scholarly journals Development and Characterization of a Novel Single-Cycle Recombinant-Virus Assay To Determine Human Immunodeficiency Virus Type 1 Coreceptor Tropism

2006 ◽  
Vol 51 (2) ◽  
pp. 566-575 ◽  
Author(s):  
Jeannette M. Whitcomb ◽  
Wei Huang ◽  
Signe Fransen ◽  
Kay Limoli ◽  
Jonathan Toma ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Load ◽  
Human Immunodeficiency Virus Type ◽  
Plasma Viral Load ◽  
Coreceptor Tropism ◽  
Immunodeficiency Virus ◽  
U87 Cells ◽  
Hiv 1

ABSTRACT Most human immunodeficiency virus type 1 (HIV-1) strains require either the CXCR4 or CCR5 chemokine receptor to efficiently enter cells. Blocking viral binding to these coreceptors is an attractive therapeutic target. Currently, several coreceptor antagonists are being evaluated in clinical trials that require characterization of coreceptor tropism for enrollment. In this report, we describe the development of an automated and accurate procedure for determining HIV-1 coreceptor tropism (Trofile) and its validation for routine laboratory testing. HIV-1 pseudoviruses are generated using full-length env genes derived from patient virus populations. Coreceptor tropism is determined by measuring the abilities of these pseudovirus populations to efficiently infect CD4+/U87 cells expressing either the CXCR4 or CCR5 coreceptor. Viruses exclusively and efficiently infecting CXCR4+/CD4+/U87 cells are designated X4-tropic. Conversely, viruses exclusively and efficiently infecting CCR5+/CD4+/U87 cells are designated R5-tropic. Viruses capable of infecting both CXCR4+/CD4+/U87 and CCR5+/CD4+/U87 cells are designated dual/mixed-tropic. Assay accuracy and reproducibility were established by evaluating the tropisms of well-characterized viruses and the variability among replicate results from samples tested repeatedly. The viral subtype, hepatitis B virus or hepatitis C virus coinfection, and the plasma viral load did not affect assay performance. Minority subpopulations with alternate tropisms were reliably detected when present at 5 to 10%. The plasma viral load above which samples can be amplified efficiently in the Trofile assay is 1,000 copies per ml of plasma. Trofile has been automated for high-throughput use; it can be used to identify patients most likely to benefit from treatment regimens that include a coreceptor inhibitor and to monitor patients on treatment for the emergence of resistant virus populations that switch coreceptor tropism.

scholarly journals Emergence of CXCR4-Using Human Immunodeficiency Virus Type 1 (HIV-1) Variants in a Minority of HIV-1-Infected Patients following Treatment with the CCR5 Antagonist Maraviroc Is from a Pretreatment CXCR4-Using Virus Reservoir

2006 ◽  
Vol 80 (10) ◽  
pp. 4909-4920 ◽  
Author(s):  
Mike Westby ◽  
Marilyn Lewis ◽  
Jeannette Whitcomb ◽  
Mike Youle ◽  
Anton L. Pozniak ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Phylogenetic Analysis ◽  
Viral Load ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Response To Treatment ◽  
Coreceptor Tropism ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Antagonists of the human immunodeficiency virus type 1 (HIV-1) coreceptor, CCR5, are being developed as the first anti-HIV agents acting on a host cell target. We monitored the coreceptor tropism of circulating virus, screened at baseline for coreceptor tropism, in 64 HIV-1-infected patients who received maraviroc (MVC, UK-427,857) as monotherapy for 10 days. Sixty-two patients harbored CCR5-tropic virus at baseline and had a posttreatment phenotype result. Circulating virus remained CCR5 tropic in 60/62 patients, 51 of whom experienced an HIV RNA reduction from baseline of >1 log10 copies/ml, indicating that CXCR4-using variants were not rapidly selected despite CCR5-specific drug pressure. In two patients, viral load declined during treatment and CXCR4-using virus was detected at day 11. No pretreatment factor predicted the emergence of CXCR4-tropic virus during maraviroc therapy in these two patients. Phylogenetic analysis of envelope (Env) clones from pre- and posttreatment time points indicated that the CXCR4-using variants probably emerged by outgrowth of a pretreatment CXCR4-using reservoir, rather than via coreceptor switch of a CCR5-tropic clone under selection pressure from maraviroc. Phylogenetic analysis was also performed on Env clones from a third patient harboring CXCR4-using virus prior to treatment. This patient was enrolled due to a sample labeling error. Although this patient experienced no overall reduction in viral load in response to treatment, the CCR5-tropic components of the circulating virus did appear to be suppressed while receiving maraviroc as monotherapy. Importantly, in all three patients, circulating virus reverted to predominantly CCR5 tropic following cessation of maraviroc.

