Self-similarity of complex networks and hidden metric spaces

We demonstrate that the self-similarity of some scale-free networks with respect to a simple degree-thresholding renormalization scheme finds a natural interpretation in the assumption that network nodes exist in hidden metric spaces. Clustering, i.e., cycles of length three, plays a crucial role in this framework as a topological reflection of the triangle inequality in the hidden geometry. We prove that a class of hidden variable models with underlying metric spaces are able to accurately reproduce the self-similarity properties that we measured in the real networks. Our findings indicate that hidden geometries underlying these real networks are a plausible explanation for their observed topologies and, in particular, for their self-similarity with respect to the degree-based renormalization.

Optimal priming of poxvirus vector (NYVAC)-based HIV vaccine regimens requires 3 DNA injections. Results of the EV03/ANRS Vac20 Phase I/II Trial

Background: Two or three DNA primes have been used in previous smaller clinical trials, but the number required for optimal priming of viral vectors has never been assessed in adequately powered clinical trials. The EV03/ANRS Vac20 phase I/II trial investigated this issue using the DNA prime/poxvirus NYVAC boost combination, both expressing a common HIV-1 clade C immunogen consisting of Env and Gag-Pol-Nef polypeptide. Methods: 147 healthy volunteers were randomly allocated through 8 European centres to either 3xDNA plus 1xNYVAC (weeks 0, 4, 8 plus 24; n¼74) or to 2xDNA plus 2xNYVAC (weeks 0, 4 plus 20, 24; n¼73), stratified by geographical region and sex. T cell responses were quantified using the interferon g Elispot assay and 8 peptide pools; samples from weeks 0, 26 and 28 (time points for primary immunogenicity endpoint), 48 and 72 were considered for this analysis. Results: 140 of 147 participants were evaluable at weeks 26 and/ or 28. 64/70 (91%) in the 3xDNA arm compared to 56/70 (80%) in the 2xDNA arm developed a T cell response (P¼0.053). 26 (37%) participants of the 3xDNA arm developed a broader T cell response (Env plus at least to one of the Gag, Pol, Nef peptide pools) versus 15 (22%) in the 2xDNA arm (P¼0.047). At week 26, the overall magnitude of responses was also higher in the 3xDNA than in the 2xDNA arm (similar at week 28), with a median of 545 versus 328 SFUs/106 cells at week 26 (P<0.001). Preliminary overall evaluation showed that participants still developed T-cell response at weeks 48 (78%, n¼67) and 72 (70%, n¼66). Conclusion: This large clinical trial demonstrates that optimal priming of poxvirus-based vaccine regimens requires 3 DNA regimens and further confirms that the DNA/NYVAC prime boost vaccine combination is highly immunogenic and induced durable T-cell responses.

CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors.

Gene transfer-based therapeutic approaches have greatly benefited from the ability of some viral vectors to efficiently integrate within the cell genome and ensure persistent transmission of newly acquired transgenes to the target cell progeny. However, integration of provirus has been associated with epigenetic repercussions that may influence the expression of both the transgene and cellular genes close to vector integration loci. The exploitation of genetic insulator elements may overcome both issues through their ability to act as barriers that limit transgene silencing and/or as enhancer-blockers preventing the activation of endogenous genes by the vector enhancer. We established quantitative plasmid-based assay systems to screen enhancer-blocker and barrier genetic elements. Short synthetic insulators that bind to nuclear factor-I protein family transcription factors were identified to exert both enhancer-blocker and barrier functions, and were compared to binding sites for the insulator protein CTCF (CCCTC-binding factor). Gamma-retroviral vectors enclosing these insulator elements were produced at titers similar to their non-insulated counterparts and proved to be less genotoxic in an in vitro immortalization assay, yielding lower activation of Evi1 oncogene expression and reduced clonal expansion of bone marrow cells.

Poxvirus vector-based HIV vaccines.

