分子靶向联合放射治疗头颈部鳞癌的实验研究
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摘要
目的:
探讨分子靶向治疗联合放射的最佳效应和多靶向联合放射的疗效,对VEGFR抗体联合放射以及VEGFR和EGFR抗体协同联合放射的作用进行了实验研究。
材料和方法:
1、建立FaDu头颈部鳞状细胞癌动物模型。头颈部鳞状细胞株FaDu来源于人咽鳞状细胞癌,从液氮中取出后复苏,配制成单细胞悬液,细胞浓度为1×10~8/ml。3—5月大小的瑞士nu—nu/Ncr雌性裸鼠被用于实验,1×10~6/10μlFaDu细胞种植于裸鼠右侧大腿肌肉内,FaDu肿瘤细胞形成移植瘤。
2、分别研究了VEGFR抗体DC101不同时相用药联合多分割放射的效应。移植瘤直径达到7mm(6.8-7.2mm)时,随机分组,包括对照组(无治疗),单纯药物组,单纯放射组,放射前用药联合放射组,放射同时用药联合放射组,放射后用药联合放射组,放射前+放射同时用药/联合放射组,放射同时+放射后用药/联合放射组,和放射前+放射同时+放射后用药/联合放射组。放射治疗为2.5Gy/次/天,连续6次。分别计算肿瘤8-12mm生长时间,比较其放射增敏效应。
3、瘤床放射后效应,用于评价放射与DC101的作用机制。将裸鼠随机分组,包括对照组(无治疗组),单纯药物组,单纯放射组,和药物+放射组。放射组对瘤床(裸鼠右侧大腿)先放射,放射剂量为2.5Gy/次/天,连续6次。24小时后各组分别种植肿瘤,48小时后开始药物治疗/无药物治疗。肿瘤直径达到7mm(6.8-7.2mm)时开始测量肿瘤大小,分别计算肿瘤8-12mm生长时间,比较其放射增敏效应。
4、DC101协同C225联合放射治愈肿瘤的疗效。移植瘤直径达到7mm(6.8-7.2mm)时,随机分组,包括对照组(单纯放射组),DC101联合放射组,C225联合放射组,和DC101协同C225联合放射组。放射治疗2.5Gy/次,2次/天,连续9天,24小时后各亚组加量照射10—40Gy。观察指标为肿瘤控制(肿瘤生长延迟和肿瘤复发),放射增益因子,肿瘤治愈和腹盆腔淋巴结转移率。
结果:
1、FaDu移植瘤成功率98%,分别选择直径6.8-7.2mm的肿瘤随机分组实验。
2、DC101放射同时和放射后用药放射增敏效益最大,增益因子分别是1.8和2.3。放射前用药无放射增敏作用;放射前加放射同时加放射后用药,产生最大的绝对生长延迟时间为18.5天,但是如果计算标准生长延迟时间,其结果与放射后用药相似,标准生长延迟时间分别为10.2天和10.5天,放射增益因子分别为2.2和2.3。
3、瘤床放射后比没有放射者明显延缓肿瘤生长,瘤床放射后DC101的增益因子为1.9,其机理是瘤床放射后增加对DC101的敏感性。
4、DC101协同C225联合放射与C225联合放射比较单纯放射能提高肿瘤的控制率,延长肿瘤复发的时间,增益因子分别是3.46和2.84;也能提高肿瘤的治愈率(P=0.004和0.005),但两者无统计学差异(P=0.433)。DC101联合放射与C225联合放射比较单纯放射能减少腹盆腔淋巴结转移率(P=0.01),DC101协同C225联合放射腹盆腔淋巴结转移率最低(P=0.0001)。
结论:
DC101能提高对头颈鳞癌FaDu的放射敏感性,DC101协同C225联合放射能提高肿瘤的局部控制率,降低腹盆腔淋巴结转移率。
Purpose:
To explore the efficacy of combination of molecular therapeutic and radiation including the optimal timing of radiation and drug and simultaneously targeting multiple signaling pathways.We investigated to optimize the combination of VEGFR-2 antibody with radiation and to integrate VEGFR-2 antibody,EGFR antibody with radiation in vivo.
Material and Methods:
1.FaDu xenografts of Head and Neck squamous cell cancer were set up.The human FaDu tumor cell line was originally derived from a human pharyngeal squamous cell carcer,which suspensions at 1×10~8/ml were prepared as monolayer in vitro.Female nude(nu-nu/Ncr) mice of 3-5 month-old were used.1×10~6/10μl tumor cells were inoculated intramuscularly into the mouse right hind legs for FaDu xenografts..
