Clinical Evaluation of Rega 8: An Updated Genotypic Interpretation System That Significantly Predicts HIV-Therapy Response

Introduction

Clinically evaluating genotypic interpretation systems is essential to provide optimal guidance in designing potent individualized HIV-regimens. This study aimed at investigating the ability of the latest Rega algorithm to predict virological response on a short and longer period.

Materials & Methods

9231 treatment changes episodes were extracted from an integrated patient database. The virological response after 8, 24 and 48 weeks was dichotomized to success and failure. Success was defined as a viral load below 50 copies/ml or alternatively, a 2 log decrease from the baseline viral load at 8 weeks. The predictive ability of Rega version 8 was analysed in comparison with that of previous evaluated version Rega 5 and two other algorithms (ANRS v2011.05 and Stanford HIVdb v6.0.11). A logistic model based on the genotypic susceptibility score was used to predict virological response, and additionally, confounding factors were added to the model. Performance of the models was compared using the area under the ROC curve (AUC) and a Wilcoxon signed-rank test.

Results

Per unit increase of the GSS reported by Rega 8, the odds on having a successful therapy response on week 8 increased significantly by 81% (OR = 1.81, CI = [1.76–1.86]), on week 24 by 73% (OR = 1.73, CI = [1.69–1.78]) and on week 48 by 85% (OR = 1.85, CI = [1.80–1.91]). No significant differences in AUC were found between the performance of Rega 8 and Rega 5, ANRS v2011.05 and Stanford HIVdb v6.0.11, however Rega 8 had the highest sensitivity: 76.9%, 76.5% and 77.2% on 8, 24 and 48 weeks respectively. Inclusion of additional factors increased the performance significantly.

Conclusion

Rega 8 is a significant predictor for virological response with a better sensitivity than previously, and with rules for recently approved drugs. Additional variables should be taken into account to ensure an effective regimen.     

Combination of Immune and Viral Factors Distinguishes Low-Risk versus High-Risk HIV-1 Disease Progression in HLA-B*5701 Subjects

HLA-B*5701 is the host factor most strongly associated with slow HIV-1 disease progression, although rates can vary within this group. Underlying mechanisms are not fully understood but likely involve both immunological and virological dynamics. The present study investigated HIV-1 in vivo evolution and epitope-specific CD8(+) T cell responses in six HLA-B*5701 patients who had not received antiretroviral treatment, monitored from early infection for up to 7 years. The subjects were classified as high-risk progressors (HRPs) or low-risk progressors (LRPs) based on baseline CD4(+) T cell counts. Dynamics of HIV-1 Gag p24 evolution and multifunctional CD8(+) T cell responses were evaluated by high-resolution phylogenetic analysis and polychromatic flow cytometry, respectively. In all subjects, substitutions occurred more frequently in flanking regions than in HLA-B*5701-restricted epitopes. In LRPs, p24 sequence diversity was significantly lower; sequences exhibited a higher degree of homoplasy and more constrained mutational patterns than HRPs. The HIV-1 intrahost evolutionary rate was also lower in LRPs and followed a strict molecular clock, suggesting neutral genetic drift rather than positive selection. Additionally, polyfunctional CD8(+) T cell responses, particularly to TW10 and QW9 epitopes, were more robust in LRPs, who also showed significantly higher interleukin-2 (IL-2) production in early infection. Overall, the findings indicate that HLA-B*5701 patients with higher CD4 counts at baseline have a lower risk of HIV-1 disease progression because of the interplay between specific HLA-linked immune responses and the rate and mode of viral evolution. The study highlights the power of a multidisciplinary approach, integrating high-resolution evolutionary and immunological data, to understand mechanisms underlying HIV-1 pathogenesis.     