scholarly journals Breadth of Neutralizing Antibody Response to Human Immunodeficiency Virus Type 1 Is Affected by Factors Early in Infection but Does Not Influence Disease Progression

2009 ◽  
Vol 83 (19) ◽  
pp. 10269-10274 ◽  
Author(s):  
Anne Piantadosi ◽  
Dana Panteleeff ◽  
Catherine A. Blish ◽  
Jared M. Baeten ◽  
Walter Jaoko ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Load ◽  
Disease Progression ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Neutralizing Antibody ◽  
Subtype B ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The determinants of a broad neutralizing antibody (NAb) response and its effect on human immunodeficiency virus type 1 (HIV-1) disease progression are not well defined, partly because most prior studies of a broad NAb response were cross-sectional. We examined correlates of NAb response breadth among 70 HIV-infected, antiretroviral-naïve Kenyan women from a longitudinal seroincident cohort. NAb response breadth was measured 5 years after infection against five subtype A viruses and one subtype B virus. Greater NAb response breadth was associated with a higher viral load set point and greater HIV-1 env diversity early in infection. However, greater NAb response breadth was not associated with a delayed time to a CD4+ T-cell count of <200, antiretroviral therapy, or death. Thus, a broad NAb response results from a high level of antigenic stimulation early in infection, which likely accounts for prior observations that greater NAb response breadth is associated with a higher viral load later in infection.

scholarly journals Screening for antiretroviral drug resistance among treatment-naive human immunodeficiency virus type 1-infected individuals in Lebanon

2014 ◽  
Vol 8 (03) ◽  
pp. 339-348
Author(s):  
Jacques M Mokhbat ◽  
Nada M. Melhem ◽  
Ziad El-Khatib ◽  
Pierre Zalloua
Keyword(s):  
Human Immunodeficiency Virus ◽  
Drug Resistance ◽  
Viral Load ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Newly Diagnosed ◽  
Reverse Transcriptase Inhibitors ◽  
Immunodeficiency Virus ◽  
Hiv 1

Introduction: Antiretroviral therapy (ART) has been successful at decreasing the morbidity and mortality associated with human immunodeficiency virus type 1 (HIV-1) infection. HIV-1 drug resistance (HIVDR) among ART-naive patients has been documented to compromise the success of initial therapy. This study was conducted to determine the prevalence of HIVDR mutations among newly diagnosed drug-naive HIV-infected individuals in Lebanon. Methodology: Plasma samples from 37 newly diagnosed participants at various stages of HIV-1 infection were used to determine HIV-1 RNA viral load, isolate viral RNA, and amplify DNA by RT-PCR. Purified PCR products were used to perform genotypic resistance tests. Results: The prevalence of resistance mutations to nucleoside reverse transcriptase inhibitors (NRTI), non-nucleoside reverse transcriptase inhibitors (NNRT), and protease inhibitors (PI) were 5.4%, 10.8%, and 8%, respectively. The major mutations detected in the study participants conferred resistance to NRTIs and NNRTIs recommended for HIV-1 treatment.  No significant relationship between HIV-1 viral load of participants and the mode of HIV-1 transmission or between the occurrence of HIVDR and the mode of transmission was found. Conclusions: To our knowledge, this is the first study on HIVDR mutations among newly diagnosed HIV-infected persons in Lebanon. The overall prevalence of HIVDR mutations detected in our study was 16%. Our results are important for evaluating the utility of the standard first-line regimens in use, determining the feasibility of HIVDR testing before the initiation of ART, as well as minimizing the emergence and transmission of HIVDR.

scholarly journals Identification and Characterization of a New Cross-Reactive Human Immunodeficiency Virus Type 1-Neutralizing Human Monoclonal Antibody

