C. Based on the result of cell experiments, PhA-loaded nanomicelles exhibit excellent PDT efficacy and low dark toxicity toward A549 cells. Thus, this supramolecular diblock copolymer enriches the methodology of constructing stimuli-responsive drug carriers and presents a great potential in PDT.

Topics: A549 Cells; Acrylamides; Acrylic Resins; Calixarenes; Chlorophyll; Drug Carriers; Humans; Methacrylates; Micelles; Nanoparticles; Nylons; Photochemotherapy; Photosensitizing Agents; Polymerization; Polymers

Both melanoma cells and tissues were allowed to interact with an identical pool of billions of human-safe phage nanofiber clones with each genetically displaying a unique 12-mer peptide at the tips, respectively, resulting in the discovery of bionanofibers displaying a melanoma cell/tissue dual-homing peptide for personalized targeted melanoma therapy.

Topics: Amino Acid Sequence; Animals; Bacteriophage M13; Cell Line, Tumor; Chlorophyll; Drug Carriers; Female; Humans; Light; Melanocytes; Melanoma; Mice, Inbred BALB C; Nanofibers; Peptide Library; Peptides; Photochemotherapy; Photosensitizing Agents; Protein Binding; Viral Proteins

Many approaches exist for stimuli-triggered cargo release from nanocarriers, but few can provide for on-demand release of multiple payloads, selectively. Here, we report the synthesis of purpurin-phospholipid (Pur-P), a lipid chromophore that has near-infrared absorbance red-shifted by 30 nm compared to a structurally similar pyropheophorbide-phospholipid (Pyr-P). Liposomes containing small amounts of either Pur-P or Pyr-P exhibited similar physical properties and fluorescence self-quenching. Loaded with distinct cargos, Pur-P and Pyr-P liposomes were mixed into a single colloidal suspension and selectively released cargo depending on irradiation wavelength. Spatiotemporal control of distinct cargo release was achieved by controlling multicolor laser placement. Using basic orange and doxorubicin anthraquinones, multidimensional cytotoxicity gradients were established to gauge efficacy against cancer cells using light-released drug. Wavelength selectivity of cargo release was maintained following intramuscular administration to mice.

Topics: Animals; Anthraquinones; Antibiotics, Antineoplastic; Cell Line, Tumor; Chlorophyll; Coloring Agents; Delayed-Action Preparations; Doxorubicin; Drug Liberation; Humans; Infrared Rays; Liposomes; Mice

Pyropheophorbide-a (Pyro) is a highly promising photosensitizer for tumor photodynamic therapy (PDT), although its very limited tumor-accumulation ability seriously restricts its clinical applications. A higher accumulation of photosensitizers is very important for the treatment of deeply seated and larger tumors. The conjugation of Pyro with tumor-homing peptide ligands could be a very useful strategy to optimize the physical properties of Pyro. Herein, we reported our studies on the conjugation of Pyro with a cyclic cRGDfK (cRGD) peptide, an integrin binding sequence, to develop highly tumor-specific photosensitizers for PDT application. To further reduce the nonspecific uptake and, thus, reduce the background distribution of the conjugates in normal tissues, we opted to add a highly hydrophilic polyethylene glycol (PEG) chain and an extra strongly hydrophilic carboxylic acid group as the linker to avoid the direct connection of the strongly hydrophobic Pyro macrocycle and cRGD ligand. We reported here the synthesis and characterization of these conjugates, and the influence of the hydrophilic modification on the biological function of the conjugates was carefully studied. The tumor-accumulation ability and photodynamic-induced cell-killing ability of these conjugates were evaluated through both in vitro cell-based experiment and in vivo distribution and tumor therapy experiments with tumor-bearing mice. Thus, the synthesized conjugate significantly improved the tumor enrichment and tumor selectivity of Pyro, as well as abolished the xenograft tumors in the murine model through a one-time PDT treatment.

Topics: Animals; Cell Line, Tumor; Chlorophyll; Female; Humans; Hydrophobic and Hydrophilic Interactions; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Peptides, Cyclic; Photochemotherapy; Photosensitizing Agents; Polyethylene Glycols; Tissue Distribution

Cancer photodynamic therapy (PDT) involves the absorption of light by photosensitizers (PSs) to generate cytotoxic singlet oxygen for killing cancer cells. The success of this method is usually limited by the lack of selective accumulation of the PS at cancer cells. Bioengineered viruses with cancer cell-targeting peptides fused on their surfaces are great drug carriers that can guide the PS to cancer cells for targeted cancer treatment. Here, we use cell-targeting fd bacteriophages (phages) as an example to describe how to chemically conjugate PSs (e.g., pyropheophorbide-a (PPa)) onto a phage particle to achieve targeted PDT.

Topics: Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Chlorophyll; Drug Carriers; Humans; Oncolytic Viruses; Peptides; Photochemotherapy; Photosensitizing Agents; Singlet Oxygen

Topics: Animals; Chlorophyll; Fluorescence; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Micelles; Particle Size; Permeability; Photochemotherapy; Photosensitizing Agents; Polymers; Polymethacrylic Acids; Theranostic Nanomedicine; Time Factors; Tissue Distribution

Conjugates of 17α-substituted testosterone (1 and 2) and 17β-substituted epitestosterone (3 and 4) with pyropheophorbide a were synthesized. The scheme consisted of synthesis of 17α-hydroxy-3-oxopregn-4-en-21-oic and 17β-hydroxy-3-oxopregn-4-en-21-oic acids, and their coupling with pyropheophorbide a by means of either ethylene diamine, or 1,5-diamino pentane linkers. Mutual influence of steroidal and macrocyclic fragments in conjugates molecules was dependent on configuration of C17 and length of linker, that was established by analysis of

Topics: Antineoplastic Agents; Cell Line, Tumor; Cell Proliferation; Cell Survival; Chlorophyll; Epitestosterone; Humans; Male; PC-3 Cells; Prostatic Neoplasms; Structure-Activity Relationship; Testosterone

In the context of prodrug nanomedicines for cancer therapy, one of the great challenges is the slow and variable release of the parent drug in tumors. Recently, many smart redox-sensitive nanocarriers have been developed to address this problem. However, due to significant tumor heterogeneity, some redox-sensitive nanomedicines still show poor selectivity in drug release. Herein, we report the design and synthesis of a ROS-triggered prodrug nanoplatform fabricated with oxidation-responsive cabazitaxel (CTX) prodrugs for synergistic chemo-photodynamic therapy, thioether-/selenoether-linked conjugates of CTX and oleic acid (OA). These prodrugs can be readily self-assembled into nanoparticles, with pyropheophorbide a (PPa) co-encapsulated into the prodrug-nanosystem for combination therapy. The dual-source ROS-responsive prodrug nanosystems selectively and rapidly release CTX not only in response to endogenous ROS overproduced in tumor cells, but also to exogenous PPa-generated ROS under laser irradiation. Moreover, the selenium-containing linkage demonstrates significant advantages over a sulfur-containing linkage in terms of ROS-triggered drug release and cytotoxicity. The prepared prodrug-nanosystems significantly prolong the systemic circulation and tumor distribution of both CTX and PPa, thereby demonstrating synergistic chemo-photodynamic therapy in vivo. All these drug delivery advantages render the nanosystem extremely promising for cancer treatment.

Topics: Animals; Cell Survival; Chlorophyll; Drug Carriers; Drug Liberation; Drug Synergism; Intracellular Space; Light; Male; Mice; Nanostructures; Oleic Acid; Photochemotherapy; Photosensitizing Agents; Prodrugs; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Taxoids; Tissue Distribution

We describe a simple and effective bioconjugation strategy to extend the plasma circulation of a low molecular weight targeted phototheranostic agent, which achieves high tumor accumulation (9.74 ± 2.26%ID/g) and high tumor-to-background ratio (10:1). Long-circulating pyropheophorbide (LC-Pyro) was synthesized with three functional building blocks: (1) a porphyrin photosensitizer for positron-emission tomography (PET)/fluorescence imaging and photodynamic therapy (PDT), (2) a urea-based prostate-specific membrane antigen (PSMA) targeting ligand, and (3) a peptide linker to prolong the plasma circulation time. With porphyrin's copper-64 chelating and optical properties, LC-Pyro demonstrated its dual-modality (fluorescence/PET) imaging potential for selective and quantitative tumor detection in subcutaneous, orthotopic, and metastatic murine models. The peptide linker in LC-Pyro prolonged its plasma circulation time about 8.5 times compared to its truncated analog. High tumor accumulation of LC-Pyro enabled potent PDT, which resulted in significantly delayed tumor growth in a subcutaneous xenograft model. This approach can be applied to improve the pharmacokinetics of existing and future targeted PDT agents for enhanced tumor accumulation and treatment efficacy.

Topics: Animals; Chlorophyll; Copper Radioisotopes; Male; Mice; Mice, Nude; Optical Imaging; Peptides; Photochemotherapy; Photosensitizing Agents; Porphyrins; Positron-Emission Tomography; Prostate-Specific Antigen; Prostatic Neoplasms; Theranostic Nanomedicine

Emerging evidence indicates that the transcription factor nuclear factor-E2-related factor 2 (Nrf2) plays an essential role in cellular defense against oxidative stress; its activation has been related to cytoprotection.. Here, we investigated the role of Nrf2 in improving the efficacy of methyl pyropheophorbide-amediated photodynamic therapy (Mppa-PDT) via the downregulation of Nrf2.. Human ovarian cancer A2780 cells and SKOV3 cells were treated with Mppa-PDT and siRNA transfection was performed to inhibit Nrf2. After treated with siRNA and Mppa-PDT, the cell viability was examined with CCK-8 assay; cell apoptosis was detected tested by flow cytometry with Annexin V-FITC/PI; the celluar reactive oxygen species (ROS) and mitochondrial membrane potential were measured with DCFHDA and JC-1 staining; expression of protein was assessed by western blot analysis.. We found that Nrf2 translocated from the cytoplasm to the nucleus in vitro and in vivo, and the expression of Nrf2 and P-Nrf2 increased through a possible mechanism regulated by mitogen-activated protein kinase (MAPK) after Mppa-PDT treatment. Furthermore, cytotoxicity and apoptosis induced by Mppa-PDT increased after Nrf2down-regulation. Nrf2 down -regulation increased reactive oxygen species (ROS) levels by attenuating antioxidants or pumping Mppa out of cells,which resulted from the inhibition of Nrf2-HO-1 or Nrf2- ABCG2 signaling. In addition, SKOV3 cells exhibited increased resistance to Mppa-PDT, and the expression levels of P-Nrf2 and ABCG2 were higher in SKOV3 cells than in A2780 cells, suggesting that Nrf2-ABCG2 signaling might be involved in the intrinsic resistanceto Mppa-PDT.. These results provided evidence that Nrf2 down-regulation can enhance the effect of Mppa-PDT.

