Mechanism of resistance to S138A substituted enfuvirtide and its application to peptide design

• Published in: The international journal of biochemistry & cell biology 2013-04, Vol.45 (4), p.908-915
• Main Authors: Izumi, Kazuki, Kawaji, Kumi, Miyamoto, Fusasko, Shimane, Kazuki, Shimura, Kazuya, Sakagami, Yasuko, Hattori, Toshio, Watanabe, Kentaro, Oishi, Shinya, Fujii, Nobutaka, Matsuoka, Masao, Kaku, Mitsuo, Sarafianos, Stefan G., Kodama, Eiichi N.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino Acid Substitution; Anti-HIV Agents - chemistry; Anti-HIV Agents - pharmacology; Binding; binding capacity; binding sites; biochemistry; cell biology; Drug Design; Drug Resistance, Viral - drug effects; Drug Resistance, Viral - genetics; Fusion inhibitor; gp41; HEK293 Cells; HIV Envelope Protein gp41 - chemistry; HIV Envelope Protein gp41 - genetics; HIV Envelope Protein gp41 - pharmacology; HIV-1; HIV-1 - drug effects; HIV-1 - genetics; HIV-1 - physiology; Human immunodeficiency virus 1; Humans; Kinetics; Molecular Sequence Data; Mutation; Peptide Fragments - chemistry; Peptide Fragments - pharmacology; Resistance; T-20; virus replication; Virus Replication - drug effects; viruses
• ISSN: 1357-2725; 1878-5875
• DOI: 10.1016/j.biocel.2013.01.015

► T-20S138A, a resistance mutation introduced peptide, blocks T20-resistant HIV-1. ► To determine how HIV-1 escapes, we induced T-20S138A-resistant HIV-1 in vitro. ► We found that a unique mutation E137K in resistant HIV-1. ► E137K introduced T-20S138A suppressed replication of T-20S138A-resistant HIV-1. ► Mutations can be consistently useful for the design of peptide inhibitors. T-20 (enfuvirtide) resistance is caused by the N43D primary resistance mutation at its presumed binding site at the N-terminal heptad repeat (N-HR) of gp41, accompanied by the S138A secondary mutation at the C-terminal HR of gp41 (C-HR). We have discovered that modifying T-20 to include S138A (T-20S138A) allows it to efficiently block wild-type and T20-resistant viruses, by a mechanism that involves improved binding of T-20S138A to the N-HR that contains the N43D primary mutation. To determine how HIV-1 in turn escapes T-20S138A we used a dose escalation method to select T-20S138A-resistant HIV-1 starting with either wild-type (HIV-1WT) or T-20-resistant (HIV-1N43D/S138A) virus. We found that when starting with WT background, I37N and L44M emerged in the N-HR of gp41, and N126K in the C-HR. However, when starting with HIV-1N43D/S138A, L33S and I69L emerged in N-HR, and E137K in C-HR. T-20S138A-resistant recombinant HIV-1 showed cross-resistance to other T-20 derivatives, but not to C34 derivatives, suggesting that T-20S138A suppressed HIV-1 replication by a similar mechanism to T-20. Furthermore, E137K enhanced viral replication kinetics and restored binding affinity with N-HR containing N43D, indicating that it acts as a secondary, compensatory mutation. We therefore introduced E137K into T-20S138A (T-20E137K/S138A) and revealed that T-20E137K/S138A moderately suppressed replication of T-20S138A-resistant HIV-1. T-20E137K/S138A retained activity to HIV-1 without L33S, which seems to be a key mutation for T-20 derivatives. Our data demonstrate that secondary mutations can be consistently used for the design of peptide inhibitors that block replication of HIV resistant to fusion inhibitors.