PURPOSE OF REVIEW: In this review, we will provide the scientific rationale for the use of poxvirus vectors in the field of HIV vaccines, the immunological profile of the vaccine-induced immune responses, an update on the current use of poxvirus vector-based vaccines in HIV vaccine clinical trials, and the development of new modified poxvirus vectors with improved immunological profile. RECENT FINDINGS: An Ad5-HIV vaccine was tested in a phase IIb clinical trial (known as the Step trial). Vaccinations in the Step trial were discontinued because the vaccine did not show any effect on acquisition of infection and on viral load. After the disappointing failure of the Step trial, the field of HIV vaccine has regained enthusiasm and vigour due to the promising protective effect observed in the phase III efficacy trial (known as RV-144) performed in Thailand which has tested a poxvirus-gp120 combination. SUMMARY: The RV-144 phase III has provided for the first time evidence that an HIV vaccine can prevent HIV infection. The results from the RV-144 trial are providing the scientific rationale for the future development of the HIV vaccine field and for designing future efficacy trials.

Improved NYVAC-based vaccine vectors.

While as yet there is no vaccine against HIV/AIDS, the results of the phase III Thai trial (RV144) have been encouraging and suggest that further improvements of the prime/boost vaccine combination of a poxvirus and protein are needed. With this aim, in this investigation we have generated derivatives of the candidate vaccinia virus vaccine vector NYVAC with potentially improved functions. This has been achieved by the re-incorporation into the virus genome of two host range genes, K1L and C7L, in conjunction with the removal of the immunomodulatory viral molecule B19, an antagonist of type I interferon action. These novel virus vectors, referred to as NYVAC-C-KC and NYVAC-C-KC-ΔB19R, have acquired relevant biological characteristics, giving higher levels of antigen expression in infected cells, replication-competency in human keratinocytes and dermal fibroblasts, activation of selective host cell signal transduction pathways, and limited virus spread in tissues. Importantly, these replication-competent viruses have been demonstrated to maintain a highly attenuated phenotype.

Rpe65-Gene Transfer Using an Integration-Deficient Lentiviral Vector

Purpose: We previously demonstrated efficient retinal rescue of RPE65 mouse models (Rpe65-/- (Bemelmans et al, 2006) and Rpe65R91W/R91W mice) using a HIV1-derived lentiviral vector encoding for the mouse RPE65 cDNA. In order to optimize a lentiviral vector as an alternative tool for RPE65-derived Leber Congenital Amaurosis clinical trials, we evaluated the efficiency of an integration-deficient lentiviral vector (IDLV) encoding the human RPE65 cDNA to restore retinal function in the Rpe65R91W/R91W mice. Methods: An HIV-1-derived lentiviral vector expressing either the hrGFPII or the human Rpe65 cDNA under the control of a 0.8 kb fragment of the human Rpe65 promoter (R0.8) was produced by transient transfection of 293T cells. A LQ-integrase mutant was used to generate the IDLV vectors. IDLV-R0.8-hRPE65 or hrGFPII were injected subretinally into 1 month-old Rpe65R91W/R91W mice. Functional rescue was assessed by ERG (1 and 3 months post-injection) and cone survival by immunohistology. Results: An increased light sensitivity was detected by scotopic ERG in animals injected with IDLV-R0.8-hRPE65 compared to hrGFPII-treated animals or untreated mice. However the improvement was delayed compared to integration-proficient LV and observed at 3 months but not 1 month post-injection. Immunolabelling of cone markers showed an increased number of cones in the transduced area compared to control groups. Conclusions: The IDLV-R0.8-hRPE65 vectors allow retinal improvement in the Rpe65R91W/R91W mice. Both rod function and cone survival were demonstrated even if there is a delay in the rescue as assessed by scotopic ERG. Integration-deficient vectors minimize insertional mutagenesis and thus are safer candidates for human application. Further experiments using large animals are now needed to validate correct gene transfer and expression of the RPE65 gene as well as tolerance of the vector after subretinal injection before envisaging a clinical trial application.

Development of a Liver-specific Tet-On Inducible System for AAV Vectors and Its Application in the Treatment of Liver Cancer.