Topics: Animals; ATP Binding Cassette Transporter, Subfamily G, Member 2; Cell Line, Tumor; Chlorophyll; Down-Regulation; Female; Heme Oxygenase (Decyclizing); Humans; MAP Kinase Signaling System; Mice; Neoplasm Proteins; NF-E2-Related Factor 2; Ovarian Neoplasms; Oxidative Stress; Photochemotherapy; Reactive Oxygen Species; Signal Transduction; Xenograft Model Antitumor Assays

The application of photodynamic therapy (PDT) for the diagnosis and treatment of cancer is hindered by the intrinsic defects of the currently available photosensitizers (PSs), such as poor water solubility and limited light-penetration depth. In this study, pH-responsive polymeric micelles that co-encapsulate therapeutic PSs and organooxotin two-photon compounds were applied for two-photon PDT (TP-PDT) against breast cancer. The TP-PDT effect of the drug-loaded micelles was "activated" when the micelles turned into aggregates at a triggering pH level. The in vitro therapeutic effect was evaluated on 4T1 murine breast cancer cells by viability assays, real-time morphology collapsing, and reactive oxygen species determination. Time-dependent ex vivo organ distribution and in vivo anticancer efficacy results suggested that the drug carriers could accumulate in tumors and suppress tumor growth by TP-PDT. The delivery system could enhance the solubility and distribution of PSs and, if administered along with a tissue-penetrating prolonged light source, could thus have good potential for cancer therapy.

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Chlorophyll; Drug Carriers; Female; Humans; Hydrogen-Ion Concentration; Mice, Inbred BALB C; Micelles; Photochemotherapy; Photons; Photosensitizing Agents; Polymers; Reactive Oxygen Species; Tissue Distribution; Xenograft Model Antitumor Assays

Here we developed the first case of pyropheophorbide-a-loaded PEGylated-hybrid carbon nanohorns (CNH-Pyro) to study tumor targeting therapy. During incubation with living cells, CNH-Pyro exhibited very intense red emissions. The intracellular imaging results were carried out by flow cytometry based on four different kinds of cell lines (including three adherent cell lines and one suspension cell line). Compared with free pyropheophorbide-a, CNH-Pyro demonstrated enhanced photodynamic tumor ablation efficiency during in vitro experiments due to improved biocompatibility of the hybrid nanomaterial and the photothermal therapy effect derived from carbon-network structure. Trypan blue staining experiments supported that the cell fate was dependent on the synergistic effects of both CNH-Pyro and laser irradiations. These results indicated that the chlorin-entrapped carbon nanohorns could provide powerful delivery vehicles for increasing photodynamic efficacy and possess early identification of the disease.

Topics: Animals; Cell Line, Tumor; Chlorophyll; Flow Cytometry; Humans; Mice; Nanoparticles; Optics and Photonics; Prodrugs; Spectrometry, Fluorescence; Spectroscopy, Near-Infrared; Staining and Labeling

Photodynamic therapy (PDT) is entering the mainstream of the cancer treatments recently. Pyropheophorbide-a (Pa), as a degradation product of chlorophyll-a, has been shown to be a potent photosensitizer in photodynamic therapy. In this paper, we investigated the in vitro photodynamic therapy of 13

Topics: Cell Survival; Chlorophyll; Dose-Response Relationship, Drug; HeLa Cells; Humans; Hydrazones; Photochemotherapy; Photosensitizing Agents; Porphyrins

Photodynamic therapy (PDT) is a highly localized and minimally invasive cancer treatment modality with many important advantages, but the lack of ideal photosensitizers (PSs) greatly restricts its clinical utility. To develop new PSs with highly efficient singlet oxygen production and high tumor-localizing ability to reduce damage to healthy adjacent tissues, we conjugated folic acid (FA) with pyropheophorbide a (Pyro), a potent PS with a very high singlet oxygen quantum yield and a high extinction coefficient. In the present work, we describe the synthesis and PDT evaluation of this FA-Pyro conjugate both in vitro and in vivo. This conjugation increased the accumulation of Pyro inside the tumors and improved the efficiency of PDT, resulting in eradication of subcutaneous xenograft KB (human mouth epidermal carcinoma) tumors after only 1 or 2 applications of external near infrared light irradiation. This outstanding PDT outcome in a tumor-bearing mouse model and the simple synthesis of the conjugate should have very good practical potential for clinical application.

Topics: Animals; Cell Line, Tumor; Cell Survival; Chlorophyll; Female; Folic Acid; Humans; Mice, Inbred BALB C; Mouth Neoplasms; Photochemotherapy; Photosensitizing Agents

A pyropheophorbide-α-based building block (Ppa-PLGVRG-Van) can be used to construct self-aggregated superstructures in vivo for highly specific and sensitive diagnosis of bacterial infection by noninvasive photoacoustic tomography. This in vivo supramolecular chemistry approach opens a new avenue for efficient, rapid, and early-stage disease diagnosis with high sensitivity and specificity.

Topics: Animals; Bacterial Infections; Chlorophyll; Contrast Media; Disease Models, Animal; Escherichia coli; Gelatinases; Mice; Models, Biological; Molecular Structure; Myositis; Nanostructures; Phantoms, Imaging; Photoacoustic Techniques; Proteus vulgaris; Staphylococcus aureus; Staphylococcus epidermidis; Tomography; Vancomycin; Water

Current approaches in use of water-insoluble photosensitizers for photodynamic therapy (PDT) of cancer often demand a nano-delivery system. Here, we report a photosensitizer-loaded biocompatible nano-delivery formulation (PPaN-20) whose size was engineered to ca. 20nm to offer improved cell/tissue penetration and efficient generation of cytotoxic singlet oxygen. PPaN-20 was fabricated through the physical assembly of all biocompatible constituents: pyropheophorbide-a (PPa, water-insoluble photosensitizer), polycaprolactone (PCL, hydrophobic/biodegradable polymer), and Pluronic F-68 (clinically approved polymeric surfactant). Repeated microemulsification/evaporation method resulted in a fine colloidal dispersion of PPaN-20 in water, where the particulate PCL matrix containing well-dispersed PPa molecules inside was stabilized by the Pluronic corona. Compared to a control sample of large-sized nanoparticles (PPaN-200) prepared by a conventional solvent displacement method, PPaN-20 revealed optimal singlet oxygen generation and efficient cellular uptake by virtue of the suitably engineered size and constitution, leading to high in vitro phototoxicity against cancer cells. Upon administration to tumor-bearing mice by peritumoral route, PPaN-20 showed efficient tumor accumulation by the enhanced cell/tissue penetration evidenced by in vivo near-infrared fluorescence imaging. The in vivo PDT treatment with peritumorally administrated PPaN-20 showed significantly enhanced suppression of tumor growth compared to the control group, demonstrating great potential as a biocompatible photosensitizing agent for locoregional PDT treatment of cancer.

Topics: Animals; Biocompatible Materials; Chlorophyll; Drug Delivery Systems; Flow Cytometry; HeLa Cells; Humans; Male; Mice; Mice, Nude; Nanoparticles; Nanotechnology; Neoplasms; NIH 3T3 Cells; Particle Size; Photobleaching; Photochemotherapy; Photosensitizing Agents; Polyesters; Polymers; Singlet Oxygen

Advanced colorectal cancer is one of the deadliest cancers, with a 5-year survival rate of only 12% for patients with the metastatic disease. Checkpoint inhibitors, such as the antibodies inhibiting the PD-1/PD-L1 axis, are among the most promising immunotherapies for patients with advanced colon cancer, but their durable response rate remains low. We herein report the use of immunogenic nanoparticles to augment the antitumour efficacy of PD-L1 antibody-mediated cancer immunotherapy. Nanoscale coordination polymer (NCP) core-shell nanoparticles carry oxaliplatin in the core and the photosensitizer pyropheophorbide-lipid conjugate (pyrolipid) in the shell (NCP@pyrolipid) for effective chemotherapy and photodynamic therapy (PDT). Synergy between oxaliplatin and pyrolipid-induced PDT kills tumour cells and provokes an immune response, resulting in calreticulin exposure on the cell surface, antitumour vaccination and an abscopal effect. When combined with anti-PD-L1 therapy, NCP@pyrolipid mediates regression of both light-irradiated primary tumours and non-irradiated distant tumours by inducing a strong tumour-specific immune response.