2004 ◽  
Vol 78 (17) ◽  
pp. 9233-9242 ◽  
Author(s):  
Mei-Yun Zhang ◽  
Xiaodong Xiao ◽  
Igor A. Sidorov ◽  
Vidita Choudhry ◽  
Fatim Cham ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Human Antibody ◽  
Immunodeficiency Virus ◽  
Clade C ◽  
Identification And Characterization ◽  
Hiv 1 ◽  
Soluble Cd4

ABSTRACT The identification and characterization of new human monoclonal antibodies (hMAbs) able to neutralize primary human immunodeficiency virus type 1 (HIV-1) isolates from different subtypes may help in our understanding of the mechanisms of virus entry and neutralization and in the development of entry inhibitors and vaccines. For enhanced selection of broadly cross-reactive antibodies, soluble HIV-1 envelope glycoproteins (Envs proteins) from two isolates complexed with two-domain soluble CD4 (sCD4) were alternated during panning of a phage-displayed human antibody library; these two Env proteins (89.6 and IIIB gp140s), and one additional Env (JR-FL gp120) alone and complexed with sCD4 were used for screening. An antibody with relatively long HCDR3 (17 residues), designated m14, was identified that bound to all antigens and neutralized heterologous HIV-1 isolates in multiple assay formats. Fab m14 potently neutralized selected well-characterized subtype B isolates, including JRCSF, 89.6, IIIB, and Yu2. Immunoglobulin G1 (IgG1) m14 was more potent than Fab m14 and neutralized 7 of 10 other clade B isolates; notably, although the potency was on average significantly lower than that of IgG1 b12, IgG1 m14 neutralized two of the isolates with significantly lower 50% inhibitory concentrations than did IgG1 b12. IgG1 m14 neutralized four of four selected clade C isolates with potency higher than that of IgG1 b12. It also neutralized 7 of 17 clade C isolates from southern Africa that were difficult to neutralize with other hMAbs and sCD4. IgG1 m14 neutralized four of seven primary HIV-1 isolates from other clades (A, D, E, and F) much more efficiently than did IgG1 b12; for the other three isolates, IgG b12 was much more potent. Fab m14 bound with high (nanomolar range) affinity to gp120 and gp140 from various isolates; its binding was reduced by soluble CD4 and antibodies recognizing the CD4 binding site (CD4bs) on gp120, and its footprint as defined by alanine-scanning mutagenesis overlaps that of b12. These results suggest that m14 is a novel CD4bs cross-reactive HIV-1-neutralizing antibody that exhibits a different inhibitory profile compared to the only known potent broadly neutralizing CD4bs human antibody, b12, and may have implications for our understanding of the mechanisms of immune evasion and for the development of inhibitors and vaccines.

scholarly journals Identification of Sequential Viral Escape Mutants Associated with Altered T-Cell Responses in a Human Immunodeficiency Virus Type 1-Infected Individual

2003 ◽  
Vol 77 (23) ◽  
pp. 12430-12440 ◽  
Author(s):  
Mark J. Geels ◽  
Marion Cornelissen ◽  
Hanneke Schuitemaker ◽  
Kiersten Anderson ◽  
David Kwa ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Load ◽  
T Cell ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Viral Escape ◽  
Ctl Epitopes ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Control of viremia in natural human immunodeficiency virus type 1 (HIV-1) infection in humans is associated with a virus-specific T-cell response. However, still much is unknown with regard to the extent of CD8+ cytotoxic T-lymphocyte (CTL) responses required to successfully control HIV-1 infection and to what extent CTL epitope escape can account for rises in viral load and ultimate progression to disease. In this study, we chose to monitor through full-length genome sequence of replication-competent biological clones the modifications that occurred within predicted CTL epitopes and to identify whether the alterations resulted in epitope escape from CTL recognition. From an extensive analysis of 59 biological HIV-1 clones generated over a period of 4 years from a single individual in whom the viral load was observed to rise, we identified the locations in the genome of five CD8+ CTL epitopes. Fixed mutations were identified within the p17, gp120, gp41, Nef, and reverse transcriptase genes. Using a gamma interferon ELIspot assay, we identified for four of the five epitopes with fixed mutations a complete loss of T-cell reactivity against the wild-type epitope and a partial loss of reactivity against the mutant epitope. These results demonstrate the sequential accumulation of CTL escape in a patient during disease progression, indicating that multiple combinations of T-cell epitopes are required to control viremia.

