HIV-1 Drug Resistance to Integrase Strand Transfer Inhibitors in HIV-1 non-B Subtypes

Human immunodeficiency syndrome (HIV-1) has infected over 75 million people and over 35 million have succumbed to virus related illnesses. Despite access to a variety of antiretroviral therapy (ART) options, ART programs have been disproportionally spread in the world with low-and middle-income countries (LMICs) facing challenges to access the most potent ART options. With less potent ART remaining in use in LMICs, HIV-1 drug resistance (HIVDR) presents a growing challenge in LMICs. Since approval of the first-generation integrase strand transfer inhibitor (INSTIs), Raltegravir (RAL) in 2007, INSTIs remain the best choice as a backbone of ART. Access to second generation INSTIs, Dolutegravir (DTG) and bictegravir (BIC) in LMICs is based on need and not on a full evaluation of the effectiveness of these treatments in patients infected with non-B HIV-1 subtypes. To address this challenge of limited INSTIs associated HIVDR data in non-B HIV subtypes, we first screened for the presence of INSTIs associated drug resistance mutations (DRMs) in ART naïve and experienced patients in Uganda using Sanger and Illumina sequencing. In Uganda, 47% of patients failing on RAL carry resistance to RAL-and elvitegravir (EVG), and only 4% harbor resistant virus to DTG. A panel of recombinant viruses from patient-derived HIV-1 integrases carrying resistant mutations was created and tested for susceptibility to a panel of INSTIs: EVG, RAL, DTG, BIC, and CAB. The virus carrying N155H or Y143R/S was susceptible to DTG, BIC, and CAB but highly resistant to RAL and EVG (\u3e50-fold change). Two patients, one with E138A/G140A/Q148R/G163R and one with E138K/G140A/S147G/Q148K, displayed the highest reported resistance to RAL, EVG (FC, \u3e1000) and even DTG (FC, \u3e100), BIC (FC, 60-\u3e100), and CAB (FC, 429-\u3e1000). All viruses had impaired replication fitness and \u3c50% reduction in integration capacity. We further determined potential novel polymorphisms associated with INSTI resistance in HIV-1 subtype A and D using simple vector machine analysis. The identified I208L and I203M, did not show reduced susceptibility to RAL or DTG with 1.3-1.8-fold and 1-1.4-fold observed, respectively. Further investigation is required to determine how these novel mutations influence susceptibility to INSTIs in HIV-1 subtype A and D infected patients

Addressing an HIV cure in LMIC

HIV-1 persists in infected individuals despite years of antiretroviral therapy (ART), due to the formation of a stable and long-lived latent viral reservoir. Early ART can reduce the latent reservoir and is associated with post-treatment control in people living with HIV (PLWH). However, even in post-treatment controllers, ART cessation after a period of time inevitably results in rebound of plasma viraemia, thus lifelong treatment for viral suppression is indicated. Due to the difficulties of sustained life-long treatment in the millions of PLWH worldwide, a cure is undeniably necessary. This requires an in-depth understanding of reservoir formation and dynamics. Differences exist in treatment guidelines and accessibility to treatment as well as social stigma between low- and-middle income countries (LMICs) and high-income countries. In addition, demographic differences exist in PLWH from different geographical regions such as infecting viral subtype and host genetics, which can contribute to differences in the viral reservoir between different populations. Here, we review topics relevant to HIV-1 cure research in LMICs, with a focus on sub-Saharan Africa, the region of the world bearing the greatest burden of HIV-1. We present a summary of ART in LMICs, highlighting challenges that may be experienced in implementing a HIV-1 cure therapeutic. Furthermore, we discuss current research on the HIV-1 latent reservoir in different populations, highlighting research in LMIC and gaps in the research that may facilitate a global cure. Finally, we discuss current experimental cure strategies in the context of their potential application in LMICs

Evolution of Protease Inhibitor Resistance in Human Immunodeficiency Virus Type 1 Infected Patients Failing Protease Inhibitor Monotherapy as Second-line Therapy in Low-income Countries: An Observational Analysis Within the EARNEST Randomized Trial.