Topics: Animals; Apoptosis; B7-H1 Antigen; CD8-Positive T-Lymphocytes; Cell Line, Tumor; Chlorophyll; Coordination Complexes; Cytotoxicity, Immunologic; Disease Models, Animal; Drug Compounding; Immunity; Immunotherapy; Inflammation; Lipids; Male; Mice, Inbred BALB C; Mice, Nude; Nanoparticles; Neoplasms; Organoplatinum Compounds; Oxaliplatin; Photochemotherapy; Polymers; Tissue Distribution

An effective, nontoxic, tumor-specific immunotherapy is the ultimate goal in the battle against cancer, especially the metastatic disease. Checkpoint blockade-based immunotherapies have been shown to be extraordinarily effective but benefit only the minority of patients whose tumors have been pre-infiltrated by T cells. Here, we show that Zn-pyrophosphate (ZnP) nanoparticles loaded with the photosensitizer pyrolipid (ZnP@pyro) can kill tumor cells upon irradiation with light directly by inducing apoptosis and/or necrosis and indirectly by disrupting tumor vasculature and increasing tumor immunogenicity. Furthermore, immunogenic ZnP@pyro photodynamic therapy (PDT) treatment sensitizes tumors to checkpoint inhibition mediated by a PD-L1 antibody, not only eradicating the primary 4T1 breast tumor but also significantly preventing metastasis to the lung. The abscopal effects on both 4T1 and TUBO bilateral syngeneic mouse models further demonstrate that ZnP@pyro PDT treatment combined with anti-PD-L1 results in the eradication of light-irradiated primary tumors and the complete inhibition of untreated distant tumors by generating a systemic tumor-specific cytotoxic T cell response. These findings indicate that nanoparticle-mediated PDT can potentiate the systemic efficacy of checkpoint blockade immunotherapies by activating the innate and adaptive immune systems in tumor microenvironment.

Topics: Animals; Apoptosis; Breast Neoplasms; Cell Line, Tumor; Chlorophyll; Combined Modality Therapy; Diphosphates; Humans; Immunotherapy; Light; Lipids; Lung Neoplasms; Mice; Nanoparticles; Necrosis; Neoplasm Metastasis; Photochemotherapy; Photosensitizing Agents; Zinc

We developed novel methods to convert the C3-vinyl group of a chlorophyll derivative, methyl pyropheophorbide-a, into an acetyl group, an epoxy group, and a formyl group via iodination with I2 and phenyliodine(III) bis(trifluoroacetate). Reaction of the iodinated intermediate with ethylene glycol and subsequent treatment with base led to formation of the C3-acetyl chlorin. Reaction of the iodinated intermediate with ethylenediamine afforded the C3-oxiranyl chlorin. The C3-formyl chlorin was readily derived from the epoxide without hazardous reagents such as OsO4. These reactions were facile and useful alternatives to the previous methods.

Topics: Acetylation; Chlorophyll; Epoxy Compounds; Halogenation; Iodobenzenes; Methylation; Porphyrins; Trifluoroacetic Acid

The (13(2)R/S)-methoxycarbonyl group of methyl pheophorbides a/a' (chlorophyll a/a' derivatives) was converted to methyl, ethyl, propyl, and isopropyl groups through the C13(2)-alkylation under basic conditions followed by pyrolysis in 2,4,6-collidine with lithium iodide. All the resulting products, methyl 13(2)-alkyl-pyropheophorbides a, predominantly gave the (13(2)R)-stereoisomers with about one tenth of the (13(2)S)-epimers. Their stereochemistry was determined by 1D/2D NMR and their optical properties were characterized by visible absorption and circular dichroism spectroscopy. Methyl (13(2)R)-propyl-pyropheophorbide a was converted to (13(2)R)-propyl-pyrochlorophyll a by ester exchanging and magnesium chelating reactions. The synthetic chlorophyll a analogue showed non-epimerization at the 13(2)-position in pyridine-d5 at 40°C, while naturally occurring chlorophyll a was easily epimerized under the same conditions to give its epimeric mixture.

Topics: Chlorophyll; Chlorophyll A; Models, Molecular; Molecular Structure

Upconversion nanoparticles have shown to be a promising prospect for biological detection and photodynamic therapy (PDT). The focus of this study was to develop an upconversion nanoparticle modified with a targeting peptide and photosensitizer for near-infrared photodynamic therapy. To produce a tumor-targeting nanophotosensitizer with near-infrared excitation, NaYF4:Yb/Er upconversion nanoparticles were first wrapped with O-carboxymethyl chitosan to develop an upconversion rianoplatform and then chemically conjugated with the photosensitizer pyropheophorbide-a (Ppa) and RGD peptide c(RGDyK). The nanoparticle exhibited low dark toxicity and high biocompatibility. When injected into the tail vein of tumor-bearing U87-MG mice, UCNP-Ppa-RGD revealed an enhanced tumor-specific biodistribution and successful therapeutic effect following near-infrared laser irradiation. It possessed a significantly deeper therapeutic depth compared with conventional visible light triggered PDT using Ppa. The results suggest that the nanoplatform has advantages in the spectral application, and the constructed tumor-specific nanoparticle shows high clinical potential to serve not only as a photodynamic imaging reagent but also as a therapeutic agent for the treatment of large or deeply seated tumors.

Topics: Animals; Cell Line, Tumor; Chitosan; Chlorophyll; Drug Delivery Systems; Fluorides; Humans; Mice; Mice, Nude; Nanoparticles; Photochemotherapy; Photosensitizing Agents; Spectroscopy, Near-Infrared; Tissue Distribution; Yttrium

Sunlight is the most abundant energy source on this planet. However, the ability to convert sunlight into biological energy in the form of adenosine-5'-triphosphate (ATP) is thought to be limited to chlorophyll-containing chloroplasts in photosynthetic organisms. Here we show that mammalian mitochondria can also capture light and synthesize ATP when mixed with a light-capturing metabolite of chlorophyll. The same metabolite fed to the worm Caenorhabditis elegans leads to increase in ATP synthesis upon light exposure, along with an increase in life span. We further demonstrate the same potential to convert light into energy exists in mammals, as chlorophyll metabolites accumulate in mice, rats and swine when fed a chlorophyll-rich diet. Results suggest chlorophyll type molecules modulate mitochondrial ATP by catalyzing the reduction of coenzyme Q, a slow step in mitochondrial ATP synthesis. We propose that through consumption of plant chlorophyll pigments, animals, too, are able to derive energy directly from sunlight.

Topics: Adenosine Triphosphate; Adipose Tissue; Animals; Brain; Caenorhabditis elegans; Chlorophyll; Diet; Ducks; Fluorescence; Mammals; Mice; Mice, Inbred ICR; Mitochondria, Liver; Photons; Rats; Rats, Inbred F344; Sus scrofa; Tissue Extracts; Ubiquinone

The development of complementary and/or alternative drugs for treatment of hepatitis C virus (HCV) infection is still needed. Antiviral compounds in medicinal plants are potentially good targets to study. Morinda citrifolia is a common plant distributed widely in Indo-Pacific region; its fruits and leaves are food sources and are also used as a treatment in traditional medicine. In this study, using a HCV cell culture system, it was demonstrated that a methanol extract, its n-hexane, and ethyl acetate fractions from M. citrifolia leaves possess anti-HCV activities with 50%-inhibitory concentrations (IC(50)) of 20.6, 6.1, and 6.6 μg/mL, respectively. Bioactivity-guided purification and structural analysis led to isolation and identification of pheophorbide a, the major catabolite of chlorophyll a, as an anti-HCV compound present in the extracts (IC(50) = 0.3 μg/mL). It was also found that pyropheophorbide a possesses anti-HCV activity (IC(50) = 0.2 μg/mL). The 50%-cytotoxic concentrations (CC(50)) of pheophorbide a and pyropheophorbide a were 10.0 and 7.2 μg/mL, respectively, their selectivity indexes being 33 and 36, respectively. On the other hand, chlorophyll a, sodium copper chlorophyllin, and pheophytin a barely, or only marginally, exhibited anti-HCV activities. Time-of-addition analysis revealed that pheophorbide a and pyropheophorbide a act at both entry and the post-entry steps. The present results suggest that pheophorbide a and its related compounds would be good candidates for seed compounds for developing antivirals against HCV.

Topics: Antiviral Agents; Chlorophyll; Hepacivirus; Hepatitis C; Humans; Morinda; Plant Extracts; Plant Leaves

A chlorophyll-a derivative bonded directly with epoxide at the peripheral position of the chlorin π-system was reacted with N-urethane and C-ester protected amino acids bearing an alcoholic or phenolic hydroxy group as well as a carboxy group at the residue to give chlorophyll-amino acid conjugates. The carboxy residues of N,C-protected aspartic and glutamic acids were esterified with the epoxide in high yields. The synthetic conjugates in dichloromethane had absorption bands throughout the visible region including intense red-side Qy and blue-side Soret bands. By their excitation at the visible bands, strong and efficient fluorescence emission was observed up to the near-infrared region. The chromo/fluorophores are promising for preparation of functional peptides and modification of proteins.

Topics: Amino Acids; Catalysis; Chlorophyll; Chlorophyll A; Epoxy Compounds; Esterification; Models, Molecular; Porphyrins; Proteins; Spectrophotometry, Infrared

Photodynamic efficacy of pyropheophorbide-a (PPa) is limited due to poor aqueous solubility. In the present study, organically modified silica nanoparticles (ORMOSIL) entrapping PPa and its folate receptor targeted conjugate (FR-Np-PPa) were prepared and the effect of pH on uptake and photodynamic action of plain and FR-Np-PPa in squamous cell carcinoma (Nt-8e) cells and adenocarcinoma of breast (MCF-7) cells were studied.. Nanoformulations of PPa were characterized by absorption and fluorescence spectroscopy. Dynamic light scattering was used for size measurements. The uptake of the two nanoformulations by cells incubated in media of pH 6.5 and 7.4 was studied by confocal fluorescence microscopy and spectrofluoremetrically. Phototoxicity of PPa was studied by MTT assay.. In MCF-7 and Nt-8e cells, while the uptake of PPa was observed to increase with a decrease in pH of the incubation media for folate receptor targeted Np, uptake of Np-PPa was not influenced by a change in the pH of the media. Inhibition in the uptake of PPa in presence of free folic acid for cells incubated in a medium of pH 6.5 with targeted nanoparticles was higher compared to physiological pH. Consistent with uptake studies in both the cell lines phototoxicity of PPa delivered through FR-Np-PPa was higher in medium of pH 6.5 as compared to physiological pH and phototoxicity induced by NP-PPa was independent of the pH of medium.. Acidic pH enhances the photodynamic efficacy of FR-targeted nanoformulated PPa.

