- Email: [email protected]
or Cisplatin on Cholangiocarcinoma Stem-Like Cells
Volume 90 Number 1S Supplement 2014
3467 In Vitro Determination of the Relative Biological Effectiveness of Carbon Ion Beam Irradiation in Cells Derived From Human Normal Lung Tissues N. Okano,1 T. Oike,1 J. Saitoh,1 K. Shirai,2 T. Kiyono,3 M. Enari,3 M. Isono,2 K. Torikai,2 T. Ohno,2 and T. Nakano1; 1Gunma University, Maebashi, Gunma, Japan, 2Gunma University Heavy Ion Medical Center, Gunma University, Maebashi, Gunma, Japan, 3National Cancer Center Research Institute, Tokyo, Japan Purpose/Objective(s): Radiation-induced lung injury (RILI) is a serious concern in carbon-ion (C-ion) radiation therapy (CIRT) for thoracic malignancies. However, current shortage of knowledge in biological effect of C-ion beam irradiation to normal lung tissues makes it difficult to predict RILI in CIRT. This problem is partially based on the fact that the relative biological effectiveness (RBE) in cells derived from human normal lung tissues, which lack colony-forming ability, cannot be assessed by colony formation assay. To address this issue, we tested another cell survival assay for the validity as an alternative to colony formation assay, and assessed the RBE in such cells. Materials/Methods: An immortalized small airway epithelial cells (iSAECs) originated from human bronchi, a human lung fibroblast cell line HFL-I, and a human lung adenocarcinoma cell line A549 were used. X-ray irradiation was performed by a Faxitron RX-650 (100 kVp, 1.14 Gy/min) C-ion beam irradiation was performed employing the clinically-used beam spec; 290 MeV/nucleon, the average linear energy transfer at the center of a 6 cm spread-out Bragg peak of w50 keV/mm. Radiosensitivity of iSAECs and HFL-I cells was assessed by an assay previously reported by Guzzi F, et al. (Guzzi’s assay, Oncogene 2002). Cells irradiated with X-ray or C-ion beam were cultured for 10 days, fixed with methanol, stained with crystal violet and solubilized with acetic acid. The absorbance of the solution was measured using a spectrophotometer and used to calculate relative surviving fraction (SF). Agreement between the results of Guzzi’s assay and colony formation assay was evaluated by comparing the SFs of the same set of X-ray-irradiated A549 cells showing colony-forming ability. The survival curves were generated by fitting the SFs to the linearquadratic model. RBE values were calculated at D50, the dose giving 50% SF. Results: The absorbance values obtained from Guzzi’s assay were highly correlated to the number of colonies in colony formation assay (R Z 0.905, P < 0.01 by Pearson’s correlation coefficient), resulting in strong agreement between the survival curves generated using the two assays. In Guzzi’s assay, the D50 values for X-ray and C-ion beam irradiation in iSAECs was 8.3 and 2.6 Gy, respectively, resulting in the RBE value of 3.2. The D50 values for X-ray and C-ion beam irradiation in HFL-I cells was 3.3 and 1.5 Gy, respectively, resulting in the RBE value of 2.2. Conclusions: These results suggest that Guzzi’s assay can be a viable alternative to the golden standard colony formation assay in the assessment of radiosensitivity. The RBE of C-ion beam irradiation in iSAECs and HFL-I cells obtained in the present study will help the understanding of the biological effect of CIRT on human normal lung tissues. Author Disclosure: N. Okano: None. T. Oike: None. J. Saitoh: None. K. Shirai: None. T. Kiyono: None. M. Enari: None. M. Isono: None. K. Torikai: None. T. Ohno: None. T. Nakano: None.
