aluminum-phthalocyanine-disulfonate and Fibrosarcoma

Local recurrences after inadequate surgery are a major challenge in many cancers. Photochemical internalization (PCI) is a new technology to release endocytosed macromolecules into the cytosol by a photochemical rupture of the endocytic vesicles. A recent study on an invasive human fibrosarcoma xenograft HT1080 indicated low sensitivity of the tumour periphery to disulfonated aluminium phthalocyanine (AlPcS(2a))-based photochemical treatment. The main goal of the present study was to evaluate the sensitivity of the remaining tumour after marginal resection of the HT1080 tumour to the photochemical treatment alone and PCI of bleomycin. AlPcS(2a) and bleomycin was systemically administrated 48 h and 30 min, respectively, prior to surgery and immediately followed by intra-operative light exposure at 670 nm (15 J cm(-2)). PCI of bleomycin as an adjunct to surgery did significantly delay tumour growth in contrast to the photochemical treatment alone. The results indicate that PCI of bleomycin may be an efficient intra-operative technique to eradicate cancer cells in the wound bed after inadequate surgery.

Topics: Animals; Bleomycin; Cell Line, Tumor; Disease Models, Animal; Endocytosis; Endosomes; Female; Fibrosarcoma; Humans; Indoles; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Organometallic Compounds; Photochemical Processes; Photochemotherapy; Tumor Burden

In this study we compared the photosensitizer concentration in two experimental murine tumors using an in situ fluorescence detection instrument to examine temporal and spatial variations, after intravenous versus intratumor injection. Also, the variations in the estimate as detected by large area sampling and micro-region sampling are compared, in order to determine what the inter-tissue and inter-animal variations are, and how the method of sampling affects this estimate. The latter study was carried out ex vivo in the same tumors, which had been harvested and frozen after in vivo measurements were made. The photosensitizer, disulphonated aluminum phthalocyanine (AlPcS2) was injected either intravenously (IV) or directly into the tumor (ITu), using two murine models, MTG-B (mammary adenocarcinoma) and RIF-1 (radiation-induced fibrosarcoma) grown subcutaneously on the flank. An in situ microsampling fluorescence probe was used to assess photosensitizer concentration, through real-time measurement of the remitted intensity. The photosensitizer concentration was evaluated at 8 time endpoints between 15 min and 48 h post-injection. Inter-tumor and intra-tumor variations were assessed by repeated samples from the tumor tissues. The average photosensitizer level reaches a peak between 3 to 6 h in both tumor and normal tissues using IV administration, but peaks within 1 h following ITu administration. MTG-B tumors demonstrated a factor of 2 higher uptake than RIF-1 tumors. The pharmacokinetic uptake rates of the RIF-1 tumor were 3 times faster than for MTG-B, while there was no statistical difference in their clearance rates. Preferential uptake of AlPcS2 by both tumors compared to contra-lateral flank subcutaneous normal tissue was documented, with ITu injection exceeding IV injection by a factor of 10 in the tumor to normal tissue ratio. Inter-animal standard deviation in the mean fluorescence was near 76% for both routes of administration, but estimates of the variation within tumor were near 16% standard deviation when a large sampling volume was used. In contrast, microscopic intra-tumor standard deviation in the mean estimate was near 76%, with IV injection, indicating that high heterogeneity exists in the photosensitizer concentration on a smaller distance scale. The inter-tumor variation was reduced by ITu injection, but at the expense of increasing intra-tumor variation.