Recombinant adeno-associated virus (rAAV) are effective gene delivery vehicles that can mediate long-lasting transgene expression. However, tight regulation and tissue-specific transgene expression is required for certain therapeutic applications. For regulatable expression from the liver we designed a hepatospecific bidirectional and autoregulatory tetracycline (Tet)-On system (Tet(bidir)Alb) flanked by AAV inverted terminal repeats (ITRs). We characterized the inducible hepatospecific system in comparison with an inducible ubiquitous expression system (Tet(bidir)CMV) using luciferase (luc). Although the ubiquitous system led to luc expression throughout the mouse, luc expression derived from the hepatospecific system was restricted to the liver. Interestingly, the induction rate of the Tet(bidir)Alb was significantly higher than that of Tet(bidir)CMV, whereas leakage of Tet(bidir)Alb was significantly lower. To evaluate the therapeutic potential of this vector, an AAV-Tet(bidir)-Alb-expressing interleukin-12 (IL-12) was tested in a murine model for hepatic colorectal metastasis. The vector induced dose-dependent levels of IL-12 and interferon-γ (IFN-γ), showing no significant toxicity. AAV-Tet(bidir)-Alb-IL-12 was highly efficient in preventing establishment of metastasis in the liver and induced an efficient T-cell memory response to tumor cells. Thus, we have demonstrated persistent, and inducible in vivo expression of a gene from a liver-specific Tet-On inducible construct delivered via an AAV vector and proved to be an efficient tool for treating liver cancer.