3468 Pulsed Low-Dose-Rate External Beam Radiation Therapy: An in-vivo Study on Prostate Cancer B. Wang,1 P. Zhang,2,1 D. Cvetkovic,1 X. Chen,1 C. Zhang,3,1 L. Chen,1 and C. Ma1; 1Fox Chase Cancer Center, Philadelphia, PA, 2Sichuan Cancer Hospital, Chengdu, China, 3Qiqihar Medical University, Qiqihar, China Purpose/Objective(s): Although hyper radio-sensitivity has been demonstrated at low doses below 0.4 Gy for various tumor cell lines in
Poster Viewing Abstracts S787 vitro, there was no report on in vivo studies for human prostate cancer cell lines. In our previous in vivo studies, we performed total body irradiation of nude mice with both pulsed low dose rate (PLDR) and conventional radiation therapy (CRT) techniques. The PLDR treated animal group showed significantly lower level of radiation toxicity to normal tissues than CRT, which leads to much longer survival of the irradiated animals. The purpose of this study is to investigate the local tumor control of the PLDR treatment for prostate cancer with an animal model of human prostate cancer LNCaP to explore the potential RT treatment option for recurrent cancer patients. Materials/Methods: We used an orthotopic murine model of human prostate cancer for our study and evaluated the in vivo treatment efficacy by measuring the tumor volume weekly using MRI. LNCaP cells were implanted into immune-suppressed male nude mice via surgery. The mice with tumors were randomly allocated into a PLDR group (n Z 9), a CRT group (n Z 7), and a control group (n Z 7). The mice in the PLDR and CRT groups were treated with 2Gy radiation dose for one time. For the CRT treatment, the mice received 2Gy at a dose rate of 300 MU/minute. For the PLDR treatment, the 2Gy dose was further divided into ten pulses of 0.2Gy at the same dose rate with an interval of 3 minutes between the pulses. Results: We monitored the tumor growth via MRI on a weekly basis. Sizable delay of the tumor growth after the treatments was observed for the PLDR and CRT groups. The mean normalized tumor volumes ( standard deviation of the mean) were 1.530.07, 1.530.14, and 1.810.09 at one week after treatment, 2.280.13, 2.190.16, and 3.040.25 at two weeks after treatment, and 3.310.23, 3.140.24 and 4.620.49 at three weeks after treatment, for the PLDR, CRT, and control groups, respectively. Conclusions: The PLDR and CRT treatments showed comparable efficacy in local tumor control for our animal model of LNCaP cell line. Therefore, our in vivo results imply that the PLDR treatment technique could be a viable approach for the treatment of recurrent prostate cancer after radiation therapy. Author Disclosure: B. Wang: None. P. Zhang: None. D. Cvetkovic: None. X. Chen: None. C. Zhang: None. L. Chen: None. C. Ma: None.
3469 Effects of Carbon Ion Beam Alone or in Combination With Gemcitabine and/or Cisplatin on Cholangiocarcinoma Stem-Like Cells S. Sai,1 T. Wakai,2 and T. Shirai1; 1National Institute of Radiological Sciences, Chiba, Japan, 2Division of Digestive and General Surgery, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan Purpose/Objective(s): To examine whether carbon ion beam alone or in combination with gemcitabine and/or cisplatin is effective in targeting putative human cholangiocarcinoma stem cells. Materials/Methods: Putative cancer stem cells (CSCs) sorted from HuCCT1 and Huh28 cells were treated with or without carbon ion beam, X-ray alone or in combination with gemcitabine and/or cisplatin, then colony and spheroid formation assay, FACS analysis, gamma-H2AX foci assay were performed. Results: Colony and spheroid formation assay as well as in vivo tumorigenic assay showed that CD44+/ESA+, CD133+/CD90+ cells exactly have CSC properties compared to CD44-/ESA- and CD133-/CD90- cells. The proportion of CD44+/ESA+, CD133+/CD90+ was increased by X-ray compared to carbon-ion beam alone, and it was further enhanced when combine with gemcitabine and/or cisplatin. The number of colony and spheroid formed from CSCs was remarkably reduced after carbon ion beam combined with gemcitabine and/or cisplatin compared to that of Xray or carbon ion beam alone. A more large number and large-sized gamma-H2AX foci were observed in CSCs after 24 h carbon ion beam combined with gemcitabine and/or cisplatin compared to that of X-ray or carbon ion beam alone.
