Overcoming drug resistance in HIV-1 chemotherapy: The binding thermodynamics of Amprenavir and TMC-126 to wild-type and drug-resistant mutants of the HIV-1 protease

AbstractSome of drug-resistant mutants of HIV-1 protease (PR), such as a clinically-relevant drug- resistant PR mutant (Flap+(I54V)) containing L10I, G48V, I54V and V82A mutations, produce significant changes in the balance between entropy and enthalpy of the drug-PR interactions, compared to the wild-type (WT) PR. Here, to gain a comprehensive understanding of the entropy-enthalpy compensation effects, we compared nuclear magnetic resonance (NMR), fluorescence spectroscopy and isothermal titration calorimetry (ITC) data of a WT PR with Flap+(I54V)and related mutants: (1) Flap+(I54V); (2) Flap+(I54A)which evolves from Flap+(I54V)in the continued presence of inhibitor yet does not exhibit entropy-enthalpy compensation; and (3) Flap+(I54), a control mutant that contains only L10I, G48V and V82A mutations. Our data indicate that WT and Flap+(I54A)show enthalpy-driven inhibitor-interaction, while Flap+(I54)and Flap+(I54V)exhibit entropy-driven inhibitor interaction. Interestingly, Flap+(I54A)exhibited significantly slower heat flow in the competitive ITC experiment with a strong binder, darunavir, and a weak binder, acetyl-pepstatin, but did not exhibit such slow heat flow in the direct inhibitor-titration experiments. NMR confirmed replacement of the weak binder by the strong binder in a competitive manner. This difference in the heat flow of the competitive binding experiment compared to the direct experiment can only be explained by assuming an inhibitor-bound intermediate pathway. A similar, but attenuated, tendency for slow heat flow was also detected in the competitive experiment with WT. Overall, our data suggests that an inhibitor-bound intermediate affects the entropy-enthalpy compensation of inhibitor-PR interaction.

Download Full-text

Impact of Novel Resistance Profiles in HIV-1 Reverse Transcriptase on Phenotypic Resistance to NVP

AIDS Research and Treatment ◽

10.1155/2012/637263 ◽

2012 ◽

Vol 2012 ◽

pp. 1-8

Author(s):

Liyan Jiao ◽

Hanping Li ◽

Lin Li ◽

Daomin Zhuang ◽

Yongjian Liu ◽

...

Keyword(s):

Drug Resistance ◽

Site Directed Mutagenesis ◽

Drug Resistant ◽

Wild Type ◽

Phenotypic Resistance ◽

Drug Resistant Mutation ◽

The Impact ◽

Hiv 1 ◽

Resistant Mutation ◽

Phenotypic Drug Resistance

Objective. To clarify the impact of H221Y mutation on drug resistance to NVP.Methods. 646 bp HIV-1polgene fragments (from 592 to 1237 nucleotide) with different NNRTIs mutation profiles from AIDS patients receiving antiretroviral therapy containing NVP regimens were introduced into pNL4-3 backbone plasmid. H221Y and (or) Y181C mutations were reverted to wild type amino acids by site-directed mutagenesis, then strains containing various mutation patterns were packaged. Phenotypic drug resistance was analyzed on TZM-bl cells.Results. 12 strains containing different drug-resistant mutation profiles were constructed, including the K101Q series (K101Q/Y181C/H221Y, K101Q/Y181C, K101Q/H221Y, and K101Q), the V179D series (V179D/Y181C/H221Y, V179D/Y181C, V179D/H221Y, and V179D), and the K103N series (K103N/Y181C/H221Y, K103N/Y181C, K103N/H221Y, K103N). For strains containing the mutation profiles (K101Q/Y181C, K101Q, V179D/Y181C, V179D, K103N/Y181C, and K103N), the presence of H221Y reduced NVP susceptibility by2.1±0.5to3.6±0.5fold. To the mutation profiles K101Q/H221Y, K101Q, V179D/H221Y, V179D, K103N/H221Y, and K103N, the presence of Y181C reduced NVP susceptibility by41.9±8.4to1297.0±289.1fold. For the strains containing K101Q, V179D, and K103N, the presence of Y181C/H221Y combination decreased NVP susceptibility by100.6±32.5to3444.6±834.5fold.Conclusion. On the bases of various NNRTIs mutation profiles, Y181C remarkably improved the IC50to NVP, although H221Ymutation alone just increases 2.1 ∼ 3.6-fold resistance to NVP, the mutation could improve 100.6 ∼ 3444.6-fold resistance to NVP when it copresent with Y181C, the phenotypic drug resistance fold was improved extremely. For strains containing the mutation profiles (K101Q/Y181C, K101Q, V179D/Y181C, V179D, K103N/Y181C, and K103N), the presence of H221Y reduced NVP susceptibility by2.1±0.5to3.6±0.5fold.