The role of vectors in the epidemiology of avian malaria

Studies on host-parasite relationships have commonly reported that parasitized hosts undergo changes in their behavioural and life history traits. How do these changes affect the fitness of the hosts? What are the ecological and evolutionary drivers of these changes? These open questions are crucial to predict the parasite spread amongst hosts. Surprisingly, mosquito vectors of diseases to humans and animals have long been seen as passive parasite transporters, being unaffected by the infection though they also function as hosts. Natural parasite-vector interactions are therefore poorly documented in the literature. In this thesis, we seek to address the role of wild vectors in the epidemiology of avian Plasmodium, the etiological agents of malaria in birds. We first conducted avian malaria surveys in field-caught mosquitoes to identify the natural vectors in our temperate study area. We report that ornithophilic Culex pipiens primarily act as a vector for Plasmodium vaughani in spring, this parasite species being progressively replaced by P. relictum along with the season. Season-related factors may thus shape the mosquitoes' vectorial capacity. We then used experimental approaches to determine the effect of avian malaria on wild, naturally infected C. pipiens. We show that infected mosquitoes incur unavoidable physiological costs associated with parasite exploitation, these costs being expressed as a reduced survival under nutritionally stressed conditions only. These results are of significant importance for the epidemiology of avian malaria since seasonal changes in climate may likely influence food quality and quantity available to the mosquitoes. The host-selection preferences of the vectors with respect to the malaria-infection status of their bird hosts largely determine the disease spreading. In a second laboratory experiment, we thus offered wild C. pipiens the opportunity to choose between uninfected and naturally infected great tits, Parus major. We show that host-seeking mosquitoes have innate orientation preferences for uninfected birds. This suggests that avian malaria parasites exert strong selective pressures on their vectors, pushing them to evolve anti-parasite behaviours. We lastly investigated the links between malaria-associated symptoms in birds and resulting attractiveness to the mosquitoes. We show that experimentally malaria-infected canaries, Serinus canaria, suffer severe haematocrit reduction at peak parasitaemia and reduced basal metabolic rate later in the course of the infection. However, no links between infection and bird attractiveness to the mosquitoes were shown in an experiment using canaries as live bait for mosquito trap in the field. These links may have been masked by confounding environmental factors. Using a system where the vectors, parasites and hosts co-occur in sympatry, this thesis illustrates that vectors are not always Plasmodium permissive, which opposes to the traditional view that malaria parasites should have little effect on their vectors. The way that the vectors respond to the parasite threat is largely determined by the environmental conditions. This may have major implications for the epidemiology of avian malaria. - Les études portant sur les relations hôtes-parasites mentionnent souvent que les hôtes parasités subissent des modifications de leurs traits d'histoire de vie ou bien comportementaux. Comment ces changements affectent-ils la valeur sélective des hôtes et celle de leurs parasites ? Quels sont les déterminants de ces modifications ? Ces questions sont d'un grand intérêt en épidémiologie. Pour autant, les moustiques vecteurs de maladies infectieuses ont longtemps été perçus comme de simples transporteurs de parasites, n'étant pas affectés par ces derniers. Cette thèse porte sur le rôle des vecteurs dans l'épidémiologie des Plasmodium aviaires, agents étiologiques de la malaria chez les oiseaux. Dans le but d'identifier les vecteurs naturels de malaria aviaire dans notre zone d'étude, nous avons tout d'abord collecté des moustiques sur le terrain, puis déterminé leur statut infectieux. Nous rapportons que les moustiques Culex pipiens sont principalement impliqués dans la transmission de Plasmodium vaughani au printemps, cette espèce de parasite étant progressivement remplacée par P. relictum au fil de la saison de transmission. Nous avons ensuite conduit une expérience visant à déterminer l'effet de la malaria aviaire sur des C. pipiens sauvages, naturellement infectés. Nous montrons que des coûts sont associés à l'infection pour les moustiques. Ces coûts occasionnent une diminution de la survie des vecteurs seulement lorsque ceux-ci sont privés de ressources nutritionnelles. Des changements saisonniers de climats pourraient affecter la quantité et la qualité des ressources disponibles pour les vecteurs et donc, leur aptitude à transmettre l'infection. Les traits comportementaux des moustiques vecteurs, tels que la recherche et le choix d'un hôte pour se nourrir, sont d'une importance majeure pour la dispersion de la malaria. Pour cela, nous avons offert à des C. pipiens sauvages l'opportunité de choisir simultanément entre une mésange charbonnière (Parus major) saine et une autre naturellement infectée. Nous montrons que les moustiques s'orientent préférentiellement vers des mésanges saines. Les Plasmodium aviaires exerceraient donc de fortes pressions de sélection sur leurs vecteurs, favorisant ainsi l'évolution de comportements d'évitement des parasites. Enfin nous avons cherché à identifier de potentiels liens entre symptômes de l'infection malarique chez les oiseaux et attractivité de ces derniers pour les moustiques. Nous montrons que des canaris (Serinus canaria) expérimentalement infectés sont fortement anémiés au moment du pic infectieux et que leur métabolisme basai diminue plus tard au cours de l'infection. Toutefois, aucun lien entre le statut infectieux et l'attractivité des canaris pour les moustiques n'a pu être montré lors d'une expérience réalisée en nature. Il se peut que ces liens aient été masqués par des facteurs environnementaux confondants. Dans son ensemble, cette thèse illustre que, contrairement aux idées reçues, les vecteurs de malaria aviaire ne sont pas toujours permissifs avec leurs parasites. L'environnement apparaît aussi comme un facteur déterminant dans la réponse des vecteurs face à la menace d'infection malarique. Cela pourrait fortement affecter l'épidémiologie de la malaria aviaire.

Spinor Bose-Einstein Condensates and Topological Defects

This thesis concentrates on the topological defects of spin-1 and spin-2 Bose-Einstein condensates, the ground states of spin-3 condensates, and the inert states of spinor condensates with arbitrary spin. Our work is based on the description of a spinor condensate of spin-S atoms in terms of a state vector of a spin-S particle. The results of the homotopy theory are used to study the existence and structure of the topological defects in spinor condensates. We construct examples of defects, study their energetics, and examine how their stability is affected by the presence of an external magnetic field. The ground states of spin-3 condensates are calculated using analytical and numerical means. Special emphasis is put on the ground states of a chromium condensate, whose dependence on the magnetic dipole-dipole interaction is studied. A simple geometrical method for the calculation of inert states of spinor condensates is presented. This method is used to find candidates for the ground states of spin-S condensates.