S788
International Journal of Radiation Oncology Biology Physics
Conclusions: Carbon ion beam combined with gemcitabine and/or cisplatin effectively induced prolonged and more complex DNA damage, as a result effectively destroyed cholangiocarcinoma stem cells. Author Disclosure: S. Sai: None. T. Wakai: None. T. Shirai: None.
Scientific Abstract 3471;Table examination of the lung tissue
6th week
-Normal
-Vacuoles
3470
16th week
-Identical to the counterparts of the same age
-More and larger vacuoles than 6th week
Local Tumor Control and Normal Tissue Toxicity of Pulsed LowDose-Rate Radiation Therapy (PLRT) for Recurrent Lung Cancer: An In Vivo Study P. Zhang,1 B. Wang,2 X. Chen,2 D. Cvetkovic,2 L. Chen,2 J. Lang,1 and C. Ma2; 1Department of Radiation Oncology, Sichuan Cancer Hospital, Chengdu, China, 2Radiation Oncology Department, Fox Chase Cancer Center, Philadelphia, PA Purpose/Objective(s): The purpose of this in vivo study was to investigate the tumor control and normal tissue toxicity of pulsed low dose rate radiation therapy (PLRT) for recurrent lung cancer. Materials/Methods: This study consisted of two experiments: (1) total body irradiation (TBI) to investigate the normal tissue toxicity of PLRT and (2) PLRT treatment of A549 tumor-bearing mice to investigate the tumor local control. For the TBI experiment, 20 mice were divided into two groups randomly. One group was irradiated continuously at a dose rate of 300 MU/min. The other group was irradiated with 400.2Gy pulses with a 3min internal. The total body radiation dose was 8Gy in one treatment. The weight of each mouse was measured daily. For the A549 tumor treatment experiment, 1106 A549 cells were injected subcutaneously into the left and right abdominal region. The tumor-bearing mice were randomly assigned to three groups: (1) control group (n Z 10), (2) conventional RT group (n Z 10), and (3) PLRT group (n Z 10). When the tumor volume reached approximately 80 mm3, treatment was initiated. All treatments were delivered in two fractions of 2Gy each in two consecutive days. Following the treatment, mice were MR-scanned weekly and the tumor volumes were measured using MRI (resolution: 0.2mm) for tumor growth monitoring. Results: In the TBI experiment, there was a significant difference in the weight and survival time between the mice treated with conventional RT and PLRT; the average weight of the conventional RT group decreased continuously from 26.550.93g to 20.941.68g while the PLRT group showed no change in weight, and the survival time for the conventional RT group was eight days and for the PLRT group it was twelve days. For the tumor control experiment using the A549 xenograft model, the results showed that both conventional RT and PLRT could significantly inhibited the growth of A549 tumors compared with the untreated control group (>35% difference in the mean tumor volume). The PLRT results were slightly better than the conventional RT treatment in terms of the mean tumor volume although there was no significant difference (8% difference in the mean tumor volume) between the two groups. Conclusions: This study showed that pulsed low dose rate radiation therapy (PLRT) could control A549 tumors as effectively as conventional RT with less normal tissue toxicity. Thus, PLRT would be a good modality for treating recurrent lung cancers. Author Disclosure: P. Zhang: None. B. Wang: None. X. Chen: None. D. Cvetkovic: None. L. Chen: None. J. Lang: None. C. Ma: None.