BACKGROUND: Limited viral load (VL) testing in human immunodeficiency virus (HIV) treatment programs in low-income countries often delays detection of treatment failure. The impact of remaining on failing protease inhibitor (PI)-containing regimens is unclear. METHODS: We retrospectively tested VL in 2164 stored plasma samples from 386 patients randomized to receive lopinavir monotherapy (after initial raltegravir induction) in the Europe-Africa Research Network for Evaluation of Second-line Therapy (EARNEST) trial. Protease genotypic resistance testing was performed when VL >1000 copies/mL. We assessed evolution of PI resistance mutations from virological failure (confirmed VL >1000 copies/mL) until PI monotherapy discontinuation and examined associations using mixed-effects models. RESULTS: Median post-failure follow-up (in 118 patients) was 68 (interquartile range, 48-88) weeks. At failure, 20% had intermediate/high-level resistance to lopinavir. At 40-48 weeks post-failure, 68% and 51% had intermediate/high-level resistance to lopinavir and atazanavir; 17% had intermediate-level resistance (none high) to darunavir. Common PI mutations were M46I, I54V, and V82A. On average, 1.7 (95% confidence interval 1.5-2.0) PI mutations developed per year; increasing after the first mutation; decreasing with subsequent mutations (P < .0001). VL changes were modest, mainly driven by nonadherence (P = .006) and PI mutation development (P = .0002); I47A was associated with a larger increase in VL than other mutations (P = .05). CONCLUSIONS: Most patients develop intermediate/high-level lopinavir resistance within 1 year of ongoing viral replication on monotherapy but retain susceptibility to darunavir. Viral load increased slowly after failure, driven by non-adherence and PI mutation development. CLINICAL TRIALS REGISTRATION: NCT00988039

HIV Drug Resistance Mutations in Non-B Subtypes After Prolonged Virological Failure on NNRTI-Based First-Line Regimens in Sub-Saharan Africa

OBJECTIVE: To determine drug resistance mutation (DRM) patterns in a large cohort of patients failing nonnucleoside reverse transcriptase inhibitor (NNRTI)-based first-line antiretroviral therapy regimens in programs without routine viral load (VL) monitoring and to examine intersubtype differences in DRMs. DESIGN: Sequences from 787 adults/adolescents who failed an NNRTI-based first-line regimen in 13 clinics in Uganda, Kenya, Zimbabwe, and Malawi were analyzed. Multivariable logistic regression was used to determine the association between specific DRMs and Stanford intermediate-/high-level resistance and factors including REGA subtype, first-line antiretroviral therapy drugs, CD4, and VL at failure. RESULTS: The median first-line treatment duration was 4 years (interquartile range 30-43 months); 42% of participants had VL ≥100,000 copies/mL and 63% participants had CD4 <100 cells/mm. Viral subtype distribution was A1 (40%; Uganda and Kenya), C (31%; Zimbabwe and Malawi), and D (25%; Uganda and Kenya), and recombinant/unclassified (5%). In general, DRMs were more common in subtype-C than in subtype-A and/or subtype-D (nucleoside reverse transcriptase inhibitor mutations K65R and Q151M; NNRTI mutations E138A, V106M, Y181C, K101E, and H221Y). The presence of tenofovir resistance was similar between subtypes [P (adjusted) = 0.32], but resistance to zidovudine, abacavir, etravirine, or rilpivirine was more common in subtype-C than in subtype-D/subtype-A [P (adjusted) < 0.02]. CONCLUSIONS: Non-B subtypes differ in DRMs at first-line failure, which impacts on residual nucleoside reverse transcriptase inhibitor and NNRTI susceptibility. In particular, higher rates of etravirine and rilpivirine resistance in subtype-C may limit their potential utility in salvage regimens

Efficient handover methods for vehicle-to-infrastructure communications over heterogeneous wireless networks

One of the key performance-limiting factors in achieving the Quality of Services (QoSs) in Vehicle-to-Infrastructure (V2I) communications over a heterogeneous network is the increased number of unnecessary handovers. In heterogeneous network environments, handovers consume a lot of network resources and consequently increase end-to-end network latency. Other factors affecting the handover performance are the traffic type and network loading. In this thesis, an efficient network selection method for handover is investigated. In the study, a multi-tier, multi-RATs heterogeneous network is considered. We first propose a novel algorithm for selecting the most suitable candidate network. We then propose a method for reducing the number of unnecessary handovers by considering the distance between the vehicle and the target Base Station (BS). The short-list of the potential target networks includes only those lying in the direction of movement and are determined based on the geo-location of both vehicle and the candidate network BSs. Certainly, the better network performance is achieved by reducing the scanning time of the candidate networks. The surveyed literature has shown that the network load at the target network has a significant impact on the handover performance due to an increased handover drop probability. For example, a macro-cell BS could be regarded as a good candidate network for V2I communication due to the reasonable sojourn time, however, in the case, the BS is overloaded all handover requests are dropped. The integration of mobility and network information can further improve handover performance. A network selection method that applies the knowledge of both mobility and network load information is proposed for further improving handover performance. The better performance is achieved by optimizing the parameters such as cell dwelling time, load index, and Received Signal Strength (RSS) values. The type of traffic such as safety-related applications that require stringent QoS requirements and the network usage-cost also influence the network selection for handover. The concept of multi-criteria decision making which combines cell QoS metrics and user budget is investigated and applied for developing an application- aware network selection for handover method. Five application profiles are proposed based on the QoS metrics and network usage monetary cost. It was observed that the QoS of V2I safety-related applications could be guaranteed once they are allocated to the appropriate application profile. The thesis includes three main contributions. First, the scanning time of all available networks in the direction of movement is reduced by geo-locating both the vehicle and candidate network base stations. Second, by getting the knowledge of network information and vehicle mobility we further improved the system performance. Third, the overall network performance/utilization is improved by considering traffic type and network usage-cost. The thesis concludes with network design guidelines and deployment strategies for various practical network scenarios