scholarly journals Identification and Characterization of a Peptide That Specifically Binds the Human, Broadly Neutralizing Anti-Human Immunodeficiency Virus Type 1 Antibody b12

2001 ◽  
Vol 75 (14) ◽  
pp. 6692-6699 ◽  
Author(s):  
Michael B. Zwick ◽  
Lori L. C. Bonnycastle ◽  
Alfredo Menendez ◽  
Melita B. Irving ◽  
Carlos F. Barbas ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Binding Site ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Human Monoclonal Antibody ◽  
Recombinant Polypeptide ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Human monoclonal antibody (MAb) b12 recognizes a conformational epitope that overlaps the CD-4-binding site of the human immunodeficiency virus type 1 (HIV-1) envelope. MAb b12 neutralizes a broad range of HIV-1 primary isolates and protects against primary virus challenge in animal models. We report here the discovery and characterization of B2.1, a peptide that binds specifically to MAb b12. B2.1 was selected from a phage-displayed peptide library by using immunoglobulin G1 b12 as the selecting agent. The peptide is a homodimer whose activity depends on an intact disulfide bridge joining its polypeptide chains. Competition studies with gp120 indicate that B2.1 occupies the b12 antigen-binding site. The affinity of b12 for B2.1 depends on the form in which the peptide is presented; b12 binds best to the homodimer as a recombinant polypeptide fused to the phage coat. Originally, b12 was isolated from a phage-displayed Fab library constructed from the bone marrow of an HIV-1-infected donor. The B2.1 peptide is highly specific for b12 since it selected only phage bearing b12 Fab from this large and diverse antibody library.

scholarly journals Intrapatient Diversity and Its Correlation with Viral Setpoint in Human Immunodeficiency Virus Type 1 CRF02_A/G-IbNG Infection

2002 ◽  
Vol 76 (21) ◽  
pp. 10745-10755 ◽  
Author(s):  
Indu Mani ◽  
Peter Gilbert ◽  
Jean-Louis Sankalé ◽  
Geoffrey Eisen ◽  
Souleymane Mboup ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Load ◽  
Human Immunodeficiency Virus Type ◽  
Disease Free Interval ◽  
Free Interval ◽  
Immunodeficiency Virus ◽  
Disease Free ◽  
Hiv 1 ◽  
Over Time

ABSTRACT The human immunodeficiency virus type 1 (HIV-1) viral setpoint during the disease-free interval has been strongly associated with future risk of disease progression. An awareness of the correlation between viral setpoint and HIV-1 genetic evolution over time is important in the understanding of viral dynamics and infection. We examined genetic diversity in HIV-1 CRF02_A/G-IbNG-infected seroincident women in Dakar, Senegal; determined whether a viral setpoint kinetic pattern existed for CRF02_A/G-IbNG during the disease-free interval; and correlated viral load level and diversity. Samples were drawn during the disease-free interval from consenting CRF02_A/G-IbNG-infected, antiretroviral therapy-naïve female commercial sex workers in Dakar, Senegal. Based on sequential plasma RNA values, low and high viral setpoint groups were established. Intrapatient diversity and divergence over time was determined from earlier and later time point DNA samples from each person. Most individuals followed the viral setpoint paradigm. For each 1|−|log10 copy/ml of plasma increase in viral load, intrapatient diversity increased by 1.4% (P = 0.028). A greater diversification rate was observed in the high viral setpoint group than in the low viral setpoint group (P = 0.01). Greater nucleotide (P = 0.015) and amino acid (P = 0.048) divergences and a greater nucleotide divergence rate (P = 0.03) were found in the high viral setpoint group. There was no difference between the groups in the ratio of the number of nonsynonymous substitutions per nonsynonymous site to the number of synonymous substitutions per synonymous site. The greater intrapatient diversity, divergence, and diversification rates observed in the high viral setpoint group supports the notion that diversity is driven by cycles of viral replication resulting in accumulated mutations. Recognizing diversity potential based on viral load levels in individuals may inform the design of vaccines and therapies.

Sign in / Sign up

Export Citation Format

Share Document