Treatment-Mediated Alterations in HIV Fitness Preserve CD4+ T Cell Counts but Have Minimal Effects on Viral Load

For most HIV-infected patients, antiretroviral therapy controls viral replication. However, in some patients drug resistance can cause therapy to fail. Nonetheless, continued therapy with a failing regimen can preserve or even lead to increases in CD4+ T cell counts. To understand the biological basis of these observations, we used mathematical models to explain observations made in patients with drug-resistant HIV treated with enfuvirtide (ENF/T-20), an HIV-1 fusion inhibitor. Due to resistance emergence, ENF was removed from the drug regimen, drug-sensitive virus regrown, and ENF was re-administered. We used our model to study the dynamics of plasma-viral RNA and CD4+ T cell levels, and the competition between drug-sensitive and resistant viruses during therapy interruption and re-administration. Focusing on resistant viruses carrying the V38A mutation in gp41, we found ENF-resistant virus to be 17±3% less fit than ENF-sensitive virus in the absence of the drug, and that the loss of resistant virus during therapy interruption was primarily due to this fitness cost. Using viral dynamic parameters estimated from these patients, we show that although re-administration of ENF cannot suppress viral load, it can, in the presence of resistant virus, increase CD4+ T cell counts, which should yield clinical benefits. This study provides a framework to investigate HIV and T cell dynamics in patients who develop drug resistance to other antiretroviral agents and may help to develop more effective strategies for treatment

Immune Activation, Cd4+ T Cell Counts, and Viremia Exhibit Oscillatory Patterns over Time in Patients with Highly Resistant HIV Infection

The rates of immunologic and clinical progression are lower in patients with drug-resistant HIV compared to wild-type HIV. This difference is not fully explained by viral load. It has been argued that reductions in T cell activation and/or viral fitness might result in preserved target cells and an altered relationship between the level of viremia and the rate of CD4+ T cell loss. We tested this hypothesis over time in a cohort of patients with highly resistant HIV. Fifty-four antiretroviral-treated patients with multi-drug resistant HIV and detectable plasma HIV RNA were followed longitudinally. CD4+ T cell counts and HIV RNA levels were measured every 4 weeks and T cell activation (CD38/HLA-DR) was measured every 16 weeks. We found that the levels of CD4+ T cell activation over time were a strong independent predictor of CD4+ T cell counts while CD8+ T cell activation was more strongly associated with viremia. Using spectral analysis, we found strong evidence for oscillatory (or cyclic) behavior in CD4+ T cell counts, HIV RNA levels, and T cell activation. Each of the cell populations exhibited an oscillatory behavior with similar frequencies. Collectively, these data suggest that there may be a mechanistic link between T cell activation, CD4+ T cell counts, and viremia and lends support for the hypothesis of altered predator-prey dynamics as a possible explanation of the stability of CD4+ T cell counts in the presence of sustained multi-drug resistant viremia

