scholarly journals Analysis of Human Immunodeficiency Virus Type 1 Reverse Transcriptase Subunit Structure/Function in the Context of Infectious Virions and Human Target Cells

2005 ◽  
Vol 49 (9) ◽  
pp. 3762-3769 ◽  
Author(s):  
Alok Mulky ◽  
John C. Kappes
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Structure Function ◽  
Human Immunodeficiency Virus Type ◽  
Target Cells ◽  
Subunit Structure ◽  
Immunodeficiency Virus ◽  
Relevant Context ◽  
Hiv 1

ABSTRACT The reverse transcriptase (RT) of all retroviruses is required for synthesis of the viral DNA genome. The human immunodeficiency virus type 1 (HIV-1) RT exists as a heterodimer made up of 51-kDa and 66-kDa subunits. The crystal structure and in vitro biochemical analyses indicate that the p66 subunit of RT is primarily responsible for the enzyme's polymerase and RNase H activities. Since both the p51 and p66 subunits are generated from the same coding region, as part of the Pr160Gag-Pol precursor protein, there are inherent limitations for studying subunit-specific function with intact provirus in a virologically relevant context. Our lab has recently described a novel system for studying the RT heterodimer (p51/p66) wherein a LTR-vpr-p51-IRES-p66 expression cassette provided in trans to an RT-deleted HIV-1 genome allows precise molecular analysis of the RT heterodimer. In this report, we describe in detail the specific approaches, alternative strategies, and pitfalls that may affect the application of this novel assay for analyzing RT subunit structure/function in infectious virions and human target cells. The ability to study HIV-1 RT subunit structure/function in a physiologically relevant context will advance our understanding of both RT and the process of reverse transcription. The study of antiretroviral drugs in a subunit-specific virologic context should provide new insights into drug resistance and viral fitness. Finally, we anticipate that this approach will help elucidate determinants that mediate p51-p66 subunit interactions, which is essential for structure-based drug design targeting RT heterodimerization.

scholarly journals Discovery and Characterization of Vicriviroc (SCH 417690), a CCR5 Antagonist with Potent Activity against Human Immunodeficiency Virus Type 1

2005 ◽  
Vol 49 (12) ◽  
pp. 4911-4919 ◽  
Author(s):  
Julie M. Strizki ◽  
Cecile Tremblay ◽  
Serena Xu ◽  
Lisa Wojcik ◽  
Nicole Wagner ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Calcium Flux ◽  
Ccr5 Antagonist ◽  
Target Cells ◽  
Gene Transcript ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Inhibiting human immunodeficiency virus type 1 (HIV-1) infection by blocking the host cell coreceptors CCR5 and CXCR4 is an emerging strategy for antiretroviral therapy. Currently, several novel coreceptor inhibitors are being developed in the clinic, and early results have proven promising. In this report, we describe a novel CCR5 antagonist, vicriviroc (formerly SCH-D or SCH 417690), with improved antiviral activity and pharmacokinetic properties compared to those of SCH-C, a previously described CCR5 antagonist. Like SCH-C, vicriviroc binds specifically to the CCR5 receptor and prevents infection of target cells by CCR5-tropic HIV-1 isolates. In antiviral assays, vicriviroc showed potent, broad-spectrum activity against genetically diverse and drug-resistant HIV-1 isolates and was consistently more active than SCH-C in inhibiting viral replication. This compound demonstrated synergistic anti-HIV activity in combination with drugs from all other classes of approved antiretrovirals. Competition binding assays revealed that vicriviroc binds with higher affinity to CCR5 than SCH-C. Functional assays, including inhibition of calcium flux, guanosine 5′-[35S]triphosphate exchange, and chemotaxis, confirmed that vicriviroc acts as a receptor antagonist by inhibiting signaling of CCR5 by chemokines. Finally, vicriviroc demonstrated diminished affinity for the human ether a-go-go related gene transcript ion channel compared to SCH-C, suggesting a reduced potential for cardiac effects. Vicriviroc represents a promising new candidate for the treatment of HIV-1 infection.