Topics: Cell Survival; Chlorophyll; Extracellular Fluid; Folic Acid Transporters; Hydrogen-Ion Concentration; MCF-7 Cells; Metabolic Clearance Rate; Nanocapsules; Neoplasms, Experimental; Radiation-Sensitizing Agents; Silicon Dioxide; Treatment Outcome

Ovarian cancer is the leading cause of morbidity/mortality from gynecologic malignancy. Early detection of disease is difficult due to the propensity for ovarian cancer to disseminate throughout the peritoneum. Currently, there is no single accurate test to detect primary or recurrent ovarian cancer. We report a novel clinical strategy using PPF: a multimodal, PET and optical, folate receptor (FR)-targeted agent for ovarian cancer imaging. The capabilities of PPF were evaluated in primary human ovarian cancer cells, in vivo xenografts derived from primary cells and ex vivo patient omemtum, as the heterogeneity and phenotype displayed by patients is retained. Primary cells uptake PPF in a FR-dependent manner demonstrating approximately a 5- to 25-fold increase in fluorescence. By both PET and fluorescence imaging, PPF specifically delineated FR-positive, ovarian cancer xenografts, with similar tumor-to-background ratios of 8.91±0.91 and 7.94±3.94, and micro-metastatic studding (<1mm), which demonstrated a 3.5-fold increase in PPF uptake over adjacent normal tissue. Ex vivo patient omentum demonstrated selective uptake of PFF by tumor deposits. The ability of PPF to identify metastatic deposits <1mm could facilitate more complete debulking (currently, optimal debulking is <10mm residual tumor), by providing a more sensitive imaging strategy improving treatment planning, response assessment and residual/recurrent disease detection. Therefore, PPF is a novel clinical imaging strategy that could substantially improve the prognosis of patients with ovarian cancer by allowing pre-, post- and intra-operative tumor monitoring, detection and possibly treatment throughout all stages of therapy and tumor progression.

Topics: Animals; Cell Line, Tumor; Chlorophyll; Female; Folate Receptor 1; Folic Acid; Humans; Mice; Microscopy, Fluorescence; Molecular Diagnostic Techniques; Ovarian Neoplasms; Positron-Emission Tomography; Staining and Labeling

We recently developed porphysomes as intrinsically multifunctional nanovesicles. A photosensitizer, pyropheophorbide α, was conjugated to a phospholipid and then self-assembled to liposome-like spherical vesicles. Due to the extremely high density of porphyrin in the porphyrin-lipid bilayer, porphysomes generated large extinction coefficients, structure-dependent fluorescence self-quenching, and excellent photothermal efficacy. In our formulation, porphysomes were synthesized using high pressure extrusion, and displayed a mean particle size around 120 nm. Twenty-four hr post-intravenous injection of porphysomes, the local temperature of the tumor increased from 30 °C to 62 °C rapidly upon one minute exposure of 750 mW (1.18 W/cm(2)), 671 nm laser irradiation. Following the complete thermal ablation of the tumor, eschars formed and healed within 2 weeks, while in the control groups the tumors continued to grow and all reached the defined end point within 3 weeks. These data show how porphysomes can be used as potent photothermal therapy (PTT) agents.

Topics: Ablation Techniques; Animals; Chlorophyll; Humans; Hyperthermia, Induced; KB Cells; Mice; Mice, Nude; Nanoparticles; Phospholipids; Photosensitizing Agents; Phototherapy; Xenograft Model Antitumor Assays

Near-infrared (NIR)-to-visible upconversion nanoparticle (UCNP) has shown promising prospects in photodynamic therapy (PDT) as a drug carrier or energy donor. In this work, a photosensitizer pyropheophorbide a (Ppa) and RGD peptide c(RGDyK) comodified chitosan-wrapped NaYF(4):Yb/Er upconversion nanoparticle UCNP-Ppa-RGD was developed for targeted near-infrared photodynamic therapy. The properties of UCNP-Ppa-RGD, such as morphology, stability, optical spectroscopy and singlet oxygen generation efficiency, were investigated. The results show that covalently linked pyropheophorbide a molecule not only is stable but also retains its spectroscopic and functional properties. In vitro studies confirm a stronger targeting specificity of UCNP-Ppa-RGD to integrin α(v)β(3)-positive U87-MG cells compared with that in the corresponding negative group. The photosensitizer-attached nanostructure exhibited low dark toxicity and high phototoxicity against cancer cells upon 980 nm laser irradiation at an appropriate dosage. These results represent the first demonstration of a highly stable and efficient photosensitizer modified upconversion nanostructure for targeted near-infrared photodynamic therapy of cancer cells. The novel UCNP-Ppa-RGD nanoparticle may provide a powerful alternative for near-infrared photodynamic therapy with an improved tumor targeting specificity.

Topics: Cell Line, Tumor; Cell Survival; Chitosan; Chlorophyll; Humans; Nanoparticles; Oligopeptides; Photochemotherapy; Photosensitizing Agents; Reactive Oxygen Species

Efflux pumps are vital bacterial components, and research has demonstrated that some plant compounds such as pheophorbide a (php) possess efflux pump inhibitor (EPI) activity. This study determined the quantity of php present in feces as an indicator of EPI activity. Feces were collected from different species of animals fed a variety of feeds. The chlorophyll metabolites php and pyropheophorbide a (pyp) were determined using fluorescense spectroscopy. The average concentrations [μg/g dry matter (DM) feces] of pyp/php in feces were as follows: guinea pig, 180; goat, 150; rabbit, 150; dairy cow, 120; feedlot cattle, 60; rat, <1; pig, <1; chicken, <1. These data indicate that animals consuming "green" diets will excrete feces with concentrations of php/pyp that exceed levels demonstrated to be inhibitory to bacterial efflux pumps (0.5 μg/mL). The natural presence EPIs in the gastrointestinal tract may modulate the activity of microbial efflux pumps and exert selection pressure upon resident microbial populations.

Topics: Animals; Chlorophyll; Feces; Spectrometry, Fluorescence

The C3-vinyl group of a chlorophyll derivative, methyl pyropheophorbide-a, was converted into the formyl group by a novel one-pot reaction with thiophenol at room temperature. The mild reaction can provide insight into development of 'green' catalysts displacing OsO(4) or O(3), and into elucidation of unknown biosynthetic processes of chlorophyll-d.

Topics: Catalysis; Chlorophyll; Chlorophyll A; Osmium Tetroxide; Oxidation-Reduction; Phenols; Sulfhydryl Compounds

Both bacteriopyropheophorbide-a and ring-B reduced pyropheophorbide-a on reacting with NBS (N-bromosuccinamide) undergo electrophilic bromination to provide 10-bromo analogs. The electronic nature of the substituents present at position-3 did not make any difference in the regioselective outcome of the brominated products. These relatively stable brominated chlorins and bacteriochlorins provide an easy way of introducing a wide variety of functionalities, which could be extremely useful in developing improved agents for biomedical applications and supramolecular chemistry.

Topics: Chlorophyll; Halogenation; Molecular Structure; Oxidation-Reduction; Stereoisomerism

We hypothesized that expression of nuclear estrogen receptor (ER) in hormone-sensitive breast cancer cells could be harnessed synergistically with the tumor-accumulating effect of porphyrins to selectively deliver estrogen-porphyrin conjugates into breast tumor cells, and preferentially kill tumor cells upon exposure to visible light. In this study we synthesized a conjugate of C(17α)-alkynylestradiol and pyropheophorbide and demonstrated that this conjugate is internalized by ER-positive MCF-7 cells while pyropheophorbide did not, suggesting an ER-mediated uptake and internalization of the conjugate by incipient nuclear ER in MCF-7 cells. This study is a direct demonstration of our hypothesis about ER-mediated internalization of estrogen-porphyrin conjugates.

Topics: Breast Neoplasms; Cell Line, Tumor; Chlorophyll; Estradiol; Female; Humans; Microscopy, Fluorescence; Photochemotherapy; Photosensitizing Agents; Receptors, Estrogen

Demetalation of three synthetic zinc cyclic tetrapyrroles that possess identical peripheral substituents, zinc methyl bacteriopyropheophorbide a (zinc bacteriochlorin 1), zinc methyl 3-devinyl-3-acetyl-pyropheophorbide a (zinc chlorin 2), and zinc methyl 3-devinyl-3-acetyl-protopyropheophorbide a (zinc porphyrin 3), was kinetically analyzed under acidic conditions to examine the effects of macrocyclic structures on demetalation without peripheral substitution effects. Zinc bacteriochlorin 1 exhibited much slower demetalation kinetics than zinc chlorin 2 and zinc porphyrin 3. These results indicate that the bacteriochlorin skeleton provides significant resistance to the removal of the central metal from the tetrapyrrole ligand. Comparison of demetalation kinetics of 3-acetyl zinc complexes 2 and 3 with that of 3-vinyl zinc complexes under the same reaction condition demonstrated that the relative ratio (5.0 × 10(-2)) of the demetalation rate constant of the 3-acetyl zinc chlorin 2 to that of the corresponding 3-vinyl zinc chlorin 4 resembled the case of the 3-acetyl zinc porphyrin 3 to the 3-vinyl zinc porphyrin 5 (the relative ratio was 6.8 × 10(-2)). These suggest that the electron-withdrawing 3-acetyl group slows down the demetalation from the tetrapyrrole ligands more than the 3-vinyl group and that the 3-acetyl effect is analogous in both chlorin and porphyrin π-systems.

