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Modification of oral mucositis (Mouse) by keratinocyte growth factor (KGF): effect of KGF treatment protocol
76
I. J. Radiation Oncology
● Biology ● Physics
Volume 54, Number 2, Supplement, 2002
contrast, control mice, or mice treated with adv.oriP.ncFasL or RT alone demonstrated progressive tumour growth. The healthiest appearing mice are the ones treated with the therapeutic gene. Conclusions: We are the first group to have successfully developed a novel adenoviral vector incorporating EBV-specific gene expression of a mutant FasL, which achieves significant cytotoxicity mediated primarily by apoptosis through the Fas pathway. This strategy also suppresses tumour formation, and in combination with RT, result in significant tumour regression. Most importantly, this strategy appears to be safe, and well tolerated in the host mice. This work is funded by the Canadian Institutes of Health Research, and the Elia Chair for Head/Neck Cancer Research.
127
Modification of Oral Mucositis (Mouse) by Keratinocyte Growth Factor (KGF): Effect of KGF Treatment Protocol
W. Doerr, K. Heider, S. Reichel, K. Spekl Depts. of Radiotherapy and Radiation Oncology, Technical University, Dresden, Germany Purpose/Objective: The present study was initiated to quantify the mucoprotective efficacy of keratinocyte growth factor (rHuKGF) in mouse oral mucosa with respect to growth factor treatment protocol. For this, KGF was administered in addition to daily fractionated radiation treatment. Materials/Methods: The mouse tongue model was used in the present investigation. Radiation-induced mucosal ulceration was analysed as the clinically relevant quantal endpoint. Daily fractionated irradiation with 5x3 Gy per week was given to the whole snout, followed by graded top-up doses in order to generate full dose effect curves. At least five dose groups with 8-10 animals each were used per protocol. Top-up irradiation, as well as single dose irradiation, was applied to a test area of the lower tongue surface only. rHuKGF was kindly provided by AMGEN Inc., Thousand Oaks, CA, U.S.A. All rHuKGF injections were given subcutaneously. In a first experiment, fractionation with 5x3 Gy over 1 week was followed by top-up irradiation on day 7. Based on previous studies, KGF was applied either before the onset of fractionated irradiation (day -3, day -1 or days -3 to -1) or at the end of the first treatment week (day 4 or days ⫹4 to ⫹6). KGF doses applied were 5, 15, 22.5, or 30 mg/kg in single injections or 5 mg/kg per injection in protocols with daily repeated injections. In an additional experiment, fractionated irradiation was applied over 2 weeks, with top-up irradiation on day 14. KGF (5 mg/kg per injection) was administered on days ⫹4, ⫹5, ⫹6 and ⫹11, or on days ⫹4 and ⫹11, or days ⫹5 and ⫹11. Results: In a single dose control experiment, the ED50, i.e. the dose after which ulcer induction is expected in 50% of the mice, was 10.7⫾1.0 Gy. After 5x3 Gy, the ED50 for top-up irradiation on day 7 was 5.1⫾1.9 Gy. This decrease in ED50 reflects the reduction in mucosal radiation tolerance by the fractionated radiation doses. KGF administered on day -1 resulted in ED50-values of 10.7⫾1.7 Gy and 14.1⫾1.5 Gy at doses of 5 and 15 mg/kg, respectively. Repeated injections (days -3 to -1) of 3x5 mg/kg yielded an ED50 of 7.8⫾3.3 Gy (pvs. control⫽0.01). After a single injection of 15 mg/kg on day -3, an ED50 of 12.1⫾1.3 Gy was found. KGF given on day ⫹4 resulted in ED50 values of 11.3⫾1.3 Gy, 14.1⫾1.8 Gy, 14.4⫾1.3 Gy and 13.4⫾1.4 Gy at doses of 5, 15, 22.5 and 30 mg/kg, respectively. With 3 daily injections of 5 mg/kg each, on days ⫹4 to ⫹6, an ED50 of 13.5⫾2.3 Gy was observed. With fractionated irradiation over 2 weeks, the ED50 for top-up irradiation alone was 5.7⫾1.5 Gy, indicating the high effectiveness of repopulation processes in oral mucosa. ED50-values in combination with KGF treatment were 12.3⫾1.8 Gy (days -3 to -1, ⫹4), 12.8⫾1.1 Gy (days ⫹4 to ⫹6, ⫹11), 14.0⫾0.1 Gy (days ⫹4 and ⫹11) and 14.3⫾1.4 Gy (days ⫹5 and ⫹11), respectively. Conclusions: A marked increase in oral mucosal radiation tolerance by rHuKGF in all protocols tested. With 1 week of fractionated irradiation, a clear effect of the KGF dose was observed at both time points when the dose was increased from single injections of 5 mg/kg to 15 mg/kg. At higher KGF doses, no further increase in ED50-values was seen. Compared to 3 injections of 5 mg/kg each, on days -3 to -1, single injections were more effective. In contrast, no significant difference was found between 3 injections on days ⫹4 to ⫹6 and a single injection of 15 mg/kg on day ⫹4, while a single injection of 5 mg/kg was slightly less effective. In combination with 2 weeks of radiation treatment, KGF given in two single injections at the end of week 1 was slightly more effective than 3 injections over the first weekend plus one additional injection at the end of the second week. These results indicate that single KGF injections, given before the onset of radiotherapy and at the end of treatment weeks 1 and 2, should be most effective. In the mouse model, an optimum rHuKGF-dose of 15 mg/kg per injection was defined. rHuKGF has shown no or only very minor effects on tumor cells in vitro as well as on xenograft tumors in nude mice. Therefore, a selective reduction of oral mucosal toxicity during radiation treatment of head and neck tumors can be expected.
