- Email: [email protected]
Photocytotoxicity in vivo of haematoporphyrin derivative components
111
Cancer Letters, 28 (1985) 111-118 Elsevier Scientific Publishers Ireland Ltd.
PHOTOCYTOTOXICITY IN VIVO OF HAEMATOPORPHYRIN DERIVATIVE COMPONENTS
P.A. COWLED and I.J. FORBES* Department (Australia)
of Medicine,
University
of Adelaide,
Queen Elizabeth
Hospital,
Adelaide
(Received 10 June 1985) (Accepted 13 June 1985)
SUMMARY
Phototoxicity of haematoporphyrin derivative (HPD) and some of its component porphyrins was assayed using Lewis Lung Carcinoma transplanted in C57BL mice. Skin photosensitivity was assessed by measuring percentage increase in footpad thickness after exposure to light. HPD and its aggregate fraction photosensitised the skin, was active in tumour destruction and caused fluorescence in the tumour while the nonaggregate fraction and pure haematoporphyrin were inactive, both in treatment of tumours and in sensitising the skin. The commercial product, Photofrin II was also photoactive and protoporphyrin was slightly active. It is concluded that the therapeutic activity of HPD is associated entirely with the aggregate fraction and compounds effective in tumour phototherapy also sensitise the skin.
INTRODUCTION
Photochemotherapy with HPD has been used in the treatment of malignant tumours in man [ 1,2,3,7,8]. Intravenously injected HPD is retained selectively in tumours and is activated by red light (approx. 630 nm). Tumour destruction has been attributed to release of singlet oxygen [4,5,6]. The only major side effect so far reported is skin photosensitivity. HPD is a complex mixture of porphyrins which can be separated into several fractions by polyacrylamide gel chromatography. The activity as measured in vitro [ 91 and in vivo [lo] is concentrated in a fast-running fraction. The aggregated porphyrins show greatest phototoxicity in a tissue culture assay and porphyrins with vinyl side chains also show greater phototoxicity [ 111. We now report studies of the phototoxicity and skin sensitizing capac*To whom correspondence
should be addressed.
0304-3835/85/$03.30 0 1985 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
112
ity of porphyrins in a transplantable mouse tumour model, and skin photosensitivity using a foot pad assay. MATERIALS
Mouse
AND METHODS
tumour assay
Lewis lung carcinoma cells (Dr. L. Dent, Flinders Medical Centre), were transplanted into the back of C57BL mice by subcutaneous injection of approximately lo6 cells per mouse. After 7-10 days, when the tumours were 5-7 mm in diameter, mice in groups of 10 were given porphyrins (2560 mg/kg, i.p. or i.v.). Twenty four hours later mice were anaesthetised with Sagatal (May and Baker Aust. Pty. Ltd.), the fur over the tumour shaved and a l-cm diameter area over the tumour was irradiated with red light as specified. Mice were palpated daily for recurrence of tumour. The end point was the number of days for 5 out of 10 mice to regrow palpable tumour. The assay was modified from Dougherty [lo]. F’reparation Of pOFphyFin fFUCtiOns
HPD [ 21, haematoporphyrin, protoporphyrin, HPD aggregate and HPD non-aggregate were prepared and characterised as previously described [ 111 and administered in isotonic saline. Photofrin II was supplied by Dr. T.J. Dougherty, Roswell Park Memorial Institute, The concentration was determined by measuring 0Ds9, of a solution of porphyrin diluted in 50% ethanol/ 0.1 N NaOH. Skin photosensitivity
Mice in groups of 10 were given porphyrins i.p. and the left footpad was irradiated 24 h later with a light dose of 112 J/cm’. Twenty four hours later the thickness of both feet was measured with a micrometer and percentage increase in footpad thickness was calculated using the right foot as an untreated control. Light sources
A gold metal vapour laser [12] (Q uentron Optics Pty, Ltd) with wavelength 627.8 nm was used to generate average light intensities of 400 mW, coupled to a 400 pm quartz fibre and positioned to create a l-cm diameter spot. An incandescent filament lamp [ 131 fitted with a perspex lens 10 mm in diameter and 50 mm length delivered 2.5 W uniformly over 1 cm at 620720 nm wavelength. The effective light flux at 630 nm was 890 mW, determined from the relative light flux absorbed by HPD at 630 nm (Wilksch, 1982, unpublished data). The skin of the mouse was sprayed with water and irradiated in 50-s exposures with 10-s pauses to prevent thermal effects. Fluorescence
of tumours
Tumours were removed 24 h after injection of porphyrin and snap frozen.