Topics: Chlorophyll; Kinetics; Porphyrins; Tetrapyrroles; Zinc

Photocatalytic activity of a photosensitizer (PS) in an oligodeoxyribonucleotide duplex 5'-PS~ODN1/ODN2~Q-3' is inhibited because of close proximity of a quencher Q. The ODN2 in this duplex is selected to be longer than the ODN1. Therefore, in the presence of a nucleic acid (analyte), which is fully complementary to the ODN2 strand, the duplex is decomposed with formation of an analyte/ODN2~Q duplex and a catalytically active, single stranded PS~ODN1. In this way the catalytic activity of the PS can be controlled by the specific nucleic acids. We applied this reaction earlier for the amplified detection of ribonucleic acids in live cells (Arian, D.; Cló, E.; Gothelf, K.; Mokhir, A. Chem.-Eur. J.2010, 16(1), 288). As a photosensitizer (PS) we used In(3+)(pyropheophorbide-a)chloride and as a quencher (Q)--Black-Hole-Quencher-3 (BHQ-3). The In(3+) complex is a highly active photocatalyst in aqueous solution. However, it can coordinate additional ligands containing thiols (e.g., proteins, peptides, and aminoacids), that modulate properties of the complex itself and of the corresponding bio- molecules. These possible interactions can lead to undesired side effects of nucleic acid controlled photocatalysts (PS~ODN1/ODN2∼Q) in live cells. In this work we explored the possibility to substitute the In(3+) complex for those ones of divalent metal ions, Zn(2+) and Pd(2+), which exhibit lower or no tendency to coordinate the fifth ligand. We found that one of the compounds tested (Pd(pyropheophorbide-a) is as potent and as stable photosensitizer as its In(3+) analogue, but does not coordinate additional ligands that makes it more suitable for cellular applications. When the Pd complex was introduced in the duplex PS~ODN1/ODN2~Q as a PS, its photocatalytic activity could be controlled by nucleic acids as efficiently as that of the corresponding In(3+) complex.

Topics: Catalysis; Chlorophyll; Nucleic Acid Hybridization; Nucleic Acids; Oligodeoxyribonucleotides; Organometallic Compounds; Photochemical Processes

A DNA quadruplex system that exhibits ultrafast intramolecular energy transfer is discussed.

Topics: Base Sequence; Chlorophyll; Energy Transfer; G-Quadruplexes; Nucleic Acid Conformation; Oligonucleotides

The limitation of specific delivery of photosensitizers to tumor sites, represents a significant shortcoming of photodynamic therapy (PDT) application at present. Prostate-specific membrane antigen (PSMA), a validated biomarker for prostate cancer, has attracted considerable attention as a target for imaging and therapeutic applications for prostate cancer. The present study focuses on the investigation of a PSMA inhibitor-conjugate of pyropheophorbide-a (Ppa-conjugate 2.1) for a targeted PDT application and the mechanism of its mediated-cell death in prostate cancer cells. Multiple fluorescence labeling methods were employed to monitor PDT-treated prostate cancer cells by confocal laser scanning microscopy. Our results demonstrate that Ppa-conjugate 2.1 mediated apoptosis is specific to PSMA+ (positive) LNCaP cells, but not PSMA- (negative) PC-3 cells. Furthermore, these results indicate that following PDT, the activation of caspase-8, -3, -9, cleavage of poly(ADP-ribose) polymerase (PARP) and DNA fragmentation is sequential. The appearance of cleaved beta-actin further supported involvement of caspase-3. Specific caspase inhibitor blocking studies reveal that the caspase-8/-3 cascade pathway plays a key role in apoptosis of LNCaP cells induced by Ppa-conjugate 2.1. The demonstrated selective targeting and efficacy of this agent suggests that targeted PDT could serve as an alternative treatment option for prostate cancer.

Topics: Antineoplastic Agents; Apoptosis; Blotting, Western; Caspase 3; Caspase 8; Caspase Inhibitors; Cell Line, Tumor; Cell Survival; Chlorophyll; Cysteine Proteinase Inhibitors; Dose-Response Relationship, Drug; Enzyme Activation; Fluorescent Antibody Technique; Humans; In Situ Nick-End Labeling; Inhibitory Concentration 50; Male; Microscopy, Confocal; Photochemotherapy; Photosensitizing Agents; Poly(ADP-ribose) Polymerases; Prostate-Specific Antigen; Prostatic Neoplasms; Signal Transduction; Time Factors

Photodynamic Therapy (PDT) is a minimally invasive procedure used for treating a range of neoplastic diseases, which utilises combined action of light and a PDT drug called a photosensitiser. The efficiency of this treatment depends crucially on the properties of the photosensitiser used, namely on its efficient uptake by cells or by the surrounding vasculature, intracellular localisation, minimal dark toxicity and substantial phototoxicity. In this report we compare the spectroscopic properties, cell uptake and in vitro phototoxicity of two novel hydrophilic photosensitisers derived from pyropheophorbide-a (PPa). Both new photosensitisers have the potential to form bioconjugates with antibody fragments for targeted PDT. We find that the photophysical properties of both new photosensitisers are favourable compared to the parent PPa, including enhanced absorption in the red spectral region and substantial singlet oxygen quantum yields. Both molecules show efficient cellular uptake, but display a different intracellular localisation. Both new photosensitisers exhibit no significant dark-toxicity at concentrations of up to 100 microM. The phototoxicity of the two photosensitisers is strikingly different, with one derivative being 13 times more efficient than the parent PPa and another derivative being 18 times less efficient in SKOV3 ovarian cancer cells. We investigate the reasons behind such drastic differences in phototoxicity using confocal fluorescence microscopy and conclude that intracellular localisation is a crucial factor in the photodynamic efficiency of pheophorbide derivatives. These studies highlight the underlying factors behind creating more potent photosensitisers through synthetic manipulation.

Topics: Biological Transport; Chlorophyll; Humans; KB Cells; Microscopy, Confocal; Photosensitizing Agents; Spectrometry, Fluorescence

Whole-cell patch-clamp recordings from single cultured mammalian neurons have been used to provide insight into early membrane-dependent events that result upon the intracellular photosensitized production of singlet molecular oxygen, O(2)(a(1)Δ(g)). The singlet oxygen sensitizers used, pyropheophorbide a (PPa) and protoporphyrin IX (PpIX), locate mainly in cell membranes and mitochondria, respectively. Irradiation of these sensitizers altered both passive and dynamic electrophysiological properties of the neurons in a dose-dependent manner, though the response threshold was much lower with PPa than with PpIX. In particular, notable decreases were observed in the rising and falling rates of action potentials and, at higher light fluences, plateau potentials consistent with activation of Ca(2+) channels also developed. The data suggest that singlet oxygen production specifically influences Na(+), K(+) and Ca(2+) ionophores in the cell membrane. Upon terminating sensitizer irradiation, responses evoked by PPa stabilized immediately whereas those evoked by PpIX continued to develop. These data are consistent with a spatially-resolved sphere of intracellular singlet oxygen activity. While the response to PPa irradiation appears to be membrane specific, the response to PpIX irradiation appears to be systemic and possibly part of a cascade of apoptotic events. These results should contribute to a better understanding of membrane-dependent events pertinent to cell death mediated by singlet oxygen.

Topics: Animals; Apoptosis; Calcium; Calcium Compounds; Cell Membrane; Cells, Cultured; Chlorates; Chlorophyll; Fluorescent Dyes; Hippocampus; Kinetics; Membrane Potentials; Neurons; Photosensitizing Agents; Potassium; Protoporphyrins; Rats; Rats, Wistar; Singlet Oxygen; Sodium

The lack of specific delivery of photosensitizers (PSs), represents a significant limitation of photodynamic therapy (PDT) of cancer. The biomarker prostate-specific membrane antigen (PSMA) has attracted considerable attention as a target for imaging and therapeutic applications for prostate cancer. Although recent efforts have been made to conjugate inhibitors of PSMA with imaging agents, there have been no reports on PS-conjugated PSMA inhibitors for targeted PDT of prostate cancer. The present study focuses on the use of a PSMA inhibitor-conjugate of pyropheophorbide-a (Ppa-conjugate 2) for targeted PDT to achieve apoptosis in PSMA+ LNCaP cells.. Confocal laser scanning microscopy with a combination of nuclear staining and immunofluorescence methods were employed to monitor the specific imaging and PDT-mediated apoptotic effects on PSMA-positive LNCaP and PSMA-negative (PC-3) cells.. Our results demonstrated that PDT-mediated effects by Ppa-conjugate 2 were specific to LNCaP cells, but not PC-3 cells. Cell permeability was detected as early as 2 hr by HOE33342/PI double staining, becoming more intense by 4 hr. Evidence for the apoptotic caspase cascade being activated was based on the appearance of poly-ADP-ribose polymerase (PARP) p85 fragment. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay detected DNA fragmentation 16 hr post-PDT, confirming apoptotic events.. Cell permeability by HOE33342/PI double staining as well as PARP p85 fragment and TUNEL assays confirm cellular apoptosis in PSMA+ cells when treated with PS-inhibitor conjugate 2 and subsequently irradiated. It is expected that the PSMA targeting small-molecule of this conjugate can serve as a delivery vehicle for PDT and other therapeutic applications for prostate cancer.

Topics: Apoptosis; Cell Line, Tumor; Chlorophyll; DNA Fragmentation; DNA, Neoplasm; Humans; Male; Photochemotherapy; Prostate-Specific Antigen; Prostatic Neoplasms

Comparative spectroscopic study including the photosensitizers of pyropheophorbide methyl ester (PPME) and pyropheophorbide a (PPa) was performed to study their photodynamic activity. The investigated photosensitizers in a homogeneous system of dimethylformamide (DMF) are not photostable upon irradiation. The photobleaching efficiency of PPa is higher than that of PPME. Combining these results with the data obtained by measuring the singlet oxygen quantum yield and the hydroxyl group generation, it was revealed that the photobleaching efficiency could be correlated with the singlet oxygen quantum yield and the hydroxyl group production of the photosensitizer.

