Capsid Inhibition with Lenacapavir in Multidrug-Resistant HIV-1 Infection.

BACKGROUND: Patients with multidrug-resistant human immunodeficiency virus type 1 (HIV-1) infection have limited treatment options. Lenacapavir is a first-in-class capsid inhibitor that showed substantial antiviral activity in a phase 1b study. METHODS: In this phase 3 trial, we enrolled patients with multidrug-resistant HIV-1 infection in two cohorts, according to the change in the plasma HIV-1 RNA level between the screening and cohort-selection visits. In cohort 1, patients were first randomly assigned in a 2:1 ratio to receive oral lenacapavir or placebo in addition to their failing therapy for 14 days; during the maintenance period, starting on day 15, patients in the lenacapavir group received subcutaneous lenacapavir once every 6 months, and those in the placebo group received oral lenacapavir, followed by subcutaneous lenacapavir; both groups also received optimized background therapy. In cohort 2, all the patients received open-label oral lenacapavir with optimized background therapy on days 1 through 14; subcutaneous lenacapavir was then administered once every 6 months starting on day 15. The primary end point was the percentage of patients in cohort 1 who had a decrease of at least 0.5 log10 copies per milliliter in the viral load by day 15; a key secondary end point was a viral load of less than 50 copies per milliliter at week 26. RESULTS: A total of 72 patients were enrolled, with 36 in each cohort. In cohort 1, a decrease of at least 0.5 log10 copies per milliliter in the viral load by day 15 was observed in 21 of 24 patients (88%) in the lenacapavir group and in 2 of 12 patients (17%) in the placebo group (absolute difference, 71 percentage points; 95% confidence interval, 35 to 90). At week 26, a viral load of less than 50 copies per milliliter was reported in 81% of the patients in cohort 1 and in 83% in cohort 2, with a least-squares mean increase in the CD4+ count of 75 and 104 cells per cubic millimeter, respectively. No serious adverse events related to lenacapavir were identified. In both cohorts, lenacapavir-related capsid substitutions that were associated with decreased susceptibility developed in 8 patients during the maintenance period (6 with M66I substitutions). CONCLUSIONS: In patients with multidrug-resistant HIV-1 infection, those who received lenacapavir had a greater reduction from baseline in viral load than those who received placebo. (Funded by Gilead Sciences; CAPELLA ClinicalTrials.gov number, NCT04150068.).

Efficacy and Safety of Long-Acting Subcutaneous Lenacapavir in Heavily Treatment-Experienced People with Multi-Drug Resistant HIV-1: Week 104 Results of a Phase 2/3 Trial.

BACKGROUND: Lenacapavir is a long-acting HIV-1 capsid inhibitor for treatment of HIV-1 infection. We evaluated the efficacy and safety of lenacapavir in combination with an investigator-selected optimized background regimen (OBR) after 104 weeks in adults with multidrug-resistant HIV-1. METHODS: This ongoing, international, Phase 2/3 trial at 42 sites included 72 adults living with multidrug-resistant HIV-1. Following a 2-week oral lenacapavir loading phase, participants received subcutaneous lenacapavir every 26 weeks with an OBR. HIV-1 RNA, CD4 cell counts, and adverse events were assessed over 104 weeks. One participant did not enter the extension phase. RESULTS: At Week 104, 44 of 71 participants (62%, 95% CI 50; 73) had HIV-1 RNA <50 copies/mL via US Food & Drug Administration (FDA) snapshot algorithm. When missing data (including discontinuations) were excluded, 44 of 54 participants (82%) had HIV-1 RNA <50 copies/mL at Week 104, mean CD4 cell count increased by 122 cells/µL (95% CI 80; 165), and the proportion of participants with CD4 cell count <200 cells/µL decreased from 64% (46 of 72) at Baseline to 29% (16 of 55). Fourteen participants had treatment-emergent lenacapavir resistance; seven resuppressed (HIV-1 RNA <50 copies/mL) while maintaining lenacapavir use. There were no Grade 4 or serious treatment-related adverse events. One participant discontinued study drug due to an injection site reaction. CONCLUSIONS: Treatment with subcutaneous lenacapavir in combination with an OBR was well tolerated and resulted in a high rate of virological suppression over 104 weeks. Lenacapavir represents an important treatment option in people with multidrug-resistant HIV-1.