128
Keratinocyte Growth Factor (FGF7) Does Not Affect Growth Characteristics and Radiation Response of Early Passage Human Tumor Cells
A. Hille1, M. Rave-Fraenk1, C. Damm1, W. Doerr2, O. Pradier1, C.F. Hess1, H. Schmidberger1 1 Department of Radiotherapy, University Goettingen, Goettingen, Germany, 2Department of Radiotherapy, University Dresden, Dresden, Germany Purpose/Objective: Amelioration of the radiation-induced mucositis aiming at avoiding treatment interruptions could increase the therapeutic ratio of radiotherapy in head and neck cancer (HNSCC). Keratinocyte growth factor (FGF7), a member of the fibroblast growth factor family, is produced by mesenchymal cells; its target cells are epithelial cells in a variety of tissues. FGF7 has been identified as a mediator of proliferation and modulation of migration and differentiation processes in squamous epithelia and to ameliorate the acute response to radiation in animal models. One concern with respect to the clinical use of FGF7 is that the agent not only protects normal epithelia but also the tumor from cytoablative therapy. The purpose of this study is to investigate the in vitro effect FGF7 on proliferation, clonogenic capacity, and radiation response of primary or low passage human epithelial tumor cells.
I. J. Radiation Oncology
● Biology ● Physics
Volume 54, Number 2, Supplement, 2002
contrast, control mice, or mice treated with adv.oriP.ncFasL or RT alone demonstrated progressive tumour growth. The healthiest appearing mice are the ones treated with the therapeutic gene. Conclusions: We are the first group to have successfully developed a novel adenoviral vector incorporating EBV-specific gene expression of a mutant FasL, which achieves significant cytotoxicity mediated primarily by apoptosis through the Fas pathway. This strategy also suppresses tumour formation, and in combination with RT, result in significant tumour regression. Most importantly, this strategy appears to be safe, and well tolerated in the host mice. This work is funded by the Canadian Institutes of Health Research, and the Elia Chair for Head/Neck Cancer Research.
127
Modification of Oral Mucositis (Mouse) by Keratinocyte Growth Factor (KGF): Effect of KGF Treatment Protocol
W. Doerr, K. Heider, S. Reichel, K. Spekl Depts. of Radiotherapy and Radiation Oncology, Technical University, Dresden, Germany Purpose/Objective: The present study was initiated to quantify the mucoprotective efficacy of keratinocyte growth factor (rHuKGF) in mouse oral mucosa with respect to growth factor treatment protocol. For this, KGF was administered in addition to daily fractionated radiation treatment. Materials/Methods: The mouse tongue model was used in the present investigation. Radiation-induced mucosal ulceration was analysed as the clinically relevant quantal endpoint. Daily fractionated irradiation with 5x3 Gy per week was given to the whole snout, followed by graded top-up doses in order to generate full dose effect curves. At least five dose groups with 8-10 animals each were used per protocol. Top-up irradiation, as well as single dose irradiation, was applied to a test area of the lower tongue surface only. rHuKGF was kindly provided by AMGEN Inc., Thousand Oaks, CA, U.S.A. All rHuKGF injections were given subcutaneously. In a first experiment, fractionation with 5x3 Gy over 1 week was followed by top-up irradiation on day 7. Based on previous studies, KGF was applied either before the onset of fractionated irradiation (day -3, day -1 or days -3 to -1) or at the end of the first treatment week (day 4 or days ⫹4 to ⫹6). KGF doses applied were 5, 15, 22.5, or 30 mg/kg in single injections or 5 mg/kg per injection in protocols with daily repeated injections. In an additional experiment, fractionated irradiation was applied over 2 weeks, with top-up irradiation on day 14. KGF (5 mg/kg per injection) was administered on days ⫹4, ⫹5, ⫹6 and ⫹11, or on