Specialized DNA Structures Act as Genomic Beacons for Integration by Evolutionarily Diverse Retroviruses

Retroviral integration site targeting is not random and plays a critical role in expression and long-term survival of the integrated provirus. To better understand the genomic environment surrounding retroviral integration sites, we performed a meta-analysis of previously published integration site data from evolutionarily diverse retroviruses, including new experimental data from HIV-1 subtypes A, B, C and D. We show here that evolutionarily divergent retroviruses exhibit distinct integration site profiles with strong preferences for integration near non-canonical B-form DNA (non-B DNA). We also show that in vivo-derived HIV-1 integration sites are significantly more enriched in transcriptionally silent regions and transcription-silencing non-B DNA features of the genome compared to in vitro-derived HIV-1 integration sites. Integration sites from individuals infected with HIV-1 subtype A, B, C or D viruses exhibited different preferences for common genomic and non-B DNA features. In addition, we identified several integration site hotspots shared between different HIV-1 subtypes, all of which were located in the non-B DNA feature slipped DNA. Together, these data show that although evolutionarily divergent retroviruses exhibit distinct integration site profiles, they all target non-B DNA for integration. These findings provide new insight into how retroviruses integrate into genomes for long-term survival

Specialized DNA Structures Act as Genomic Beacons for Integration by Evolutionarily Diverse Retroviruses

Retroviral integration site targeting is not random and plays a critical role in expression and long-term survival of the integrated provirus. To better understand the genomic environment surrounding retroviral integration sites, we performed a meta-analysis of previously published integration site data from evolutionarily diverse retroviruses, including new experimental data from HIV-1 subtypes A, B, C and D. We show here that evolutionarily divergent retroviruses exhibit distinct integration site profiles with strong preferences for integration near non-canonical B-form DNA (non-B DNA). We also show that in vivo-derived HIV-1 integration sites are significantly more enriched in transcriptionally silent regions and transcription-silencing non-B DNA features of the genome compared to in vitro-derived HIV-1 integration sites. Integration sites from individuals infected with HIV-1 subtype A, B, C or D viruses exhibited different preferences for common genomic and non-B DNA features. In addition, we identified several integration site hotspots shared between different HIV-1 subtypes, all of which were located in the non-B DNA feature slipped DNA. Together, these data show that although evolutionarily divergent retroviruses exhibit distinct integration site profiles, they all target non-B DNA for integration. These findings provide new insight into how retroviruses integrate into genomes for long-term survival

Absence of HIV-1 Drug Resistance Mutations Supports the Use of Dolutegravir in Uganda

To screen for drug resistance and possible treatment with Dolutegravir (DTG) in treatment-naive patients and those experiencing virologic failure during first-, second-, and third-line combined antiretroviral therapy (cART) in Uganda. Samples from 417 patients in Uganda were analyzed for predicted drug resistance upon failing a first- (N = 158), second- (N = 121), or third-line [all 51 involving Raltegravir (RAL)] treatment regimen. HIV-1 pol gene was amplified and sequenced from plasma samples. Drug susceptibility was interpreted using the Stanford HIV database algorithm and SCUEAL was used for HIV-1 subtyping. Frequency of resistance to nucleoside reverse transcriptase inhibitors (NRTIs) (95%) and non-NRTI (NNRTI, 96%) was high in first-line treatment failures. Despite lack of NNRTI-based treatment for years, NNRTI resistance remained stable in 55% of patients failing second-line or third-line treatment, and was also at 10% in treatment-naive Ugandans. DTG resistance (n = 366) was not observed in treatment-naive individuals or individuals failing first- and second-line cART, and only found in two patients failing third-line cART, while 47% of the latter had RAL- and Elvitegravir-resistant HIV-1. Secondary mutations associated with DTG resistance were found in 2%-10% of patients failing third-line cART. Of 14 drugs currently available for cART in Uganda, resistance was readily observed to all antiretroviral drugs (except for DTG) in Ugandan patients failing first-, second-, or even third-line treatment regimens. The high NNRTI resistance in first-line treatment in Uganda even among treatment-naive patients calls for the use of DTG to reach the UNAIDS 90:90:90 goals

Antiretroviral APOBEC3 cytidine deaminases alter HIV-1 provirus integration site profiles

Antiretroviral APOBEC3 may contribute to HIV-1 latency. In this study, Ajoge and Renner et al. identify a previously undescribed function of human APOBEC3 proteins in redirecting integrations of HIV-1 DNA into more transcriptionally inactive regions of the genome