Hepatitis C viral evolution in genotype 1 treatment-naïve and treatment-experienced patients receiving telaprevir-based therapy in clinical trials

Background: In patients with genotype 1 chronic hepatitis C infection, telaprevir (TVR) in combination with peginterferon and ribavirin (PR) significantly increased sustained virologic response (SVR) rates compared with PR alone. However, genotypic changes could be observed in TVR-treated patients who did not achieve an SVR. Methods: Population sequence analysis of the NS3•4A region was performed in patients who did not achieve SVR with TVR-based treatment. Results: Resistant variants were observed after treatment with a telaprevir-based regimen in 12% of treatment-naïve patients (ADVANCE; T12PR arm), 6% of prior relapsers, 24% of prior partial responders, and 51% of prior null responder patients (REALIZE, T12PR48 arms). NS3 protease variants V36M, R155K, and V36M+R155K emerged frequently in patients with genotype 1a and V36A, T54A, and A156S/T in patients with genotype 1b. Lower-level resistance to telaprevir was conferred by V36A/M, T54A/S, R155K/T, and A156S variants; and higher-level resistance to telaprevir was conferred by A156T and V36M+R155K variants. Virologic failure during telaprevir treatment was more common in patients with genotype 1a and in prior PR nonresponder patients and was associated with higher-level telaprevir-resistant variants. Relapse was usually associated with wild-type or lower-level resistant variants. After treatment, viral populations were wild-type with a median time of 10 months for genotype 1a and 3 weeks for genotype 1b patients. Conclusions: A consistent, subtype-dependent resistance profile was observed in patients who did not achieve an SVR with telaprevir-based treatment. The primary role of TVR is to inhibit wild-type virus and variants with lower-levels of resistance to telaprevir. The complementary role of PR is to clear any remaining telaprevir-resistant variants, especially higher-level telaprevir-resistant variants. Resistant variants are detectable in most patients who fail to achieve SVR, but their levels decline over time after treatment

Inhibitie van cytokinenproduktie door een humane urinaire proteine

SIGLEKULeuven Campusbibliotheek Exacte Wetenschappen / UCL - Université Catholique de LouvainBEBelgiu

Binding Kinetics of Darunavir to Human Immunodeficiency Virus Type 1 Protease Explain the Potent Antiviral Activity and High Genetic Barrier▿

The high incidence of cross-resistance between human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs) limits their sequential use. This necessitates the development of PIs with a high genetic barrier and a broad spectrum of activity against PI-resistant HIV, such as tipranavir and darunavir (TMC114). We performed a surface plasmon resonance-based kinetic study to investigate the impact of PI resistance-associated mutations on the protease binding of five PIs used clinically: amprenavir, atazanavir, darunavir, lopinavir, and tipranavir. With wild-type protease, the binding affinity of darunavir was more than 100-fold higher than with the other PIs, due to a very slow dissociation rate. Consequently, the dissociative half-life of darunavir was much higher (>240 h) than that of the other PIs, including darunavir's structural analogue amprenavir. The influence of protease mutations on the binding kinetics was tested with five multidrug-resistant (MDR) proteases derived from clinical isolates harboring 10 to 14 PI resistance-associated mutations with a decreased susceptibility to various PIs. In general, all PIs bound to the MDR proteases with lower binding affinities, caused mainly by a faster dissociation rate. For amprenavir, atazanavir, lopinavir, and tipranavir, the decrease in affinity with MDR proteases resulted in reduced antiviral activity. For darunavir, however, a nearly 1,000-fold decrease in binding affinity did not translate into a weaker antiviral activity; a further decrease in affinity was required for the reduced antiviral effect. These observations provide a mechanistic explanation for darunavir's potent antiviral activity and high genetic barrier to the development of resistance

Crystal Structure of Lysine Sulfonamide Inhibitor Reveals the Displacement of the Conserved Flap Water Molecule in Human Immunodeficiency Virus Type 1 Protease▿

Human immunodeficiency virus type 1 (HIV-1) protease has been continuously evolving and developing resistance to all of the protease inhibitors. This requires the development of new inhibitors that bind to the protease in a novel fashion. Most of the inhibitors that are on the market are peptidomimetics, where a conserved water molecule mediates hydrogen bonding interactions between the inhibitors and the flaps of the protease. Recently a new class of inhibitors, lysine sulfonamides, was developed to combat the resistant variants of HIV protease. Here we report the crystal structure of a lysine sulfonamide. This inhibitor binds to the active site of HIV-1 protease in a novel manner, displacing the conserved water and making extensive hydrogen bonds with every region of the active site

Modeling viral evolutionary dynamics after telaprevir-based treatment.