3471 The Effect of Dose Rate per Fraction on Lung Toxicity P. Hurmuz,1 H. Kivanc,1 A. Canbolat,1 G. Yavas,2 D. Yuce,1 S. Onder,1 M. Sargon,1 and F. Yildiz1; 1Hacettepe Universitesi Tip Fakultesi, Ankara, Turkey, 2Selcuk Universitesi Tip Fakultesi, Konya, Turkey Purpose/Objective(s): The aim of this study is to evaluate the effect of increasing dose rate (DR) per fraction in lung tissue for external radiation therapy. Materials/Methods: The rats included in the study were randomly grouped into three groups containing 16 rats each: Group (G) 1 was defined as
Group 1
The summary of the electron microscopic Group 2
Group 3 -Vacuoles -Mild fibrosis -Larger vacuoles compared to 6th week -Prominent fibrosis -Severe intercellular edema
control group, and in this group rats were sham irradiated. G2 was the group receiving a single dose of 12 Gy in DR of 300 monitor unit (MU)/ minute. G3 was the group receiving a single dose of 12 Gy in DR of 600 MU/minute. Radiation therapy (RT) was applied under general anesthesia with 6 MV photon beams to both lungs. Total monitor units were delivered in 8 fragments with 2 minutes of intervals in order to simulate the duration of intensity modulated radiation therapy treatment fraction. Thus the total fraction durations in G2 (300 MU) and in G3 (600 MU) were 18 minutes and 16 minutes, respectively At the 6th and 16th weeks of the RT animals from each group were sacrificed for light and electron microscopy evaluation. Results: Light microscopic evaluation revealed significant changes in G2 and G3 compared to G1. In G2 there were significant increases in the perivascular fibrosis (p Z 0.018), interstitial fibrosis (p Z 0.002), total inflammation (p Z 0.040) and total fibrosis (p Z 0.003) scores in 16th week compared to 6th weeek. In G3, we found statistically significant increases in perivascular fibrosis (p Z 0.001), interstitial fibrosis (p Z 0.002) and total fibrosis scores (p Z 0.029) by time. There was no significant difference in the total inflammation score in G3 (p Z 0.225). When we compared G2 and G3 in the 6th week we found significant increase in the interstitial thickening (p Z 0.039) and total inflammation (p Z 0.035) scores in G3. Electron microscopic evaluation showed vacuoles in G2 and G3 however there was also mild fibrosis in G3 in the 6th week. 16th week examination demonstrated more and larger vacuoles than the 6th week in G2 and G3 plus prominent fibrosis and severe intercellular edema in G3 (Table 1). Conclusions: DR per fraction may have an impact on normal tissue toxicity. The prominent effect of increased DR in lung tissue is fibrosis which should be kept in mind especially in cases where higher doses per fraction are used. Author Disclosure: P. Hurmuz: None. H. Kivanc: None. A. Canbolat: None. G. Yavas: None. D. Yuce: None. S. Onder: None. M. Sargon: None. F. Yildiz: None.