113
1 01 40
25
50
60
HPD Dose (mg/kg)
Fig. 1. Relationship between ‘PC,, and HPD dose. Mice were given 25-60 mg/kg HPD either i.v. (0) or i.p. (e). Light dose was 225 J/cm’. Each point represents the mean of 2 experiments.
Sections (6 pm) were cut at -14°C on an IEC microtome and examined under a Zeiss fluorescence microscope with excitation wavelength 420490 nm.
1
Light
.
.
Dose
(Joules/sq.cm.)
Fig. 2. Relatronshrp between ‘PCS0and light dose. Mice were given 50 mg/kg HPD i.p. and tumours irradiated with increasing light doses from either the gold laser (e) or the incandescent filament lamp ( n ). Each point represents the mean of 2 experiments.
114 RESULTS
Relationship between TC,, and HPD dose (Fig. 1). There was a linear relationship between TCs, and HPD dose. The TC5,, for HPD given i.v. was 1 day greater than when given i.p. for all doses tested. Since the difference was not large, i.p. injections were used routinely. Relationship between TC5,, and light dose (Fig. 2) There was a linear relationship between TCSo and light dose, with either lamp or gold laser. The gold laser is slightly more efficient than the lamp. Histology Tumours 24 h after treatment with HPD and light showed extensive necrosis (Fig. 3a) and isolated viable cells. In contrast, tumours treated with HPD or light alone or untreated showed viable cells with small areas of necrosis (Fig. 3b). Fluorescence studies Twenty four hours after i.p. injection of porphyrin, frozen sections of tumours from animals given HPD, HPD-aggregate and Photofrin II showed extensive red fluorescence. Tumours from animals receiving HPD non-aggregate fluoresced weakly (Table 1). Protoporphyrin and haematoporphyrin did not result in any fluorescence in the tumours. Relative efficiencies of components of HPD in vivo (Table 1) Haematoporphyrin and HPD non-aggregate caused no tumour phototoxicity under the conditions tested. Increasing the light dose to 300 J/cm2 also gave no response. HPD aggregate and Photofrin II caused the same phototoxicity as HPD. Protoporphyrin i.p. and i.v. showed very little photoactivity. The photoactivity corresponded with the fluorescence of porphyrin within the tumours. Skin photosensitivity (Table 1) HPD, HPD aggregate and Photofrin II all induced approximately the same photosensitivity while HPD non-aggregate, haematoporphyrin and protoporphyrin did not photosensitise the skin. DISCUSSION
This paper describes the measurement of phototoxicity of HPD and some of its component porphyrins in vivo, using Lewis lung carcinoma transplanted subcutaneously into C57BL mice, measuring the time to 50% recurrence of tumours after treatment (TC,,). The TCSo was proportional to both the light dose and the porphyrin dose. Studies with SMT-F mammary tumour in BALB/c mice have shown that
115
Fig. 3. (a) Hiitological section of Lewis Lung carcinoma before treatment with HPD and light (400x magnification). (b) Histological section of tumour 24 h after treatment with 50 mg/kg HPD and 200 s irradiation with the filament lamp (400x magnification).
116 TABLE 1
Porphyrin
T&l (days)
% increase in footpad thickness
Tumour fluorescence
HPD HPD Aggregate HPD Non-aggregate Haematoporphyrin Protoporphyrin Photofrin II
5 5 0 0 0 6
79_+21 69 + 9.9 0.8 f. 6.7 0 22.0 -0.2 + 3.0 55.4 _+11.5
Positive Positive Weak positive Negative Negative Positive
All mice were injected with 50 mg/kg porphyrin i.p. and tumours were irradiated with 225 J/cm’ light from the filament lamp, 24 h post injection. For skin photosensitivity, the left footpad was irradiated with 110 J/cm’. Results are mean values of at least 2 experiments.