Topics: Absorption; Algorithms; Chlorophyll; Dimethylformamide; Kinetics; Photobleaching; Photosensitizing Agents; Porphyrins; Singlet Oxygen; Spectrometry, Fluorescence

Potential sex and/or gametogenic stage differences in the metabolism of chlorophyll-a and carotenoids in the brown mussel Perna perna of southern Brazil were studied using high performance liquid chromatography (HPLC). Carotenoids derived directly from diet (phytoplankton) were fucoxanthin plus diatoxanthin (diatoms), alloxanthin (cryptophytes) and zeaxanthin (mainly cyanobacteria). Females accumulated carotenoid-diols and epoxides (~3-4 mg/g-dry wt.) while males had much lower concentrations (~0.7 mg/g-dry wt.). An antioxidant/free radical scavenging role is proposed for carotenoids in females. Mean ratios of chlorophyll plus derivatives (Chlns-a) to carotenoids for male and female P. perna were 50:1 and 4:1, respectively. The higher ratio in males relates to both higher carotenoid contents in females plus higher total Chlns-a in males (~22 mg/g-dry wt.), relative to the females (~4 mg/g-dry wt.). Chlorophyll-a metabolism in both sexes followed two distinct pathways. First, cyclization of pyropheophorbide-a gave 13(2), 17(3)-cyclopheophorbide-a-enol (CPPaE) which was further oxidized to hydroxy-chlorophyllone. Second, chlorophyll-a derivatives retaining the 13(2)-carbomethoxy moiety were oxidized to purpurin-18 which was hydrolyzed to chlorin-p(6). In both cases, metabolism of dietary chlorophyll-a was oxidative and derivatives could either serve as antioxidants or merely be the results of non-specific digestive processes.

Topics: Animals; Carotenoids; Chlorophyll; Chlorophyll A; Chromatography, High Pressure Liquid; Epoxy Compounds; Female; Male; Molecular Structure; Perna; Phytoplankton; Porphyrins; Xanthophylls; Zeaxanthins

Formation of pyropheophorbide (PyroPheid) during chlorophyll metabolism in some higher plants has been shown to involve the enzyme pheophorbidase (PPD). This enzyme catalyzes the conversion of pheophorbide (Pheid) a to a precursor of PyroPheid, C-13(2)-carboxylPyroPheid a, by demethylation, and then the precursor is decarboxylated non-enzymatically to yield PyroPheid a. In this study, expression, purification, and biochemical characterization of recombinant PPD from radish (Raphanus sativus L.) were performed, and its properties were compared with those of highly purified native PPD. Recombinant PPD was produced using a glutathione S-transferase (GST) fusion system. The PPD and GST genes were fused to a pGEX-2T vector and expressed in Escherichia coli under the control of a T7 promoter as a fusion protein. The recombinant PPD-GST was expressed as a 55 kDa protein as measured by SDS-PAGE and purified by single-step affinity chromatography through a GSTrap FF column. PPD-GST was purified to homogeneity with a yield of 0.42 mg L(-1) of culture. The protein purified by this method was confirmed to be PPD by measuring its activity. The purified PPD-GST fusion protein revealed potent catalytic activity for demethylation of the methoxycarbonyl group of Pheid a and showed a pH optimum, substrate specificity, and thermal stability quite similar to the native enzyme purified from radish, except for the Km values toward Pheid a: 95.5 microM for PPD-GST and about 15 microM for native PPDs.

Topics: Biocatalysis; Blotting, Western; Chlorophyll; Electrophoresis, Polyacrylamide Gel; Esterases; Glutathione Transferase; Hydrolysis; Raphanus; Recombinant Fusion Proteins; Time Factors

The photophysical properties of the novel hexapyropheophorbide a (P6), and hexakis (pyropheophorbide a)-C60 (FP6) were studied and compared with those of hexakis (pyropheophorbide a)-fullerene [5:1] hexaadduct (FHP6). It was found that after light absorption the pyropheophorbide a molecules in all three compounds undergo very efficient energy transfer as well as partly excitonic interactions. The last process results in the formation of energy traps, which could be resolved experimentally. For P6, due to shorter distances between neighboring dye molecules, stronger interactions between pyropheophorbide a units than for FHP6 were observed. As a consequence, the excitation energy is delivered rapidly to traps formed by stacked pyropheophorbide a molecules resulting in the reduction of fluorescence, intersystem crossing, and singlet oxygen quantum yields compared to the values of FHP6. For FP6 the reduction of these values is much stronger due to an additional fast and efficient deactivation process, namely photoinduced electron transfer from pyropheophorbide a to the fullerene moiety. Consequently, FP6 can be considered as a combination of a light-harvesting system consisting of several separate pyropheophorbide a molecules and a charge-separating center.

Topics: Chlorophyll; Electrons; Energy Transfer; Fullerenes; Models, Molecular; Molecular Structure; Photochemistry; Sensitivity and Specificity; Spectrophotometry, Ultraviolet

DNA sequence-controlled on-and-off switching of a singlet oxygen sensitizer has been developed and demonstrated. The singlet oxygen photosensitizer pyropheophorbide-a (P) was attached to a 15-mer nucleotide sequence. A molecule that could quench the sensitizer, the so-called "black hole quencher 3" (Q), was attached to a complementary nucleotide strand. Upon hybridization of the two conjugates, singlet oxygen production from P was completely shut down. Upon the addition of a third DNA sequence that can displace and release the P-DNA conjugate from the P-Q pair, up to 85% of the singlet oxygen production was recovered. This system is a model for a benign drug that becomes active only in the presence of a specific targeted nucleotide sequence.

Topics: Chlorophyll; DNA; Oligonucleotides; Photosensitizing Agents; Singlet Oxygen

We hypothesized that estrogen receptor (ER) in hormone-sensitive breast cancer cells could be targeted for selective photodynamic killing of tumor cell with antiestrogen-porphyrin conjugates by combining the over-expression of ER in hormone-sensitive breast cancer cells and tumor-retention property of porphyrin photosensitizers. In this study we describe that a tamoxifen (TAM)-pyropheophorbide conjugate that specifically binds to ER alpha, caused selective cell-kill in MCF-7 breast cancer cells upon light exposure. Therefore, it is a potential candidate for ER-targeted photodynamic therapy of cancers (PDT) of tissues and organs that respond to estrogens/antiestrogens.

Topics: Antineoplastic Agents, Hormonal; Breast Neoplasms; Cell Line, Tumor; Cell Survival; Chlorophyll; Estrogen Receptor alpha; Female; Humans; Molecular Structure; Photochemotherapy; Porphyrins; Selective Estrogen Receptor Modulators; Tamoxifen

In photodynamic therapy (PDT), a tumor-selective photosensitizer is administered followed by activation of the photosensitizer by exposure to a light source of a given wavelength. This, in turn, generates reactive oxygen species that induce cellular apoptosis and necrosis in tumor tissue. Based on our earlier finding that the photosensitizer pheophorbide a is an ABCG2 substrate, we explored the ability of ABCG2 to transport photosensitizers with a structure similar to that of pheophorbide a. ABCG2-overexpressing NCI-H1650 MX50 bronchoalveolar carcinoma cells were found to have reduced intracellular accumulation of pyropheophorbide a methyl ester and chlorin e6 compared to parental cells as measured by flow cytometry. The ABCG2 inhibitor fumitremorgin C was found to abrogate ABCG2-mediated transport. Intracellular fluorescence of hematoporphyrin IX, meso-tetra(3-hydroxyphenyl)porphyrin, and meso-tetra(3-hydroxyphenyl)chlorin was not substantially affected by ABCG2. ABCG2-overexpressing cells also displayed decreased intracellular fluorescence of protoporphyrin IX generated by exogenous application of 5-aminolevulinic acid. Mutations at amino acid 482 in the ABCG2 protein known to affect substrate specificity were not found to impact transport of the photosensitizers. In cytotoxicity assays, ABCG2-transfected HEK-293 cells were 11-fold, 30-fold, 4-fold, and >7-fold resistant to PDT with pheophorbide a, pyropheophorbide a methyl ester, chlorin e6, and 5-aminolevulinic acid, respectively. ABCG2-transfected cells were not resistant to PDT with meso-tetra(3-hydroxyphenyl) chlorin. Neither multidrug resistance-associated protein 1 expression nor P-glycoprotein expression appreciably decreased the intracellular fluorescence of any of the photosensitizers examined as determined by flow cytometry. The results presented here implicate ABCG2 as a possible cause for cellular resistance to photodynamic therapy.

Topics: Adenocarcinoma, Bronchiolo-Alveolar; Aminolevulinic Acid; ATP Binding Cassette Transporter, Subfamily B, Member 1; ATP Binding Cassette Transporter, Subfamily G, Member 2; ATP-Binding Cassette Transporters; Biological Transport; Cell Proliferation; Chlorophyll; Chlorophyllides; Flow Cytometry; Fluorescence; Humans; Indoles; Kidney; Lung Neoplasms; Multidrug Resistance-Associated Proteins; Neoplasm Proteins; Photochemotherapy; Photosensitizing Agents; Porphyrins; Protoporphyrins; Tumor Cells, Cultured

A series of pyropheophorbide-a and bacteriopurpurinimides were investigated to understand the correlation between HSA (site II) binding affinity and in vivo photosensitizing activity. In our study, photosensitizers that bound to site II of HSA produced a significant difference in the circular dichroism spectra of the corresponding complexes, especially at Soret band region of the photosensitizers. Our results suggest that CD spectroscopy of the photosensitizer-HSA complexes could be a valuable tool in screening new photosensitizers before evaluating them for in vivo efficacy.

