Phenotypic co-receptor tropism and Maraviroc sensitivity in HIV-1 subtype C from East Africa

Abu Bakar Siddik,Md Shanawazur Rahman,Shambhu Ganeshappa Aralaguppe,Joelle Bader,Wondwossen Amogne,Thomas Klimkait,Ujjwal Neogi,Alexandra Haas

doi:10.1038/s41598-018-20814-2

Abstract

Genotypic tropism testing (GTT) for co-receptor usage is a recommended tool for clinical practice before administration of the CCR5-antagonist maraviroc. For some isolates, phenotypic tropism testing (PTT) revealed discordant results with GTT. In this study, we performed a comparative study between GTT and PTT in HIV-1C from East Africa (HIV-1CEA) and compared the data with HIV-1B and 01_AE and described the maraviroc susceptibility in the CCR5-tropic strains. Patient-derived HIV-1 envgp120 region was cloned into a modified pNL4-3 plasmid expressing the luciferase gene. rPhenotyping dissected single clones from 31 HIV-1CEA infected patients and four strains with known phenotype. Additionally, 68 clones from 18 patients (HIV-1B: 5, 01_AE: 7, HIV-1CEA: 6) were used to determine the PTT in GHOST cell line. The respective V3-sequences were used for GTT. R5-tropic strains from HIV-1CEA (n = 20) and non-C (n = 12) were tested for maraviroc sensitivity in TZMbl cell line. The GTT falsely called a higher proportion of X4-tropic strains in HIV-1CET compared to PTT by both rPhenotyping and the GHOST-cell assay. When multiple clones were tested in a subset of patients’ samples, both dual-tropic and R5-tropic strains were identified for HIV-1C. Relatively higher EC50 values were observed in HIV-1C strains than the non-C strains (p = 0.002).

Highlights

Predicted genotypic co-receptor tropism testing (GTT) is based on the analysis of human immunodeficiency virus type 1 (HIV-1) envelop V3-loop sequences and is a comparatively inexpensive, rapid and accessible alternative approach to phenotypic tropism testing (PTT) of the HIV-1 tropism in routine clinical practice[1]
Clonal PTT using the virus produced using the individual clones in GHOST cell lines were performed with 68 individual clones which were infectious from 180 clones tested, obtained from 18 patients samples infected with HIV-1B (n = 5), HIV-1C (n = 6) and 01_AE (n = 7) subtypes
The clonal Genotypic tropism testing (GTT) by sequencing individual clones falsely identified a higher proportion of X4-tropism in HIV-1C compared to phenotypic tropism testing by Clonal PTT (cPTT) (Fig. 1A)

Summary

Introduction

Predicted genotypic co-receptor tropism testing (GTT) is based on the analysis of human immunodeficiency virus type 1 (HIV-1) envelop (env) V3-loop sequences and is a comparatively inexpensive, rapid and accessible alternative approach to phenotypic tropism testing (PTT) of the HIV-1 tropism in routine clinical practice[1]. V3-loop sequences can be derived with clonal (i.e., virus sequences are cloned) or population-based methods (i.e., bulk population sequencing of the entire viral quasispecies), and predicted co-receptor tropism can be determined in silico using bioinformatics tools Numerous such algorithms are available to predict the genotypic tropism of HIV-1 based on the V3-loop sequence. Several recent studies from countries, where HIV-1 subtype HIV-1C is dominating, indicated an increase in predicted X4-tropic strains over time[8,9] Most of these genotypic and phenotypic tropism correlation studies were performed on HIV-1C sequences from Southern Africa or India. Given the higher heterogeneity among the East African strains[10], which can significantly affect the sequence-based tropism prediction[1,4,11], we hypothesized in this study that the current GTT tools for subtype C overestimate the X4-tropism in HIV-1CEA. We sought to study Maraviroc susceptibility among the phenotypically determined R5-tropic stains

Methods

Results

Conclusion