Lentiviral vectors efficiently transduce the EGFP gene into cardiomyocytes in vivo : immunohistology and Elisa quantification of the transgene

RESUME Les maladies cardio-vasculaires représentent la cause la plus importante de mortalité et de morbidité dans les pays occidentaux. La thérapie génique offre une nouvelle approche au traitement de ces maladies. L'expression de gènes protecteurs dans le myocarde par des technologies de transfert génique peut améliorer la fonction ventriculaire lors de l'insuffisance cardiaque ou stimuler la formation de nouveaux vaisseaux dans la maladie coronarienne. Etant donné qu'une majorité des maladies cardiaques sont des maladies chroniques, l'expression durable du gène thérapeutique introduit dans le coeur est souhaitable dans de nombreux cas. Malheureusement, l'utilité des vecteurs de transfert génique les plus utilisés en thérapie génique cardiovasculaire est limitée par une performance faible (ADN plasmidique) et une courte durée d'expression (adénovirus). Récemment, des vecteurs de transfert génique dérivés des lentivirus, une sous-famille des rétrovirus, ont retenu l'attention de la communauté scientifique en raison de leur capacité à exprimer des gènes à long terme. Contrairement aux vecteurs rétroviraux traditionnels, les vecteurs lentiviraux transduisent des gènes même dans des cellules qui ne se divisent pas, ce qui est le cas des cardiomyocytes adultes. Ces vecteurs présentent un profil de biosécurité comparable à celui des vecteurs rétroviraux traditionnels. Nous avons donc décidé de tester l'utilité des vecteurs lentiviraux pour le transfert génique dans des cardiomyocytes de rat adulte in vitro et in vivo. Plusieurs versions de vecteurs lentiviraux contenant différent promoteurs ont été construites. Ces vecteurs contenant le gène marqueur EGFP (enhanced green fluorescent protein) ont été testés dans des cardiomyocytes de rat in vitro, ainsi que dans des coeurs de rat in vivo. Le but de ces expériences était de déterminer la durée de l'expression du transgène après injection intramyocardique chez le rat. Pour ce faire, nous avons développé une technique ELISA pour détecter la protéine EGFP dans des extraits de tissu cardiaque. Les résultats ont montré que la protéine EGFP était encore présente à des niveaux significatifs jusqu'à dix semaines après l'injection de vecteurs lentiviraux, alors que l'expression transgénique obtenue avec un vecteur adénoviral traditionnel a été plus limitée dans le temps. Ces résultats démontrent la capacité des vecteurs lentiviraux à exprimer des gènes d'intérêt de manière performante et stable dans le cœur de rat adulte in vivo. SUMMARY Cardiovascular diseases are the first cause of morbidity and mortality in Western countries. Gene therapy offers a new approach to these diseases. Expression of therapeutic genes in the myocardium by gene transfer technologies can improve ventricular function in heart failure and stimulate neovascularization in coronary disease. Chronic heart diseases likely require sustained expression of the therapeutic gene within the heart itself. Unfortunately, the most commonly used vectors in cardiovascular gene therapy, i.e. plasmid DNA and recombinant adenovirus vectors, are limited by poor DNA uptake and transient transgene expression, respectively. Recently, lentivirus-derived vectors have attracted much interest because of their ability to achieve long-term transgene expression. In contrast to traditional retroviral vectors, lentiviral vectors are also able to transduce non- dividing cells, while presenting a comparable biosafety profile. Adult cardiomyocytes are terminally differentiated cells that do not divide under normal conditions. For these reasons, we have decided to evaluate the efficiency of lentiviral vectors for gene-transduction of adult cardiomyocytes both in vitro and in vivo. We constructed various types of lentiviral vectors containing various promoters. Vectors encoding EGFP as a reporter gene were tested in rat cardiomyocytes in vitro and in rat hearts in vivo. The aim of the experiments involved in this thesis work was to determine the duration of the expression of the transgene after rat intramyocardial injection using a quantitative assay. Therefore, an ELISA technique was set up to measure the EGFP protein in rat heart tissue extracts. Our results showed that the EGFP protein was still present at significant levels at ten weeks after lentiviral vector injection, whereas the duration of expression with adenoviral vectors was shorter. These results demonstrate that lentiviral vectors efficiently deliver genes and achieve sustained transgene expression in adult rat cardiomyocytes in vivo.