doses of 3.5-10.0 mg/kg HPD were effective [lo]. We found 50 mg/kg was necessary to obtain a satisfactory tumour response, in agreement with Ellingsen [14] who successfully treated Lewis lung carcinoma in B6Dz mice, using a regime of 50 mg/kg HPD and 160-360 J light at 630 nm. The aggregated fraction of HPD contained the photocytotoxic activity in vivo. Non-aggregated HPD and haematoporphyrin had no activity. This confirms reports by other authors [ 10,151. The activity correlated with the uptake of porphyrin as demonstrated by fluorescence of frozen sections. Similarly, skin photosensitivity was only demonstrated with porphyrins that also caused tumour fluorescence and destruction. This suggests that the critical requirement for in vivo phototoxicity is uptake into the tumours, and HPD non-aggregate, haematoporphyrin and protoporphyrin cannot be taken up into the tumours. Protoporphyrin is photoactive in vitro, [ 11,161 implying it can be taken up into cells in tissue culture but the mechanism for the selective uptake and retention in the tumour must be lacking in mice. Photofrin II has been postulated to be the active component of HPD [lo]. It exists in aqueous solution in a very highly aggregated form, demonstrated by gel chromatography [lo] and spectroscopic data [ 171. Dougherty [lo] reported a greater tumour phototoxicity using Photofrin II than HPD. We found Photofrin II had similar phototoxic activity to both HPD and HPD aggregate prepared by ultrafiltration. This discrepancy may reflect the different tumour systems used. HPD aggregate and Photofrin II also caused similar skin photosensitivity to HPD. Gomer and Razum [18] also reported that HPD and Photofrin II caused comparable acute skin damage. Carpenter [ 191 reported stronger fluorescence and greater tumour selectivity when HPD was injected i.v. compared to i.p. in mice with transplanted mammary adenocarcinoma. Tomio [ 201 reported equivalent tumour responses in Yoshida ascites hepatomas in rats with intravenous or intraperitoneal
117
injection of commercial haematoporphyrin. In our studies intravenous HPD had a slightly greater photocytotoxic effect than intraperitoneal HPD, although no marked difference was detected in the fluorescence in the tumours. This probably reflects insensitivity of visual detection of fluorescence. ACKNOWLEDGEMENTS
We wish to thank Dr. A.G. Swincer for the preparation of haematoporphyrin and protoporphyrin. REFERENCES 1 Dougherty, T.J., Kaufman, J.E., Goldfarb,
2
3 4
8 9 10
11
12
13 14
15
A., Weishaupt, K.R., Boyle, D. and Mittelman, A. (1978) Photoradiation therapy for the treatment of malignant tumours. Cancer Res., 38, 2628-2635. Forbes, I.J., Cowled, P.A., Leong, A.&Y., Ward, A.D., Black, R.B., Blake, A.J. and Jacka, F.J. (1980) Phototherapy of human tumours using haematoporphyrin derivative. Med. J. Aust., 2, 489-493. Dahlman, A., Wile, A.G., Burns, R.G., Mason, G.R., Johnson, F.M. and Berns, M.W. (1983) Laser photoradiation therapy of cancer. Cancer Res., 43,430-434. Weishaupt, K.R., Gomer, C.J. and Dougherty, T.J. (1976) Identification of singlet oxygen as the cytotoxic agent in photoinactivation of a murine tumor. Cancer Res., 36,2326-2329. Moan, J. and Sommer, S. (1981) Fluorescence and absorbtion properties of the components of hematoporphyrin derivative. Photobiochem. Photobiophys., 3, 93-103. Lee See, K., Forbes, I.J. and Betts, W.H. (1984) Oxygen dependency of photocytotoxicity with haematoporphyrin derivative. Photochem. Photobiol., 39, 631-634. Hayata, Y., Kato, H., Konaka, C., Ono, J. and Takizawa, N. (1982) Hematoporphyrin derivative and laser photoradiation in the treatment of lung cancer. Chest, 81, 269277. Cortese, D.A. and Kinsey, J.H. (1982) Endoscopic management of lung cancer with hematoporphyrin derivative phototherapy. Mayo Clin. Proc., 57, 543-547. Moan, J. and Sommer, S. (1983) Uptake of the components of hematoporphyrin derivative by cells and tumours. Cancer Letters, 21, 167-174. Dougherty, T.J., Boyle, D.G., Weishaupt, K.R., Henderson, B.A., Potter, W.R., Bellnier, D.A. and Wityk, K.E. (1983) Photoradiation therapy: clinical and drug advances. Adv. Exp. Med. Biol., 160, 3-13. Cowled, P.A., Forbes, I.J., Swincer, A.G., Trenerry, V.C. and Ward, A.D. (1985) Separation and phototoxicity in vitro of some of the components of Haematoporphyrin derivative. Photochem. Photobiol., in press. Cowled, P.A., Grace, J.R. and Forbes, I.J. (1984) Comparison of the efficacy of pulsed and continuous wave red laser light in induction of photocytotoxicity by haematoporphyrin derivative. Photochem. Photobiol., 39, 115-117. Jacka, F. and Blake, A.J. (1983) A lamp for cancer phototherapy. Aust. J. Phys., 36, 221-226. Ellingsen, R., Svaasand, L.O. and Odegard, A. (1984) Photoradiation therapy (PRT) of Lewis Lung carcinoma in B,D, mice, dosimetry considerations. In: Porphyrins in Tumour Phototherapy, pp. 213-220. Editors: A. Andreoni and R. Cubeddu. Plenum Press, New York. Moan, J., Christensen, T. and Sommer, S. (1982) The main photosensitising components of hematoporphyrin derivative. Cancer Letters, 15.161-166.