Report of the NIH Panel to Define Principles of Therapy of HIV Infection

Report of the NIH Panel to Define Principles of Therapy of HIV Infection: Recent research advances have afforded substantially improved understanding of the biology of human immunodeficiency virus (HIV) infection and the pathogenesis of the acquired immunodeficiency syndrome (AIDS). With the advent of sensitive tools for monitoring HIV replication in infected persons, the risk of disease progression and death can be assessed accurately and the efficacy of anti-HIV therapies can be determined directly. Furthermore, when used appropriately, combinations of newly available, potent antiviral therapies can effect prolonged suppression of detectable levels of HIV replication and circumvent the inherent tendency of HIV to generate drug-resistant viral variants. However, as antiretroviral therapy for HIV infection has become increasingly effective, it has also become increasingly complex. Familiarity with recent research advances is needed to ensure that newly available therapies are used in ways that most effectively improve the health and prolong the lives of HIV-infected persons. To enable practitioners and HIV-infected persons to best use rapidly accumulating new information about HIV disease pathogenesis and treatment, the Office of AIDS Research of the National Institutes of Health sponsored the NIH Panel to Define Principles of Therapy of HIV Infection. This Panel was asked to define essential scientific principles that should be used to guide the most effective use of antiretroviral therapies and viral load testing in clinical practice. Based on detailed consideration of the most current data, the Panel delineated eleven principles that address issues of fundamental importance for the treatment of HIV infection. These principles provide the scientific basis for the specific treatment recommendations made by the Panel on Clinical Practices for the Treatment of HIV Infection sponsored by the Department of Health and Human Services and the Henry J. Kaiser Family Foundation. The reports of both of these panels are provided in this publication. Together, they summarize new dta and provide both the scientific basis and specific guidelines for the treatment of HIV-infected persons. This information will be of interest to health-care providers, HIV-infected persons, HIV/AIDS educators, public health educators, public health authorities, and all organizations that fund medical care of HIV-infected persons.Guidelines for the use of antiretroviral agents in HIV-infected adults and adolescents: With the development and FDA approval of an increasing number of antiretroviral agents, decisions regarding the treatment of HIV-infected persons have become complex; and the field continues to evolve rapidly. In 1996, the Department of Health and Human Services and the Henry J. Kaiser Family Foundation convened the Panel on Clinical Practices for the Treatment of HIV to develop guidelines for the clinical management of HIV-infected persons. This report includes the guidelines developed by the Panel regarding the use of laboratory testing in initiating and managing antiretroviral therapy, considerations for initiating therapy, whom to treat, what regimen of antiretroviral agents to use, when to change the antiretroviral regimen, treatment of the acutely HIV-infected person, special considerations in adolescents, and special considerations in pregnant women. Viral load and CD4+ T cell testing should ideally be performed twice before initiating or changing an antiretroviral treatment regimen. All patients who have advanced or symptomatic HIV disease should receive aggressive antiretroviral therapy. Initiation of therapy in the asymptomatic person is more complex and involves consideration of multiple virologic, immunologic, and psychosocial factors. In general, persons who have 500 CD4+ T cells per mm3 can be observed or can be offered therapy; again, risk of progression to AIDS, as determined by HIV RNA viremia and CD4+ T cell count, should guide the decision to treat. Once the decision to initiate antiretroviral therapy has been made, treatment should be aggressive with the goal of maximal viral suppression. In general, a protease inhibitor and two nucleoside [corrected] reverse transcriptase inhibitors should be used initially. Other regimens may be utilized but are considered less than optimal Many factors, including reappearance of previously undetectable HIV RNA, may indicate treatment failure. Decisions to change therapy and decisions regarding new regimens must be carefully considered; there are minimal clinical data to guide these decisions. Patients with acute HIV infection should probably be administered aggressive antiretroviral therapy; once initiated, duration of treatment is unknown and will likely need to continue for several years, if not for life. Special considerations apply to adolescents and pregnant women and are discussed in detail.Report of the NIH Panel to Define Principles of Therapy of HIV Infection / the material in this report was prepared for publication by: Mark B. Feinberg, Office of AIDS Research, National Institutes of Health, in collaboration with Jonathan E. Kaplan, Division of AIDS, STD, and TB Laboratory Research, National Center for Infectious Diseases and Division of HIV/AIDS Prevention\ue2\u20ac\u201cSurveillance, and Epidemiology, National Center for HIV, STD, and TB Prevention -- Guidelines for the use of antiretroviral agents in HIV-infected adults and adolescents / the material in this report was prepared for publication by Sharilyn K. Stanley, National Institute of Allergy and Infectious Diseases, National Institutes of Health, in collaboration with Jonathan E. Kaplan, Division of AIDS, STD, and TB Laboratory Research, National Center for Infectious Diseases and Division of HIV/AIDS Prevention\ue2\u20ac\u201cSurveillance, and Epidemiology, National Center for HIV, STD, and TB Prevention.April 24, 1998.Includes bibliographical references (p. 27-32 and p. 63-65)

Characterization of HIV-2 susceptibility to protease and entry inhibitors and identification of envelope determinants of coreceptor usage, cell tropism and antibody neutralization