A Topological Approach to Fuzzy Sets

This thesis presents a topological approach to studying fuzzy setsby means of modifier operators. Modifier operators are mathematical models, e.g., for hedges, and we present briefly different approaches to studying modifier operators. We are interested in compositional modifier operators, modifiers for short, and these modifiers depend on binary relations. We show that if a modifier depends on a reflexive and transitive binary relation on U, then there exists a unique topology on U such that this modifier is the closure operator in that topology. Also, if U is finite then there exists a lattice isomorphism between the class of all reflexive and transitive relations and the class of all topologies on U. We define topological similarity relation "≈" between L-fuzzy sets in an universe U, and show that the class LU/ ≈ is isomorphic with the class of all topologies on U, if U is finite and L is suitable. We consider finite bitopological spaces as approximation spaces, and we show that lower and upper approximations can be computed by means of α-level sets also in the case of equivalence relations. This means that approximations in the sense of Rough Set Theory can be computed by means of α-level sets. Finally, we present and application to data analysis: we study an approach to detecting dependencies of attributes in data base-like systems, called information systems.

Multigenic lentiviral vectors for combined and tissue-specific expression of miRNA- and protein-based antiangiogenic factors.

Lentivirus-based gene delivery vectors carrying multiple gene cassettes are powerful tools in gene transfer studies and gene therapy, allowing coexpression of multiple therapeutic factors and, if desired, fluorescent reporters. Current strategies to express transgenes and microRNA (miRNA) clusters from a single vector have certain limitations that affect transgene expression levels and/or vector titers. In this study, we describe a novel vector design that facilitates combined expression of therapeutic RNA- and protein-based antiangiogenic factors as well as a fluorescent reporter from back-to-back RNApolII-driven expression cassettes. This configuration allows effective production of intron-embedded miRNAs that are released upon transduction of target cells. Exploiting such multigenic lentiviral vectors, we demonstrate robust miRNA-directed downregulation of vascular endothelial growth factor (VEGF) expression, leading to reduced angiogenesis, and parallel impairment of angiogenic pathways by codelivering the gene encoding pigment epithelium-derived factor (PEDF). Notably, subretinal injections of lentiviral vectors reveal efficient retinal pigment epithelium-specific gene expression driven by the VMD2 promoter, verifying that multigenic lentiviral vectors can be produced with high titers sufficient for in vivo applications. Altogether, our results suggest the potential applicability of combined miRNA- and protein-encoding lentiviral vectors in antiangiogenic gene therapy, including new combination therapies for amelioration of age-related macular degeneration.