3472 Experimental Radiation (2400 MU/min/50cGy) In-Vitro Achieved Significant Increase in Apoptosis on Melanoma Cells With Superior Survival of Normal Cells When Compared to 400 MU/min S. Sarojini, N. Milinovikj, A. Pecora, J. Barbiere, M. Tuna, S. Jeong, J. Jun, L. Catello, J. Jiang, A. Ndlovu, A. Ingenito, and K.S. Suh; Hackensack University Medical Center, Hackensack, NJ Purpose/Objective(s): Purpose of this study was to investigate apoptotic effects of 10MV x-rays at high dose rate (2400 MU/min) combined with unconventional low total dose (50cGy) in malignant melanoma cells in vitro. Concurrently, the preservation of cell survival on normal melanocytes (HEM), keratinocytes (HEK) and stromal cells (HEF) and radiosensitization using mitochondrial inhibitors were also studied. Materials/Methods: Cell growth condition and density of cells were optimized by customizing culture media, cell confluence and supplemental factors. The cells were radiated using a linear accelerator with 400 or 2400 MU/min dose rates and total doses from 25-100 cGy. Cell counting, Colony Assays, Mitotracker fluorescent and bright fileld imaging, and
3467 In Vitro Determination of the Relative Biological Effectiveness of Carbon Ion Beam Irradiation in Cells Derived From Human Normal Lung Tissues N. Okano,1 T. Oike,1 J. Saitoh,1 K. Shirai,2 T. Kiyono,3 M. Enari,3 M. Isono,2 K. Torikai,2 T. Ohno,2 and T. Nakano1; 1Gunma University, Maebashi, Gunma, Japan, 2Gunma University Heavy Ion Medical Center, Gunma University, Maebashi, Gunma, Japan, 3National Cancer Center Research Institute, Tokyo, Japan Purpose/Objective(s): Radiation-induced lung injury (RILI) is a serious concern in carbon-ion (C-ion) radiation therapy (CIRT) for thoracic malignancies. However, current shortage of knowledge in biological effect of C-ion beam irradiation to normal lung tissues makes it difficult to predict RILI in CIRT. This problem is partially based on the fact that the relative biological effectiveness (RBE) in cells derived from human normal lung tissues, which lack colony-forming ability, cannot be assessed by colony formation assay. To address this issue, we tested another cell survival assay for the validity as an alternative to colony formation assay, and assessed the RBE in such cells. Materials/Methods: An immortalized small airway epithelial cells (iSAECs) originated from human bronchi, a human lung fibroblast cell line HFL-I, and a human lung adenocarcinoma cell line A549 were used. X-ray irradiation was performed by a Faxitron RX-650 (100 kVp, 1.14 Gy/min) C-ion beam irradiation was performed employing the clinically-used beam spec; 290 MeV/nucleon, the average linear energy transfer at the center of a 6 cm spread-out Bragg peak of w50 keV/mm. Radiosensitivity of iSAECs and HFL-I cells was assessed by an assay previously reported by Guzzi F, et al. (Guzzi’s assay, Oncogene 2002). Cells irradiated with X-ray or C-ion beam were cultured for 10 days, fixed with methanol, stained with crystal violet and solubilized with acetic acid. The absorbance of the solution was measured using a spectrophotometer and used to calculate relative surviving fraction (SF). Agreement between the results of Guzzi’s assay and colony formation assay was evaluated by comparing the SFs of the same set of X-ray-irradiated A549 cells showing colony-forming ability. The survival curves were generated by fitting the SFs to the linearquadratic model. RBE values were calculated at D50, the dose giving 50% SF. Results: The absorbance values obtained from Guzzi’s assay were highly correlated to the number of colonies in colony formation assay (R Z 0.905, P < 0.01 by Pearson’s correlation coefficient), resulting in strong agreement between the survival curves generated using the two assays. In Guzzi’s assay, the D50 values for X-ray and C-ion beam irradiation in iSAECs was 8.3 and 2.6 Gy, respectively, resulting in the RBE value of 3.2. The D50 values for X-ray and C-ion beam irradiation in HFL-I cells was 3.3 and 1.5 Gy, respectively, resulting in the RBE value of 2.2. Conclusions: These results suggest that Guzzi’s assay can be a viable alternative to the golden standard colony formation assay in the assessment of radiosensitivity. The RBE of C-ion beam irradiation in iSAECs and HFL-I cells obtained in the present study will help the understanding of the biological effect of CIRT on human normal lung tissues. Author Disclosure: N. Okano: None. T. Oike: None. J. Saitoh: None. K. Shirai: None. T. Kiyono: None. M. Enari: None. M. Isono: None. K. Torikai: None. T. Ohno: None. T. Nakano: None.