118 16 Kessel, D., (1982) Components of hematoporphyrin derivative and their tumour localising capacity. Cancer Res., 42,1703-1706. 17 Poletti, A., Murgia, S.M., Pasqua, A., Reddi, E. and Jori, G. (1984) Photophysical and photosensitising properties of Photofrin II. In: Porphyrins in Tumour Phototherapy, pp. 37-43. Editors: A. Andreoni and R. Cuheddu. Plenum Press, New York. 18 Gomer, C.J. and Razum, N.J. (1984) Acute skin responses in albino mice following porphyrin photosensitisation under oxic and anoxic conditions. Photochem. Photobiol., 40,435-440. 19 Carpenter, R.J., Ryan, R.J., Neel, H.B. and Sanderson, D.R. (1977) Tumor fluorescence with hematoporphyrin derivative. Ann. Otol., 86, 661-666. 20 Tomio, L., Reddi, E., Jori, G., Zorat, P.L., Pizzi, G.B. and Calzavara, F. (1980) Hematoporphyrin as a sensitizer in tumor phototherapy: effect of medium polarity on the photosensitizing efficiency and role of administration pathway on the distribution in normal and tumor bearing rats. In: Lasers in Photomedicine and Photobiology: Springer Series in Optical Sciences, Vol. 22, pp. 7682. Editors: R. Pratesi and C.A. Sacchi. Springer-Verlag, Berlin, Heidelberg, New York.
Cancer Letters, 28 (1985) 111-118 Elsevier Scientific Publishers Ireland Ltd.
PHOTOCYTOTOXICITY IN VIVO OF HAEMATOPORPHYRIN DERIVATIVE COMPONENTS
P.A. COWLED and I.J. FORBES* Department (Australia)
of Medicine,
University
of Adelaide,
Queen Elizabeth
Hospital,
Adelaide
(Received 10 June 1985) (Accepted 13 June 1985)
SUMMARY
Phototoxicity of haematoporphyrin derivative (HPD) and some of its component porphyrins was assayed using Lewis Lung Carcinoma transplanted in C57BL mice. Skin photosensitivity was assessed by measuring percentage increase in footpad thickness after exposure to light. HPD and its aggregate fraction photosensitised the skin, was active in tumour destruction and caused fluorescence in the tumour while the nonaggregate fraction and pure haematoporphyrin were inactive, both in treatment of tumours and in sensitising the skin. The commercial product, Photofrin II was also photoactive and protoporphyrin was slightly active. It is concluded that the therapeutic activity of HPD is associated entirely with the aggregate fraction and compounds effective in tumour phototherapy also sensitise the skin.
INTRODUCTION
Photochemotherapy with HPD has been used in the treatment of malignant tumours in man [ 1,2,3,7,8]. Intravenously injected HPD is retained selectively in tumours and is activated by red light (approx. 630 nm). Tumour destruction has been attributed to release of singlet oxygen [4,5,6]. The only major side effect so far reported is skin photosensitivity. HPD is a complex mixture of porphyrins which can be separated into several fractions by polyacrylamide gel chromatography. The activity as measured in vitro [ 91 and in vivo [lo] is concentrated in a fast-running fraction. The aggregated porphyrins show greatest phototoxicity in a tissue culture assay and porphyrins with vinyl side chains also show greater phototoxicity [ 111. We now report studies of the phototoxicity and skin sensitizing capac*To whom correspondence
should be addressed.
