Development of Low Molecular Weight HIV Protease Inhibitor for Oral Use

人类免疫缺陷病毒（HIV）编码已知对逆转录病毒成熟和复制至关重要的天冬氨酸蛋白酶。HIV蛋白酶可以识别Phe-Pro和Tyr-Pro序列作为病毒特异性切割位点。基于底物过渡态，我们设计合成了一类含非天然氨基酸（2S，3S）-3-氨基-2-羟基-4-苯基丁酸的新型HIV蛋白酶抑制剂，命名为别苯基去甲他汀（Allophenylnorstatine，Apns）。其中，构象受限的三肽kynostatin（KNI）-272及其低分子量的二肽类似物是高度选择性和超强效的HIV蛋白酶抑制剂。例如，KNI-272对AZT敏感和不敏感的临床HIV-1分离株以及具有低细胞毒性的HIV-2表现出有效的抗病毒活性。i.d.给药后，KNI-272在大鼠中的生物利用度为42.3%。此外，KNI-272及其低分子量类似物在人PBMC-SCID小鼠体内表现出抗HIV活性。我们发现，基于“O-N分子内酰基迁移”的前药在水中具有良好的溶解性，在PBS（pH7.4）中可转化为活性形式。这些结果表明，前药可以通过静脉或口服给药，而无需特殊技术。

The human immunodeficiency virus (HIV) codes for an aspartic portease known to be essential for retroviral maturation and replication. The HIV protease can recognize Phe-Pro and Tyr-Pro sequences as the virus-specific cleavage site. These features provided a basis for the rational design of selective HIV protease-targeted drugs for the treatment of acquired immunodeficiency syndrome (AIDS).Based on the substrate transition state, we designed and synthesized a novel class of HIV protease inhibitors containing an unnatural amino acid, (2S,3S)-3-amino-2-hydroxy-4-phenylbutyric acid, named allophenylnorstatine (Apns) with a hydroxymethylcarbonyl (HMC) isostere. Among them, the conformationally constrained tripeptide kynostatin (KNI)-272 and its low molecular weight dipeptide analogs were highly selective and superpotent HIV protease inhibitors. For example, KNI-272 exhibited potent antiviral activities against both AZT-sensitive and-insensitive clinical HIV-1 isolates as well as HIV-2 with low cytotoxicity. After i.d.administration, bioavailability of KNI-272 was 42.3% in rats. Also, KNI-272 and its low molecular weight analogs exhibited in vivo anti-HIV activities in human PBMC-SCID mice.We then converted the above inhibitors into soluble prodrugs in order to improve the solubility in water. We found that the prodrugs based on "O-N intramolecular acyl migration" had good solubility in water and could be converted into active form in PBS (pH7.4). These results show that the prodrugs can be administrated by intravenously or oraly without special techniques.