3468 Pulsed Low-Dose-Rate External Beam Radiation Therapy: An in-vivo Study on Prostate Cancer B. Wang,1 P. Zhang,2,1 D. Cvetkovic,1 X. Chen,1 C. Zhang,3,1 L. Chen,1 and C. Ma1; 1Fox Chase Cancer Center, Philadelphia, PA, 2Sichuan Cancer Hospital, Chengdu, China, 3Qiqihar Medical University, Qiqihar, China Purpose/Objective(s): Although hyper radio-sensitivity has been demonstrated at low doses below 0.4 Gy for various tumor cell lines in
Poster Viewing Abstracts S787 vitro, there was no report on in vivo studies for human prostate cancer cell lines. In our previous in vivo studies, we performed total body irradiation of nude mice with both pulsed low dose rate (PLDR) and conventional radiation therapy (CRT) techniques. The PLDR treated animal group showed significantly lower level of radiation toxicity to normal tissues than CRT, which leads to much longer survival of the irradiated animals. The purpose of this study is to investigate the local tumor control of the PLDR treatment for prostate cancer with an animal model of human prostate cancer LNCaP to explore the potential RT treatment option for recurrent cancer patients. Materials/Methods: We used an orthotopic murine model of human prostate cancer for our study and evaluated the in vivo treatment efficacy by measuring the tumor volume weekly using MRI. LNCaP cells were implanted into immune-suppressed male nude mice via surgery. The mice with tumors were randomly allocated into a PLDR group (n Z 9), a CRT group (n Z 7), and a control group (n Z 7). The mice in the PLDR and CRT groups were treated with 2Gy radiation dose for one time. For the CRT treatment, the mice received 2Gy at a dose rate of 300 MU/minute. For the PLDR treatment, the 2Gy dose was further divided into ten pulses of 0.2Gy at the same dose rate with an interval of 3 minutes between the pulses. Results: We monitored the tumor growth via MRI on a weekly basis. Sizable delay of the tumor growth after the treatments was observed for the PLDR and CRT groups. The mean normalized tumor volumes ( standard deviation of the mean) were 1.530.07, 1.530.14, and 1.810.09 at one week after treatment, 2.280.13, 2.190.16, and 3.040.25 at two weeks after treatment, and 3.310.23, 3.140.24 and 4.620.49 at three weeks after treatment, for the PLDR, CRT, and control groups, respectively. Conclusions: The PLDR and CRT treatments showed comparable efficacy in local tumor control for our animal model of LNCaP cell line. Therefore, our in vivo results imply that the PLDR treatment technique could be a viable approach for the treatment of recurrent prostate cancer after radiation therapy. Author Disclosure: B. Wang: None. P. Zhang: None. D. Cvetkovic: None. X. Chen: None. C. Zhang: None. L. Chen: None. C. Ma: None.
3469 Effects of Carbon Ion Beam Alone or in Combination With Gemcitabine and/or Cisplatin on Cholangiocarcinoma Stem-Like Cells S. Sai,1 T. Wakai,2 and T. Shirai1; 1National Institute of Radiological Sciences, Chiba, Japan, 2Division of Digestive and General Surgery, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan Purpose/Objective(s): To examine whether carbon ion beam alone or in combination with gemcitabine and/or cisplatin is effective in targeting putative human cholangiocarcinoma stem cells. Materials/Methods: Putative cancer stem cells (CSCs) sorted from HuCCT1 and Huh28 cells were treated with or without carbon ion beam, X-ray alone or in combination with gemcitabine and/or cisplatin, then colony and spheroid formation assay, FACS analysis, gamma-H2AX foci assay were performed. Results: Colony and spheroid formation assay as well as in vivo tumorigenic assay showed that CD44+/ESA+, CD133+/CD90+ cells exactly have CSC properties compared to CD44-/ESA- and CD133-/CD90- cells. The proportion of CD44+/ESA+, CD133+/CD90+ was increased by X-ray compared to carbon-ion beam alone, and it was further enhanced when combine with gemcitabine and/or cisplatin. The number of colony and spheroid formed from CSCs was remarkably reduced after carbon ion beam combined with gemcitabine and/or cisplatin compared to that of Xray or carbon ion beam alone. A more large number and large-sized gamma-H2AX foci were observed in CSCs after 24 h carbon ion beam combined with gemcitabine and/or cisplatin compared to that of X-ray or carbon ion beam alone.
