Helle Anholt

Fractionated treatment of CaD2 tumors in mice sensitized with aluminium phthlocyanine tetrasulfonate

CaD2 mammary carcinomas transplanted into the feet of mice were treated with tetrasulfonated phthalocyanine (AlPcS4) and laser light at 680 nm. A light dose of 135 J/cm2 was either given as continuous radiation (15 min) or fractionated with 15 s exposure, 15 s darkness, 15 s exposure and so on for 30 min. The CaD2 tumors were found to respond better to a fractionated exposure than to the same energy given in one exposure. The reason for this is assumed to be a relocalization of the dye upon illumination, seen as a rapid decrease in fluorescence. When the laser light was turned off, the fluorescence returned to almost the initial value.

Photobiological properties of hematoporphyrin diesters: Evaluation for possible application in photochemotherapy of cancer

The dimethyl, diethyl, dipropyl, dibutyl, diamyl, dihexyl and diheptyl esters of hematoporphyrin (Hp) were synthesized and shown to be more strongly retained on a reverse phase (C18) high performance liquid chromatography column than most components of Photofrin II (PII) - the sensitizer used for photochemical treatment of cancer in the clinic. The Hp diesters were found to be less efficient than PII in sensitizing cells to photoinactivation. This was partly due to de-esterification of the Hp diesters by esterase activity in the serum. The de-esterification of the Hp diesters was highly dependent on the ester group, with Hp dimethyl ester (t1/2 for conversion to Hp monomethyl ester was 6 min) being de-esterified with a rate 500 times faster than that for Hp diheptyl ester. Incubation of NHIK 3025 cells with these dyes showed that the Hp diesters were all partly located in extranuclear spots and partly diffusely distributed in the cytoplasm. The fluorescing spots may be due to lysosomally located Hp diesters, since the lysosomal marker enzyme beta-Nacetyl-D-glucosaminidase was partly inactivated by Hp diesters and light.

PHTHALOCYANINE FLUORESCENCE IN TUMORS DURING PDT

Abstract— Athymic nude mice with human tumors transplanted to one of the hind legs were given aluminium phthalocyanine disulfonate (AlPcS2) intraperitoneally. Twenty‐four hours after the injection the mice were placed with the tumor in the sample position in a fluorescence spectrometer with modulated excitation. Exposure of the tumors to laser light at a fluence rate of50–200 mW/cm2 led to a rapid transient reduction by up to 50% of the phthalocyanine fluorescence of the tumor. After the laser irradiation the fluence rate of the fluorescence increased almost up to the initial value within a few minutes. This finding should be taken into account when optimal fluence rates and dose fractionation schemes are sought for photodynamic therapy.

Inhibiting effects of antioxidants on drug-induced phototoxicity in cell cultures. Investigations with sulphonamide-derived oral antidiabetics and diuretics.

The sulphonamide-derived oral antidiabetics chlorpropamide, glibenclamide, glipizide, gliquidone, glymidine, tolazamide and tolbutamide, and the diuretics bemetizide, bendroflumethiazide, benzylhydrochlorothiazide, bumetanide, butizide, chlortalidone, furosemide, hydrochlorothiazide, hydroflumethiazide, indapamide, piretanide, polythiazide, trichlormethiazide and xipamide were investigated for phototoxicity in a cell culture model. Cell death dependent on ultraviolet A (UVA) radiation fluence and test substance concentration was observed in the presence of the oral antidiabetics glibenclamide and gliquidone, as well as the diuretics bemetizide, bendroflumethiazide, benzylhydrochlorothiazide, bumetanide, butizide, hydrochlorothiazide, hydroflumethiazide, piretanide, polythiazide and trichlormethiazide. Bendroflumethiazide was phototoxic at concentrations of 0.05 mM and above; bemetizide, benzylhydrochlorothiazide, bumetanide and hydroflumethiazide were phototoxic at concentrations of 0.25 mM or more; the oral antidiabetics glibenclamide and gliquidone, as well as the diuretics butizide, hydrochlorothiazide, piretanide, polythiazide and trichlormethiazide were phototoxic at concentrations of 0.5 mM. To evaluate the effects of antioxidants, ascorbic acid, alpha-tocopherol, beta-carotene or ubiquinone was added to the tissue culture flasks before irradiation. The phototoxic inhibition of the colony-forming ability was largely reduced by the addition of ascorbic acid and alpha-tocopherole, indicating the involvement of reactive oxygen species in the phototoxic process.