Tese de doutoramento, Farmácia (Microbiologia), Universidade de Lisboa, Faculdade de Farmácia, 2018The main aim of this work was to characterize the susceptibility of HIV-2 to protease and entry inhibitors and to identify viral determinants of coreceptor usage, cellular tropism and antibody neutralization. The specific objectives were: 1) to determine the contribution of amino acids residues in the V3 loop involved in CCR5 and CXCR4 use, susceptibility to antibody neutralization and cellular tropism; 2) to develop a genotypic method for the prediction of HIV-2 coreceptor usage based on V3 loop; 3) to evaluate the antiviral activity of a new short-peptide fusion inhibitor in HIV-2 and 4) to characterize the evolution and diversity of protease (PR) in HIV-2 infected patients treated and untreated with protease inhibitors (PIs). In the first study (Chapter 3), site-directed mutagenesis was used to create amino acid substitutions in residues 18 and/or 29 and/or single deletions at positions 23 and 24 in V3 loop of pROD10, an infectious molecular clone of HIV-2ROD, the reference X4 isolate. Cellular assays demonstrated that: 1) conversion from X4 to R5 phenotype in HIV-2ROD10 requires H18L substitution and the deletion Δ(23,24); 2) H18L and H23Δ + Y24Δ mutants are more easily neutralized than HIV-2ROD and other mutated viruses by plasma from HIV-2 infected individuals; on the other hand, K29T substitution seems to contribute to increase resistance to neutralization; 3) K29T mutants acquire macrophage tropism without compromising replicative capacity in CD4+ T lymphocytes; 4) H18L + Δ(23,24) and (23,24) mutants gained the ability to replicate in macrophages albeit at the cost of some capacity to replicate in CD4+T cells. Structural analysis by homology modelling showed that: 1) H18L substitution disrupts the interaction of histidine with methionine at position 15 and with phenylalanine at position 20; 2) deletion of H23 and Y24 leads to the elimination of the parallel β sheets presented in the V3 loop and the loss of the aromatic system which can compromise the binding of cellular coreceptors or other molecules (e.g. antibodies); 3) K29T substitution reduces the charge of V3 and leads to the loss of the interactions with isoleucine at position 27. Collectively, these results demonstrated that V3 is an important determinant in HIV-2 coreceptor usage, susceptibility to antibody neutralization and replication capacity on CD4+ T cells and macrophages and that these phenotypic characteristics can be modulated by a single amino acid change in V3. These results support an important role for V3 in the pathogenesis of HIV-2 infection. In the second study (Chapter 4), a genotypic method was developed for the prediction of HIV-2 coreceptor usage from the V3 loop, similar to an existing tool created for HIV-1 (geno2pheno [coreceptor-hiv2]). The development and validation of this tool was based on a data set of 126 samples from HIV-2 infected patients, most of them from Portugal, with phenotypic coreceptor usage annotations. Predictive accuracy was also validated based on the V3 mutants produced and phenotypically characterized in the previous chapter. Overall, these findings indicated that geno2pheno [coreceptor-hiv2] can be a useful tool in clinical practice, allowing better management of HIV-2 infected patients eligible for maraviroc (MVC). In the third study (Chapter 5) a short-peptide named 2P23 was produced by combining a M-T hook structure, HIV-2 sequences and ‘salt-bridge’-based strategies. This peptide showed a potent antiviral activity against HIV-2 and HIV-1 isolates (mean 50% inhibitory concentration- IC50: 20.17 nM and 5.57 nM, respectively) and SIV (IC50: 1.8 nM for SIVpbj and 3.29 for SIV239). This new fusion inhibitor also demonstrated a strong activity against the V3 variants (Chapter 3) (IC50:15.38 nM), irrespectively of the coreceptor phenotype. Thus, 2P23 is an ideal candidate for further clinical development due to its broad antiviral activity against several HIV-2 isolates, with different coreceptor tropism. The last study (Chapter 6), involved the characterization of PR diversity and genotypic resistance to PIs of HIV-2 infected individuals living in Portugal and the evaluation of the impact of resistance mutations to PIs in treatment outcome eight years post-therapy. A high prevalence of PR mutations (e.g. I54M, I82F, L90M) associated to saquinavir (SQV), darunavir (DRV) and lopinavir (LPV) resistance, were detected in proviral DNA from these patients at baseline. Eight years after study entry, the genotypic analysis identified: 1) loss of resistance mutations in two patients, that were initially detected at baseline, presumably as a consequence of treatment interruption; 2) long term persistence of resistance mutations in one individual as a result of virologic and immunologic failure, which might raise concern about transmission of drug resistance in the future and 3) development of new resistance mutations in three patients due to previous treatment failures. The analysis of genetic diversity in PR showed an increase in this parameter in two treated patients, with undetectable viral loads and higher CD4+ T counts, comparing with the baseline. On the other hand, a reduction in PR genetic diversity was exhibited in three patients (two treated and one untreated), who presented detectable viral loads in at least one time point during the follow up. Due to small sample size it was not possible to investigate a potential relationship between PR genetic diversity and CD4+ T cell counts, presence of resistance mutations or/and treatment status. However, these results seem to indicate a persistent viral replication during long term highly active antiretroviral therapy (HAART), regardless of plasma viral load. The maintenance of viral replication can act as a source of new proviral quasispecies, resulting in the gradual substitution of the ancestral variants over time. Most importantly, we found two potential cases of transmitted drug resistance. However, due to the small sample size, additional studies with a higher number of patients are required to determine if primary drug resistance is a major problem in HIV-2 infected patients in Portugal. Our findings suggest that proviral DNA may be useful in resistance testing in HIV-2 patients with low or suppressed viremia and in untreated patients, and that early resistance analysis of these archived viruses may predict treatment response