Topics: Animals; Binding Sites; Chlorophyll; Circular Dichroism; Drug Carriers; Humans; Mice; Mice, Inbred C3H; Photochemotherapy; Photosensitizing Agents; Protein Binding; Purinones; Serum Albumin; Xenograft Model Antitumor Assays

The synthesis and photophysical studies of a fullerene [6:0]-hexaadduct that carries 12 pyropheophorbide a units are reported. The synthesis started with the malonate 1, which was coupled under template conditions to C(60)() to give the hexaadduct 2. After removal of the protecting group with acid the dodecakis amino-substituted precursor compound 3 was generated. 3 was not isolated but directly reacted with the N-succinimid ester 4 of pyropheophorbide a (5), which delivered the desired fullerene [6:0]-hexaadduct 6 in excellent yield. The photophysical properties of 6 were studied and compared with those of the fullerene [5:1]-hexaadduct 7 with six pyropheophorbide a groups and the bispyropheophorbide a-fullerene [5:1]-hexaadduct 8. The pyropheophorbide a units in 6 undergo after light absorption very efficient energy transfer as well as partly excitonic interaction. The last process results in formation of energy traps, which could be resolved experimentally. Compared to the reference compounds 7 and 8, 6 has a higher probability of trap formation due to a higher local concentration of dye molecules and shorter distances between them. As a consequence, the excitation energy is delivered rapidly (within 23 ps) to the traps, resulting in decreases of the fluorescence, intersystem crossing, and singlet oxygen quantum yields in comparison with the values of the reference compounds.

Topics: Biomimetic Materials; Chlorophyll; Drug Delivery Systems; Fullerenes; Models, Molecular; Photosynthetic Reaction Center Complex Proteins; Spectrometry, Fluorescence

The main challenge in searching for new photosensitizers is to improve their specificity for target cells to avoid toxicity towards normal cells. New modular drug delivery systems were proposed consisting of a multiplying unit with the property of carrying several drug moieties and an addressing unity with high selectivity for target cells. Following this concept, two new fullerene-bis-pyropheophorbide a derivatives were synthesized: a mono-(FP1) and a hexa-adduct (FHP1). The photophysical characterization of the compounds revealed significantly different parameters related to the number of addends at the fullerene core. In this study, the derivatives were tested with regard to their intracellular uptake and photosensitizing activity towards human leukemia T-lymphocytes (Jurkat cells) in comparison with the free sensitizer, pyropheophorbide a. The C(60)-hexa-adduct FHP1 resulted to have a significative phototoxic activity (58% dead cell, after a dose of 400 mJ/cm(2), 688 nm) while the mono-adduct FP1 had a very low phototoxicity and only at higher light doses. The photosensitizing activity of the fullerene hexa-adduct, FHP1, resulted to be lower than that of pyropheophorbide a. The lesser intracellular concentration reached by the C(60)-hexa-adduct FHP1 is probably the reason for its lower phototoxicity with respect to pyropheophorbide a.

Topics: Cell Survival; Chlorophyll; Fullerenes; Humans; Jurkat Cells; Light; Photosensitizing Agents

In an outbreak of food poisoning involving a dried purple laver product (called nori), four persons had allergic-like symptoms such as inflammation and red rash on their face, mouth and belly. The causative nori was extracted and smeared on the arm-skin of five volunteers. Three out of five volunteers had a slight allergic reaction after 5 to 30 min when they were exposed to sunlight. The levels of the chlorophyll derivatives, pheophorbide a and pyropheophorbide a, measured by HPLC were 851-906 and 5,460-5,624 microg/g, respectively, in the causative samples. Judging from the high contents of pyropheophorbide a and pheophorbide a and the symptoms of patients and volunteers, the causative agents were concluded to be the photosensitizers pyropheophorbide a and pheophorbide a.

Topics: Adult; Chlorophyll; Chromatography, High Pressure Liquid; Disease Outbreaks; Food Hypersensitivity; Foodborne Diseases; Humans; Male; Photosensitizing Agents; Seaweed

Multi-targeting strategies improve the efficacy of antibody and immunotoxin therapies but have not yet been thoroughly explored for HER2-based cancer treatments. We investigated multi-epitope HER2 targeting to boost photosensitizer immunoconjugate uptake as a way of enhancing photoimmunotherapy. Photoimmunotherapy may allow targeted photodynamic destruction of malignancies and may also potentiate anticancer antibodies. However, one obstacle preventing its clinical use is the delivery of enough photosensitizer immunoconjugates to target cells. Anti-HER2 photosensitizer immunoconjugates were constructed from two monoclonal antibodies (mAb), HER50 and HER66, using a novel method originally developed to label photosensitizer immunoconjugates with the photosensitizer, benzoporphyrin derivative verteporfin. Photosensitizer immunoconjugates were labeled instead with a promising alternative photosensitizer, pyropheophorbide-a (PPa), which required only minor changes to the conjugation procedure. Uptake and phototoxicity experiments using human cancer cells were conducted with the photosensitizer immunoconjugates and, for comparison, with free PPa. SK-BR-3 and SK-OV-3 cells served as HER2-overexpressing target cells. MDA-MB-468 cells served as HER2-nonexpressing control cells. Photosensitizer immunoconjugates with PPa/mAb molar ratios up to approximately 10 specifically targeted and photodynamically killed HER2-overexpressing cells. On a per mole basis, photosensitizer immunoconjugates were less phototoxic than free PPa, but photosensitizer immunoconjugates were selective for target cells whereas free PPa was not. Multiepitope targeted photoimmunotherapy with a HER50 and HER66 photosensitizer immunoconjugate mixture was significantly more effective than single-epitope targeted photoimmunotherapy with a single anti-HER2 photosensitizer immunoconjugate, provided photosensitizer immunoconjugate binding was saturated. This study shows that multiepitope targeting enhances HER2-targeted photoimmunotherapy and maintains a high degree of specificity. Consequently, it seems that multitargeted photoimmunotherapy should also be useful against cancers that overexpress other receptors.

Topics: Breast Neoplasms; Cell Line, Tumor; Chlorophyll; Epitopes; Female; Humans; Immunotherapy; Immunotoxins; Ovarian Neoplasms; Photochemotherapy; Photosensitizing Agents; Receptor, ErbB-2

We report a new concept for type-II photosensitization, based on incorporating the photosensitizer (PS) and a singlet-oxygen (1O2) quenching/scavenging molecule onto a disease-targeting linker, such that the PS becomes activatable by light only when targeting has occurred. In this first proof-of-concept report, a model photosensitizing beacon was synthesized containing a pyropheophorbide as the PS and a carotenoid as the 1O2 quencher. These were kept in close proximity by the self-folding of a caspase-3-specific peptide sequence. Upon caspase-3-induced cleavage, the 1O2 production increased markedly, as measured directly by 1O2 near-infrared luminescence and lifetime measurements.

Topics: Carotenoids; Caspase 3; Caspase Inhibitors; Caspases; Chlorophyll; Luminescent Measurements; Oligopeptides; Photosensitizing Agents; Singlet Oxygen

The ethylene ketal of pyropheophorbides, chlorophylls possessing the 13-keto carbonyl group and lacking the 13(2)-methoxycarbonyl group, reacted with H(2)(18)O (ca. 95% 18O atom) by acidic hydrolysis to give efficiently and regioselectively 13(1)-18O-oxo-labelled compounds (ca. 92% 18O). The resulting 18O-labelled chlorin was modified by several chemical reactions to afford some derivatives with little loss of the 18O atom. Following the same procedures, 3(1),13(1)-doubly-18O-labelled pyrochlorophyll derivatives were also prepared. All the synthetic 18O-labelled compounds were identified by FAB-mass and vibrational spectra. Especially, in the vibrational spectroscopic results including IR and resonance Raman spectra, an about 30 cm(-1) wavenumber down-shift of the 3- and/or 13-C[double bond]O stretching vibrational bands was observed by exchanging 3(1)- or 13(1)-oxo-oxygen atom from 16O to 18O.

Topics: Acetals; Chlorophyll; Chlorophyll A; Dioxolanes; Hydrogen-Ion Concentration; Hydrolysis; Isotope Labeling; Magnetic Resonance Spectroscopy; Molecular Structure; Oxidation-Reduction; Oxygen; Oxygen Isotopes; Porphyrins; Spectroscopy, Fourier Transform Infrared

A study has been carried out to define the importance of the peripheral benzodiazepine receptor (PBR) as a binding site for a series of chlorin-type photosensitizers, pyropheophorbide-a ethers, the subject of a previous quantitative structure-activity relationship study by us. The effects of the PBR ligand PK11195 on the photodynamic activity have been determined in vivo for certain members of this series of alkyl-substituted ethers: two of the most active derivatives (hexyl and heptyl), the least active derivative (dodecyl [C12]) and one of intermediate activity (octyl [C8]). The photodynamic therapy (PDT) effect was inhibited by PK11195 for both of the most active derivatives, but no effect on PDT activity was found for the less active C12 or C8 ethers. The inhibitory effects of PK11195 were predicted by the binding of only the active derivatives to the benzodiazepine site on albumin, ie. human serum albumin (HSA)-Site II. Thus, as with certain other types of photosensitizers, it has been demonstrated with this series of pyropheophorbide ethers that in vitro binding to HSA-Site II is a predictor of both optimal in vivo activity and binding to the PBR in vivo.

