Crankshaw Daune L, Berkeley Lorelle I, Cohen Jonathan F, Shirota Frances N, Nagasawa Herbert T
Medical Research Laboratories, DVA Medical Center, Minneapolis, MN 55417, USA.
J Biochem Mol Toxicol. 2002;16(5):235-44. doi: 10.1002/jbt.10044.
A series of double-prodrugs of L-cysteine, designed to release L-cysteine in vivo and stimulate the biosynthesis of glutathione (GSH), were synthesized. To evaluate the hepatoprotective effectiveness of these double-prodrugs, male Swiss-Webster mice were administered acetaminophen (ACP) (2.45 mmol/kg (360 mg/kg), intraperitoneally (i.p.)). Prodrug (2.50 mmol/kg, i.p. or 1.25 mmol/kg, i.p., depending on the protocol) was administered 1 h before ACP as a priming dose. A supplementary dose of prodrug (2.5 mmol/kg, i.p. or 1.25 mmol/kg, i.p. depending on the protocol) was administered 0.5 h after ACP. The plasma alanine amino transferase (ALT) values, 24 h after ACP administration were transformed to logs and the 95% and 99% confidence intervals of the log values were plotted and compared for each group. Hepatoprotection was assessed by the degree of attenuation of plasma ALT levels. With these multiple dose schedules, the use of 2% carboxymethylcellulose as vehicle for the prodrugs was found to be detrimental; therefore, the prodrugs were dissolved in dilute aqueous base and the pH adjusted for administration. When a priming dose was given 1 h before ACP followed by a supplementary dose 0.5 h after ACP, only N,S-bis-acetyl-L-cysteine, where both the sulfhydryl and amino groups of L-cysteine were functionalized with the acetyl group, was found to be effective in protecting mice against the hepatotoxic effects of ACP. This suggests that these acetyl groups were rapidly hydrolyzed in vivo to liberate L-cysteine. In contrast, N-acetylation of 2(R,S)-methylthiazolidine-4(R)-carboxylic acid (MTCA) and its 2-n-propyl analog (PTCA), or N-acetylation of 2-oxothiazolidine-4-carboxylic acid (OTCA), reduced the hepatoprotective effects relative to the parent MTCA, PTCA, and OTCA, indicating that the release of L-cysteine in vivo from these N-acetylated thiazolidine prodrugs was metabolically unfavorable. The carbethoxy group, whether functionalized on the sulfhydryl or on the amino group of L-cysteine, or on the secondary amino group of MTCA, appears to be a poor "pro-moiety," since these carbethoxylated double-prodrugs of L-cysteine did not protect mice from ACP-induced hepatotoxicity.
合成了一系列L-半胱氨酸双前药，旨在使其在体内释放L-半胱氨酸并刺激谷胱甘肽（GSH）的生物合成。为了评估这些双前药的肝保护效果，给雄性瑞士韦伯斯特小鼠腹腔注射对乙酰氨基酚（ACP）（2.45 mmol/kg（360 mg/kg））。根据实验方案，在注射ACP前1小时给予前药（2.50 mmol/kg，腹腔注射或1.25 mmol/kg，腹腔注射）作为起始剂量。在注射ACP后0.5小时给予补充剂量的前药（2.5 mmol/kg，腹腔注射或1.25 mmol/kg，腹腔注射，取决于实验方案）。将注射ACP后24小时的血浆丙氨酸氨基转移酶（ALT）值转化为对数，并绘制每组对数的95%和99%置信区间并进行比较。通过血浆ALT水平的衰减程度评估肝保护作用。采用这些多剂量方案时，发现使用2%羧甲基纤维素作为前药的载体是有害的；因此，将前药溶解在稀碱水溶液中并调节pH值用于给药。当在注射ACP前1小时给予起始剂量，随后在注射ACP后0.5小时给予补充剂量时，发现只有N,S-双乙酰-L-半胱氨酸（其中L-半胱氨酸的巯基和氨基均用乙酰基官能化）能有效保护小鼠免受ACP的肝毒性作用。这表明这些乙酰基在体内迅速水解以释放L-半胱氨酸。相比之下，2(R,S)-甲基噻唑烷-4(R)-羧酸（MTCA）及其2-正丙基类似物（PTCA）的N-乙酰化，或2-氧代噻唑烷-4-羧酸（OTCA）的N-乙酰化，相对于母体MTCA、PTCA和OTCA降低了肝保护作用，表明这些N-乙酰化噻唑烷前药在体内释放L-半胱氨酸在代谢上是不利的。乙氧羰基，无论其在L-半胱氨酸的巯基、氨基上官能化，还是在MTCA 的仲氨基上官能化，似乎都是一种较差的“前体部分”，因为这些L-半胱氨酸的乙氧羰基化双前药不能保护小鼠免受ACP诱导的肝毒性。