Better-quasi-order : ideals and spaces

This thesis deals with combinatorics, order theory and descriptive set theory. The first contribution is to the theory of well-quasi-orders (wqo) and better-quasi-orders (bqo). The main result is the proof of a conjecture made by Maurice Pouzet in 1978 his thèse d'état which states that any wqo whose ideal completion remainder is bqo is actually bqo. Our proof relies on new results with both a combinatorial and a topological flavour concerning maps from a front into a compact metric space. The second contribution is of a more applied nature and deals with topological spaces. We define a quasi-order on the subsets of every second countable To topological space in a way that generalises the Wadge quasi-order on the Baire space, while extending its nice properties to virtually all these topological spaces. The Wadge quasi-order of reducibility by continuous functions is wqo on Borei subsets of the Baire space, this quasi-order is however far less satisfactory for other important topological spaces such as the real line, as Hertling, Ikegami and Schlicht notably observed. Some authors have therefore studied reducibility with respect to some classes of discontinuous functions to remedy this situation. We propose instead to keep continuity but to weaken the notion of function to that of relation. Using the notion of admissible representation studied in Type-2 theory of effectivity, we define the quasi-order of reducibility by relatively continuous relations. We show that this quasi-order both refines the classical hierarchies of complexity and is wqo on the Borei subsets of virtually every second countable To space - including every (quasi-)Polish space. -- Cette thèse se situe dans les domaines de la combinatoire, de la théorie des ordres et de la théorie descriptive. La première contribution concerne la théorie des bons quasi-ordres (wqo) et des meilleurs quasi-ordres (bqo). Le résultat principal est la preuve d'une conjecture, énoncée par Pouzet en 1978 dans sa thèse d'état, qui établit que tout wqo dont l'ensemble des idéaux non principaux ordonnés par inclusion forme un bqo est alors lui-même un bqo. La preuve repose sur de nouveaux résultats, qui allient la combinatoire et la topologie, au sujet des fonctions d'un front vers un espace métrique compact. La seconde contribution de cette thèse traite de la complexité topologique dans le cadre des espaces To à base dénombrable. Dans le cas de l'espace de Baire, le quasi-ordre de Wadge est un wqo sur les sous-ensembles Boréliens qui a suscité énormément d'intérêt. Cependant cette relation de réduction par fonctions continues s'avère bien moins satisfaisante pour d'autres espaces d'importance tels que la droite réelle, comme l'ont fait notamment remarquer Hertling, Schlicht et Ikegami. Nous proposons de conserver la continuité et d'affaiblir la notion de fonction pour celle de relation. Pour ce faire, nous utilisons la notion de représentation admissible étudiée en « Type-2 theory of effectivity » initiée par Weihrauch. Nous introduisons alors le quasi-ordre de réduction par relations relativement continues et montrons que celui-ci à la fois raffine les hiérarchies classiques de complexité topologique et forme un wqo sur les sous-ensembles Boréliens de chaque espace quasi-Polonais.

Interleukin-1- and type I interferon-dependent enhanced immunogenicity of an NYVAC-HIV-1 Env-Gag-Pol-Nef vaccine vector with dual deletions of type I and type II interferon-binding proteins.

UNLABELLED: NYVAC, a highly attenuated, replication-restricted poxvirus, is a safe and immunogenic vaccine vector. Deletion of immune evasion genes from the poxvirus genome is an attractive strategy for improving the immunogenic properties of poxviruses. Using systems biology approaches, we describe herein the enhanced immunological profile of NYVAC vectors expressing the HIV-1 clade C env, gag, pol, and nef genes (NYVAC-C) with single or double deletions of genes encoding type I (ΔB19R) or type II (ΔB8R) interferon (IFN)-binding proteins. Transcriptomic analyses of human monocytes infected with NYVAC-C, NYVAC-C with the B19R deletion (NYVAC-C-ΔB19R), or NYVAC-C with B8R and B19R deletions (NYVAC-C-ΔB8RB19R) revealed a concerted upregulation of innate immune pathways (IFN-stimulated genes [ISGs]) of increasing magnitude with NYVAC-C-ΔB19R and NYVAC-C-ΔB8RB19R than with NYVAC-C. Deletion of B8R and B19R resulted in an enhanced activation of IRF3, IRF7, and STAT1 and the robust production of type I IFNs and of ISGs, whose expression was inhibited by anti-type I IFN antibodies. Interestingly, NYVAC-C-ΔB8RB19R induced the production of much higher levels of proinflammatory cytokines (tumor necrosis factor [TNF], interleukin-6 [IL-6], and IL-8) than NYVAC-C or NYVAC-C-ΔB19R as well as a strong inflammasome response (caspase-1 and IL-1β) in infected monocytes. Top network analyses showed that this broad response mediated by the deletion of B8R and B19R was organized around two upregulated gene expression nodes (TNF and IRF7). Consistent with these findings, monocytes infected with NYVAC-C-ΔB8RB19R induced a stronger type I IFN-dependent and IL-1-dependent allogeneic CD4(+) T cell response than monocytes infected with NYVAC-C or NYVAC-C-ΔB19R. Dual deletion of type I and type II IFN immune evasion genes in NYVAC markedly enhanced its immunogenic properties via its induction of the increased expression of type I IFNs and IL-1β and make it an attractive candidate HIV vaccine vector. IMPORTANCE: NYVAC is a replication-deficient poxvirus developed as a vaccine vector against HIV. NYVAC expresses several genes known to impair the host immune defenses by interfering with innate immune receptors, cytokines, or interferons. Given the crucial role played by interferons against viruses, we postulated that targeting the type I and type II decoy receptors used by poxvirus to subvert the host innate immune response would be an attractive approach to improve the immunogenicity of NYVAC vectors. Using systems biology approaches, we report that deletion of type I and type II IFN immune evasion genes in NYVAC poxvirus resulted in the robust expression of type I IFNs and interferon-stimulated genes (ISGs), a strong activation of the inflammasome, and upregulated expression of IL-1β and proinflammatory cytokines. Dual deletion of type I and type II IFN immune evasion genes in NYVAC poxvirus improves its immunogenic profile and makes it an attractive candidate HIV vaccine vector.