Download Full-text

Tackling the problem of HIV drug resistance

Postępy Biochemii ◽

10.18388/pb.2016_26 ◽

2016 ◽

Vol 62 (3) ◽

pp. 273-279

Author(s):

Irene T. Weber ◽

Robert W. Harrison

Keyword(s):

Drug Resistance ◽

Experimental Studies ◽

Hiv Infections ◽

Initial Step ◽

Wild Type ◽

Wild Type Enzyme ◽

Practical Applications ◽

Inhibitory Compounds ◽

Resistant Mutants ◽

Hiv 1

The virally-encoded HIV-1 protease is an effective target for antiviral drugs, however, treatment for HIV infections is limited by the prevalence of drug resistant viral mutants. In this review, we describe our three-pronged approach to analyze and combat drug resistance. Understanding the molecular basis for resistance due to protease inhibitors is a key initial step in this approach. This knowledge is being employed for the design of new, improved inhibitors with high affinity for resistant mutants as well as wild type enzyme. In parallel with experimental studies of diverse mutants and inhibitory compounds, we are developing efficient algorithms to predict drug resistance phenotype from genotype data. This approach has important practical applications in the clinic where genotyping is recommended for individuals with new infections.

Download Full-text

Enantioselective binding of second generation pyrrolobenzoxazepinones to the catalytic ternary complex of HIV-1 RT wild-type and L100I and K103N drug resistant mutants

Bioorganic & Medicinal Chemistry Letters ◽

10.1016/j.bmcl.2011.05.020 ◽

2011 ◽

Vol 21 (13) ◽

pp. 3935-3938 ◽

Cited By ~ 3

Author(s):

Stefania Butini ◽

Sandra Gemma ◽

Margherita Brindisi ◽

Giuseppe Borrelli ◽

Isabella Fiorini ◽

...

Keyword(s):

Ternary Complex ◽

Second Generation ◽

Drug Resistant ◽

Wild Type ◽

Resistant Mutants ◽

Hiv 1

Download Full-text

Competitive Fitness of Nevirapine-Resistant Human Immunodeficiency Virus Type 1 Mutants

Journal of Virology ◽

10.1128/jvi.78.2.603-611.2004 ◽

2004 ◽

Vol 78 (2) ◽

pp. 603-611 ◽

Cited By ~ 69

Author(s):

Jennifer A. Collins ◽

M. Gregory Thompson ◽

Elijah Paintsil ◽

Melisa Ricketts ◽

Joanna Gedzior ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Reverse Transcriptase ◽

Drug Resistant ◽

Wild Type ◽

Competitive Fitness ◽

Immunodeficiency Virus ◽

Resistant Mutants ◽

Nevirapine Resistance ◽

Hiv 1

ABSTRACT Determining the fitness of drug-resistant human immunodeficiency virus type 1 (HIV-1) strains is necessary for the development of population-based studies of resistance patterns. For this purpose, we have developed a reproducible, systematic assay to determine the competitive fitness of HIV-1 drug-resistant mutants. To demonstrate the applicability of this assay, we tested the fitness of the five most common nevirapine-resistant mutants (103N, 106A, 181C, 188C, and 190A), with mutations in HIV-1 reverse transcriptase (RT), singly and in combination (for a total of 31 variants) in a defined HIV-1 background. For these experiments, the 27 RT variants that produced viable virus were cocultured with wild-type virus without nevirapine. The ratios of the viral species were determined over time by utilization of a quantitative real-time RT-PCR-based assay. These experiments revealed that all of the viable variants were less fit than the wild type and demonstrated that the order of relative fitness of the single mutants tested was as follows: 103N > 181C > 190A > 188C > 106A. This order correlated with the commonality of these mutants as a result of nevirapine monotherapy. These investigations also revealed that, on average, the double mutants were less fit than the single mutants and the triple mutants were less fit than the double mutants. However, the fitness of the single and double mutants was often not predictive of the fitness of the derivative triple mutants, suggesting the presence of complex interactions between the closely aligned residues that confer nevirapine resistance. This complexity was also evident from the observation that all three of the replication-competent quadruple mutants were fitter than most of the triple mutants, and in some cases, even the double mutants. Our data suggest that, in many cases, viral fitness is the determining factor in the evolution of nevirapine-resistant mutants in vivo, that interactions between the residues that confer nevirapine resistance are complex, and that these interactions substantially affect reverse transcriptase structure and/or function.