0304-3835/85/$03.30 0 1985 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
112
ity of porphyrins in a transplantable mouse tumour model, and skin photosensitivity using a foot pad assay. MATERIALS
Mouse
AND METHODS
tumour assay
Lewis lung carcinoma cells (Dr. L. Dent, Flinders Medical Centre), were transplanted into the back of C57BL mice by subcutaneous injection of approximately lo6 cells per mouse. After 7-10 days, when the tumours were 5-7 mm in diameter, mice in groups of 10 were given porphyrins (2560 mg/kg, i.p. or i.v.). Twenty four hours later mice were anaesthetised with Sagatal (May and Baker Aust. Pty. Ltd.), the fur over the tumour shaved and a l-cm diameter area over the tumour was irradiated with red light as specified. Mice were palpated daily for recurrence of tumour. The end point was the number of days for 5 out of 10 mice to regrow palpable tumour. The assay was modified from Dougherty [lo]. F’reparation Of pOFphyFin fFUCtiOns
HPD [ 21, haematoporphyrin, protoporphyrin, HPD aggregate and HPD non-aggregate were prepared and characterised as previously described [ 111 and administered in isotonic saline. Photofrin II was supplied by Dr. T.J. Dougherty, Roswell Park Memorial Institute, The concentration was determined by measuring 0Ds9, of a solution of porphyrin diluted in 50% ethanol/ 0.1 N NaOH. Skin photosensitivity
Mice in groups of 10 were given porphyrins i.p. and the left footpad was irradiated 24 h later with a light dose of 112 J/cm’. Twenty four hours later the thickness of both feet was measured with a micrometer and percentage increase in footpad thickness was calculated using the right foot as an untreated control. Light sources
A gold metal vapour laser [12] (Q uentron Optics Pty, Ltd) with wavelength 627.8 nm was used to generate average light intensities of 400 mW, coupled to a 400 pm quartz fibre and positioned to create a l-cm diameter spot. An incandescent filament lamp [ 131 fitted with a perspex lens 10 mm in diameter and 50 mm length delivered 2.5 W uniformly over 1 cm at 620720 nm wavelength. The effective light flux at 630 nm was 890 mW, determined from the relative light flux absorbed by HPD at 630 nm (Wilksch, 1982, unpublished data). The skin of the mouse was sprayed with water and irradiated in 50-s exposures with 10-s pauses to prevent thermal effects. Fluorescence
of tumours
Tumours were removed 24 h after injection of porphyrin and snap frozen.
113
1 01 40
25
50
60
HPD Dose (mg/kg)
Fig. 1. Relationship between ‘PC,, and HPD dose. Mice were given 25-60 mg/kg HPD either i.v. (0) or i.p. (e). Light dose was 225 J/cm’. Each point represents the mean of 2 experiments.
Sections (6 pm) were cut at -14°C on an IEC microtome and examined under a Zeiss fluorescence microscope with excitation wavelength 420490 nm.
1
Light
.
.
Dose
(Joules/sq.cm.)
Fig. 2. Relatronshrp between ‘PCS0and light dose. Mice were given 50 mg/kg HPD i.p. and tumours irradiated with increasing light doses from either the gold laser (e) or the incandescent filament lamp ( n ). Each point represents the mean of 2 experiments.
114 RESULTS
Relationship between TC,, and HPD dose (Fig. 1). There was a linear relationship between TCs, and HPD dose. The TC5,, for HPD given i.v. was 1 day greater than when given i.p. for all doses tested. Since the difference was not large, i.p. injections were used routinely. Relationship between TC5,, and light dose (Fig. 2) There was a linear relationship between TCSo and light dose, with either lamp or gold laser. The gold laser is slightly more efficient than the lamp. Histology Tumours 24 h after treatment with HPD and light showed extensive necrosis (Fig. 3a) and isolated viable cells. In contrast, tumours treated with HPD or light alone or untreated showed viable cells with small areas of necrosis (Fig. 3b). Fluorescence studies Twenty four hours after i.p. injection of porphyrin, frozen sections of tumours from animals given HPD, HPD-aggregate and Photofrin II showed extensive red fluorescence. Tumours from animals receiving HPD non-aggregate fluoresced weakly (Table 1). Protoporphyrin and haematoporphyrin did not result in any fluorescence in the tumours. Relative efficiencies of components of HPD in vivo (Table 1) Haematoporphyrin and HPD non-aggregate caused no tumour phototoxicity under the conditions tested. Increasing the light dose to 300 J/cm2 also gave no response. HPD aggregate and Photofrin II caused the same phototoxicity as HPD. Protoporphyrin i.p. and i.v. showed very little photoactivity. The photoactivity corresponded with the fluorescence of porphyrin within the tumours. Skin photosensitivity (Table 1) HPD, HPD aggregate and Photofrin II all induced approximately the same photosensitivity while HPD non-aggregate, haematoporphyrin and protoporphyrin did not photosensitise the skin. DISCUSSION
This paper describes the measurement of phototoxicity of HPD and some of its component porphyrins in vivo, using Lewis lung carcinoma transplanted subcutaneously into C57BL mice, measuring the time to 50% recurrence of tumours after treatment (TC,,). The TCSo was proportional to both the light dose and the porphyrin dose. Studies with SMT-F mammary tumour in BALB/c mice have shown that
115
Fig. 3. (a) Hiitological section of Lewis Lung carcinoma before treatment with HPD and light (400x magnification). (b) Histological section of tumour 24 h after treatment with 50 mg/kg HPD and 200 s irradiation with the filament lamp (400x magnification).