Head-to-Head Comparison of Poxvirus NYVAC and ALVAC Vectors Expressing Identical HIV-1 Clade C Immunogens in Prime-Boost Combination with Env Protein in Nonhuman Primates.

UNLABELLED: We compared the HIV-1-specific cellular and humoral immune responses elicited in rhesus macaques immunized with two poxvirus vectors (NYVAC and ALVAC) expressing the same HIV-1 antigens from clade C, Env gp140 as a trimeric cell-released protein and a Gag-Pol-Nef polyprotein as Gag-induced virus-like particles (VLPs) (referred to as NYVAC-C and ALVAC-C). The immunization protocol consisted of two doses of the corresponding poxvirus vector plus two doses of a combination of the poxvirus vector and a purified HIV-1 gp120 protein from clade C. This immunogenicity profile was also compared to that elicited by vaccine regimens consisting of two doses of the ALVAC vector expressing HIV-1 antigens from clades B/E (ALVAC-vCP1521) plus two doses of a combination of ALVAC-vCP1521 and HIV-1 gp120 protein from clades B/E (similar to the RV144 trial regimen) or clade C. The results showed that immunization of macaques with NYVAC-C stimulated at different times more potent HIV-1-specific CD4(+) T-cell responses and induced a trend toward higher-magnitude HIV-1-specific CD8(+) T-cell immune responses than did ALVAC-C. Furthermore, NYVAC-C induced a trend toward higher levels of binding IgG antibodies against clade C HIV-1 gp140, gp120, or murine leukemia virus (MuLV) gp70-scaffolded V1/V2 and toward best cross-clade-binding IgG responses against HIV-1 gp140 from clades A, B, and group M consensus, than did ALVAC-C. Of the linear binding IgG responses, most were directed against the V3 loop in all immunization groups. Additionally, NYVAC-C and ALVAC-C also induced similar levels of HIV-1-neutralizing antibodies and antibody-dependent cellular cytotoxicity (ADCC) responses. Interestingly, binding IgA antibody levels against HIV-1 gp120 or MuLV gp70-scaffolded V1/V2 were absent or very low in all immunization groups. Overall, these results provide a comprehensive survey of the immunogenicity of NYVAC versus ALVAC expressing HIV-1 antigens in nonhuman primates and indicate that NYVAC may represent an alternative candidate to ALVAC in the development of a future HIV-1 vaccine. IMPORTANCE: The finding of a safe and effective HIV/AIDS vaccine immunogen is one of the main research priorities. Here, we generated two poxvirus-based HIV vaccine candidates (NYVAC and ALVAC vectors) expressing the same clade C HIV-1 antigens in separate vectors, and we analyzed in nonhuman primates their immunogenicity profiles. The results showed that immunization with NYVAC-C induced a trend toward higher HIV-1-specific cellular and humoral immune responses than did ALVAC-C, indicating that this new NYVAC vector could be a novel optimized HIV/AIDS vaccine candidate for human clinical trials.