scholarly journals Novel Nonnucleoside Inhibitors That Select Nucleoside Inhibitor Resistance Mutations in Human Immunodeficiency Virus Type 1 Reverse Transcriptase

2006 ◽  
Vol 50 (8) ◽  
pp. 2772-2781 ◽  
Author(s):  
Zhijun Zhang ◽  
Michelle Walker ◽  
Wen Xu ◽  
Jae Hoon Shim ◽  
Jean-Luc Girardet ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Immunodeficiency Virus ◽  
Nonnucleoside Inhibitors ◽  
Rt Inhibitors ◽  
Hiv 1 ◽  
M184v Mutation

ABSTRACT Mutations in and around the catalytic site of the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1) are associated with resistance to nucleoside RT inhibitors (NRTIs), whereas changes in the hydrophobic pocket of the RT are attributed to nonnucleoside RT inhibitor (NNRTI) resistance. In this study, we report a novel series of nonnucleoside inhibitors of HIV-1, exemplified by VRX-329747 and VRX-413638, which inhibit both NNRTI- and NRTI-resistant HIV-1 isolates. Enzymatic studies indicated that these compounds are HIV-1 RT inhibitors. Surprisingly, however, following prolonged (6 months) tissue culture selection, this series of nonnucleoside inhibitors did not select NNRTI-resistant mutations in HIV-1 RT. Rather, four mutations (M41L, A62T/V, V118I, and M184V) known to cause resistance to NRTIs and two additional novel mutations (S68N and G112S) adjacent to the catalytic site of the enzyme were selected. Although the M184V mutation appears to be the initial mutation to establish resistance, this mutation alone confers only a two- to fourfold decrease in susceptibility to VRX-329747 and VRX-413638. At least two additional mutations must accumulate for significant resistance. Moreover, while VRX-329747-selected viruses are resistant to lamivudine and emtricitabine due to the M184V mutation, they remain susceptible to zidovudine, stavudine, dideoxyinosine, abacavir, tenofovir, and efavirenz. These results directly demonstrate that VRX-329747 and VRX-413638 are novel nonnucleoside inhibitors of HIV-1 RT with the potential to augment current therapies.

scholarly journals The Thiocarboxanilide Nonnucleoside Inhibitor UC781 Restores Antiviral Activity of 3′-Azido-3′-Deoxythymidine (AZT) against AZT-Resistant Human Immunodeficiency Virus Type 1

1999 ◽  
Vol 43 (2) ◽  
pp. 259-263 ◽  
Author(s):  
Gadi Borkow ◽  
Dominique Arion ◽  
Mark A. Wainberg ◽  
Michael A. Parniak
Keyword(s):  
Human Immunodeficiency Virus ◽  
Antiviral Activity ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Wild Type Virus ◽  
Wild Type ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT N-[4-Chloro-3-(3-methyl-2-butenyloxy)phenyl]-2-methyl-3-furancarbothioamide (UC781) is an exceptionally potent nonnucleoside inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase. We found that a 1:1 molar combination of UC781 and 3′-azido-3′-deoxythymidine (AZT) showed high-level synergy in inhibiting the replication of AZT-resistant virus, implying that UC781 can restore antiviral activity to AZT against AZT-resistant HIV-1. Neither the nevirapine plus AZT nor the 2′,5′-bis-O-(t-butyldimethylsilyl)-3′-spiro-5"-(4"-amino-1",2"-oxathiole-2",2"-dioxide plus AZT combinations had this effect. Studies with purified HIV-1 reverse transcriptase (from a wild type and an AZT-resistant mutant) showed that UC781 was a potent inhibitor of the pyrophosphorolytic cleavage of nucleotides from the 3′ end of the DNA polymerization primer, a process that we have proposed to be critical for the phenotypic expression of AZT resistance. Combinations of UC781 plus AZT did not act in synergy to inhibit the replication of either wild-type virus or UC781-resistant HIV-1. Importantly, the time to the development of viral resistance to combinations of UC781 plus AZT is significantly delayed compared to the time to the development of resistance to either drug alone.

