Sonodynamic therapy with photofrin II on AH130 solid tumor. Pharmacokinetics, tissue distribution and sonodynamic antitumoral efficacy of photofrin II.

BACKGROUND: The pharmacokinetics and tissue distribution of photofrin II (PF) and its efficacy in sonodynamic therapy were studied in rats bearing AH130 solid tumors. MATERIALS AND METHODS: In order to find the optimum timing of the ultrasound exposure after administration of PF, the PF concentrations in plasma, skin, muscle and tumor were measured and pharmacokinetically analyzed. Antitumor effects were estimated by measuring tumor size. RESULTS: Since the highest concentration of PF in tumors occurred 24 h after administration, ultrasound administration 24 h after the intravenous administration of PF was chosen. Ultrasound alone showed a slight antitumor effect, which became increasingly significant as the dose of PF was increased, while PF alone showed no significant effect. CONCLUSIONS: PF significantly sensitized solid tumors to the antitumor effect of ultrasound in a synergistic manner.

Pharmacokinetic study of a gallium-porphyrin photo- and sono-sensitizer, ATX-70, in tumor-bearing mice.

The tissue distribution of a gallium-porphyrin photo- and sono-sensitizer, 7,12-bis(1-decyloxyethyl)-Ga(III)-3,8,13,17-tetramethylporphyrin-2,18-dipropionyldiaspartic acid, ATX-70, was pharmacokinetically examined in tumor-bearing mice. The drug was administered intravenously to CDF(1) mice implanted with Colon 26 carcinoma. Blood and tissue samples were collected for up to 72 h after administration. The drug concentration was determined by high-performance liquid chromatography (HPLC) with fluorescence detection. ATX-70 was found to accumulate in tumors at a relatively high concentration that peaked between 2 h and 6 h after administration. However, modest concentrations of ATX-70 also remained in healthy tissues for up to 6 h. We examined the distribution of ATX-70 in the tumor in comparison with other tissues from the viewpoint of minimizing possible side effects of laser or ultrasound exposure while maintaining the treatment effect. About 24 h after administration, the tumor / plasma concentration ratio peaked, and relatively high tumor / skin and tumor / muscle concentration ratios were seen.

Sonodynamically induced antitumor effect of Photofrin II on colon 26 carcinoma.

The sonodynamically induced antitumor effect of Photofrin II (PF), was evaluated in mice bearing colon 26 carcinoma. In order to find the optimum timing for ultrasonic exposure after the administration of PF, the PF concentrations in the plasma, skin, muscle, and tumor were measured. The antitumor effect was estimated by measuring the tumor size. Since the highest concentration of PF in the tumor was observed 24 h after administration, an ultrasonic exposure timing of 24 h after the intravenous administration of PF was chosen. When used alone, ultrasound showed a slight antitumor effect, which became increasingly significant as the dose of PF was increased, while use of PF alone showed no significant effect. From these results, it is concluded that PF significantly sensitizes solid tumors to ultrasound, demonstrating a synergistic antitumor effect.

Ultrasonically induced cell damage enhanced by photofrin II: mechanism of sonodynamic activation.

UNLABELLED: Ultrasonically induced cell damage and active oxygen generation with photofrin II (PF) were compared in the same in vitro insonation setup. MATERIALS AND METHODS: Sarcoma 180 cells suspended in air-saturated PBS were exposed to ultrasound for up to 60 seconds in the presence and absence of PF. The viability was determined by the Trypan Blue exclusion test. Ultrasonically induced active oxygen generation in the presence and absence of PF in air-saturated aqueous solutions of 50 mM 2,2,6,6-tetramethyl-4-piperidone was detected by electron spin resonance (ESR) spectrum. RESULTS: Significant enhancement of the rates of both ultrasonically induced cell damage and nitroxide generation was demonstrated with 20-80 micrograms/ml PF. Both rates correlated very well. The enhancement of both rates with PF was suppressed by 10 mM histidine. CONCLUSION: These results may suggest that ultrasonically generated active oxygen plays a primary role in ultrasonically induced cell damage in the presence of PF.