Topics: Animals; Binding Sites; Chlorophyll; Humans; In Vitro Techniques; Isoquinolines; Mice; Mice, Inbred C3H; Photochemotherapy; Photosensitizing Agents; Receptors, GABA-A; Sarcoma, Experimental; Serum Albumin

Amphiphilic sensitizers self-associate in aqueous environments and form aggregated species that exhibit no or only negligible photodynamic activity. However, amphiphilic photosensitizers number among the most potent agents of photodynamic therapy. The processes by which these sensitizers are internalized into tumor cells have yet to be fully elucidated and thus remain the subject of debate. In this study the uptake of photosensitizer aggregates into tumor cells was examined directly using subcellular time-resolved fluorescence spectroscopy with a high temporal resolution (20-30 ps) and high sensitivity (time-correlated single-photon counting). The investigations were performed on selected sensitizers that exhibit short fluorescence decay times (< 50 ps) in aggregated form. Derivatives of pyropheophorbide-a ether and chlorin e6 with varying lipophilicity were used for the study. The characteristic fluorescence decay times and spectroscopic features of the sensitizer aggregates measured in aqueous solution also could be observed in A431 human endothelial carcinoma cells administered with these photosensitizers. This shows that tumor cells can internalize sensitizers in aggregated form. Uptake of aggregates and their monomerization inside cells were demonstrated directly for the first time by means of fluorescence lifetime imaging with a high temporal resolution. Internalization of the aggregates seems to be endocytosis mediated. The degree of their monomerization in tumor cells is strongly influenced by the lipophilicity of the compounds.

Topics: Chlorophyll; Chlorophyllides; Fluorescence; Humans; Kinetics; Light; Molecular Structure; Photons; Photosensitizing Agents; Porphyrins; Solutions; Spectrometry, Fluorescence; Time Factors; Tumor Cells, Cultured

Chlorophyll degradation was investigated in cells of a chlorophyll b-less mutant of Chlamydomonas reinhardtii under aerobic and anaerobic conditions. During degradation of chlorophyll under anaerobic conditions, chlorophyll catabolite P535, an open-tetrapyrrole, was not excreted, but pyropheophorbide a was accumulated as the end product with a transient accumulation of chlorophyllide a and pheophorbide a in cells, in contrast to the breakdown under aerobic conditions. It is likely that in the absence of oxygen, degradation of chlorophyll a proceeds to pyropheophorbide a by three consecutive reactions, dephytylation, metal-releasing and demethoxycarbonylation, and then stops due to a limitation of the oxygen that the monooxygenase reaction requires for bilin formation. A novel enzyme catalyzing demethoxycarbonylation of pheophorbide a was partially purified. The enzyme activity increased dependent on the age of cells, and its increase was completely suppressed by cycloheximide. Production of P535 was also dependent on cytoplasmic protein synthesis.

Topics: Anaerobiosis; Animals; Chlamydomonas reinhardtii; Chlorophyll; Mutation; Protein Synthesis Inhibitors

Canine hemangiopericytomas are a commonly occurring neoplasm with a clinical course of recurrence after surgical removal. This study sought to evaluate Photochlor (HPPH) photodynamic therapy (HPPH-PDT) as an adjuvant therapy to prevent recurrence of tumor after surgical removal.. Sixteen dogs with naturally occurring hemangiopericytomas were treated with surgical removal of the tumor followed by PDT using Photochlor as the photosensitizer. Photochlor was injected intravenously at a dose of 0.3 mg/kg. Forty-eight hours later the treatment consisted of surgical removal of the tumor followed by HPPH-PDT.. Nine dogs (56%) had recurrence of tumor from 2 to 29 (median 9) months after treatment. These results are comparable or not as good as other forms of therapy.. Photochlor photodynamic therapy applied after surgery appears to have no advantage over other forms of therapy in regards to preventing recurrence. Delayed wound healing and infections are problematic and make HPPH-PDT an undesirable addition to surgery for the treatment of this tumor type.

Topics: Animals; Chlorophyll; Dog Diseases; Dogs; Female; Forelimb; Hemangiopericytoma; Hindlimb; Male; Neoplasm Recurrence, Local; Photochemotherapy; Postoperative Care

Pyropheophorbide-a methyl ester (PPME) is a second generation of photosensitizers used in photodynamic therapy. We demonstrated that PPME photosensitization activated NF-kappaB transcription factor in colon cancer cells. Unexpectedly, this activation occurred in two separate waves, i.e. a rapid and transient one and a second slower but sustained phase. The former was due to photosensitization by PPME localized in the cytoplasmic membrane which triggered interleukin-1 receptor internalization and the transduction pathways controlled by the interleukin-1 type I receptor. Indeed, TRAF6 dominant negative mutant abolished NF-kappaB activation by PPME photosensitization, and TRAF2 dominant negative mutant was without any effect, and overexpression of IkappaB kinases increased gene transcription controlled by NF-kappaB. Oxidative stress was not likely involved in the activation. On the other hand, the slower and sustained wave could be the product of the release of ceramide through activation of the acidic sphingomyelinase. PPME localization within the lysosomal membrane could explain why ceramide acted as second messenger in NF-kappaB activation by PPME photosensitization. These data will allow a better understanding of the molecular basis of tumor eradication by photodynamic therapy, in particular the importance of the host cell response in the treatment.

Topics: Chlorophyll; Colonic Neoplasms; Humans; Lysosomes; Models, Chemical; NF-kappa B; Photochemotherapy; Proteins; Receptors, Interleukin-1; Signal Transduction; Sphingomyelin Phosphodiesterase; TNF Receptor-Associated Factor 2; Transcriptional Activation; Tumor Cells, Cultured

An open three-compartment pharmacokinetic model was applied to the in vivo quantitative structure-activity relationship (QSAR) data of a homologous series of pyropheophorbide photosensitizers for photodynamic therapy (PDT). The physical model was a lipid compartment sandwiched between two identical aqueous compartments. The first compartment was assumed to clear irreversibly at a rate K0. The measured octanol-water partition coefficients, P(i) (where i is the number of carbons in the alkyl chain) and the clearance rate K0 determined the clearance kinetics of the drugs. Solving the coupled differential equations of the three-compartment model produced clearance kinetics for each of the sensitizers in each of the compartments. The third compartment was found to contain the target of PDT. This series of compounds is quite lipophilic. Therefore these drugs are found mainly in the second compartment. The drug level in the third compartment represents a small fraction of the tissue level and is thus not accessible to direct measurement by extraction. The second compartment of the model accurately predicted the clearance from the serum of mice of the hexyl ether of pyropheophorbide a, one member of this series of compounds. The diffusion and clearance rate constants were those found by fitting the pharmacokinetics of the third compartment to the QSAR data. This result validated the magnitude and mechanistic significance of the rate constants used to model the QSAR data. The PDT response to dose theory was applied to the kinetic behavior of the target compartment drug concentration. This produced a pharmacokinetic-based function connecting PDT response to dose as a function of time postinjection. This mechanistic dose-response function was fitted to published, single time point QSAR data for the pheophorbides. As a result, the PDT target threshold dose together with the predicted QSAR as a function of time postinjection was found.

Topics: Animals; Chlorophyll; Metabolic Clearance Rate; Mice; Models, Biological; Neoplasms, Experimental; Photochemotherapy; Photosensitizing Agents; Structure-Activity Relationship

To determine if subcellular localization is important to photodynamic therapy (PDT) efficacy, an in vitro fluorescence microscopy study was conducted with a congeneric series of pyropheophorbide-a derivatives in human pharyngeal squamous cell carcinoma (FaDu) cells and murine radiation-induced fibrosarcoma (RIF) mutant cells. In the FaDu cells the octyl, decyl and dodecyl ether derivatives localized to the lysosomes at extracellular concentrations less than needed to produce a 50% cell kill (LD50). At extracellular concentrations equal or greater than the LD50 the compounds localized mainly to mitochondria. The propyl, pentyl, hexyl and heptyl ether derivatives localized mainly to the mitochondria at all concentrations studied. This suggested that mitochondria are a sensitive PDT target for these derivatives. Similar experiments were performed with two Photofrin-PDT resistant RIF cell lines, one of which was found to be resistant to hexyl ether derivative (C6) mediated-PDT and the other sensitive to C6-PDT relative to the parent line. At extracellular concentrations of C6 below the LD50 of each cell line, the mutants exhibited lysosomal localization. At concentrations above these values the patterns shifted to a mainly mitochondrial pattern. In these cell lines mitochondrial localization also correlated with PDT sensitivity. Localization to mitochondria or lysosomes appeared to be affected by the aggregation state of the congeners, all of which are highly aggregated in aqueous medium. Monomers apparently were the active fraction of these compounds because equalizing the extracellular monomer concentrations produced equivalent intracellular concentrations, photoxicity and localization patterns. Compounds that were mainly aggregates localized to the lysosomes where they were rendered less active. Mitochondria appear to be a sensitive target for pyropheophorbide-a-mediated photodamage, and the degree of aggregation seems to be a determinant of the localization site.

Topics: Animals; Chlorophyll; Humans; Lethal Dose 50; Mice; Microscopy, Fluorescence; Mutation; Photochemotherapy; Photosensitizing Agents; Subcellular Fractions; Tumor Cells, Cultured

Antiviral activity-guided isolation studies on the leaves of Atalantia monophylla (ROXB.) CORR. (Rutaceae) led to the identification of pyropheophorbide a (1), a simple chlorin derivative, from the chloroform extract (fr. B) as a possible antiviral active principle against herpes simplex virus type 2 (HSV-2). Pyropheophorbide a methyl ester (2) was also isolated from the hexane extract (fr. A).

Topics: Animals; Antiviral Agents; Chlorocebus aethiops; Chlorophyll; Herpesvirus 2, Human; Plant Extracts; Vero Cells; Viral Plaque Assay

Topics: Animals; Chlorophyll; Male; Photosensitivity Disorders; Rats

Topics: Chlorella; Chlorophyll; Diterpenes; Gas Chromatography-Mass Spectrometry; Hydrolysis; Phytol; Vegetables