S788
International Journal of Radiation Oncology Biology Physics
Conclusions: Carbon ion beam combined with gemcitabine and/or cisplatin effectively induced prolonged and more complex DNA damage, as a result effectively destroyed cholangiocarcinoma stem cells. Author Disclosure: S. Sai: None. T. Wakai: None. T. Shirai: None.
Scientific Abstract 3471;Table examination of the lung tissue
6th week
-Normal
-Vacuoles
3470
16th week
-Identical to the counterparts of the same age
-More and larger vacuoles than 6th week
Local Tumor Control and Normal Tissue Toxicity of Pulsed LowDose-Rate Radiation Therapy (PLRT) for Recurrent Lung Cancer: An In Vivo Study P. Zhang,1 B. Wang,2 X. Chen,2 D. Cvetkovic,2 L. Chen,2 J. Lang,1 and C. Ma2; 1Department of Radiation Oncology, Sichuan Cancer Hospital, Chengdu, China, 2Radiation Oncology Department, Fox Chase Cancer Center, Philadelphia, PA Purpose/Objective(s): The purpose of this in vivo study was to investigate the tumor control and normal tissue toxicity of pulsed low dose rate radiation therapy (PLRT) for recurrent lung cancer. Materials/Methods: This study consisted of two experiments: (1) total body irradiation (TBI) to investigate the normal tissue toxicity of PLRT and (2) PLRT treatment of A549 tumor-bearing mice to investigate the tumor local control. For the TBI experiment, 20 mice were divided into two groups randomly. One group was irradiated continuously at a dose rate of 300 MU/min. The other group was irradiated with 400.2Gy pulses with a 3min internal. The total body radiation dose was 8Gy in one treatment. The weight of each mouse was measured daily. For the A549 tumor treatment experiment, 1106 A549 cells were injected subcutaneously into the left and right abdominal region. The tumor-bearing mice were randomly assigned to three groups: (1) control group (n Z 10), (2) conventional RT group (n Z 10), and (3) PLRT group (n Z 10). When the tumor volume reached approximately 80 mm3, treatment was initiated. All treatments were delivered in two fractions of 2Gy each in two consecutive days. Following the treatment, mice were MR-scanned weekly and the tumor volumes were measured using MRI (resolution: 0.2mm) for tumor growth monitoring. Results: In the TBI experiment, there was a significant difference in the weight and survival time between the mice treated with conventional RT and PLRT; the average weight of the conventional RT group decreased continuously from 26.550.93g to 20.941.68g while the PLRT group showed no change in weight, and the survival time for the conventional RT group was eight days and for the PLRT group it was twelve days. For the tumor control experiment using the A549 xenograft model, the results showed that both conventional RT and PLRT could significantly inhibited the growth of A549 tumors compared with the untreated control group (>35% difference in the mean tumor volume). The PLRT results were slightly better than the conventional RT treatment in terms of the mean tumor volume although there was no significant difference (8% difference in the mean tumor volume) between the two groups. Conclusions: This study showed that pulsed low dose rate radiation therapy (PLRT) could control A549 tumors as effectively as conventional RT with less normal tissue toxicity. Thus, PLRT would be a good modality for treating recurrent lung cancers. Author Disclosure: P. Zhang: None. B. Wang: None. X. Chen: None. D. Cvetkovic: None. L. Chen: None. J. Lang: None. C. Ma: None.