Evolutionary dynamics in an SI epidemic model with phenotype-structured susceptible compartment

When modeling infectious diseases, it is common to assume that infection-derived immunity is either (1) non-existent or (2) perfect and lifelong. However there are many diseases in which infection-derived immunity is known to be present but imperfect. There are various ways in which infection-derived immunity can fail, which can ultimately impact the probability that an individual be reinfected by the same pathogen, as well as the long-run population-level prevalence of the pathogen. Here we discuss seven different models of imperfect infection-derived immunity, including waning, leaky and all-or-nothing immunity. For each model we derive the probability that an infected individual becomes reinfected during their lifetime, given that the system is at endemic equilibrium. This can be thought of as the impact that each of these infection-derived immunity failures have on reinfection. This measure is useful because it provides us with a way to compare different modes of failure of infection-derived immunity.

     

AKTIP interacts with ESCRT I and is needed for the recruitment of ESCRT III subunits to the midbody

To complete mitosis, the bridge that links the two daughter cells needs to be cleaved. This step is carried out by the endosomal sorting complex required for transport (ESCRT) machinery. AKTIP, a protein discovered to be associated with telomeres and the nuclear membrane in interphase cells, shares sequence similarities with the ESCRT I component TSG101. Here we present evidence that during mitosis AKTIP is part of the ESCRT machinery at the midbody. AKTIP interacts with the ESCRT I subunit VPS28 and forms a circular supra-structure at the midbody, in close proximity with TSG101 and VPS28 and adjacent to the members of the ESCRT III module CHMP2A, CHMP4B and IST1. Mechanistically, the recruitment of AKTIP is dependent on MKLP1 and independent of CEP55. AKTIP and TSG101 are needed together for the recruitment of the ESCRT III subunit CHMP4B and in parallel for the recruitment of IST1. Alone, the reduction of AKTIP impinges on IST1 and causes multinucleation. Our data altogether reveal that AKTIP is a component of the ESCRT I module and functions in the recruitment of ESCRT III components required for abscission.     

Species distribution models as a tool to estimate reproductive parameters: a case study with a passerine bird species

1.?Correlative species distribution models (SDMs) assess relationships between species distribution data and environmental features, to evaluate the environmental suitability (ES) of a given area for a species, by providing a measure of the probability of presence. If the output of SDMs represents the relationships between habitat features and species performance well, SDM results can be related also to other key parameters of populations, including reproductive parameters. To test this hypothesis, we evaluated whether SDM results can be used as a proxy of reproductive parameters (breeding output, territory size) in red-backed shrikes (Lanius collurio). 2.?The distribution of 726 shrike territories in Northern Italy was obtained through multiple focused surveys; for a subset of pairs, we also measured territory area and number of fledged juveniles. We used Maximum Entropy modelling to build a SDM on the basis of territory distribution. We used generalized least squares and spatial generalized mixed models to relate territory size and number of fledged juveniles to SDM suitability, while controlling for spatial autocorrelation. 3.?Species distribution models predicted shrike distribution very well. Territory size was negatively related to suitability estimated through SDM, while the number of fledglings significantly increased with the suitability of the territory. This was true also when SDM was built using only spatially and temporally independent data. 4.?Results show a clear relationship between ES estimated through presence-only SDMs and two key parameters related to species' reproduction, suggesting that suitability estimated by SDM, and habitat quality determining reproduction parameters in our model system, are correlated. Our study shows the potential use of SDMs to infer important fitness parameters; this information can have great importance in management and conservation.     

Analysis of a clonal lineage of HIV-1 envelope V2/V3 conformational epitope-specific broadly neutralizing antibodies and their inferred unmutated common ancestors