Combination effect of photodynamic and sonodynamic therapy on experimental skin squamous cell carcinoma in C3H/HeN mice.

We studied a combination of photodynamic therapy (PDT) and sonodynamic therapy (SDT) for improving tumoricidal effects in a transplantable mouse squamous cell carcinoma (SCC) model. Two sensitizers were utilized: the pheophorbide-a derivative PH-1126, which is a newly developed photosensitizer, and the gallium porphyrin analogue ATX-70, a commonly used sonosensitizer. Mice were injected with either PH-1126 or ATX-70 i.p. at doses of 5 or 10 mg/kg.bw. At 24 (ATX-70) or 36 hr (PH-1126) (time of optimum drug concentration in the tumor) after injection, SCCs underwent laser light irradiation (88 J/cm2 of 575 nm for ATX-70; 44J/cm2 of 650 nm for PH-1126) (PDT), ultrasound irradiation (0.51 W/cm2 at 1.0 MHz for 10 minutes) (SDT), or a combination of the two treatments. The combination of PDT and SDT using either PH-1126 or ATX-70 as a sensitizer resulted in significantly improved inhibition of tumor growth (92-98%) (additive effect) as compared to either single treatment (27-77%). The combination using PH-1126 resulted in 25% of the treated mice being tumor free at 20 days after treatment. Moreover, the median survival period (from irradiation to death) of PDT + SDT-treated mice (> 120 days) was significantly greater than that in single treatment groups (77-95 days). Histological changes revealed that combination therapy could induce tumor necrosis 2-3 times as deep as in either of the single modalities. The combination of PDT and SDT could be very useful for treatment of non-superficial or nodular tumors.

Sonodynamically induced antitumor effect of 4-formyloximethylidene-3-hydroxy-2-vinyl-deuterio-porphynyl(IX)-6,7-dia spartic acid (ATX-S10).

The sonodynamically induced antitumor effect of 4-formyloximethylidene-3-hydroxy-2-vinyl-deuterio-porphyn yl(IX)-6,7-diaspartic acid (ATX-S10) was investigated. Both in vitro and in vivo antitumor effects were tested in combination with ultrasound at 2 MHz. The rate of ultrasonically induced damage to isolated sarcoma 180 cells in air-saturated suspension was enhanced two-fold with 80 microM ATX-S10. This enhancement was significantly inhibited by histidine, which may suggest that it was mediated by ultrasonically induced oxidation. The coadministraion of 25 mg/kg ATX-S10 followed by ultrasonic exposure at 2 MHz stopped the growth of implanted colon 26 tumors at an intensity at which ultrasound alone showed only a slight antitumor effect.

Sonodynamically induced cell damage with 4'-O-tetrahydropyranyladriamycin, THP.

BACKGROUND: 4'-O-tetrahydropyranyladriamycin (THP) is a novel anthracycline derivative with cardiotoxicity significantly lower than ADM. In this study, the ultrasonically induced in vitro cell damaging effect of THP was investigated. MATERIALS AND METHODS: Sarcoma 180 cells suspended in air-saturated PBS were exposed to ultrasound for up to 60 s in the presence and absence of THP. The viability of the isolated cells was determined by staining of the cells with Trypan Blue dye. RESULTS: The rate of inducing cell damage with ultrasound was doubled with 80 microM THP, while no cell damage was observed with THP alone. CONCLUSION: The enhancement of ultrasonically induced in vitro cell damage was demonstrated with THP. This enhancement was significantly inhibited by histidine, which may suggest a sonochemical mechanism.

Sonodynamically induced effect of rose bengal on isolated sarcoma 180 cells.