2.Different schedules of VEGFR-2 antibody DC101 and fractionated radiation were tested,such as neoadjuvnant,concurrent,adjuvant,neoadjuvnant+concurrent, concurrent+adjuvant,and neoadjuvnant+concurrent+adjuvant DC101.When the tumors had grown to 7 mm(6.8-7.2 mm) in diameter,the mice would be randomized into control(no treatmeat),different DC101 alone,radiation only,and different DC101 with radiation groups.The daily-fractionated dose of 2.5 Gy on 6 consecutive days(6×2.5Gy,15 Gy) was given.Time of tumor growth from 8-12mm was calculated respectively.Enhancement factors was compared.
3.A specifically designed tumor bed experiment of DC101 was performed to assess the mechanism of optimized interaction.The mice would be randomized into control(no treatmeat),DC101 only,radiation only,DC101 with radiation groups.The tumor bed(mouse right hind leg) was selectively radiated with 2.5 Gy in 6 fractions (6×2.5Gy,15 Gy).Tumor cells were injected intramuscularly at 24 hours after the last radiation dose,also for the unradiated mice at the same time.DC101 treatment started at 48 hours after tumor bed radiation.When tumors grew to 7mm(6.8-7.2 mm),the calculation was started.Time of tumor growth from 8-12mm was calculated respectively.Enhancement factors was compared.
4.An optimized schedule of DC101 was then integrated together with C225 and radiation in tumor cure assay.When tumors had grown to 7 mm(6.8-7.2 mm) in diameter,the mice would be randomized into control(radiation only),DC101 with radiation,C225 with radiation,or DC101 plus C225 with radiation groups.
The fractionated radiation was given twice daily at 2.5 Gy per fraction over 9 consecutive days(9×2×2.5Gy,45 Gy).One day after the last fractionated dose a range of single top-up doses up 10 to 40 Gy was given.Tumor control(tumor growth delay and tumor recurrence),radiation enhancement factor,tumor curability and incidence of abdominal and pelvic lymph node metastases were evaluated.
Results:
1.The survival rate of FaDu xenografts was 98%.The tumors between 6.8-7.2mm in diameter were used for randomization.
2.Concurrent and adjuvant DC101 with radiation were most effective in enhancing radiation-induced growth control(enhancement factors 1.8 and 2.3, respectively).Treatment with neoadjuvant DC101 did not result in radiation enhancement.When the extended DC101(neoadjuvant+ concurrent +adjuvant) was combined with radiation,this resulted in the longest absolute growth delay 18.5 days. However,if normalized against drug treatment,this induced similar radiation enhancement as adjuvant DC101(Normalized growth delays:radiation+extended DC101 schedule,10.2 days;radiation+adjuvant DC101,10.5 days;EF=2.2 for radiation+extended DC101 schedule;EF=2.3 for radiation+adjuvant DC101).
3.The tumor bed experiment resulted in significant enhancement of tumor growth delay in radiated tumor bed in comparison to unradiated tumor bed.Radiation enhancement was achieved for the fractionated tumor bed radiation(EF=1.9) The mechanism of interaction was increased sensitivity of radiated tumor bed to DC101.
4.Concurrent and adjuvant DC101 plus C225 with radiation promoted tumor control(enhancement factor 3.46 versus 2.84 of C225 with radiation),decreased the time to recurrence.Concurrent and adjuvant C225 with radiation improved tumor cure(P=0.005).Also,concurrent and adjuvant DC101 plus C225 with radiation improved tumor cure(P=0.004),but was not superior to C225 with radiation(P= 0.433).DC101 or C225 with radiation reduced the incidence of abdominal and pelvic lymph node metastasis(P=0.01,respectively).The lowest incidence of lymph node metastasis was achieved when both compounds were combined with radiation(P= 0.0001).
Conclusion:
DC101 enhances FaDu xenograft radioresponse.The combination of DC101 plus C225 with radiation improves tumor control and reduces abdominal and pelvic lymph node metastasis in FaDu xenografts.