scholarly journals Impaired Processing and Presentation of Cytotoxic-T-Lymphocyte (CTL) Epitopes Are Major Escape Mechanisms from CTL Immune Pressure in Human Immunodeficiency Virus Type 1 Infection

2004 ◽  
Vol 78 (3) ◽  
pp. 1324-1332 ◽  
Author(s):  
Yoshiyuki Yokomaku ◽  
Hideka Miura ◽  
Hiroko Tomiyama ◽  
Ai Kawana-Tachikawa ◽  
Masafumi Takiguchi ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Target Cells ◽  
Ctl Epitopes ◽  
Field Isolates ◽  
Ctl Epitope ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Investigating escape mechanisms of human immunodeficiency virus type 1 (HIV-1) from cytotoxic T lymphocytes (CTLs) is essential for understanding the pathogenesis of HIV-1 infection and developing effective vaccines. To study the processing and presentation of known CTL epitopes, we prepared Epstein-Barr virus-transformed B cells that endogenously express the gag gene of six field isolates by adopting an env/nef-deletion HIV-1 vector pseudotyped with vesicular stomatitis virus G protein and then tested them for the recognition by Gag epitope-specific CTL lines or clones. We observed that two field variants, SLFNTVAVL and SVYNTVATL, of an A*0201-restricted Gag CTL epitope SLYNTVATL, and three field variants, KYRLKHLVW, QYRLKHIVW, and RYRLKHLVW, of an A24-restricted Gag CTL epitope KYKLKHIVW escaped from being killed by the CTL lines, despite the fact that they were recognized when the synthetic peptides corresponding to these variant sequences were exogenously loaded onto the target cells. Thus, their escape is likely due to the changes that occur during the processing and presentation of epitopes in the infected cells. Mutations responsible for this mode of escape were located within the epitope regions rather than the flanking regions, and such mutations did not influence the virus replication. The results suggest that the impaired antigen processing and presentation often occur in HIV-1 field isolates and thus are one of the major mechanisms that enable HIV-1 to escape from CTL recognition. We emphasize the importance of testing HIV-1 variants in an endogenous expression system.

An Approach towards the Synthesis of Potential Metal-Chelating TSAO-T Derivatives as Bidentate Inhibitors of Human Immunodeficiency virus Type 1 Reverse Transcriptase

1998 ◽  
Vol 9 (5) ◽  
pp. 412-421 ◽  
Author(s):  
C Chamorro ◽  
M-J Camarasa ◽  
M-J Pérez-Pérez ◽  
E de Clercq ◽  
J Balzarini ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Parent Compound ◽  
Immunodeficiency Virus ◽  
Anti Hiv ◽  
Hiv 1

Novel derivatives of the potent human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) inhibitor TSAO-T have been designed, synthesized and tested for their in vitro antiretro-viral activity against HIV. These TSAO-T derivatives have been designed as potential bidentate inhibitors of HIV-1 RT, which combine in their structure the functionality of a non-nucleoside RT inhibitor (TSAO-T) and a bivalent ion-chelating moiety (a β-diketone moiety) linked through an appropriate spacer to the N-3 of thymine of TSAO-T . Some of the new compounds have an anti-HIV-1 activity comparable to that of the parent compound TSAO-T, but display a markedly increased antiviral selectivity. There was a clear relationship between antiviral activity and the length of the spacer group that links the TSAO molecule with the chelating moiety. A shorter spacer invariably resulted in increased antiviral potency. None of the TSAO-T derivatives were endowed with anti-HIV-2 activity.

scholarly journals The M184V Mutation Reduces the Selective Excision of Zidovudine 5′-Monophosphate (AZTMP) by the Reverse Transcriptase of Human Immunodeficiency Virus Type 1