Disease progression in Human Immunodeficiency Virus type 1 infected viraemic controllers

PhDBackground: The mechanism of CD4+ T-cell decline in Human Immunodeficiency Virus-1 (HIV-1) infection is unclear, but the association with plasma HIV-1 RNA-load suggests viral replication is involved. Viraemic controller patients with low HIV-1 RNA-loads (<2000 copies/ml) typically maintain good CD4+ T-cell counts (>450 CD4+ T-cells/mm3). However, within a cohort of 86 viraemic controllers, a subgroup (18 ‘discord controllers’) was identified with low CD4+ T-cell counts (<450 CD4+ T-cells/mm3) which present clinical uncertainty. The underlying mechanism accounting for CD4+ T-cell decline in the face of low or undetectable HIV-1 RNAloads is unknown. The objective of the work described in this thesis was to investigate the virological and host immune system dynamics in discord controllers compared with typical controllers. Method Epidemiological features, HIV-1 subtype, cellular HIV-1 DNA-load, T-cell populations (CD4+/CD8+ naïve/ central-memory/ effector-memory subsets; CD45RA/RO ± CD62L) and Tcell activation markers (CD38, HLA-DR) were examined for discord controllers and typical controllers as well as progressors with HIV-1 RNA-load >10000 copies/ml, <450 CD4+ Tcells/ mm3. Results Discord controllers and typical controllers were similar, based on epidemiological features and viral subtype distribution. They resembled progressors, showing high HIV-1 DNA-load, depletion of naïve CD4+ T-cells and higher activation in all CD4+ T-cell subsets. However, the CD8+ T-cell compartment in discord controllers was similar to typical controllers with preserved naïve CD8+ T-cells and low level CD8+ T-cells activation. Conclusion The data presented in this thesis is consistent with a relationship between CD4+ T-cell activation, HIV-1 DNA-load and disease progression but not HIV-1 RNA-load. This suggests that in viraemic controllers, HIV-1 DNA-load may be a better marker of viral replication and disease progression than HIV-1 RNA-load. Furthermore, low level CD8+ T-cell activation correlate with low plasma HIV-1 RNA-load but not with HIV-1 DNA-load.Barts and the London Charity Grant(MMBG1E7R); BHIVA SpR Research Grant (MMBG1F2R); MRC Senior Non-Clinical Fellowship awarded to supervisor A.M. (G117/547); Wellcome Trust grant awarded to supervisor A.M. (WT075853MA)

Anti-apoptotic effects of Interleukin 7 in Human Immunodeficiency Virus Type 1 infection: Mechanism and implications for immune reconstitution strategies