PURPOSE: The ultrasonically induced effect of rose bengal (RB) on isolated tumor cells was investigated. METHODS: Sarcoma 180 cells were suspended in air-saturated phosphate-buffered saline and exposed to ultrasound in standing wave mode for up to 60 s in the presence and absence of RB. Cell viability was determined by the ability to exclude trypan blue. RESULTS: The rate of inducing cell damage by ultrasound was enhanced two to three times with 160 microM RB. while no cell damage was observed with RB alone. This enhancement was significantly inhibited by histidine. CONCLUSIONS: Ultrasonically induced in vitro cell damage was significantly enhanced by RB. A sonochemical mechanism may be suggested since the enhancement was significantly inhibited by an active oxygen scavenger.

Antitumor effect sonodynamically induced by focused ultrasound in combination with Ga-porphyrin complex.

The antitumor effect of focused ultrasound in combination with Ga-porphyrin complex, 7,12-bis(1-decyloxyethyl)-Ga(III)-3,8,13,17 tetramethylporphyrin-2,18-dipropionyl diaspartic acid (ATX-70), on the growth of experimental murine tumors was examined. Walker 256 tumors implanted in rat kidneys were exposed in a progressive wave field for 5 min to focused ultrasound at 500 kHz with the second harmonic (at 1 MHz) superimposed after administration of ATX-70 to the rats. A significant antitumor effect was obtained at a focal-spot average acoustic intensity of 12 W/cm2 or higher with an ATX-70 dose of not less than 2.5 mg/kg. When the acoustic intensity was 8 W/cm2 or lower, no significant effect was observed even at an ATX-70 dose of 2.5 mg/kg. Also when the ATX-70 dose was 1.0 mg/kg or lower, no significant effect was observed even at a focal-spot average acoustic intensity of 40 W/cm2.

Sonodynamically-induced cell damage with fluorinated anthracycline derivative, FAD104.

The ultrasonically-induced in vitro cell damaging effect of fluorine-containing anthracycline derivative (FAD104) was investigated. Sarcoma 180 cells suspended in air-saturated PBS were exposed to ultrasound for up to 60 s in the presence and absence of FAD104. The rate of inducing cell damage with ultrasound was doubled with 80 microM FAD104, while no cell damage was observed with FAD104 alone. This enhancement was significantly inhibited by histidine, which may suggest a sonochemical mechanism.

Sonodynamically induced antitumor effect of gallium-porphyrin complex by focused ultrasound on experimental kidney tumor.

The antitumor effect of a gallium-porphyrin complex, ATX-70, induced by focused ultrasound, on colon 26 carcinoma implanted in a mouse kidney was investigated. Colon 26 tumors were exposed to focused ultrasound at 500 kHz and 1 MHz in a progressive wave mode. Both frequency components were superimposed onto each other in the focal zone to efficiently produce cavitation in the tumor. ATX-70 was administered intravenously at the dose of 2.5 mg/kg, 24 h before the ultrasonic exposure. Antitumor effects were evaluated by histological observation 7 days after the exposure. The destruction of tumor tissue was observed with the ultrasonic treatment in combination with ATX-70, while neither the treatment with ATX-70 alone nor that with ultrasound alone caused any necrosis. These results first demonstrated that antitumor effects of a porphyrin compound can be induced by focused ultrasound in a progressive wave mode.

Sonodynamically-induced in vitro cell damage enhanced by adriamycin.

Sonodynamically-induced cell damage and active oxygen generation enhanced by adriamycin (ADM) were compared in the same in vitro insonation set-up. Significant enhancement of the rates of both ultrasonically-induced cell damage and nitroxide generation was demonstrated with 40-160 microM ADM. Both rates correlated very well resulting in a correlation coefficient of more than 0.99. The enhancement of both rates was suppressed by 10 mM histidine. These results are consistent with the hypothesis that ultrasonically-generated active oxygen plays a major role in the sonodynamically-induced cell damage enhanced by ADM.

Sonodynamically induced antitumor effect of pheophorbide a.