Phototoxicity due to sulphonamide derived oral antidiabetics and diuretics: investigations in a cell culture model

A number of sulphonamide‐derived oral antidiabetics (chlorpropamide, glibenclamide, glipizide, gliquidone, glymidine, tolazamide and tolbutamide) and diuretics (bemetizide, bendroflumethiazide, benzylhydrochlorothiazide, bumetanide, butizide, chlortalidone, furosemide, hydrochlorothiazide, hydroflumethiazide, indapamide, piretanide, polythiazide, trichlormethiazide and xipamide) were investigated for phototoxicity in a cell culture model. Cell death dependent on ultraviolet A fluence and test substance concentration was observed in the presence of the oral antidiabetics glibenclamide and gliquidone, as well as the diuretics bemetizide, bendroflumethiazide, benzyl‐hydrochlorothiazide, bumetanide, butizide, hydrochlorothiazide, hydroflumethiazide, piretanide, polythiazide and trichlormethiazide. Bendroflumethiazide was phototoxic at 5times10−5 M and higher concentrations, bemetizide, benzylhydrochlorothiazide, bumetanide and hydroflumethiazide were phototoxic at 2.5times10−4 M and higher concentrations, and the oral antidiabetics glibenclamide and gliquidone as well as the diuretics butizide, hydrochlorothiazide, piretanide, polythiazide and trichlormethiazide were phototoxic at 5−4 M and higher concentrations. Electron microscopic investigations showed swelling of mitochondria and endoplasmic reticulum as well as aggregation of euchromatin when the cells were irradiated in the presence of photosensitizers***

A transient reduction of the fluorescence of aluminium phthalocyanine tetrasulphonate in tumours during photodynamic therapy

Phthalocyanines are a group of sensitizers that possess promising properties for future use in photochemotherapy of cancer [ 1 7 1. Several of these compounds show some selectivity with respect to tumour uptake or retention. Furthermore, they have a strong absorption in the wavelength region above 600 nm and a significant photosensitizing efficiency, in vitro as well as in uiuo . In this paper, we report a surprising behaviour of the fluorescence of aluminium phthalocyanine tetrasulphonate (A1PcS4) in tumours during laser exposure under conditions relevant for photodynamic therapy (PDT). This behaviour should be taken into consideration when optimal exposure schemes are sought for PDT with this dye.

Uptake, localization and effect of laser treatment on CaD2 tumours sensitized with aluminium phthalocyanine tetrasulfonate (AIPcS4)

C3D2/F1 mice bearing a mammary carcinoma (CaD2) transplanted subcutaneously on the foot were sensitized with 10 mg kg−1 aluminium phthalocyanine tetrasulfonate (AlPcS4). The sensitizer was injected i.p. from 1 to 72 h before tissue samples from the tumour, liver and muscle were collected and their concentration of AlPcS4 were determined. The highest dye concentration in tumours and muscles was found at 1 h, which was the first point of measurement. The dye concentration reached a peak after 6 h in liver. At 2, 24 and 48 h after sensitization tumours were exposed to laser light (680 nm), 135 J cm−2. Growth curves showed the largest growth delay of tumours irradiated at 2h after sensitization compared with tumours irradiated at 24 or 48 h. Fluorescence micrographs of tumours taken at 2, 24 and 48 h after AlPcS4 injection were also obtained. They showed that AlPcS4 was localized in the vascular system in the tumour to a higher degree at 2 h than at 24–48 h after injection.

Pressure against the tumor can reduce the efficiency of photochemotherapy

C3D2/F1 mice with mammary carcinoma tumors growing subcutaneously on their right foot were given 10 mg/kg aluminium phthalocyanine tetrasulfonate (AlPcS4) by intraperitoneal injection. Twenty-four hours later these tumors were exposed to light at 680 nm. The size of the tumors was measured daily. An exposure of 135 J/cm2 (150 mW/cm2) reduced the tumor growth rate so that the time needed for the tumors to reach a volume five times larger than that at the time of exposure increased from 4 days for control tumors to 15 days. However, when a glass plate was gently pressed against the tumor surface during irradiation, the effect of an identical exposure was significantly smaller. In agreement with this, microscopic studies showed that tumors exposed to laser light without any pressure applied during irradiation were more damaged than tumors receiving a slight pressure. Thus, pressure against the tumor can obviously reduce the oxygen concentration in this tumor enough to reduce the efficiency of the treatment.

Phototoxicity to sulphonamide-derived oral antidiabetics and diuretics: comparative in-vitro and in-vivo investigations

Seven oral antidiabetics (chlorpropamide, glibenclamide, glipizide, gliquidone, glymidine, tolazamide, and tolbutamide), and 14 diuretics (bemetizide, bendroflumethiazide, benzylhydrochlorothiazide, bumetanide, butizide, chlortalidone, furosemide, hydrochlorothiazide, hydroflumethiazide, indapamide, piretanide, polythiazide, trichlormethiazide, and xipamide) were investigated for potential phototoxicity in vitro using a cell culture model and in vivo in hairless mice. After exposure to broad band UVA, the majority of the substances tested in vitro yielded phototoxic action leading to loss of culture forming ability. In vivo, all tested substances induced edema or ulceration, and lead to a significant increase in skin fold thickness of the mouse skin. In all a number of substances not described to induce clinical photosensitivity nor phototoxicity in vitro or in vivo were detected in our testing. In determining potential photosensitizers, it seems important to utilize different test methods, as not all substances will exhibit action in a given assay.