Su Yang, Chong Huihiui, Xiong Shengwen, Qiao Yuanyuan, Qiu Zonglin, He Yuxian
MOH Key Laboratory of Systems Biology of Pathogens and AIDS Research Center, Institute of Pathogen Biology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
MOH Key Laboratory of Systems Biology of Pathogens and AIDS Research Center, Institute of Pathogen Biology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China
J Virol. 2015 Dec;89(24):12467-79. doi: 10.1128/JVI.01741-15. Epub 2015 Oct 7.
The peptide drug enfuvirtide (T20) is the only HIV-1 fusion inhibitor in clinical use, but it easily induces drug resistance, calling for new strategies for developing effective drugs. On the basis of the M-T hook structure, we recently developed highly potent short-peptide HIV-1 fusion inhibitors (MTSC22 and HP23), which mainly target the conserved gp41 pocket and possess high genetic barriers to resistance. Here, we focused on the selection and characterization of HIV-1 escape mutants of MTSC22, which revealed new resistance pathways and mechanisms. Two mutations (E49K and L57R) located at the inhibitor-binding site and two mutations (N126K and E136G) located at the C-terminal heptad repeat region of gp41 were identified as conferring high resistance either singly or in combination. While E49K reduced the C-terminal binding of inhibitors via an electrostatic repulsion, L57R dramatically disrupted the N-terminal binding of M-T hook structure and pocket-binding domain. Unlike E49K and N126K, which enhanced the stability of the endogenous viral six-helical bundle core (6-HB), L57R and E136G conversely destabilized the 6-HB structure. We also demonstrated that both primary and secondary mutations caused the structural changes in 6-HB and severely impaired the capability for HIV-1 entry. Collectively, our data provide novel insights into the mechanisms of short-peptide fusion inhibitors targeting the gp41 pocket site and help increase our understanding of the structure and function of gp41 and HIV-1 evolution.
The deep pocket on the N-trimer of HIV-1 gp41 has been considered an ideal drug target because of its high degree of conservation and essential role in viral entry. Short-peptide fusion inhibitors, which contain an M-T hook structure and mainly target the pocket site, show extremely high binding and inhibitory activities as well as high genetic barriers to resistance. In this study, the HIV-1 mutants resistant to MTSC22 were selected and characterized, which revealed that the E49K and L57R substitutions at the inhibitor-binding site and the N126K and E136G substitutions at the C-terminal heptad repeat region of gp41 critically determine the resistance phenotype. The data provide novel insights into the mechanisms of action of the M-T hook structure-based fusion inhibitors which will help further our understanding of the structure-function relationship of gp41 and molecular pathways of HIV-1 evolution and eventually facilitate the development of new anti-HIV drugs.
肽类药物恩夫韦肽（T20）是临床上唯一使用的HIV-1融合抑制剂，但它很容易诱导耐药性，因此需要开发有效药物的新策略。基于M-T钩结构，我们最近开发了高效的短肽HIV-1融合抑制剂（MTSC22和HP23），它们主要靶向保守的gp41口袋，并且具有较高的耐药基因屏障。在这里，我们专注于MTSC22的HIV-1逃逸突变体的筛选和表征，这揭示了新的耐药途径和机制。位于抑制剂结合位点的两个突变（E49K和L57R）以及位于gp41 C末端七肽重复区域的两个突变（N126K和E136G）被确定为单独或联合时都赋予高耐药性。虽然E49K通过静电排斥降低了抑制剂的C末端结合，但L57R极大地破坏了M-T钩结构和口袋结合域的N末端结合。与增强内源性病毒六螺旋束核心（6-HB）稳定性的E49K和N126K不同，L57R和E136G反而使6-HB结构不稳定。我们还证明，初级和次级突变都会导致6-HB的结构变化，并严重损害HIV-1进入的能力。总体而言，我们的数据为靶向gp41口袋位点的短肽融合抑制剂的作用机制提供了新的见解，并有助于加深我们对gp41的结构和功能以及HIV-1进化的理解。
由于HIV-1 gp41 N三聚体上的深口袋具有高度保守性且在病毒进入中起关键作用，因此被认为是理想的药物靶点。含有M-T钩结构且主要靶向口袋位点的短肽融合抑制剂显示出极高的结合和抑制活性以及较高的耐药基因屏障。在本研究中，对MTSC22耐药的HIV-1突变体进行了筛选和表征，结果表明，位于抑制剂结合位点的E49K和L57R取代以及位于gp41 C末端七肽重复区域的N126K和E136G取代对耐药表型起关键决定作用。这些数据为基于M-T钩结构的融合抑制剂的作用机制提供了新的见解，这将有助于进一步加深我们对gp41的结构-功能关系以及HIV-1进化的分子途径的理解，并最终促进新型抗HIV药物的开发。
