Nicole Brasseur

BIOLOGICAL ACTIVITIES OF PHTHALOCYANINES—IV. TYPE II SENSITIZED PHOTOOXIDATION OF L-TRYPTOPHAN AND CHOLESTEROL BY SULFONATED METALLO PHTHALOCYANINES

Abstract— Sulfonated phthalocyanine and a series of its metal chelates in combination with red light irradiation led to the degradation of L‐tryptophan in oxygenated aqueous solution. The photoproducts and the rate of transformation of L‐tryptophan are compared with hematoporphyrin and rose bengal sensitized photooxidation. In all cases the primary photoproducts are characterized as cis and trans‐3a‐hydroperoxy‐l,2,3,3a,8,8a‐hexahydropyrrolo[2,3‐b]indole‐2‐carboxylic acid. Support for the involvement of singlet excited oxygen is obtained from azide inhibition and the formation of the specific singlet oxygen product with cholesterol. We observed the contribution of another pathway in the case of the manganese complex.

BIOLOGICAL ACTIVITIES OF PHTHALOCYANINES-X. SYNTHESES AND ANALYSES OF SULFONATED PHTHALOCYANINES

Abstract— Synthetic methods to obtain selectively sulfonated metallo phthalocyanines are compared. Both condensation and direct sulfonation procedures lead to mixtures of mono‐ to tetrasulfonated products which are resolved by reverse phase liquid chromatography in buffered aqueous‐methanol. The proportion of sulfonated derivatives is examined as a function of the starting reagents in the case of the condensation method, and as a function of the temperature and reaction time in the case of the direct sulfonation procedure. The number of sulfonate groups per phthalocyanine molecule is determined by oxidative degradation of the phthalocyanine ring followed by quantitative chromatographic analysis of the sulfophthalamide and phthalamide fragments.

Preparation and characterization of ph-responsive polymeric micelles for the delivery of photosensitizing anticancer drugs

pH-responsive polymeric micelles (PM) consisting of random copolymers of N-isopropylacrylamide (NIPA), methacrylic acid (MAA), and octadecyl acrylate (ODA) were prepared and characterized. The critical aggregation concentration, as determined by a fluorescence probe technique, was approximately 10 mg/L in water and phosphate-buffered saline. Phase transition pH was estimated at 5.7. The decrease in pH was accompanied by the destruction of hydrophobic clusters. Micelle size was dependent on temperature and the nature of the aqueous medium. The micelles were successfully loaded with a substantial amount of a photoactive anticancer drug, namely, aluminum chloride phthalocyanine (AlClPc). pH-responsive PM loaded with AlClPc were found to exhibit higher cytotoxicity against EMT-6 mouse mammary cells in vitro than control Cremophor EL formulation. These results show the potential of poly(NIPA-co-MAA-co-ODA) for in vivo administration of water-insoluble, photosensitizing anticancer drugs.

BIOLOGICAL ACTIVITIES OF PHTHALOCYANINES—VI. PHOTOOXIDATION OF L‐TRYPTOPHAN BY SELECTIVELY SULFONATED GALLIUM PHTHALOCYANINES: SINGLET OXYGEN YIELDS AND EFFECT OF AGGREGATION

Abstract— The photosensitized oxidation of L‐tryptophan by gallium phthalocyanines sulfonated to different degrees is studied as a function of both substrate and sensitizer concentrations in water and 50% MeOH‐H2O solutions. The maximum quantum yield of singlet oxygen was found to be nearly 0.5 for all sulfonated gallium complexes. The effect of adding sulfonate groups in the phthalocyanine backbone is to change the tendency of dye molecules to dimerize or aggregate in a particular solvent. A shift in the chemical equilibrium away from the monomeric state, which occurs at high dye concentrations and at lower degrees of dye sulfonation, results in a reduced photochemical yield. The variation of quantum yields in different solvent systems and at several wavelengths is similarly accounted for by the fraction of light absorbed by the productive monomer state.

BIOLOGICAL ACTIVITIES OF PHTHALOCYANINES‐IX. PHOTOSENSITIZATION OFV–79 CHINESE HAMSTER CELLS ANDEMT–6 MOUSE MAMMARY TUMOR BY SELECTIVELY SULFONATED ZINC PHTHALOCYANINES

Abstract— Zinc phthalocyanines sulfonated to different degrees are tested for their ability to sensitizeV–79 Chinese hamster cells andEMT–6 mouse mammary tumors to red light. In vitro, the lower sulfonated derivatives were the most active with the exception of the poorly water‐soluble monosulfonated dye. An isomeric mixture of tetrasulfonated derivatives obtained via direct sulfonation was ten times more active than the homogeneous tetrasulfo derivative prepared via the condensation of sulfophthalic acid. In vivo, the latter dye was completely inactive, whereas the remainder of the sulfonated preparations exhibited a similar structure‐activity pattern as observed with theV–79 cells in vitro. The disulfonated zinc phthalocyanine showed the best tumoricidal activity in the series and also appeared to be a more efficient photosensitizer of cell inactivation and tumor cure than the aluminum or gallium complexes as well as hematoporphyrin derivative preparations. No significant differences in skin phototoxicity were observed among the various dyes.