3471 The Effect of Dose Rate per Fraction on Lung Toxicity P. Hurmuz,1 H. Kivanc,1 A. Canbolat,1 G. Yavas,2 D. Yuce,1 S. Onder,1 M. Sargon,1 and F. Yildiz1; 1Hacettepe Universitesi Tip Fakultesi, Ankara, Turkey, 2Selcuk Universitesi Tip Fakultesi, Konya, Turkey Purpose/Objective(s): The aim of this study is to evaluate the effect of increasing dose rate (DR) per fraction in lung tissue for external radiation therapy. Materials/Methods: The rats included in the study were randomly grouped into three groups containing 16 rats each: Group (G) 1 was defined as
Group 1
The summary of the electron microscopic Group 2
Group 3 -Vacuoles -Mild fibrosis -Larger vacuoles compared to 6th week -Prominent fibrosis -Severe intercellular edema
control group, and in this group rats were sham irradiated. G2 was the group receiving a single dose of 12 Gy in DR of 300 monitor unit (MU)/ minute. G3 was the group receiving a single dose of 12 Gy in DR of 600 MU/minute. Radiation therapy (RT) was applied under general anesthesia with 6 MV photon beams to both lungs. Total monitor units were delivered in 8 fragments with 2 minutes of intervals in order to simulate the duration of intensity modulated radiation therapy treatment fraction. Thus the total fraction durations in G2 (300 MU) and in G3 (600 MU) were 18 minutes and 16 minutes, respectively At the 6th and 16th weeks of the RT animals from each group were sacrificed for light and electron microscopy evaluation. Results: Light microscopic evaluation revealed significant changes in G2 and G3 compared to G1. In G2 there were significant increases in the perivascular fibrosis (p Z 0.018), interstitial fibrosis (p Z 0.002), total inflammation (p Z 0.040) and total fibrosis (p Z 0.003) scores in 16th week compared to 6th weeek. In G3, we found statistically significant increases in perivascular fibrosis (p Z 0.001), interstitial fibrosis (p Z 0.002) and total fibrosis scores (p Z 0.029) by time. There was no significant difference in the total inflammation score in G3 (p Z 0.225). When we compared G2 and G3 in the 6th week we found significant increase in the interstitial thickening (p Z 0.039) and total inflammation (p Z 0.035) scores in G3. Electron microscopic evaluation showed vacuoles in G2 and G3 however there was also mild fibrosis in G3 in the 6th week. 16th week examination demonstrated more and larger vacuoles than the 6th week in G2 and G3 plus prominent fibrosis and severe intercellular edema in G3 (Table 1). Conclusions: DR per fraction may have an impact on normal tissue toxicity. The prominent effect of increased DR in lung tissue is fibrosis which should be kept in mind especially in cases where higher doses per fraction are used. Author Disclosure: P. Hurmuz: None. H. Kivanc: None. A. Canbolat: None. G. Yavas: None. D. Yuce: None. S. Onder: None. M. Sargon: None. F. Yildiz: None.
3472 Experimental Radiation (2400 MU/min/50cGy) In-Vitro Achieved Significant Increase in Apoptosis on Melanoma Cells With Superior Survival of Normal Cells When Compared to 400 MU/min S. Sarojini, N. Milinovikj, A. Pecora, J. Barbiere, M. Tuna, S. Jeong, J. Jun, L. Catello, J. Jiang, A. Ndlovu, A. Ingenito, and K.S. Suh; Hackensack University Medical Center, Hackensack, NJ Purpose/Objective(s): Purpose of this study was to investigate apoptotic effects of 10MV x-rays at high dose rate (2400 MU/min) combined with unconventional low total dose (50cGy) in malignant melanoma cells in vitro. Concurrently, the preservation of cell survival on normal melanocytes (HEM), keratinocytes (HEK) and stromal cells (HEF) and radiosensitization using mitochondrial inhibitors were also studied. Materials/Methods: Cell growth condition and density of cells were optimized by customizing culture media, cell confluence and supplemental factors. The cells were radiated using a linear accelerator with 400 or 2400 MU/min dose rates and total doses from 25-100 cGy. Cell counting, Colony Assays, Mitotracker fluorescent and bright fileld imaging, and