Antiviral Activity of Lenacapavir Against HIV-2 Isolates and Drug-resistant HIV-2 Mutants.

The activity of lenacapavir against HIV-1 has been extensively evaluated in vitro, but comparable data for HIV-2 are scarce. We determined the anti-HIV-2 activity of lenacapavir using single-cycle infections of MAGIC-5A cells and multicycle infections of a T cell line. Lenacapavir exhibited low-nanomolar activity against HIV-2, but was 11- to 14-fold less potent against HIV-2 in comparison to HIV-1. Mutations in HIV-2 that confer resistance to other antiretrovirals did not confer cross-resistance to lenacapavir. Although lenacapavir-containing regimens might be considered for appropriate patients with HIV-2, more frequent viral load and/or CD4 testing may be needed to assess clinical response.

Antiviral activity of tenofovir alafenamide (TAF) against HIV-1 clinical isolates harboring K65R.

Tenofovir alafenamide (TAF) is a prodrug of the nucleoside reverse transcriptase (RT) inhibitor tenofovir (TFV). Compared to the earlier TFV prodrug, TFV disoproxil fumarate (TDF), TAF achieves more than fourfold-higher intracellular levels of its active metabolite TFV diphosphate (TFV-DP) in clinical studies, while achieving a significant reduction of TFV systemic exposure. Resistance to TFV has been well established, with the K65R mutation in RT as the signature mutation. Here we evaluated the in vitro activity of TAF and TDF in patient-derived HIV-1 isolates harboring the K65R mutation. Clinical isolates containing K65R were cloned into the pXXLAI construct (n = 42). In vitro phenotypic susceptibility of the constructs to TAF and TDF was evaluated in an MT-2 cell HIV assay and in viral breakthrough assays modeling physiological concentrations of TAF and TDF. TAF and TDF susceptibility were highly correlated in K65R-containing mutants, ranging from 2.7- to 3.0-fold (K65R alone) and 1.2- to 27.6-fold (K65R+ other RT mutations) relative to wild-type. In viral breakthrough assays mimicking differences in physiological concentrations, TAF inhibited breakthrough of 40 of 42 clinical isolates, while the TDF equivalent only inhibited 32 of 42 isolates tested. TAF displayed a higher barrier to resistance than TDF in this panel of K65R-containing clinical isolates.

Rapid HIV-1 genotyping assay for the detection of capsid mutations.

Human immunodeficiency virus (HIV) capsid is one of the most recent viral proteins successfully targeted for the development of antiretrovirals. Lenacapavir is a first in class HIV-1 capsid inhibitor that was recently approved for the treatment of highly treatment-experienced people with HIV in combination with other anti-HIV drugs. Owing to the novelty of the viral target, methods to characterize the potential resistance-associated mutations present in capsid upon treatment failure have not been fully established yet. Here, we describe a rapid and simple method to amplify capsid fragments and to determine their sequence from various clinical samples including diverse HIV-1 subtypes. These methods could easily be implemented in laboratories, including hospital laboratories often caring for this patient population.

Resistance Analyses in Highly Treatment-Experienced People With Human Immunodeficiency Virus (HIV) Treated With the Novel Capsid HIV Inhibitor Lenacapavir.