2002 ◽  
Vol 76 (7) ◽  
pp. 3248-3256 ◽  
Author(s):  
Paul L. Boyer ◽  
Stefan G. Sarafianos ◽  
Edward Arnold ◽  
Stephen H. Hughes
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Published Data ◽  
Immunodeficiency Virus ◽  
Azt Resistance ◽  
Hiv 1 ◽  
M184v Mutation

ABSTRACT The M184V mutation in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) causes resistance to lamivudine, but it also increases the sensitivity of the virus to zidovudine (3′-azido-3′-deoxythymidine; AZT). This sensitization to AZT is seen both in the presence and the absence of the mutations that confer resistance to AZT. AZT resistance is due to enhanced excision of AZT 5′-monophosphate (AZTMP) from the end of the primer by the RT of the resistant virus. Published data suggest that the excision reaction involves pyrophosphorolysis but that the likely in vivo pyrophosphate donor is not pyrophosphate but ATP. The mutations that lead to AZT resistance enhance ATP binding and, in so doing, enhance pyrophosphorolysis. The excision reaction is specific for AZT because HIV-1 RT, which can form a closed complex with a dideoxy-terminated primer and an incoming deoxynucleoside triphosphate (dNTP), does not form the closed complex with an AZTMP-terminated primer and an incoming dNTP. This means that an AZTMP-terminated primer has better access to the site where it can be excised. The M184V mutation alters the polymerase active site in a fashion that specifically interferes with ATP-mediated excision of AZTMP from the end of the primer strand. The M184V mutation does not affect the incorporation of AZT 5′-triphosphate (AZTTP), either in the presence or the absence of mutations that enhance AZTMP excision. However, in the presence of ATP, the M184V mutation does decrease the ability of HIV-1 RT to carry out AZTMP excision. Based on these results, and on the results of other excision experiments, we present a model to explain how the M184V mutation affects AZTMP excision.

scholarly journals The Human Immunodeficiency Virus Type 1 Nonnucleoside Reverse Transcriptase Inhibitor Resistance Mutation I132M Confers Hypersensitivity to Nucleoside Analogs

2009 ◽  
Vol 83 (8) ◽  
pp. 3826-3833 ◽  
Author(s):  
Zandrea Ambrose ◽  
Brian D. Herman ◽  
Chih-Wei Sheen ◽  
Shannon Zelina ◽  
Katie L. Moore ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Nucleoside Analogs ◽  
Replication Capacity ◽  
Immunodeficiency Virus ◽  
Viral Replication Capacity ◽  
Rt Inhibitors ◽  
Hiv 1

ABSTRACT We previously identified a rare mutation in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT), I132M, which confers high-level resistance to the nonnucleoside RT inhibitors (NNRTIs) nevirapine and delavirdine. In this study, we have further characterized the role of this mutation in viral replication capacity and in resistance to other RT inhibitors. Surprisingly, our data show that I132M confers marked hypersusceptibility to the nucleoside analogs lamivudine (3TC) and tenofovir at both the virus and enzyme levels. Subunit-selective mutagenesis studies revealed that the mutation in the p51 subunit of RT was responsible for the increased sensitivity to the drugs, and transient kinetic analyses showed that this hypersusceptibility was due to I132M decreasing the enzyme's affinity for the natural dCTP substrate but increasing its affinity for 3TC-triphosphate. Furthermore, the replication capacity of HIV-1 containing I132M is severely impaired. This decrease in viral replication capacity could be partially or completely compensated for by the A62V or L214I mutation, respectively. Taken together, these results help to explain the infrequent selection of I132M in patients for whom NNRTI regimens are failing and furthermore demonstrate that a single mutation outside of the polymerase active site and inside of the p51 subunit of RT can significantly influence nucleotide selectivity.