The sonodynamically induced antitumor effect of pheophorbide a (Ph-a) was investigated. Both in vitro and in vivo effects on sarcoma 180 were tested in combination with ultrasound at 2 MHz. The rate of ultrasonically induced cell damage in air-saturated suspension was enhanced by twice with 80 microM Ph-a. This enhancement was significantly inhibited by histidine, which may suggest it was mediated by ultrasonically induced oxidation. For mice, 5 mg/kg Ph-a was administered before the insonation, and ultrasound stopped the tumor growth at an intensity with which ultrasound alone showed only a slight antitumor effect.

Sonodynamically induced antitumor effect of a gallium-porphyrin complex, ATX-70.

The sonodynamically induced antitumor effect of a gallium-porphyrin complex, ATX-70, was evaluated in mice bearing colon 26. In order to find the optimum timing for the ultrasonic exposure after the administration of ATX-70, the ATX-70 concentrations in the plasma, skin, and tumor were measured and analyzed. Antitumor effect was estimated by measuring the tumor size. When used alone, ultrasound showed a slight antitumor effect, which became increasingly significant as the dose of ATX-70 was increased, while use of ATX-70 alone had no significant effect. At an ATX-70 dose of 2.5 mg/kg or higher, the average tumor size decreased to smaller than a half by three days after the ultrasonic exposure. This was smaller than a third of the size of the untreated tumors on the same day. From these results, it is concluded that ATX-70 significantly sensitizes tumors to ultrasound, demonstrating a synergistic antitumor effect.

Membrane lipid peroxidation as a mechanism of sonodynamically induced erythrocyte lysis.

Sonodynamically induced lipid peroxidation with haematoporphyrin (Hp) was studied using rat erythrocytes. Both suspensions of erythrocyte ghosts and of intact erythrocytes were exposed to ultrasound in the same way in the presence and absence of Hp (80 microM). The lipid peroxidation in erythrocyte ghost membranes was estimated by measuring the amount of reactive substance produced from thiobarbituric acid added immediately after the exposure. Haematoporphyrin multiplied the ultrasonically induced lipid peroxidation by three to five times, while Hp alone showed no peroxidation. A 24-h interval between the exposure and the preparation for measurement did not increase the measured amount of peroxide. In the presence of Hp the estimated peroxidation rate and the rate of erythrocyte lysis correlated quite well for each acoustic condition and for each chemical condition such as the presence or absence of active oxygen scavengers in the suspension. The sonodynamically induced lipid peroxidation with Hp was doubled by deuterium oxide substitution for suspension medium and was significantly reduced by histidine, by sodium azide, and also by nitrogen substitution for saturation gas, whereas superoxide dismutase and mannitol showed no significant inhibitory effect. These results are consistent with a hypothesis that lipid peroxidation in membranes by singlet oxygen is the primary mechanism of sonodynamically induced erythrocyte lysis with Hp.

Sonochemical activation of hematoporphyrin: an ESR study.

The production of 2,2,6,6-tetramethyl-4-piperidone-N-oxyl by reaction of 2,2,6,6-tetramethyl-4-piperidone (TMPone) with ultrasonically generated active species in oxygenated solutions of hematoporphyrin (Hp) was studied by electron spin resonance spectroscopy. The nitroxide production rate in air-saturated TMPone solutions in phosphate-buffered saline of pH 9.0 was significantly higher in the presence of Hp than in its absence. The enhancement of nitroxide production by Hp was significantly inhibited in the presence of sodium azide or histidine in the solution. The production rate with Hp was doubled by substitution of deuterium oxide, while the rate without Hp increased only modestly. These results suggest that a substantial amount of active oxygen can be generated by ultrasound in aqueous solutions of Hp. Since the production rate was not reduced by mannitol and no nitroxide was produced in nitrogen-saturated solutions, it appears that hydroxyl radicals do not account for a major portion of the active oxygen species which reacted with TMPone to yield a nitroxide.

Enhancement of ultrasonically induced cell damage by a gallium-porphyrin complex, ATX-70.