探讨分子靶向治疗联合放射的最佳效应和多靶向联合放射的疗效,对VEGFR抗体联合放射以及VEGFR和EGFR抗体协同联合放射的作用进行了实验研究。
材料和方法:
1、建立FaDu头颈部鳞状细胞癌动物模型。头颈部鳞状细胞株FaDu来源于人咽鳞状细胞癌,从液氮中取出后复苏,配制成单细胞悬液,细胞浓度为1×10~8/ml。3—5月大小的瑞士nu—nu/Ncr雌性裸鼠被用于实验,1×10~6/10μlFaDu细胞种植于裸鼠右侧大腿肌肉内,FaDu肿瘤细胞形成移植瘤。
2、分别研究了VEGFR抗体DC101不同时相用药联合多分割放射的效应。移植瘤直径达到7mm(6.8-7.2mm)时,随机分组,包括对照组(无治疗),单纯药物组,单纯放射组,放射前用药联合放射组,放射同时用药联合放射组,放射后用药联合放射组,放射前+放射同时用药/联合放射组,放射同时+放射后用药/联合放射组,和放射前+放射同时+放射后用药/联合放射组。放射治疗为2.5Gy/次/天,连续6次。分别计算肿瘤8-12mm生长时间,比较其放射增敏效应。
3、瘤床放射后效应,用于评价放射与DC101的作用机制。将裸鼠随机分组,包括对照组(无治疗组),单纯药物组,单纯放射组,和药物+放射组。放射组对瘤床(裸鼠右侧大腿)先放射,放射剂量为2.5Gy/次/天,连续6次。24小时后各组分别种植肿瘤,48小时后开始药物治疗/无药物治疗。肿瘤直径达到7mm(6.8-7.2mm)时开始测量肿瘤大小,分别计算肿瘤8-12mm生长时间,比较其放射增敏效应。
4、DC101协同C225联合放射治愈肿瘤的疗效。移植瘤直径达到7mm(6.8-7.2mm)时,随机分组,包括对照组(单纯放射组),DC101联合放射组,C225联合放射组,和DC101协同C225联合放射组。放射治疗2.5Gy/次,2次/天,连续9天,24小时后各亚组加量照射10—40Gy。观察指标为肿瘤控制(肿瘤生长延迟和肿瘤复发),放射增益因子,肿瘤治愈和腹盆腔淋巴结转移率。
结果:
1、FaDu移植瘤成功率98%,分别选择直径6.8-7.2mm的肿瘤随机分组实验。
2、DC101放射同时和放射后用药放射增敏效益最大,增益因子分别是1.8和2.3。放射前用药无放射增敏作用;放射前加放射同时加放射后用药,产生最大的绝对生长延迟时间为18.5天,但是如果计算标准生长延迟时间,其结果与放射后用药相似,标准生长延迟时间分别为10.2天和10.5天,放射增益因子分别为2.2和2.3。
3、瘤床放射后比没有放射者明显延缓肿瘤生长,瘤床放射后DC101的增益因子为1.9,其机理是瘤床放射后增加对DC101的敏感性。
4、DC101协同C225联合放射与C225联合放射比较单纯放射能提高肿瘤的控制率,延长肿瘤复发的时间,增益因子分别是3.46和2.84;也能提高肿瘤的治愈率(P=0.004和0.005),但两者无统计学差异(P=0.433)。DC101联合放射与C225联合放射比较单纯放射能减少腹盆腔淋巴结转移率(P=0.01),DC101协同C225联合放射腹盆腔淋巴结转移率最低(P=0.0001)。
结论:
DC101能提高对头颈鳞癌FaDu的放射敏感性,DC101协同C225联合放射能提高肿瘤的局部控制率,降低腹盆腔淋巴结转移率。
Purpose:
To explore the efficacy of combination of molecular therapeutic and radiation including the optimal timing of radiation and drug and simultaneously targeting multiple signaling pathways.We investigated to optimize the combination of VEGFR-2 antibody with radiation and to integrate VEGFR-2 antibody,EGFR antibody with radiation in vivo.
Material and Methods:
1.FaDu xenografts of Head and Neck squamous cell cancer were set up.The human FaDu tumor cell line was originally derived from a human pharyngeal squamous cell carcer,which suspensions at 1×10~8/ml were prepared as monolayer in vitro.Female nude(nu-nu/Ncr) mice of 3-5 month-old were used.1×10~6/10μl tumor cells were inoculated intramuscularly into the mouse right hind legs for FaDu xenografts..
2.Different schedules of VEGFR-2 antibody DC101 and fractionated radiation were tested,such as neoadjuvnant,concurrent,adjuvant,neoadjuvnant+concurrent, concurrent+adjuvant,and neoadjuvnant+concurrent+adjuvant DC101.When the tumors had grown to 7 mm(6.8-7.2 mm) in diameter,the mice would be randomized into control(no treatmeat),different DC101 alone,radiation only,and different DC101 with radiation groups.The daily-fractionated dose of 2.5 Gy on 6 consecutive days(6×2.5Gy,15 Gy) was given.Time of tumor growth from 8-12mm was calculated respectively.Enhancement factors was compared.