Topics: Adenocarcinoma; Animals; Disease Models, Animal; Female; Fibrosarcoma; Fluorescent Dyes; Indoles; Kinetics; Mammary Neoplasms, Experimental; Mice; Mice, Inbred C3H; Organometallic Compounds; Photosensitizing Agents; Spectrometry, Fluorescence

Time-resolved reflectance spectroscopy was performed on tumor-bearing mice, administered with disulphonated aluminum phthalocyanine (AlS(2)Pc, 5 mg/kg body weight), before, during and after photodynamic therapy. This allowed us to evaluate the absorption spectrum of AlS(2)Pc in vivo from 610 to 700 nm, and to investigate how the therapeutic irradiation affects it. Two tumor locations (intraderma on the back and intramuscular in the leg), and two uptake times (3 and 12 h) were considered. As already observed previously, the absorption spectrum of AlS(2)Pc in vivo is centered at 680-685 nm. The irradiation causes a blue-shift of the measured line shape, more or less marked depending on the experimental conditions. A reduction in absorption is also often observed upon illumination with therapeutic light doses.

Topics: Absorption; Animals; Cytosol; Disease Models, Animal; Fibrosarcoma; Indoles; Lysosomes; Mice; Organometallic Compounds; Photochemotherapy; Radiation-Sensitizing Agents; Spectrophotometry

The absorption spectrum of aluminum phthalocyanine with an average disulphonation of 2.1 (hereafter called disulphonated aluminum phthalocyanine, A1S2Pc) was measured in vivo in a murine tumour model by means of time-resolved reflectance. Mice bearing the L1210 leukaemia were administered 2.5 or 5 mg/kg body weight (b.w.) of A1S2Pc intraperitoneally. Reflectance measurements were performed in the 650-695 nm range before and 1, 4 and 7 h after the drug administration. Fitting of the data with the diffusion theory allowed us to assess the absorption coefficient in both conditions (i.e. before and after). As a difference between the latter and the former data, the in vivo absorption spectrum of A1S2Pc was evaluated. 1 h after the administration of 2.5 mg/kg b.w. A1S2Pc, the absorption peak was centred at 685 nm, red-shifted about 15 nm with respect to the spectrum in aqueous solution. For the lower dose, the absorption line shapes 4 and 7 h after the administration remained very similar. The red shift of the absorption spectrum is consistent with the therapeutic efficacy of the photodynamic therapy which was measured at 672, 685 and 695 nm, and proved to be maximum at 685 nm for both the L1210 leukaemia and the MS-2 fibrosarcoma. With the higher drug dose, the absorption spectra taken from different animals showed significant differences. In particular, in some mice the line shape was similar to that measured with 2.5 mg/kg b.w., while in other subjects it showed a broadening or a second peak at shorter wavelengths. Measurements on some animals were performed also 18 and 24 h after the injection of 5 mg/kg b.w., leading to no time evolution or to a progressive line shape narrowing.

Topics: Animals; Argon; Disease Models, Animal; Female; Fibrosarcoma; Indoles; Lasers; Leukemia L1210; Male; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Organometallic Compounds; Photochemotherapy; Radiation-Sensitizing Agents; Spectrophotometry

Photodynamic therapy (PDT) is systemic administration of tumor localizing photosensitizers and subsequent irradiation with light of the appropriate wavelength. The combination of drug uptake in malignant tissues and selective delivery of laser-generated light provides effective therapy, with efficient tumor cytotoxicity and minimal normal tissue damage. There have been few studies of the effects of photoactivated photosensitizers on the host immune response. Since immunity is important in the control of tumor growth and spread, we have examined, in our laboratory, the effects of photoactivated phthalocyanines on the antitumor immune response. Immunosuppressed and normal mice bearing the MS-2 fibrosarcoma treated with 5 mg/kg of aluminum disulfonated phthalocyanine (AIS2Pc) and then the tumor mass exposed to laser light (100 mW/cm2 x 10 min) or treated with surgical excision of the tumor survived indefinitely, with no difference between the different groups. The survivors, tumor-free 100 days after the treatment modalities described above, were rechallenged with the parental MS-2. Some groups of surviving animals were immunosuppressed with cyclophosphamide before the injection of the tumor. Resistance to rechallenge was evidenced only in normal surviving animals cured by PDT, while the immunodepressed surviving animals and animals cured by surgery died of tumor. Finally, mice, cured by PDT and tumor-free, rechallenged with L1210 and P388 murine leukemias did not survive. These results suggest that a potential and specific 'antitumor immunity' is induced by PDT with photoactivated AIS2Pc.