BACKGROUND: Lenacapavir (LEN) is a first-in-class inhibitor of human immunodeficiency virus type 1 (HIV-1) capsid function in clinical development for the treatment of heavily treatment-experienced (HTE) people with HIV (PWH) harboring multidrug resistance (MDR) in combination with an optimized background regimen (OBR). Here we describe resistance analyses conducted in the pivotal phase 2/3 CAPELLA study. METHODS: CAPELLA enrolled viremic HTE PWH with resistance to ≥3 of 4 of the main antiretroviral (ARV) classes and resistance to ≥2 ARV drugs per class. Baseline resistance analyses used commercial assays (HIV-1 protease, reverse transcriptase, integrase genotypic/phenotypic tests). Postbaseline resistance was evaluated in participants experiencing virologic failure. RESULTS: At baseline, 46% of participants had resistance to the 4 main ARV drug classes, with one-third of participants having exhausted all drugs from ≥3 of the 4 main ARV classes. Treatment with LEN + OBR for 26 weeks led to viral suppression in 81% of participants. Postbaseline resistance mutations to lenacapavir occurred in 8 participants (6 with M66I, 1 with K70H, 1 with Q67H + K70R) who were receiving unintended functional LEN monotherapy at the time of resistance selection. CONCLUSIONS: LEN added to OBR led to high efficacy in this HTE patient population with MDR but could select for resistance when used unintentionally as functional monotherapy.

Phenotypic resistance to lenacapavir and monotherapy efficacy in a proof-of-concept clinical study.

BACKGROUND: Lenacapavir in vitro resistance selections identified seven mutations in HIV-1 capsid protein (CA) associated with reduced susceptibility. OBJECTIVES: To analyse lenacapavir activity against lenacapavir-associated resistance mutations in multiple assays. We also report Day 10 resistance analyses conducted in a Phase 1b study of lenacapavir (Study 4072) in people with HIV (PWH). METHODS: Mutations were inserted in a proviral DNA clone by site-directed mutagenesis, and viruses (n = 12) were generated by transfection. Sequences were used to generate single-cycle (SC) test vectors that were evaluated in a Gag-Pro assay, and replicative viruses were tested in a multicycle (MC) MT-2 assay to determine lenacapavir susceptibility. Study 4072 was a Phase 1b, double-blinded, placebo-controlled, dose-ranging, randomized study of lenacapavir in untreated PWH. Participants received a single dose of lenacapavir (up to 750 mg) or placebo (10 day monotherapy). CA resistance was characterized using genotypic and/or phenotypic assays. RESULTS: Lenacapavir susceptibility in the SC assay showed an inverse relationship between replication capacity and resistance. In Study 4072, all 29 participants receiving lenacapavir showed a robust virological response with no rebound. At baseline, no participant had resistance mutations to lenacapavir, and all had WT susceptibility to lenacapavir. Post-monotherapy analyses revealed the emergence of CA mutation Q67H at Day 10 in two participants. CONCLUSIONS: In vitro assays confirmed that increased resistance to lenacapavir was associated with decreased replication capacity of mutant viruses. In the clinical study no pre-existing lenacapavir resistance was detected. Emergence of Q67H occurred at exposures below the dose used in current Phase 2/3 studies. These results support development of lenacapavir as an antiretroviral agent.

Absence of Lenacapavir (GS-6207) Phenotypic Resistance in HIV Gag Cleavage Site Mutants and in Isolates with Resistance to Existing Drug Classes.