BIOLOGICAL ACTIVITIES OF PHTHALOCYANINES—V. PHOTODYNAMIC THERAPY OF EMT‐6 MAMMARY TUMORS IN MICE WITH SULFONATED PHTHALOCYANINES

Abstract— We tested water‐soluble sulfonated phthalocyanine and three metal chelate derivatives for their tumoricidal effect on the EMT‐6 mammary tumor in mice exposed to red light. The metal‐free sulfophthalocyanine had little effect, whereas the aluminum complex and the lower sulfonated fraction of the gallium complex exhibited tumoricidal activity similar to hematoporphyrin‐based photosensitizer (Photofrin II). The higher sulfonated fractions of the gallium complex were less active as compared to the lower sulfonated fraction. The cerium complex was the most active sensitizer in terms of dye and light doses required to induce tumor necrosis and cure but also showed the highest phototoxicity towards healthy skin. These results suggest that sulfonated phthalocyanines will offer a new alternative in photodynamic therapy of light‐accessible neoplasms.

PEG-coated Poly(lactic acid) Nanoparticles for the Delivery of Hexadecafluoro Zinc Phthalocyanine to EMT-6 Mouse Mammary Tumours

Hexadecafluoro zinc phthalocyanine (ZnPcF16), a second generation sensitizer for the photodynamic therapy of cancer, was incorporated in three vehicles: poly(d,l‐lactic acid) (PLA) nanoparticles, polyethylene glycol (PEG)‐coated nanoparticles and a Cremophor EL (CRM) oil‐water emulsion. Nanoparticles were prepared by the salting‐out procedure. Biodistribution of the dye was assessed by fluorescence in EMT‐6 mammary tumour bearing mice after intravenous injection of 1 μmol kg−1 ZnPcF16.

Water-soluble aluminium phthalocyanine-polymer conjugates for PDT: photodynamic activities and pharmacokinetics in tumour-bearing mice.

SummaryThe potential use of unsubstituted aluminium phthalocyanine (AlClPc) as a sensitizer for photodynamic therapy (PDT) of cancer has not been fully exploited in spite of its higher efficiency as compared to the sulphonated derivatives. This is largely due to the strong hydrophobic character of AlClPc which renders the material difficult to formulate for in vivo administration. We prepared two water-soluble derivatives of AlClPc by axial coordination of polyethyleneglycol (PEG, MW 2000) or polyvinylalcohol (PVA, MW 13 000–23 000) to the central aluminium ion. Their photodynamic activities were evaluated in vitro against the EMT-6 mouse mammary tumour cells and in vivo against the EMT-6 and the colon carcinoma Colo-26 tumours implanted intradermally in Balb/c mice. Pharmacokinetics were studied in the EMT-6 tumour-bearing mice. After 1 h incubation, the light dose required to kill 90% of cells (LD90) was at least three times less for AlClPc (Cremophor emulsion) as compared to AlPc–PEG and AlPc–PVA, while after 24 h incubation all three preparations were highly phototoxic. All three dye preparations induced complete EMT-6 tumour regression in 75–100% of animals at a low drug dose (0.25 μmol kg–1) following PDT (400 J cm–2, 650–700 nm) at 24 h pi. Complete tumour regression in the Colo-26 tumour model was obtained in 30% of mice at a dose of 2 μmol kg–1. In the non-cured animals, AlPc–PVA induced the most significant tumour growth delay. This dye showed a prolonged plasma half-life (6.8 h) as compared to AlClPc (2.6 h) and AlPc–PEG (23 min), lower retention by liver and spleen and higher tumour-to-skin and tumour-to-muscle ratios. Our data demonstrate that addition of hydrophilic axial ligands to AlPc, while modifying in vitro and in vivo kinetics, does not reduce the PDT efficiency of the parent molecule. Moreover, in the case of the polyvinylalcohol derivative, axial coordination confers advantageous pharmacokinetics to AlPc, which makes this photosensitizer a valuable, water soluble candidate drug for clinical PDT of cancer.

Photodynamic therapy of tumours with hexadecafluoro zinc phthalocyanine formulated in PEG-coated poly(lactic acid) nanoparticles

Hexadecafluoro zinc phthalocyanine (ZnPcF16), a second‐generation sensitizer for the photodynamic therapy (PDT) of cancer, was formulated in polyethylene‐glycol‐coated poly(lactic acid) nanoparticles (PEG‐coated PLA‐NP) and tested in EMT‐6 tumour‐bearing mice for its photodynamic activity. The tumour response was compared to that induced by the same dye formulated as a Cremophor EL (CRM) emulsion. Formulation in the biodegradable NP improved PDT response of the tumour while providing prolonged tumour sensitivity towards PDT.

Biological activities of phthalocyanines. III: Photoinactivation of V-79 chinese hamster cells by tetrasulfophthalocyanines

Abstract— Tetrasulfophthalocyanine and a series of its metal chelates were tested for their ability to photoinactivate V‐79 Chinese hamster cells. Incubation of cells for 1 h with tetrasulfophthalocyanine at 5 μM effectively sensitized cells towards red light. At the 1% survival level, the dye was 4 x more efficient than hematoporphyrin, efficiency being defined in terms of drug concentration in the medium and incident light fluence rather than on the basis of quanta absorbed. Chelation of the dye with metal ions resulted in most cases in a greatly diminished photosensitizing effect, except for cerium. The cerium complex was about 5 x more effective for cell killing than the metal free tetrasulfophthalocyanine and 20 x more efficient as compared to hematoporphyrin. Hypoxic conditions resulted in total loss of photoactivity indicating the involvement of oxygen in the action mechanism. The inactivation by near‐UV light by these drugs was also investigated. The potential of sulfonated phthalocyanines as novel photosensitizers for photodynamic cancer therapy is discussed.