Current V3 genotyping algorithms are inadequate for predicting X4 co-receptor usage in clinical isolates

Low, Andrew; Dong, Winnie; Chan, Dennison; Sing, Tobias; Swanstrom, Ronald; Jensen, Mark A.; Pillai, Satish K.; Good, Benjamin M.; Harrigan, P. Richard

doi:10.1097/qad.0b013e3282ef81ea

Cited by 126 publications

(107 citation statements)

References 25 publications

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“…However, many brainderived HIV-1 strains have been characterized by V3 sequence rather than traditional phenotypic assays (10,42,47,56,(59)(60)(61)64). Recent studies suggest V3 sequence-based algorithms may underestimate the frequency of CXCR4 usage by R5X4 HIV-1 strains (46,48), and in fact, the reported brain-derived HIV-1 strains that have the R5X4 phenotype in HIV-1 entry assays are scored as "R5-like" viruses using these algorithms (48). Therefore, it is likely that the frequency of R5X4 HIV-1 variants in the brain has been underestimated.…”

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confidence: 99%

Tissue-Specific Sequence Alterations in the Human Immunodeficiency Virus Type 1 Envelope Favoring CCR5 Usage Contribute to Persistence of Dual-Tropic Virus in the Brain

Gray

Roche

Churchill

et al. 2009

J Virol

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Most human immunodeficiency virus type 1 (HIV-1) strains isolated from the brain use CCR5 for entry into macrophages and microglia. Strains that use both CCR5 and CXCR4 for entry (R5X4 strains) have been identified in the brains of some individuals, but mechanisms underlying the persistence of R5X4 viruses compartmentalized between the brain and other tissue reservoirs are unknown. Here, we characterized changes in the HIV-1 envelope (Env) that enhance the tropism of R5X4 variants for brain or lymphoid tissue. R5X4 Envs derived from the brains of two individuals had enhanced CCR5 usage in fusion assays compared to R5X4 Envs derived from matched spleen or blood, which was associated with reduced dependence on specific residues in the CCR5 N terminus and extracellular loop 1 (ECL1) and ECL3 regions. In contrast, spleen/blood-derived Envs had enhanced CXCR4 usage compared to brain-derived Envs, which was associated with reduced dependence on residues in the CXCR4 N terminus and ECL2 region. Consequently, brain-derived Envs had preferential CCR5 usage for HIV-1 entry into the JC53 cell line, could use either CCR5 or CXCR4 for entry into monocyte-derived macrophages (MDM), and could use CCR5 (albeit inefficiently) for entry into peripheral blood mononuclear cells (PBMC), whereas the entry of spleen-derived Envs was CXCR4 dependent in all three cell types. Mutagenesis studies of Env amino acid variants influencing coreceptor usage showed that S306 in the gp120 V3 region of brain-derived Envs reduces dependence on the CCR5 N terminus and enhances CCR5 usage for HIV-1 entry into PBMC and MDM, whereas R306 in spleen-derived Envs reduces dependence on the CXCR4 N terminus and confers the CXCR4 restricted phenotype. These results identify mechanisms underlying R5X4 HIV-1 persistence in different tissue reservoirs. Tissue-specific changes in the gp120 V3 region that increase the efficiency of CCR5 or CXCR4 usage, and thereby influence coreceptor preference, may enhance the tropism of R5X4 strains for CCR5-expressing macrophage lineage cells in the brain and CXCR4-expressing T cells in lymphoid tissues, respectively.

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Tissue-Specific Sequence Alterations in the Human Immunodeficiency Virus Type 1 Envelope Favoring CCR5 Usage Contribute to Persistence of Dual-Tropic Virus in the Brain

Gray

Roche

Churchill

et al. 2009

J Virol

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“…The 11/25 charge rule is the simplest algorithm based on the presence of basic amino acids at positions 11 and/or 25 of the V3 loop (positions 1 and 35 are invariant cysteines that form a disulfide bond with each other, creating the intervening loop). However, the 11/25 rule lacks sensitivity in detecting X4 variants compared with phenotypic assays (9), with an Arg residue at position 25 being the least predictive. The position-specific scoring matrix (PSSM) analyzes the entire V3 sequence and predicts X4 variants based on a scoring of the probability of the amino acid at each position being overrepresented among X4 sequences versus R5 sequences (10).…”

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confidence: 99%

Deep Sequencing of the HIV-1 env Gene Reveals Discrete X4 Lineages and Linkage Disequilibrium between X4 and R5 Viruses in the V1/V2 and V3 Variable Regions

