Modelling and in vitro testing of the HIV-1 Nef fitness landscape

Abstract An effective vaccine is urgently required to curb the HIV-1 epidemic. We have previously described an approach to model the fitness landscape of several HIV-1 proteins, and have validated the results against experimental and clinical data. The fitness landscape may be used to identify mutation patterns harmful to virus viability, and consequently inform the design of immunogens that can target such regions for immunological control. Here we apply such an analysis and complementary experiments to HIV-1 Nef, a multifunctional protein which plays a key role in HIV-1 pathogenesis. We measured Nef-driven replication capacities as well as Nef-mediated CD4 and HLA-I down-modulation capacities of thirty-two different Nef mutants, and tested model predictions against these results. Furthermore, we evaluated the models using 448 patient-derived Nef sequences for which several Nef activities were previously measured. Model predictions correlated significantly with Nef-driven replication and CD4 down-modulation capacities, but not HLA-I down-modulation capacities, of the various Nef mutants. Similarly, in our analysis of patient-derived Nef sequences, CD4 down-modulation capacity correlated the most significantly with model predictions, suggesting that of the tested Nef functions, this is the most important in vivo. Overall, our results highlight how the fitness landscape inferred from patient-derived sequences captures, at least in part, the in vivo functional effects of mutations to Nef. However, the correlation between predictions of the fitness landscape and measured parameters of Nef function is not as accurate as the correlation observed in past studies for other proteins. This may be because of the additional complexity associated with inferring the cost of mutations on the diverse functions of Nef.

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The Fitness Landscape of HIV-1 Gag: Advanced Modeling Approaches and Validation of Model Predictions by In Vitro Testing

PLoS Computational Biology ◽

10.1371/journal.pcbi.1003776 ◽

2014 ◽

Vol 10 (8) ◽

pp. e1003776 ◽

Cited By ~ 85

Author(s):

Jaclyn K. Mann ◽

John P. Barton ◽

Andrew L. Ferguson ◽

Saleha Omarjee ◽

Bruce D. Walker ◽

...

Keyword(s):

Fitness Landscape ◽

In Vitro Testing ◽

Modeling Approaches ◽

Model Predictions ◽

Hiv 1

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Blocking HIV-1 Infection by Chromosomal Integrative Expression of Human CD4 on the Surface of Lactobacillus acidophilus ATCC 4356

Journal of Virology ◽

10.1128/jvi.01830-18 ◽

2019 ◽

Vol 93 (8) ◽

Cited By ~ 3

Author(s):

Wenzhong Wei ◽

Joshua Wiggins ◽

Duoyi Hu ◽

Vladimir Vrbanac ◽

Dane Bowder ◽

...

Keyword(s):

Mouse Model ◽

Lactobacillus Acidophilus ◽

Sexual Transmission ◽

Effective Vaccine ◽

Humanized Mouse ◽

Humanized Mouse Model ◽

Antiviral Proteins ◽

Hiv 1

ABSTRACT Lactobacillus bacteria are potential delivery vehicles for biopharmaceutical molecules because they are well-recognized as safe microorganisms that naturally inhabit the human body. The goal of this study was to employ these lactobacilli to combat human immunodeficiency virus type 1 (HIV-1) infection and transmission. By using a chromosomal integration method, we engineered Lactobacillus acidophilus ATCC 4356 to display human CD4, the HIV-1 receptor, on the cell surface. Since human CD4 can bind to any infectious HIV-1 particles, the engineered lactobacilli can potentially capture HIV-1 of different subtypes and prevent infection. Our data demonstrate that the CD4-carrying bacteria are able to adsorb HIV-1 particles and reduce infection significantly in vitro and also block intrarectal HIV-1 infection in a humanized mouse model in preliminary tests in vivo. Our results support the potential of this approach to decrease the efficiency of HIV-1 sexual transmission. IMPORTANCE In the absence of an effective vaccine, alternative approaches to block HIV-1 infection and transmission with commensal bacteria expressing antiviral proteins are being considered. This report provides a proof-of-concept by using Lactobacillus bacteria stably expressing the HIV-1 receptor CD4 to capture and neutralize HIV-1 in vitro and in a humanized mouse model. The stable expression of antiviral proteins, such as CD4, following genomic integration of the corresponding genes into this Lactobacillus strain may contribute to the prevention of HIV-1 sexual transmission.