Topics: Animals; Cell Division; Fibrosarcoma; Indoles; Laser Therapy; Leukemia L1210; Leukemia P388; Male; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Neoplasm Transplantation; Neoplasms, Experimental; Organometallic Compounds; Photochemotherapy; Radiation-Sensitizing Agents

Levels of disulfonated and tetrasulfonated aluminum phthalocyanines (AlPcS2,4) were measured in cells derived from FsaR tumors (murine fibrosarcoma) using a fluorescence-activated cell sorter (FACS). The tumors were excised from animals injected with the sensitizer 24 h earlier and enzymatically dissociated. Before flow cytometry, the cells were stained with fluorescein isothiocyanate-conjugated anti-mouse monoclonal antibodies to specific immune cell membrane markers (Mac1, Fc receptor (FcR) or CD45). Staining to FcR and CD45 was combined with a DNA stain Hoechst 33342. This enabled concomitant discrimination to be made by the FACS between different populations of tumor-infiltrating host cells and malignant cells. The results showed on average 1.49 times higher AlPcS2 levels and 1.16 times higher AlPcS4 levels in Mac1-positive (Mac1+) compared with Mac1-negative (Mac1-) tumor cell populations. The same type of experiments performed with SCCVII tumor (squamous cell carcinoma) gave average Mac1+/Mac1- ratios of 1.75 and 1.45 for AlPcS2 and AlPcS4 respectively. The data using other antibodies and DNA staining are consistent with the conclusion that, based on average per cell content, elevated levels of AlPcS2, and to a lesser extent AlPcS4, are retained in tumor-associated macrophages (TAM). The levels of these photosensitizers in other leukocytes and in non-immune host cells were not substantially different from those in malignant tumor cells. It is also shown that elevated levels of AlPcS2 and AlPcS4 are not localized in all TAM, but rather in a fraction of this cell population characterized by extremely high photosensitizer content.

Topics: Animals; Benzimidazoles; Biomarkers; Female; Fibrosarcoma; Flow Cytometry; Fluorescein-5-isothiocyanate; Indoles; Macrophage-1 Antigen; Mice; Mice, Inbred C3H; Organometallic Compounds; Radiation-Sensitizing Agents; Sarcoma, Experimental; Staining and Labeling; Tissue Distribution

The action spectrum of disulfonated aluminum phthalocyanine (AIS2-Pc) as a photosensitizer for photodynamic therapy (PDT) was evaluated in vivo from 640 to 710 nm on two murine tumors with different biological properties. Mice bearing MS-2 fibrosarcoma and mice bearing the pigmented tumor B16 melanoma were injected with 5 mg/kg of AIS2Pc and irradiated with a light dose of 50 mW for 10 min for the MS-2 fibrosarcoma and of 100 mW for 10 min for the B16 melanoma. The action spectrum for both tumors presents a red shift with respect to the absorption spectrum of AIS2Pc in saline. The effectiveness is limited for short wavelengths (less than or equal to 655 nm for MS-2 and less than or equal to 660 for B16), whereas it increases at longer wavelengths and reaches its maximum at a peak (672 nm). For wavelengths beyond 672 nm the photodynamic activity remains up to 710 nm despite the significant decrease in absorption. The results obtained for both murine tumors seem therefore to indicate that an appreciable modification of the absorption spectrum takes place when AIS2Pc is incorporated into tissues following systematic administration.

Topics: Animals; Fibrosarcoma; Indoles; Male; Melanoma, Experimental; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Neoplasm Invasiveness; Organometallic Compounds; Photochemotherapy; Pigmentation; Radiation-Sensitizing Agents; Spectrophotometry