Friedman H S, Dolan M E, Moschel R C, Pegg A E, Felker G M, Rich J, Bigner D D, Schold S C
Department of Pediatrics, Duke University Medical Center, Durham, N.C. 27710.
J Natl Cancer Inst. 1992 Dec 16;84(24):1926-31. doi: 10.1093/jnci/84.24.1926.
Although chemotherapy offers promise of increased survival for children with medulloblastoma and glioblastoma multiforme, drug resistance occurs frequently, resulting in tumor progression and death. Resistance to nitrosoureas and methylating agents, which damage DNA, can be mediated by a DNA repair protein, O6-alkylguanine-DNA alkyltransferase (AGAT). Depletion of this protein with alkylguanines or methylating agents, however, restores tumor cell sensitivity to the cytotoxicity of chloroethylnitrosoureas (e.g., carmustine [BCNU]).
This study was designed to determine whether resistance to the activity of nitrosourea (the drug BCNU) in BCNU-resistant human medulloblastoma (D341 Med) and human glioblastoma multiforme (D-456 MG) can be reversed by the methylating agent streptozocin and the O6-substituted guanines O6-methylguanine and O6-benzylguanine.
Xenografts were grown subcutaneously in athymic BALB/c mice. BCNU was administered as a single intraperitoneal injection at doses of 100 mg/m2, 75 mg/m2, or 38 mg/m2--i.e., 1.0, 0.75, or 0.38, respectively, of the dose lethal to 10% of treated animals (LD10). Mice were treated intraperitoneally with a single dose of O6-benzylguanine or O6-methylguanine (240 mg/m2) or with streptozocin (600 mg/m2) daily for 4 days. Response was assessed by tumor growth delay and tumor regression. AGAT activity in the xenografts was measured at 1 and 6 hours after pretreatment, at the time tumors were excised.
Pretreatment with O6-benzylguanine, O6-methylguanine, or streptozocin reduced AGAT activity to 4%, 25%, and 95% of control values, respectively, in D341 Med and 0%, 0%, and 25% of control values, respectively, in D-456 MG 1 hour after injection. After 6 hours, levels changed to 7%, 61%, and 116% of control values in D341 Med and 0%, 79%, and 21% of control values in D-456 MG, respectively. Both D341 Med and D-456 MG xenografts were completely resistant to BCNU at its LD10. Pretreatment with O6-benzylguanine increased BCNU sensitivity in both types of xenograft. In contrast, treatment with BCNU plus O6-methylguanine or streptozocin did not produce growth delays substantially different from those produced by BCNU alone, reflecting the more efficient depletion of AGAT by O6-benzylguanine. Following therapy with BCNU plus O6-benzylguanine at 0.38 LD10, tumor regressions were seen in eight of 10 D341 Med and in all 10 D-456 MG xenografts.
We recommend comprehensive clinical toxicologic evaluation of combination therapy with O6-benzylguanine plus BCNU, which would allow subsequent design of phase I clinical trials.
尽管化疗有望提高髓母细胞瘤和多形性胶质母细胞瘤患儿的生存率，但耐药现象频繁发生，导致肿瘤进展和死亡。对损伤DNA的亚硝基脲类和甲基化剂的耐药性可由一种DNA修复蛋白O6-烷基鸟嘌呤-DNA烷基转移酶（AGAT）介导。然而，用烷基鸟嘌呤或甲基化剂使该蛋白耗竭可恢复肿瘤细胞对氯乙基亚硝基脲（如卡莫司汀[BCNU]）细胞毒性的敏感性。
本研究旨在确定甲基化剂链脲佐菌素以及O6-取代鸟嘌呤O6-甲基鸟嘌呤和O6-苄基鸟嘌呤是否能逆转人髓母细胞瘤（D341 Med）和人多形性胶质母细胞瘤（D-456 MG）对亚硝基脲（药物BCNU）活性的耐药性。
将异种移植物皮下接种于无胸腺BALB/c小鼠。BCNU以100 mg/m2、75 mg/m2或38 mg/m2的剂量腹腔内单次注射，即分别为对10%受试动物致死剂量（LD10）的1.0、0.75或0.38。小鼠腹腔内每日单次注射剂量为240 mg/m2的O6-苄基鸟嘌呤或O6-甲基鸟嘌呤，或链脲佐菌素（600 mg/m2），共4天。通过肿瘤生长延迟和肿瘤消退评估反应。在预处理后1小时和6小时（即切除肿瘤时）测量异种移植物中的AGAT活性。
注射后1小时，在D341 Med中，用O6-苄基鸟嘌呤、O6-甲基鸟嘌呤或链脲佐菌素预处理分别使AGAT活性降至对照值的4%、25%和95%，在D-456 MG中分别降至对照值的0%、0%和25%。6小时后，在D341 Med中，水平分别变为对照值的7%、61%和116%，在D-456 MG中分别变为对照值的0%、79%和21%。D341 Med和D-456 MG异种移植物对LD10剂量的BCNU均完全耐药。用O６-苄基鸟嘌呤预处理可增加两种异种移植物对BCNU的敏感性。相比之下，BCNU加O6-甲基鸟嘌呤或链脲佐菌素治疗产生的生长延迟与单独使用BCNU产生的生长延迟没有显著差异，这反映了O6-苄基鸟嘌呤对AGAT的更有效耗竭。在以0.38 LD10的剂量用BCNU加O6-苄基鸟嘌呤治疗后，10个D341 Med异种移植物中有8个出现肿瘤消退，10个D-456 MG异种移植物全部出现肿瘤消退。
我们建议对O6-苄基鸟嘌呤加BCNU的联合治疗进行全面的临床毒理学评估，这将有助于随后开展I期临床试验的设计。