scholarly journals High Potency of Indolyl Aryl Sulfone Nonnucleoside Inhibitors towards Drug-Resistant Human Immunodeficiency Virus Type 1 Reverse Transcriptase Mutants Is Due to Selective Targeting of Different Mechanistic Forms of the Enzyme

2005 ◽  
Vol 49 (11) ◽  
pp. 4546-4554 ◽  
Author(s):  
Reynel Cancio ◽  
Romano Silvestri ◽  
Rino Ragno ◽  
Marino Artico ◽  
Gabriella De Martino ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Mechanism Of Action ◽  
Drug Resistant ◽  
Wild Type ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Indolyl aryl sulfone (IAS) nonnucleoside inhibitors have been shown to potently inhibit the growth of wild-type and drug-resistant human immunodeficiency virus type 1 (HIV-1), but their exact mechanism of action has not been elucidated yet. Here, we describe the mechanism of inhibition of HIV-1 reverse transcriptase (RT) by selected IAS derivatives. Our results showed that, depending on the substitutions introduced in the IAS common pharmacophore, these compounds can be made selective for different enzyme-substrate complexes. Moreover, we showed that the molecular basis for this selectivity was a different association rate of the drug to a particular enzymatic form along the reaction pathway. By comparing the activities of the different compounds against wild-type RT and the nonnucleoside reverse transcriptase inhibitor-resistant mutant Lys103Asn, it was possible to hypothesize, on the basis of their mechanism of action, a rationale for the design of drugs which could overcome the steric barrier imposed by the Lys103Asn mutation.

scholarly journals Identification of novel thiocarboxanilide derivatives that suppress a variety of drug-resistant mutant human immunodeficiency virus type 1 strains at a potency similar to that for wild-type virus.

1996 ◽  
Vol 40 (6) ◽  
pp. 1454-1466 ◽  
Author(s):  
J Balzarini ◽  
W G Brouwer ◽  
D C Dao ◽  
E M Osika ◽  
E De Clercq
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Effective Concentration ◽  
Mutant Virus ◽  
Wild Type ◽  
Immunodeficiency Virus ◽  
Virus Strains ◽  
Hiv 1

A large variety of carboxanilide and thiocarboxanilide derivatives in which the original oxathiin or aliphatic moieties present in the prototype compounds UC84 and UC38 were replaced by an (un) substituted furanyl, thienyl, phenyl, or pyrrole entity have been evaluated for activity against wild-type human immunodeficiency virus type 1 strain IIIB [HIV-1 (IIIB)] and a series of mutant virus strains derived thereof. The mutant viruses contained either the Leu-100-->Ile, Lys-103-->Asn, Val-106-->Ala, Glu-138-->Lys, Tyr-181-->Cys, or Tyr-188-->Leu mutation in their reverse transcriptase. Several 3-(2-methylfuranyl)- and 3-(2-methylthienyl)-thiocarboxanilide ester, (thio)ether, and oxime ether derivatives showed exquisitely potent antiviral activity against wild-type HIV-1 (50% effective concentration, 0.009 to 0.021 microM). The pentenylethers of the 2-methylfuranyl and 2-methylthienyl derivatives (i.e., 313, N-[4-chloro-3-(3-methyl-2-butenyloxy)phenyl]- 2-methyl-3-furancarbothioamide or UC-781, and 314, N-[4-chloro-3-(3-methyl-2-butenyloxy)phenyl] -2-methyl-3-thiophenecarbothioamide or UC-82) proved virtually equally inhibitory for wild-type and the Ile-100, Ala-106, and Lys-138 mutant virus strains (50% effective concentration, 0.015 to 0.021 microM). Their inhibitory effect against the Asn-103 and Cys-181 reverse transcriptase mutant virus strains was decreased only four- to sevenfold compared with wildtype virus. UC-781 and UC-82 should be considered potential candidate drugs for the treatment of HIV-1-infected individuals.

Sign in / Sign up

Export Citation Format

Share Document