Lenacapavir (LEN; GS-6207) is a potent first-in-class inhibitor of HIV-1 capsid with long-acting properties and the potential for subcutaneous dosing every 3 months or longer. In the clinic, a single subcutaneous LEN injection (20 mg to 750 mg) in people with HIV (PWH) induced a strong antiviral response, with a >2.3 mean log10 decrease in HIV-1 RNA at day 10. HIV-1 Gag mutations near protease (PR) cleavage sites have emerged with the use of protease inhibitors (PIs). Here, we have characterized the activity of LEN in mutants with Gag cleavage site mutations (GCSMs) and mutants resistant to other drug classes. HIV mutations were inserted into the pXXLAI clone, and the resulting mutants (n = 70) were evaluated using a 5-day antiviral assay. LEN EC50 fold change versus the wild type ranged from 0.4 to 1.9 in these mutants, similar to that for the control drug. In contrast, reduced susceptibility to PIs and maturation inhibitors (MIs) was observed. Testing of isolates with resistance against the 4 main classes of drugs (n = 40) indicated wild-type susceptibility to LEN (fold change ranging from 0.3 to 1.1), while reduced susceptibility was observed for control drugs. HIV GCSMs did not impact the activity of LEN, while some conferred resistance to MIs and PIs. Similarly, LEN activity was not affected by naturally occurring variations in HIV Gag, in contrast to the reduced susceptibility observed for MIs. Finally, the activity of LEN was not affected by the presence of resistance mutations to the 4 main antiretroviral (ARV) drug classes. These data support the evaluation of LEN in PWH with multiclass resistance.

Antiviral Activity of Tenofovir Alafenamide against HIV-1 with Thymidine Analog-Associated Mutations and M184V.

Tenofovir alafenamide (TAF) and tenofovir disoproxil fumarate (TDF) are prodrugs of the HIV-1 nucleotide reverse transcriptase inhibitor tenofovir (TFV). In vivo, TAF achieves >4-fold-higher intracellular levels of TFV diphosphate (TFV-DP) compared to TDF. Since thymidine analog-associated mutations (TAMs) in HIV-1 confer reduced TFV susceptibility, patients with TAM-containing HIV-1 may benefit from higher TFV-DP levels delivered by TAF. Moreover, the presence of the M184V mutation increases TFV susceptibility during TDF- or TAF-based therapy. The susceptibilities to antiviral drugs of site-directed mutants (SDMs) and patient-derived mutants containing combinations of TAMs (M41L, D67N, K70R, L210W, T215Y, and K219Q) with or without the M184V mutation (TAMs±M184V) were evaluated using either 5-day multicycle (MC; n = 110) or 2-day single-cycle (SC; n = 96) HIV assays. The presence of M184V in TAM-containing HIV-1 SDMs (n = 48) significantly increased TAF sensitivity compared to SDMs without M184V (n = 48). The comparison of TAF and TDF resistance profiles was further assessed in viral breakthrough (VB) experiments mimicking clinically relevant drug concentrations. A total of 68 mutants were assayed at physiological concentration in VB experiments, with 15/68 mutants breaking through with TDF (TFV, the in vitro equivalent of TDF, was used in these experiments), and only 3 of 68 mutants breaking through under TAF treatment. Overall, in the VB assay mimicking the 4-fold-higher intracellular levels of TFV-DP observed clinically with TAF versus TDF, TAF inhibited viral breakthrough of most TAM-containing HIV-1, whereas TDF did not. These results indicate that TAF has a higher resistance threshold than TDF and suggest that higher resistance cutoffs should be applied for TAF compared to TDF in genotypic and phenotypic resistance algorithms.

Differential detection of M184V/I between plasma historical HIV genotypes and HIV proviral DNA from PBMCs.

BACKGROUND: The M184V/I reverse transcriptase mutation, which confers major resistance to lamivudine and emtricitabine, is still quite frequent in people living with HIV. The underlying presence of the M184V/I mutation may undermine virological outcomes of ART, particularly in the context of proposed treatment with two-drug combinations that include drugs affected by M184V, such as lamivudine. In suppressed patients for whom historical data are seldom available, resistance assays evaluating integrated viral DNA can help select a fully active switch regimen. OBJECTIVES: To compare detectability of M184V/I in historical HIV-1 RNA analyses versus HIV-1 DNA sequencing. METHODS: We analysed the detection of the M184V/I mutation in a prospective study and compared HIV historical genotypes (plasma) versus integrated HIV DNA (PBMCs) obtained via a validated commercial proviral HIV DNA assay. Eligible participants had HIV-1 RNA <50 copies/mL for ≥6 consecutive months prior to screening. A plasma historical genotypic report (HGR) showing the presence of M184V/I was required for all participants and proviral HIV DNA analysis was conducted prior to enrolment. RESULTS: All 84 participants had evidence of M184V or M184I in their HGR (100%), whereas the mutation was detected in only 40/84 participants by proviral HIV DNA sequencing analysis (48%). Differential detection of M184V/I was not associated with timing differences between the HGR and proviral HIV DNA sampling, the overall duration of ART, or CD4 cell counts and HIV-1 viral load at baseline. CONCLUSIONS: Our results suggest that undetected M184V/I should be considered when switching virologically suppressed patients to new regimens, particularly two-drug lamivudine- or emtricitabine-containing combinations.