days ⫹4 and ⫹11, or days ⫹5 and ⫹11. Results: In a single dose control experiment, the ED50, i.e. the dose after which ulcer induction is expected in 50% of the mice, was 10.7⫾1.0 Gy. After 5x3 Gy, the ED50 for top-up irradiation on day 7 was 5.1⫾1.9 Gy. This decrease in ED50 reflects the reduction in mucosal radiation tolerance by the fractionated radiation doses. KGF administered on day -1 resulted in ED50-values of 10.7⫾1.7 Gy and 14.1⫾1.5 Gy at doses of 5 and 15 mg/kg, respectively. Repeated injections (days -3 to -1) of 3x5 mg/kg yielded an ED50 of 7.8⫾3.3 Gy (pvs. control⫽0.01). After a single injection of 15 mg/kg on day -3, an ED50 of 12.1⫾1.3 Gy was found. KGF given on day ⫹4 resulted in ED50 values of 11.3⫾1.3 Gy, 14.1⫾1.8 Gy, 14.4⫾1.3 Gy and 13.4⫾1.4 Gy at doses of 5, 15, 22.5 and 30 mg/kg, respectively. With 3 daily injections of 5 mg/kg each, on days ⫹4 to ⫹6, an ED50 of 13.5⫾2.3 Gy was observed. With fractionated irradiation over 2 weeks, the ED50 for top-up irradiation alone was 5.7⫾1.5 Gy, indicating the high effectiveness of repopulation processes in oral mucosa. ED50-values in combination with KGF treatment were 12.3⫾1.8 Gy (days -3 to -1, ⫹4), 12.8⫾1.1 Gy (days ⫹4 to ⫹6, ⫹11), 14.0⫾0.1 Gy (days ⫹4 and ⫹11) and 14.3⫾1.4 Gy (days ⫹5 and ⫹11), respectively. Conclusions: A marked increase in oral mucosal radiation tolerance by rHuKGF in all protocols tested. With 1 week of fractionated irradiation, a clear effect of the KGF dose was observed at both time points when the dose was increased from single injections of 5 mg/kg to 15 mg/kg. At higher KGF doses, no further increase in ED50-values was seen. Compared to 3 injections of 5 mg/kg each, on days -3 to -1, single injections were more effective. In contrast, no significant difference was found between 3 injections on days ⫹4 to ⫹6 and a single injection of 15 mg/kg on day ⫹4, while a single injection of 5 mg/kg was slightly less effective. In combination with 2 weeks of radiation treatment, KGF given in two single injections at the end of week 1 was slightly more effective than 3 injections over the first weekend plus one additional injection at the end of the second week. These results indicate that single KGF injections, given before the onset of radiotherapy and at the end of treatment weeks 1 and 2, should be most effective. In the mouse model, an optimum rHuKGF-dose of 15 mg/kg per injection was defined. rHuKGF has shown no or only very minor effects on tumor cells in vitro as well as on xenograft tumors in nude mice. Therefore, a selective reduction of oral mucosal toxicity during radiation treatment of head and neck tumors can be expected.
128
Keratinocyte Growth Factor (FGF7) Does Not Affect Growth Characteristics and Radiation Response of Early Passage Human Tumor Cells
A. Hille1, M. Rave-Fraenk1, C. Damm1, W. Doerr2, O. Pradier1, C.F. Hess1, H. Schmidberger1 1 Department of Radiotherapy, University Goettingen, Goettingen, Germany, 2Department of Radiotherapy, University Dresden, Dresden, Germany Purpose/Objective: Amelioration of the radiation-induced mucositis aiming at avoiding treatment interruptions could increase the therapeutic ratio of radiotherapy in head and neck cancer (HNSCC). Keratinocyte growth factor (FGF7), a member of the fibroblast growth factor family, is produced by mesenchymal cells; its target cells are epithelial cells in a variety of tissues. FGF7 has been identified as a mediator of proliferation and modulation of migration and differentiation processes in squamous epithelia and to ameliorate the acute response to radiation in animal models. One concern with respect to the clinical use of FGF7 is that the agent not only protects normal epithelia but also the tumor from cytoablative therapy. The purpose of this study is to investigate the in vitro effect FGF7 on proliferation, clonogenic capacity, and radiation response of primary or low passage human epithelial tumor cells.