In vitro resistance selection with doravirine (MK-1439), a novel nonnucleoside reverse transcriptase inhibitor with distinct mutation development pathways

Abstract

Doravirine (DOR, formerly known as MK-1439) is a human immunodeficiency type 1 virus (HIV-1) nonnucleoside reverse transcriptase inhibitor (NNRTI) that is currently in phase 2b clinical trials. In vitro resistance selection of subtype B virus (MT4-green fluorescent protein [GFP] cells), as well as subtype A and C viruses (MT4-GFP/CCR5 cells) was conducted with DOR, rilpivirine (RPV), and efavirine (EFV) under low-multiplicity-of-infection conditions in a 96-well format. Resistance selection was performed with escalating concentrations of the NNRTIs ranging from the 95% effective concentration (1 × EC(95)) to 1,000 × EC(95) in the presence of 10% fetal bovine serum. In the resistance selection of subtype B virus with DOR, a V106A mutant virus led to two mutation pathways, followed by the emergence separately of either F227L or L234I. In the resistance selection of subtype A and C viruses, similar mutation development pathways were detected, in which a V106A or V106M mutant was also the starting virus in the pathways. Mutations that are commonly associated with RPV and EFV in clinical settings were also identified in subtype B viruses such as the E138K and K103N mutants, respectively, in this in vitro resistance selection study. The susceptibility of subtype B mutant viruses selected by DOR, RPV, and EFV to NNRTIs was evaluated. Results suggest that mutant viruses selected by DOR are susceptible to RPV and EFV and mutants selected by RPV and EFV are susceptible to DOR. When the replication capacity of the V106A mutant was compared with that of the wild-type (WT) virus, the mutant virus was 4-fold less fit than the WT virus.

FIG 1

The progress of viral breakthrough from low to high concentrations of compounds. Compound concentrations escalate from column 11 (lowest) to column 1 (highest). Panels: A, EFV; B, RPV; C, DOR. Gray suggests that there is no viral breakthrough. Black indicates that there may be a minor portion of breakthrough viruses but the green fluorescence is too weak to be detected. Dark green indicates a high level of viral breakthrough. Light green indicates a medium level of viral breakthrough or high level of viral breakthrough causing some extent of cell death.

FIG 2

Relative prevalence of WT and V106A mutant viruses in cultures from the replication capacity study. Solid diamonds represent the V106A mutant virus, and solid squares represent the WT virus. The experiment was conducted in triplicate, and the error bars represent standard deviations.

FIG 3

Susceptibilities of selected mutant viruses to NNRTIs. (A) Mutants selected by DOR. (B) Mutants selected by EFV. (C) Mutants selected by RPV. The x axis shows the sequential appearance of mutant viruses selected by the NNRTIs.

FIG 4

X-ray structure of the RT-DOR complex. Residues that are selected by DOR during resistance selection are shown. A red dashed line indicates the formation of a hydrogen bond between DOR and residue K103. Highlighted in yellow is V106, which is the major residue involved in the development of resistance to DOR.