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Design and crystal structure of a native-like HIV-1 envelope trimer that engages multiple broadly neutralizing antibody precursors in vivo

Journal of Experimental Medicine ◽

10.1084/jem.20161160 ◽

2017 ◽

Vol 214 (9) ◽

pp. 2573-2590 ◽

Cited By ~ 74

Author(s):

Max Medina-Ramírez ◽

Fernando Garces ◽

Amelia Escolano ◽

Patrick Skog ◽

Steven W. de Taeye ◽

...

Keyword(s):

Crystal Structure ◽

B Cells ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Effective Vaccine ◽

Major Advance ◽

Cd4 Binding Site ◽

Hiv 1

Induction of broadly neutralizing antibodies (bNAbs) by HIV-1 envelope glycoprotein immunogens would be a major advance toward an effective vaccine. A critical step in this process is the activation of naive B cells expressing germline (gl) antibody precursors that have the potential to evolve into bNAbs. Here, we reengineered the BG505 SOSIP.664 glycoprotein to engage gl precursors of bNAbs that target either the trimer apex or the CD4-binding site. The resulting BG505 SOSIP.v4.1-GT1 trimer binds multiple bNAb gl precursors in vitro. Immunization experiments in knock-in mice expressing gl-VRC01 or gl-PGT121 show that this trimer activates B cells in vivo, resulting in the secretion of specific antibodies into the sera. A crystal structure of the gl-targeting trimer at 3.2-Å resolution in complex with neutralizing antibodies 35O22 and 9H+109L reveals a native-like conformation and the successful incorporation of design features associated with binding of multiple gl-bNAb precursors.

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Combinatorial Approaches to the Prevention and Treatment of HIV-1 Infection

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00976-10 ◽

2011 ◽

Vol 55 (5) ◽

pp. 1831-1842 ◽

Cited By ~ 59

Author(s):

Vanessa Pirrone ◽

Nina Thakkar ◽

Jeffrey M. Jacobson ◽

Brian Wigdahl ◽

Fred C. Krebs

Keyword(s):

Clinical Trials ◽

Highly Active Antiretroviral Therapy ◽

Effective Vaccine ◽

Combination Therapies ◽

Preclinical Development ◽

Content Type ◽

Cell Counts ◽

Hiv 1

ABSTRACTThe discovery of the human immunodeficiency virus type 1 (HIV-1) in 1982 soon led to the identification and development of antiviral compounds to be used in treatment strategies for infected patients. Early in the epidemic, drug monotherapies frequently led to treatment failures because the virus quickly developed resistance to the single drug. Following the advent of highly active antiretroviral therapy (HAART) in 1995, dramatic improvements in HIV-1-infected patient health and survival were realized as more refined combination therapies resulted in reductions in viral loads and increases in CD4+T-cell counts. In the absence of an effective vaccine, prevention of HIV-1 infection has also gained traction as an approach to curbing the pandemic. The development of compounds as safe and effective microbicides has intensified and has focused on blocking the transmission of HIV-1 during all forms of sexual intercourse. Initial preclinical investigations and clinical trials of microbicides focused on single compounds effective against HIV-1. However, the remarkable successes achieved using combination therapy to treat systemic HIV-1 infection have subsequently stimulated the study and development of combination microbicides that will simultaneously inhibit multiple aspects of the HIV-1 transmission process by targeting incoming viral particles, virus-infected cells, and cells susceptible to HIV-1 infection. This review focuses on existing and developing combination therapies, covering preclinical development,in vitroandin vivoefficacy studies, and subsequent clinical trials. The shift in focus within the microbicide development field from single compounds to combination approaches is also explored.