116 TABLE 1
Porphyrin
T&l (days)
% increase in footpad thickness
Tumour fluorescence
HPD HPD Aggregate HPD Non-aggregate Haematoporphyrin Protoporphyrin Photofrin II
5 5 0 0 0 6
79_+21 69 + 9.9 0.8 f. 6.7 0 22.0 -0.2 + 3.0 55.4 _+11.5
Positive Positive Weak positive Negative Negative Positive
All mice were injected with 50 mg/kg porphyrin i.p. and tumours were irradiated with 225 J/cm’ light from the filament lamp, 24 h post injection. For skin photosensitivity, the left footpad was irradiated with 110 J/cm’. Results are mean values of at least 2 experiments.
doses of 3.5-10.0 mg/kg HPD were effective [lo]. We found 50 mg/kg was necessary to obtain a satisfactory tumour response, in agreement with Ellingsen [14] who successfully treated Lewis lung carcinoma in B6Dz mice, using a regime of 50 mg/kg HPD and 160-360 J light at 630 nm. The aggregated fraction of HPD contained the photocytotoxic activity in vivo. Non-aggregated HPD and haematoporphyrin had no activity. This confirms reports by other authors [ 10,151. The activity correlated with the uptake of porphyrin as demonstrated by fluorescence of frozen sections. Similarly, skin photosensitivity was only demonstrated with porphyrins that also caused tumour fluorescence and destruction. This suggests that the critical requirement for in vivo phototoxicity is uptake into the tumours, and HPD non-aggregate, haematoporphyrin and protoporphyrin cannot be taken up into the tumours. Protoporphyrin is photoactive in vitro, [ 11,161 implying it can be taken up into cells in tissue culture but the mechanism for the selective uptake and retention in the tumour must be lacking in mice. Photofrin II has been postulated to be the active component of HPD [lo]. It exists in aqueous solution in a very highly aggregated form, demonstrated by gel chromatography [lo] and spectroscopic data [ 171. Dougherty [lo] reported a greater tumour phototoxicity using Photofrin II than HPD. We found Photofrin II had similar phototoxic activity to both HPD and HPD aggregate prepared by ultrafiltration. This discrepancy may reflect the different tumour systems used. HPD aggregate and Photofrin II also caused similar skin photosensitivity to HPD. Gomer and Razum [18] also reported that HPD and Photofrin II caused comparable acute skin damage. Carpenter [ 191 reported stronger fluorescence and greater tumour selectivity when HPD was injected i.v. compared to i.p. in mice with transplanted mammary adenocarcinoma. Tomio [ 201 reported equivalent tumour responses in Yoshida ascites hepatomas in rats with intravenous or intraperitoneal
117
injection of commercial haematoporphyrin. In our studies intravenous HPD had a slightly greater photocytotoxic effect than intraperitoneal HPD, although no marked difference was detected in the fluorescence in the tumours. This probably reflects insensitivity of visual detection of fluorescence. ACKNOWLEDGEMENTS
We wish to thank Dr. A.G. Swincer for the preparation of haematoporphyrin and protoporphyrin. REFERENCES 1 Dougherty, T.J., Kaufman, J.E., Goldfarb,
2
3 4
8 9 10
11
12
13 14
15