Enhancement of ultrasonically induced cell damage by a gallium-porphyrin complex [ATX-70, 2,4-bis(1-decyloxyethyl)-Ga(III)-1,3,5,8- tetramethylporphryin-6,7-dipropionyl diaspartic acid] was investigated. The rate of damage to isolated sarcoma 180 cells in air-saturated suspension induced by 2 MHz ultrasound irradiation was enhanced more than four times by 80 microM ATX-70 in contrast to only twice by the same concentration of hematoporphyrin (Hp). The enhancement was almost completely inhibited in the presence of 10 mM histidine in the suspension, but not at all by 100 mM mannitol, which suggests that the enhanced cell damage was mostly mediated by singlet oxygen. Ultrasonically induced active oxygen generation in an air-saturated aqueous solution of ATX-70 was studied by detecting the electron spin resonance signals of 2,2,6,6,-tetramethyl-4-piperidone-N-oxyl produced by the reaction of 2,2,6,6-tetramethyl-4-piperidone with the generated active oxygen species. The rate of ultrasonically induced nitroxide generation was enhanced five times by 80 microM ATX-70 in contrast to only twice by Hp. The enhancement was inhibited significantly in the presence of 10 mM histidine in the suspension, but not at all by 100 mM mannitol. The singlet oxygen generation in air-saturated aqueous solution was further confirmed by the bleaching of N,N-dimethyl-4-nitrosoaniline in the presence of imidazole. The ultrasonically induced bleaching rate was enhanced six times by ATX-70, in contrast to only twice by Hp.

Mechanism of cell damage by ultrasound in combination with hematoporphyrin.

The mechanism of cell damage by ultrasound in combination with hematoporphyrin was studied. Mouse sarcoma 180 cell suspensions were exposed to ultrasound for up to 60 s in the presence and absence of hematoporphyrin, with and without active oxygen scavengers. The cell damage enhancement by hematoporphyrin was suppressed by adding histidine but not by mannitol. The enhancement was doubled in rate by substitution of deuterium oxide medium for normal water. Sonoluminescence was produced in a saline solution under similar acoustic conditions and observed to have spectral components that can excite hematoporphyrin molecules. These results suggest that cell damage enhancement is probably mediated via singlet oxygen generated by ultrasonically activated hematoporphyrin.

Synergistic effect of ultrasound and hematoporphyrin on sarcoma 180.

The antitumor effects of combined use of ultrasound (US) and a photosensitizer, hematoporphyrin (Hp), were determined in mice bearing sarcoma 180. In order to find the optimum timing of the US irradiation after the administration of Hp, the Hp concentrations in the tumor and in the plasma were determined and were analyzed pharmacokinetically. Antitumor effects were evaluated by measuring the tumor size and the tumor weight. Hp alone showed no antitumor effect but US alone showed a slight antitumor effect. The combined treatment with US and Hp showed marked synergistic effects on sarcoma 180 (inhibition ratio was 74% of the control). From these results, the enhancement of antitumor effect is thought to be caused by the sensitization of tumor cells to US mediated by Hp.

Hematoporphyrin as a sensitizer of cell-damaging effect of ultrasound.

Mouse sarcoma 180 or rat ascites hepatoma (AH) 130 cells were exposed to ultrasound (US; 1.27, 2.21 and 3.18 W/cm2; 1.92 MHz) for up to 60 s in vitro in the presence or absence of hematoporphyrin (Hp; 10, 25 and 50 micrograms/ml). The cell-damaging effects of treatments were determined by means of the Trypan Blue dye exclusion test. Hp alone did not show any cell-damaging effect, whereas US alone damaged 30 and 50% of sarcoma and AH 130 cells, respectively, at the maximum intensity for 60 s. In the presence of 50 micrograms/ml Hp, US damaged 99 and 95% of the above tumor cells, respectively. These results show that Hp increased the sensitivity of tumor cells to US.