Two distinct diazo precursors, imidazotetrazine and nitrous amide, were explored as promoieties in designing prodrugs of 6-diazo-5-oxo-l-norleucine (DON), a glutamine antagonist. As a model for an imidazotetrazine-based prodrug, we synthesized (S)-2-acetamido-6-(8-carbamoyl-4-oxoimidazo[5,1-d][1,2,3,5]tetrazin-3(4H)-yl)-5-oxohexanoic acid (4) containing the entire scaffold of temozolomide, a precursor of the DNA-methylating agent clinically approved for the treatment of glioblastoma multiforme. For a nitrous amide-based prodrug, we synthesized 2-acetamido-6-(((benzyloxy)carbonyl)(nitroso)amino)-5-oxohexanoic acid (5) containing a N-nitrosocarbamate group, which can be converted to a diazo moiety via a mechanism similar to that of streptozotocin, a clinically approved diazomethane-releasing drug containing an N-nitrosourea group. Preliminary characterization confirmed formation of N-acetyl DON (6), also known as duazomycin A, from compound 4 in a pH-dependent manner while compound 5 did not exhibit sufficient stability to allow further characterization. Taken together, our model studies suggest that further improvements are needed to translate this prodrug approach into glutamine antagonist-based therapy.

在设计谷氨酰胺拮抗剂 6-diazo-5-oxo-l-norleucine (DON) 的前药时，探索了两种不同的重氮前体，即咪唑四嗪和亚硝胺。作为基于咪唑四嗪的前药模型，我们合成了 ( S )-2-acetamido-6-(8-carbamoyl-4-oxoimidazo[5,1- d ][1,2,3,5]tetrazin-3( 4 H )-yl)-5-oxohexanoic acid ( 4 ) 包含替莫唑胺的整个支架，替莫唑胺是临床上批准用于治疗多形性胶质母细胞瘤的 DNA 甲基化剂的前体。对于基于亚硝酰胺的前药，我们合成了含有N-亚硝基氨基甲酸酯基团，可通过类似于链脲佐菌素的机制转化为重氮基团，链脲佐菌素是一种临床批准的含有N-亚硝基脲基团的重氮甲烷释放药物。初步表征证实了从化合物4以 pH 依赖性方式形成N-乙酰 DON ( 6)，也称为 duazomycin A，而化合物5没有表现出足够的稳定性以允许进一步表征。总之，我们的模型研究表明，需要进一步改进才能将这种前药方法转化为基于谷氨酰胺拮抗剂的治疗。