3.A specifically designed tumor bed experiment of DC101 was performed to assess the mechanism of optimized interaction.The mice would be randomized into control(no treatmeat),DC101 only,radiation only,DC101 with radiation groups.The tumor bed(mouse right hind leg) was selectively radiated with 2.5 Gy in 6 fractions (6×2.5Gy,15 Gy).Tumor cells were injected intramuscularly at 24 hours after the last radiation dose,also for the unradiated mice at the same time.DC101 treatment started at 48 hours after tumor bed radiation.When tumors grew to 7mm(6.8-7.2 mm),the calculation was started.Time of tumor growth from 8-12mm was calculated respectively.Enhancement factors was compared.
4.An optimized schedule of DC101 was then integrated together with C225 and radiation in tumor cure assay.When tumors had grown to 7 mm(6.8-7.2 mm) in diameter,the mice would be randomized into control(radiation only),DC101 with radiation,C225 with radiation,or DC101 plus C225 with radiation groups.
The fractionated radiation was given twice daily at 2.5 Gy per fraction over 9 consecutive days(9×2×2.5Gy,45 Gy).One day after the last fractionated dose a range of single top-up doses up 10 to 40 Gy was given.Tumor control(tumor growth delay and tumor recurrence),radiation enhancement factor,tumor curability and incidence of abdominal and pelvic lymph node metastases were evaluated.
Results:
1.The survival rate of FaDu xenografts was 98%.The tumors between 6.8-7.2mm in diameter were used for randomization.
2.Concurrent and adjuvant DC101 with radiation were most effective in enhancing radiation-induced growth control(enhancement factors 1.8 and 2.3, respectively).Treatment with neoadjuvant DC101 did not result in radiation enhancement.When the extended DC101(neoadjuvant+ concurrent +adjuvant) was combined with radiation,this resulted in the longest absolute growth delay 18.5 days. However,if normalized against drug treatment,this induced similar radiation enhancement as adjuvant DC101(Normalized growth delays:radiation+extended DC101 schedule,10.2 days;radiation+adjuvant DC101,10.5 days;EF=2.2 for radiation+extended DC101 schedule;EF=2.3 for radiation+adjuvant DC101).
3.The tumor bed experiment resulted in significant enhancement of tumor growth delay in radiated tumor bed in comparison to unradiated tumor bed.Radiation enhancement was achieved for the fractionated tumor bed radiation(EF=1.9) The mechanism of interaction was increased sensitivity of radiated tumor bed to DC101.
4.Concurrent and adjuvant DC101 plus C225 with radiation promoted tumor control(enhancement factor 3.46 versus 2.84 of C225 with radiation),decreased the time to recurrence.Concurrent and adjuvant C225 with radiation improved tumor cure(P=0.005).Also,concurrent and adjuvant DC101 plus C225 with radiation improved tumor cure(P=0.004),but was not superior to C225 with radiation(P= 0.433).DC101 or C225 with radiation reduced the incidence of abdominal and pelvic lymph node metastasis(P=0.01,respectively).The lowest incidence of lymph node metastasis was achieved when both compounds were combined with radiation(P= 0.0001).
Conclusion:
DC101 enhances FaDu xenograft radioresponse.The combination of DC101 plus C225 with radiation improves tumor control and reduces abdominal and pelvic lymph node metastasis in FaDu xenografts.
引文
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46. Runkel S, Hunter NMilas L An intradermal assay for quantification and kinetics studies of tumor angiogenesis in mice. Radiat Res, 126: 237-243,1991.
47. Murata R, Nishimura YHiraoka M An antiangiogenic agent (TNP-470) inhibited reoxygenation during fractionated radiotherapy of murine mammary carcinoma. Int J Radiat Oncol Biol Phys, 37: 1107-1113,1997.
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49. Bruns CJ, Harbison MT, Davis DW, et al. Epidermal growth factor receptor blockade with C225 plus gemcitabine results in regression of human pancreatic carcinoma growing orthotopically in nude mice by antiangiogenic mechanisms. Clin Cancer Res 2000;6(5): 1936-48.
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51. Jung YD, Mansfield PF, Akagi M, et al. Effects of combination anti-vascular endothelial growth factor receptor and anti-epidermal growth factor receptor therapies on the growth of gastric cancer in a nude mouse model. Eur J Cancer 2002;38(8):1133-40.
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