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ECONOMIC PERSPECTIVES OF BIOTECHNOLOGIES USING IN ANIMAL FARMING

Vestnik of the Russian agricultural science ◽

10.30850/vrsn/2019/5/57-60 ◽

1970 ◽

pp. 57-60

Author(s):

I. F. Gorlov ◽

A. A. Mosolov ◽

G. V. Komlatskiy ◽

M. A. Nesterenko ◽

K. D. Nimbona ◽

...

Keyword(s):

Embryo Transfer ◽

Economic Effect ◽

Dairy Herd ◽

State Level ◽

Modern Level ◽

Economic Perspectives ◽

The Cost ◽

Short Time

The article presents materials on the study of the possibility of reproduction and increase in the herd of highly productive cows through the use of embryo transplantation technology. The classical (in vivo) and more modern, developing (in vitro) methods of embryotransfer, their positive and negative sides are considered in detail. The possibility of accelerating the breeding process by using the method of transplantation, in which from one cow can be obtained from 10 to 100 calves, which will allow for 4-5 years, almost any herd (of any size and breed) with the help of biotechnology to turn into a cattle-breeding enterprise of the most modern level. At the same time, heifers obtained from unproductive cows can be used as "surrogate" mothers who are transplanted with the best donor embryos, which allows to obtain a full-fledged offspring adapted to local environmental conditions. A detailed scheme of obtaining, evaluation, storage, as well as the cost and economic effect of embryo transplantation was calculated, the market was evaluated, the required annual volume of transplants and the number of donor cows for large livestock farms were determined. As a positive example of "Scientific-production enterprise "Centre of biotechnology and embryo transfer" in 2014, implemented a project for accelerated replacement and genetic improvement of the dairy herd, engraftment averaged 57-69%, and the economic effect of the enterprise from getting a single animal by the method of embryo transfer, compared with imports of similar close in quality, ranged from 60 to 100 thousand rubles on his head. It is shown that it is necessary to organize at the state level a developed service for embryo transplantation to reduce the cost of embryo transfer and the possibility of creating in a short time in the country's own highly productive breeding nucleus of dairy and beef cattle, which will reduce, and in the future completely eliminate, import dependence on cattle products.

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Mutational analysis of human immunodeficiency virus type 1 (HIV-1) accessory genes: requirement of a site in the nef gene for HIV-1 replication in activated CD4+ T cells in vitro and in vivo.

Journal of Virology ◽

10.1128/jvi.71.11.8456-8466.1997 ◽

1997 ◽

Vol 71 (11) ◽

pp. 8456-8466 ◽

Cited By ~ 36

Author(s):

Y Kawano ◽

Y Tanaka ◽

N Misawa ◽

R Tanaka ◽

J I Kira ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

T Cells ◽

Human Immunodeficiency Virus Type ◽

Mutational Analysis ◽

Immunodeficiency Virus ◽

A Site ◽

Hiv 1

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1592U89, a novel carbocyclic nucleoside analog with potent, selective anti-human immunodeficiency virus activity.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.41.5.1082 ◽

1997 ◽

Vol 41 (5) ◽

pp. 1082-1093 ◽

Cited By ~ 276

Author(s):

S M Daluge ◽

S S Good ◽

M B Faletto ◽

W H Miller ◽

M H St Clair ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Oral Bioavailability ◽