V2/V3 conformational epitope antibodies that broadly neutralize HIV-1 (PG9 and PG16) have been recently described. Since an elicitation of previously known broadly neutralizing antibodies has proven elusive, the induction of antibodies with such specificity is an important goal for HIV-1 vaccine development. A critical question is which immunogens and vaccine formulations might be used to trigger and drive the development of memory B cell precursors with V2/V3 conformational epitope specificity. In this paper we identified a clonal lineage of four V2/V3 conformational epitope broadly neutralizing antibodies (CH01 to CH04) from an African HIV-1-infected broad neutralizer and inferred their common reverted unmutated ancestor (RUA) antibodies. While conformational epitope antibodies rarely bind recombinant Env monomers, a screen of 32 recombinant envelopes for binding to the CH01 to CH04 antibodies showed monoclonal antibody (MAb) binding to the E.A244 gp120 Env and to chronic Env AE.CM243; MAbs CH01 and CH02 also bound to transmitted/founder Env B.9021. CH01 to CH04 neutralized 38% to 49% of a panel of 91 HIV-1 tier 2 pseudoviruses, while the RUAs neutralized only 16% of HIV-1 isolates. Although the reverted unmutated ancestors showed restricted neutralizing activity, they retained the ability to bind to the E.A244 gp120 HIV-1 envelope with an affinity predicted to trigger B cell development. Thus, E.A244, B.9021, and AE.CM243 Envs are three potential immunogen candidates for studies aimed at defining strategies to induce V2/V3 conformational epitope-specific antibodies.     

Isolation of chitin synthetase from Saccharomyces cerevisiae. Purification of an enzyme by entrapment in the reaction product

Chitin synthetase, in the zymogen form, was extracted with digitonin from a particulate fraction from Saccharomyces cerevisiae and converted into active form by treatment with immobilized trypsin. When the activated enzyme was incubated with UDP-GlcNAc and other components of an assay mixture, a chitin precipitate formed, trapping a large portion of the synthetase. The enzyme was easily extracted frm the chitin gel with a recovery of approximately 50% and an enrichment of approximately 100-fold. Further purification was obtained by repeating the chitin step. After polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, the purified synthetase showed a major band corresponding to Mr 63,000, a weaker band at Mr 74,000, and some other minor bands. Under nondenaturing conditions, an Mr of 570,000 was calculated for the enzyme from Stokes radius and sedimentation coefficient determinations. After electrophoresis in a nondenaturing gel and incubation with the components of the standard assay, chitin was formed and precipitated in the gel, yielding an opaque band. Soluble oligosaccharides were not precursors for insoluble chitin, suggesting that synthesis of chitin chains takes place by a processive mechanism. N-Acetylglucosamine stimulated the purified synthetase only slightly and did not participate as a primer in the reaction. The same chain length, somewhat more than 100 units of GlcNAc, was determined in samples of chitin that had been synthesized either in vivo, or with a membrane preparation or with purified synthetase. These results suggest that chitin synthetase itself is capable both of initiating chitin chains without a primer and of determining their length.     

Alpine bird distributions along elevation gradients: the consistency of climate and habitat effects across geographic regions

Many species have shown recent shifts in their distributions in response to climate change. Patterns in species occurrence or abundance along altitudinal gradients often serve as the basis for detecting such changes and assessing future sensitivity. Quantifying the distribution of species along altitudinal gradients acts as a fundamental basis for future studies on environmental change impacts, but in order for models of altitudinal distribution to have wide applicability, it is necessary to know the extent to which altitudinal trends in occurrence are consistent across geographically separated areas. This was assessed by fitting models of bird species occurrence across altitudinal gradients in relation to habitat and climate variables in two geographically separated alpine regions, Piedmont and Trentino. The ten species studied showed non-random altitudinal distributions which in most cases were consistent across regions in terms of pattern. Trends in relation to altitude and differences between regions could be explained mostly by habitat or a combination of habitat and climate variables. Variation partitioning showed that most variation explained by the models was attributable to habitat, or habitat and climate together, rather than climate alone or geographic region. The shape and position of the altitudinal distribution curve is important as it can be related to vulnerability where the available space is limited, i.e. where mountains are not of sufficient altitude for expansion. This study therefore suggests that incorporating habitat and climate variables should be sufficient to construct models with high transferability for many alpine species.