Mechanism of resistance to S138A substituted enfuvirtide and its application to peptide design.
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.
Laboratory of Virus Control, Institute for Virus Research, Kyoto University, 53 Shogoin Kawaramachi, Kyoto 606-8507, Japan.
Kazuki
K
Izumi
Kumi
K
Kawaji
Fusasko
F
Miyamoto
Kazuki
K
Shimane
Kazuya
K
Shimura
Yasuko
Y
Sakagami
Toshio
T
Hattori
Kentaro
K
Watanabe
Shinya
S
Oishi
Nobutaka
N
Fujii
Masao
M
Matsuoka
Mitsuo
M
Kaku
Stefan G
SG
Sarafianos
Eiichi N
EN
Kodama
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D019380:Anti-HIV Agents / Q000737:chemistry / Q000494:pharmacology*; D015195:Drug Design; D024882:Drug Resistance, Viral / Q000187:drug effects / Q000235:genetics*; D000077560:Enfuvirtide; D057809:HEK293 Cells; D015700:HIV Envelope Protein gp41 / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D014779:Virus Replication / Q000187:drug effects
D019380:Anti-HIV Agents; D015700:HIV Envelope Protein gp41; D010446:Peptide Fragments; C514787:gp41 protein, Human immunodeficiency virus 1; C407149:peptide C34; D000077560:Enfuvirtide