Zhou

Bednar

Sturdevant

et al. 2016

J Virol

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HIV-1 requires the CD4 receptor and a coreceptor (CCR5 [R5 phenotype] or CXCR4 [X4 phenotype]) to enter cells. Coreceptor tropism can be assessed by either phenotypic or genotypic analysis, the latter using bioinformatics algorithms to predict tropism based on the env V3 sequence. We used the Primer ID sequencing strategy with the MiSeq sequencing platform to reveal the structure of viral populations in the V1/V2 and C2/V3 regions of the HIV-1 env gene in 30 late-stage and 6 early-stage subjects. We also used endpoint dilution PCR followed by cloning of env genes to create pseudotyped virus to explore the link between genotypic predictions and phenotypic assessment of coreceptor usage. We found out that the most stringently sequence-based calls of X4 variants (Geno2Pheno false-positive rate [FPR] of <2%) formed distinct lineages within the viral population, and these were detected in 24 of 30 late-stage samples (80%), which was significantly higher than what has been seen previously by using other approaches. Non-X4 lineages were not skewed toward lower FPR scores in X4-containing populations. Phenotypic assays showed that variants with an intermediate FPR (2 to 20%) could be either X4/dual-tropic or R5 variants, although the X4 variants made up only about 25% of the lineages with an FPR of <10%, and these variants carried a distinctive sequence change. Phylogenetic analysis of both the V1/V2 and C2/V3 regions showed evidence of recombination within but very little recombination between the X4 and R5 lineages, suggesting that these populations are genetically isolated. IMPORTANCEPrimer ID sequencing provides a novel approach to study genetic structures of viral populations. X4 variants may be more prevalent than previously reported when assessed by using next-generation sequencing (NGS) and with a greater depth of sampling than single-genome amplification (SGA). Phylogenetic analysis to identify lineages of sequences with intermediate FPR values may provide additional information for accurately predicting X4 variants by using V3 sequences. Limited recombination occurs between X4 and R5 lineages, suggesting that X4 and R5 variants are genetically isolated and may be replicating in different cell types or that X4/R5 recombinants have reduced fitness. H uman immunodeficiency virus type 1 (HIV-1) requires the CD4 receptor and a coreceptor to infect host cells. Entry is mediated by the viral envelope protein (Env), which is processed to give two associated subunits, gp120 and gp41, that assemble into a trimeric structure embedded in the viral envelope/membrane. The binding of gp120 to CD4 triggers a structural change that exposes its variable region 3 (V3) loop, which is part of the coreceptor domain (1, 2). The majority of transmitted/founder (T/F) viruses and viruses isolated from the clinically latent stage in HIV-infected subjects use CCR5 as the coreceptor, making the virus CCR5 tropic (or an R5 virus). The virus can evolve to use CXCR4 (CXCR4 tropic [or an X4 virus]), and viruses that have switched core...

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“…However, these methods generally are developed by fitting a model onto the respective training set, and might not perform as well in independent or unseen datasets [13]. Moreover, as previously reported [14], some of these methods were trained on clonal sequences, and may not be adequate for tropism testing in clinical isolates that are often heterogeneous and have high levels of sequence ambiguity.…”

Section: Introductionmentioning

confidence: 99%

A Case-Based Reasoning System for Genotypic Prediction of Hiv-1 Co-Receptor Tropism

Evans¹,

Paquet²,

Huang³

et al. 2013

J. Bioinform. Comput. Biol.

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Accurate co-receptor tropism (CRT) determination is critical for making treatment decisions in HIV management. We created a genotypic tropism prediction tool by utilizing the case-based reasoning (CBR) technique that attempts to solve new problems through applying the solution from similar past problems. V3 loop sequences from 732 clinical samples with diverse characteristics were used to build a case library. Additional sequence and molecular properties of the V3 loop were examined and used for similarity assessment. A similarity metric was defined based on each attribute's frequency in the CXCR4-using viruses. We implemented three other genotype-based tropism predictors, support vector machines (SVM), position specific scoring matrices (PSSM), and the 11/25 rule, and evaluated their performance as the ability to predict CRT compared to Monogram's enhanced sensitivity Trofile® assay (ESTA). Overall concordance of the CBR based tropism prediction algorithm was 81%, as compared to ESTA. Sensitivity to detect CXCR4 usage was 90% and specificity was at 73%. In comparison, sensitivity of the SVM, PSSM, and the 11/25 rule were 85%, 81% and 36% respectively while achieving a specificity of 90% by SVM, 75% by PSSM, and 97% by the 11/25 rule. When we evaluated these predictors in an unseen dataset, higher sensitivity was achieved by the CBR algorithm (87%), compared to SVM (82%), PSSM (76%), and the 11/25 rule (33%), while maintaining similar level of specificity. Overall this study suggests that CBR can be utilized as a genotypic tropism prediction tool, and can achieve improved performance in independent datasets compared to model or rule based methods.

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Current V3 genotyping algorithms are inadequate for predicting X4 co-receptor usage in clinical isolates

Cited by 126 publications

References 25 publications

Tissue-Specific Sequence Alterations in the Human Immunodeficiency Virus Type 1 Envelope Favoring CCR5 Usage Contribute to Persistence of Dual-Tropic Virus in the Brain

Tissue-Specific Sequence Alterations in the Human Immunodeficiency Virus Type 1 Envelope Favoring CCR5 Usage Contribute to Persistence of Dual-Tropic Virus in the Brain

Deep Sequencing of the HIV-1 env Gene Reveals Discrete X4 Lineages and Linkage Disequilibrium between X4 and R5 Viruses in the V1/V2 and V3 Variable Regions

A Case-Based Reasoning System for Genotypic Prediction of Hiv-1 Co-Receptor Tropism

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