Human Peripheral Blood ◽

Cross Resistance ◽

Immunodeficiency Virus ◽

Carbocyclic Nucleoside ◽

Hiv 1 ◽

In Cells

1592U89, (-)-(1S,4R)-4-[2-amino-6-(cyclopropylamino)-9H-purin-9-yl]-2-cyclo pentene-1-methanol, is a carbocyclic nucleoside with a unique biological profile giving potent, selective anti-human immunodeficiency virus (HIV) activity. 1592U89 was selected after evaluation of a wide variety of analogs containing a cyclopentene substitution for the 2'-deoxyriboside of natural deoxynucleosides, optimizing in vitro anti-HIV potency, oral bioavailability, and central nervous system (CNS) penetration. 1592U89 was equivalent in potency to 3'-azido-3'-deoxythymidine (AZT) in human peripheral blood lymphocyte (PBL) cultures against clinical isolates of HIV type 1 (HIV-1) from antiretroviral drug-naive patients (average 50% inhibitory concentration [IC50], 0.26 microM for 1592U89 and 0.23 microM for AZT). 1592U89 showed minimal cross-resistance (approximately twofold) with AZT and other approved HIV reverse transcriptase (RT) inhibitors. 1592U89 was synergistic in combination with AZT, the nonnucleoside RT inhibitor nevirapine, and the protease inhibitor 141W94 in MT4 cells against HIV-1 (IIIB). 1592U89 was anabolized intracellularly to its 5'-monophosphate in CD4+ CEM cells and in PBLs, but the di- and triphosphates of 1592U89 were not detected. The only triphosphate found in cells incubated with 1592U89 was that of the guanine analog (-)-carbovir (CBV). However, the in vivo pharmacokinetic, distribution, and toxicological profiles of 1592U89 were distinct from and improved over those of CBV, probably because CBV itself was not appreciably formed from 1592U89 in cells or animals (<2%). The 5'-triphosphate of CBV was a potent, selective inhibitor of HIV-1 RT, with Ki values for DNA polymerases (alpha, beta, gamma, and epsilon which were 90-, 2,900-, 1,200-, and 1,900-fold greater, respectively, than for RT (Ki, 21 nM). 1592U89 was relatively nontoxic to human bone marrow progenitors erythroid burst-forming unit and granulocyte-macrophage CFU (IC50s, 110 microM) and human leukemic and liver tumor cell lines. 1592U89 had excellent oral bioavailability (105% in the rat) and penetrated the CNS (rat brain and monkey cerebrospinal fluid) as well as AZT. Having demonstrated an excellent preclinical profile, 1592U89 has progressed to clinical evaluation in HIV-infected patients.

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G2-S16 Polyanionic Carbosilane Dendrimer Can Reduce HIV-1 Reservoir Formation by Inhibiting Macrophage Cell to Cell Transmission

International Journal of Molecular Sciences ◽

10.3390/ijms22168366 ◽

2021 ◽

Vol 22 (16) ◽

pp. 8366

Author(s):

Ignacio Relaño-Rodríguez ◽

María de la Sierra Espinar-Buitrago ◽

Vanessa Martín-Cañadilla ◽

Rafael Gómez-Ramírez ◽

María Ángeles Muñoz-Fernández

Keyword(s):

T Cells ◽

Developed Countries ◽

Main Role ◽

Viral Particles ◽

Carbosilane Dendrimer ◽

Science Community ◽

New Compounds ◽

Hiv 1

Human immunodeficiency virus (HIV-1) is still a major problem, not only in developing countries but is also re-emerging in several developed countries, thus the development of new compounds able to inhibit the virus, either for prophylaxis or treatment, is still needed. Nanotechnology has provided the science community with several new tools for biomedical applications. G2-S16 is a polyanionic carbosilane dendrimer capable of inhibiting HIV-1 in vitro and in vivo by interacting directly with viral particles. One of the main barriers for HIV-1 eradication is the reservoirs created in primoinfection. These reservoirs, mainly in T cells, are untargetable by actual drugs or immune system. Thus, one approach is inhibiting HIV-1 from reaching these reservoir cells. In this context, macrophages play a main role as they can deliver viral particles to T cells establishing reservoirs. We showed that G2-S16 dendrimer is capable of inhibiting the infection from infected macrophages to healthy T CD4/CD8 lymphocytes by eliminating HIV-1 infectivity inside macrophages, so they are not able to carry infectious particles to other body locations, thus preventing the reservoirs from forming.

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