A., Weishaupt, K.R., Boyle, D. and Mittelman, A. (1978) Photoradiation therapy for the treatment of malignant tumours. Cancer Res., 38, 2628-2635. Forbes, I.J., Cowled, P.A., Leong, A.&Y., Ward, A.D., Black, R.B., Blake, A.J. and Jacka, F.J. (1980) Phototherapy of human tumours using haematoporphyrin derivative. Med. J. Aust., 2, 489-493. Dahlman, A., Wile, A.G., Burns, R.G., Mason, G.R., Johnson, F.M. and Berns, M.W. (1983) Laser photoradiation therapy of cancer. Cancer Res., 43,430-434. Weishaupt, K.R., Gomer, C.J. and Dougherty, T.J. (1976) Identification of singlet oxygen as the cytotoxic agent in photoinactivation of a murine tumor. Cancer Res., 36,2326-2329. Moan, J. and Sommer, S. (1981) Fluorescence and absorbtion properties of the components of hematoporphyrin derivative. Photobiochem. Photobiophys., 3, 93-103. Lee See, K., Forbes, I.J. and Betts, W.H. (1984) Oxygen dependency of photocytotoxicity with haematoporphyrin derivative. Photochem. Photobiol., 39, 631-634. Hayata, Y., Kato, H., Konaka, C., Ono, J. and Takizawa, N. (1982) Hematoporphyrin derivative and laser photoradiation in the treatment of lung cancer. Chest, 81, 269277. Cortese, D.A. and Kinsey, J.H. (1982) Endoscopic management of lung cancer with hematoporphyrin derivative phototherapy. Mayo Clin. Proc., 57, 543-547. Moan, J. and Sommer, S. (1983) Uptake of the components of hematoporphyrin derivative by cells and tumours. Cancer Letters, 21, 167-174. Dougherty, T.J., Boyle, D.G., Weishaupt, K.R., Henderson, B.A., Potter, W.R., Bellnier, D.A. and Wityk, K.E. (1983) Photoradiation therapy: clinical and drug advances. Adv. Exp. Med. Biol., 160, 3-13. Cowled, P.A., Forbes, I.J., Swincer, A.G., Trenerry, V.C. and Ward, A.D. (1985) Separation and phototoxicity in vitro of some of the components of Haematoporphyrin derivative. Photochem. Photobiol., in press. Cowled, P.A., Grace, J.R. and Forbes, I.J. (1984) Comparison of the efficacy of pulsed and continuous wave red laser light in induction of photocytotoxicity by haematoporphyrin derivative. Photochem. Photobiol., 39, 115-117. Jacka, F. and Blake, A.J. (1983) A lamp for cancer phototherapy. Aust. J. Phys., 36, 221-226. Ellingsen, R., Svaasand, L.O. and Odegard, A. (1984) Photoradiation therapy (PRT) of Lewis Lung carcinoma in B,D, mice, dosimetry considerations. In: Porphyrins in Tumour Phototherapy, pp. 213-220. Editors: A. Andreoni and R. Cubeddu. Plenum Press, New York. Moan, J., Christensen, T. and Sommer, S. (1982) The main photosensitising components of hematoporphyrin derivative. Cancer Letters, 15.161-166.
118 16 Kessel, D., (1982) Components of hematoporphyrin derivative and their tumour localising capacity. Cancer Res., 42,1703-1706. 17 Poletti, A., Murgia, S.M., Pasqua, A., Reddi, E. and Jori, G. (1984) Photophysical and photosensitising properties of Photofrin II. In: Porphyrins in Tumour Phototherapy, pp. 37-43. Editors: A. Andreoni and R. Cuheddu. Plenum Press, New York. 18 Gomer, C.J. and Razum, N.J. (1984) Acute skin responses in albino mice following porphyrin photosensitisation under oxic and anoxic conditions. Photochem. Photobiol., 40,435-440. 19 Carpenter, R.J., Ryan, R.J., Neel, H.B. and Sanderson, D.R. (1977) Tumor fluorescence with hematoporphyrin derivative. Ann. Otol., 86, 661-666. 20 Tomio, L., Reddi, E., Jori, G., Zorat, P.L., Pizzi, G.B. and Calzavara, F. (1980) Hematoporphyrin as a sensitizer in tumor phototherapy: effect of medium polarity on the photosensitizing efficiency and role of administration pathway on the distribution in normal and tumor bearing rats. In: Lasers in Photomedicine and Photobiology: Springer Series in Optical Sciences, Vol. 22, pp. 7682. Editors: R. Pratesi and C.A. Sacchi. Springer-Verlag, Berlin, Heidelberg, New York.