For patients infected with hepatitis C virus (HCV), the combination of the direct-acting antiviral agent telaprevir, pegylated-interferon alfa (Peg-IFN), and ribavirin (RBV) significantly increases the chances of sustained virologic response (SVR) over treatment with Peg-IFN and RBV alone. If patients do not achieve SVR with telaprevir-based treatment, their viral population is often significantly enriched with telaprevir-resistant variants at the end of treatment. We sought to quantify the evolutionary dynamics of these post-treatment resistant variant populations. Previous estimates of these dynamics were limited by analyzing only population sequence data (20% sensitivity, qualitative resistance information) from 388 patients enrolled in Phase 3 clinical studies. Here we add clonal sequence analysis (5% sensitivity, quantitative) for a subset of these patients. We developed a computational model which integrates both the qualitative and quantitative sequence data, and which forms a framework for future analyses of drug resistance. The model was qualified by showing that deep-sequence data (1% sensitivity) from a subset of these patients are consistent with model predictions. When determining the median time for viral populations to revert to 20% resistance in these patients, the model predicts 8.3 (95% CI: 7.6, 8.4) months versus 10.7 (9.9, 12.8) months estimated using solely population sequence data for genotype 1a, and 1.0 (0.0, 1.4) months versus 0.9 (0.0, 2.7) months for genotype 1b. For each individual patient, the time to revert to 20% resistance predicted by the model was typically comparable to or faster than that estimated using solely population sequence data. Furthermore, the model predicts a median of 11.0 and 2.1 months after treatment failure for viral populations to revert to 99% wild-type in patients with HCV genotypes 1a or 1b, respectively. Our modeling approach provides a framework for projecting accurate, quantitative assessment of HCV resistance dynamics from a data set consisting of largely qualitative information

Kaplan-Meier curves for time-to-20% determined by population sequencing, model-predicted time-to-20%, and model-predicted time-to-1%.

<p>Results for patients with HCV subtypes 1a and 1b are shown in plots (A) and (B), respectively. Hash marks (∧) denote the censored observations indicating the time of the last visit for patients with virus that did not revert to <20% resistant. For clarity, these patients are explicitly denoted on the population sequence (“Pop. Seq.”) and model-predicted time-to-20% resistance curves only.</p

Comparison of the population sequence- and model-predicted time-to-20% for HCV subtypes 1a (A) and 1b (B).

<p>The X-axis represents the inferred time-to-loss of detectable resistance by population sequencing and reflects the first visit wherein the patient did not have detectable resistant variants. The Y-axis relies on the algorithms defined here, wherein the rate of loss is modeled continuously for each patient. The majority of the data points fall to the right of the unity line, indicating that the model predicts more rapid times-to-20% than those estimated from population sequence data.</p

Hypothetical viral dynamics for a patient with viral breakthrough during telaprevir-based treatment.

<p>(A) Dynamics for the total viral load (Total, green), wild-type virus (WT, blue), and a telaprevir-resistant variant (Resistant, red) during and after treatment with telaprevir-based treatment. LOD is the limit of detection for the ‘total’ viral load quantification, and Seq. LOD is the limit of detection above which sequencing can be reliably performed (1000 IU/ml). The treatment phase is shown by the gray bar. (B) Corresponding percent resistance dynamics on a linear scale. Viral sequencing can be performed when the total viral load exceeds the sequencing assay LOD (solid red curve). The dashed lines at 20% and 5% show the limits of detection for population and clonal sequence data, respectively.</p

Model fit for patients with genotype 1b HCV.

<p>(A): Histogram of the log₁₀ objective function values (φ; see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003772#pcbi.1003772.e021" target="_blank">Equation 5</a>) for all patients with genotype 1b. Dashed lines and numbers show quantile information for the fits. Also shown are representative fits for patients whose objective function values fall in the (B) 70%, (C) 90%, (D) 95%, and (E) 100% quantiles. Solid lines represent the model predictions, solid points represent the clonal sequence data, and error bars show the range for population sequence results.</p

Population sequence-based and model-predicted median reversion times.

1<p>The 95% CI assumes a single value for each patient and does not incorporate uncertainity of individual predictions (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003772#pcbi.1003772.s005" target="_blank">Text S1</a>).</p