Download Full-text

The inheritance of drug resistance and compatibility type in Phytophthora drechsleri

Genetics Research ◽

10.1017/s0016672300014671 ◽

1974 ◽

Vol 23 (1) ◽

pp. 75-86 ◽

Cited By ~ 21

Author(s):

Ikram A. Khaki ◽

D. S. Shaw

Keyword(s):

Drug Resistance ◽

Mating Type ◽

Drug Resistant ◽

Dominant Allele ◽

Wild Type ◽

Phytophthora Drechsleri ◽

Resistant Mutants ◽

Sexual Progeny ◽

Selection Of ◽

As If

SummaryMethods have been developed for the selection of mutants resistant to p-fluorophenylalanine, chloramphenicol and actidione. Resistance to p-fluorophenylalanine and to chloramphenicol was inherited as if determined in each case by a single dominant allele present in diploid somatic nuclei. Inheritance of mating type was anomalous in crosses involving drug-resistant mutants. Presumptive mating-type heterokaryons were generated in a few per cent of sexual progeny from a wild-type cross.

Download Full-text

A microfluidic device enabling drug resistance analysis of leukemia cells via coupled dielectrophoretic detection and impedimetric counting

Scientific Reports ◽

10.1038/s41598-021-92647-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yağmur Demircan Yalçın ◽

Taylan Berkin Töral ◽

Sertan Sukas ◽

Ender Yıldırım ◽

Özge Zorlu ◽

...

Keyword(s):

Drug Resistance ◽

K562 Cells ◽

Leukemia Cells ◽

Drug Resistant ◽

Wild Type ◽

Gene Expressions ◽

Before And After ◽

The Difference ◽

Selection Of ◽

Resistant Cells

AbstractWe report the development of a lab-on-a-chip system, that facilitates coupled dielectrophoretic detection (DEP-D) and impedimetric counting (IM-C), for investigating drug resistance in K562 and CCRF-CEM leukemia cells without (immuno) labeling. Two IM-C units were placed upstream and downstream of the DEP-D unit for enumeration, respectively, before and after the cells were treated in DEP-D unit, where the difference in cell count gave the total number of trapped cells based on their DEP characteristics. Conductivity of the running buffer was matched the conductivity of cytoplasm of wild type K562 and CCRF-CEM cells. Results showed that DEP responses of drug resistant and wild type K562 cells were statistically discriminative (at p = 0.05 level) at 200 mS/m buffer conductivity and at 8.6 MHz working frequency of DEP-D unit. For CCRF-CEM cells, conductivity and frequency values were 160 mS/m and 6.2 MHz, respectively. Our approach enabled discrimination of resistant cells in a group by setting up a threshold provided by the conductivity of running buffer. Subsequent selection of drug resistant cells can be applied to investigate variations in gene expressions and occurrence of mutations related to drug resistance.

Download Full-text

Factors Influencing HIV Drug Resistance among Pregnant Women in Luanda, Angola: Findings from a Cross-Sectional Study

Tropical Medicine and Infectious Disease ◽

10.3390/tropicalmed6010029 ◽

2021 ◽

Vol 6 (1) ◽

pp. 29

Author(s):

Cruz S. Sebastião ◽

Joana Morais ◽

Miguel Brito

Keyword(s):

Drug Resistance ◽

Pregnant Women ◽

Rural Areas ◽

Cross Sectional Study ◽

Drug Resistant ◽

Sectional Study ◽

Cross Sectional ◽

Hiv Drug Resistance ◽

Hiv 1 ◽

High Educational

The increase in HIV infection and drug-resistant strains is an important public health concern, especially in resource-limited settings. However, the identification of factors related to the propagation of infectious diseases represents a crucial target offering an opportunity to reduce health care costs as well as deepening the focus on preventing infection in high-risk groups. In this study, we investigate the factors related to drug resistance among HIV-infected pregnant women in Luanda, the capital city of Angola. This was a part of a cross-sectional study conducted with 42 HIV-positive pregnant women. A blood sample was collected, and HIV-1 genotyping was carried out using an in-house method. Multivariate analyses were performed to determine the interaction between sociodemographic characteristics and drug resistance. HIV drug resistance was detected in 44.1% of the studied population. High probabilities of drug resistance were observed for HIV-infected pregnant women living in rural areas (AOR: 2.73; 95% CI: 0.50–14.9) with high educational level (AOR: 6.27; 95% CI: 0.77–51.2) and comorbidities (AOR: 5.47; 95% CI: 0.28–106) and infected with a HIV-1 non-B subtype other than subtype C (AOR: 1.60; 95% CI: 0.25–10.3). The present study reports high HIV drug resistance. Furthermore, older-age, rural areas, high educational levels, unemployed status, having comorbidities, and HIV-1 subtypes were factors related to drug resistance. These factors impact on drug susceptibility and need to be urgently addressed in order to promote health education campaigns able to prevent the spread of drug-resistant HIV strains in Angola.

Download Full-text