Although HAART has introduced great improvements in the clinical outcome of HIV-1- infected individuals, the current protocols often fail in completely restoring the CD4+T-cell lymphopenia. Therefore, innovative approaches based on the use of immune adjuvants to HAART, including cytokines like IL-2, IL-15 and IL-7, are being explored. The present thesis aims at investigating the immunomodulatory effects of IL- 7 in HIV /SIV infection, focusing on its pro-survival properties. The first part of this work describes the results of a study in vitro to evaluate the effects of IL-7 on T cells from HIV -l-infected individuals, and shows that this cytokine strongly protects both CD4+ and CD8+T cells from spontaneous apoptosis. IL-7-mediated apoptosis reduction ex vivo inversely correlated with the CD4+T-cell count of the patient in vivo, suggesting that IL-7 treatment could be useful also for patients at with advanced disease. Moreover, the protective effect of IL-7 was not associated with the induction of cellular proliferation or viral replication. The second part of this work describes the results of a study in vivo of IL-7 administration during the acute phase of SIV infection in rhesus macaques, the pathogenic animal model for AIDS. IL-7 treatment prevented the depletion of circulating naive and memory CD4+T cells that typically occurs within the very first weeks of infection, primarily by reducing the levels of apoptosis. Moreover, IL-7 treatment also induced sustained increases in all subsets of circulating CD8+T cells. Of note, treatment with IL-7 did not have any effect on SIV plasma viral load nor on the content of SIV DNA provirus. Finally, IL-7-treated animals developed earlier and stronger CD8+ and CD4+T-cell responses as compared to untreated animals, although overall no protective effects on disease progression were observed. Taken together, these data further support the use of IL-7 as an immunomodulatory adjuvant to HAART in HIV-I-infected individuals, and suggest that this cytokine may be a useful tool to preserve or restore the CD4+T-cell pool during both the acute and chronic phases of the infection

EVALUATION OF DUAL-SEROTYPE ADENOVIRUS-BASED VACCINE-INDUCED CELLULAR IMMUNITY FOLLOWING PREVENTATIVE AND THERAPEUTIC IMMUNIZATION AGAINST SIMIAN IMMUNODEFICIENCY VIRUS

A vaccine capable of preventing or therapeutically limiting human immunodeficiency virus (HIV) pathogenesis is urgently needed to contain the acquired immunodeficiency syndrome (AIDS) pandemic. Recombinant adenovirus (Ad)-based vectors are being explored as vaccine candidates due to their potent induction of cell-mediated immunity. To circumvent the limitations of vector-specific humoral immunity, novel Ad serotypes impervious to pre-existing immunity against conventional vectors have been developed. Utilizing the nonhuman primate model of HIV infection, we evaluated the immunogenicity of conventional Ad serotype 5- (Ad5) and novel serotype 35- (Ad35) based vaccinations against simian immunodeficiency virus (SIV) infection. In a preventative, proof-of-concept vaccination regimen, immunization against the SIV Gag protein proved highly efficacious, demonstrating robust boosting of Ad5-based vaccine-induced cellular immunity by Ad35-based vectors. Ad5/Ad35-based vaccination induced durable, high-frequency effector T cell responses that were later recalled upon heterologous SIV challenge. Vaccination resulted in modest reductions in SIV viremia, notable given the limited scope of immunization. We then tested the capacity of Ad5/Ad35-based vaccination targeting the SIV Gag, Env, and Nef proteins, with or without IL-15 augmentation, to promote cellular immunity during antiretroviral-treated chronic SIV infection with the goal of limiting rebound viremia following cessation of antiretroviral therapy (ART). Vaccination enhanced both systemic and mucosal antigen-specific cell-mediated immunity, increasing the breadth and strength over innate response to infection. Ad-induced immunity consisted of CD4+ and CD8+ T lymphocyte TH1 cytokine production of a predominantly monofunctional nature. Furthermore, vaccination enhanced both central and effector memory CD4+ and CD8+ T cell populations without augmenting niave T cell responses. Although Ad-based immunotherapy transiently restored the systemic central memory CD4+ T cell compartment, vaccination failed to salvage effector memory or mucosal CD4+ T cells. Therapeutic intervention was associated with transient containment of rebound viremia upon ART cessation which vaccination failed to augment. An effective vaccination against HIV represents the most efficient method to end the AIDS pandemic, and is of considerable public health significance. The findings presented herein provide evidence to support the continued evaluation of Ad-based vectors in novel treatment strategies against HIV infection, representing an incremental advancement in the field of HIV vaccine development

A Novel Methodology for Isolating Broadly Neutralizing HIV-1 Human Monoclonal Antibodies

Abstract also published in AIDS Research and Human Retroviruses. November 2013, 29(11): A-53. doi:10.1089/aid.2013.1500Poster presentationpublished_or_final_versio

Current Perspectives in HIV Infection

This book gives a comprehensive overview of HIV and AIDS including NeuroAIDS, as well as general concepts of pathology, immunity and immunopathology, diagnosis, treatment, epidemiology and etiology to current clinical recommendations in management of HIV/AIDS including NeuroAIDS, highlighting the ongoing issues, recent advances and future directions in diagnostic approaches and therapeutic strategies