Frequency of capsid substitutions associated with GS-6207 in vitro resistance in HIV-1 from antiretroviral-naive and -experienced patients.

BACKGROUND: GS-6207 is a first-in-class HIV capsid inhibitor, targeting several functions of the HIV capsid in the viral cycle, including viral particle assembly, capsid formation and nuclear entry. GS-6207 has demonstrated picomolar potency in vitro, activity confirmed by high potency in a Phase 1 clinical study, with a long-acting antiretroviral profile with potential dosing every 6 months. In vitro resistance selections previously conducted with increasing doses of GS-6207 have identified capsid variants with reduced susceptibility to GS-6207. OBJECTIVES: To study the prevalence of capsid mutations associated with in vitro resistance to GS-6207 in people living with HIV (PLWH). METHODS: Plasma samples from ART-naive or -experienced PLWH, including PI-experienced people, were sequenced and analysed for the presence of capsid variants identified during in vitro resistance selection: L56I, M66I, Q67H, K70N, N74D, N74S and T107N. RESULTS: Among the samples from the 1500 patients studied, none of the seven GS-6207 resistance mutations identified during in vitro selection experiments was detected, regardless of HIV subtype or PLWH treatment history. CONCLUSIONS: Out of the seven HIV capsid substitutions previously selected in vitro and shown to confer phenotypic resistance to GS-6207, none of these seven mutations was observed in this large dataset, suggesting that neither PLWH with previous PI failure nor PLWH with emergence of PI resistance mutations are anticipated to impact GS-6207 activity in these diverse HIV-infected populations.

Development of an HIV-1 integrase genotyping assay for HIV-1 subtype AE.

An human immunodeficiency virus (HIV)-1 integrase (IN) genotyping assay was developed to evaluate clinical samples from patients infected with HIV-1 subtype AE, a subtype highly prevalent in Asia. The HIV-1 IN gene was amplified from plasma-derived HIV-1 viral RNA via reverse transcription polymerase chain reaction followed by population sequencing. Assay sensitivity was also evaluated using serially diluted plasma samples. The genotyping assay successfully amplified the IN gene from patient plasma samples with HIV-1 RNA as low as 500 copies/mL. This assay is suitable for IN genotyping to identify IN drug resistance mutations in HIV-1 patients harboring subtype AE virus.

Antiviral activity of HIV-1 integrase strand-transfer inhibitors against mutants with integrase resistance-associated mutations and their frequency in treatment-naïve individuals.

The development of resistance to human immunodeficiency virus 1 (HIV-1) integrase strand-transfer inhibitors (INSTI) has been documented; however, knowledge of the impact of pre-existing integrase (IN) mutations on INSTI resistance (INSTI-R) is still evolving. The frequency of HIV-1 IN mutations in 2177 treatment-naïve subjects was investigated, along with the INSTI susceptibility of site-directed mutant viruses containing major and minor INSTI-R mutations. Total 6 of 39 minor INSTI-R mutations (M50I, S119P/G/T/R, and E157Q) were found in >1% of IN-treatment-naïve subjects with no impact on INSTI susceptibility. When each combined with major INSTI-R mutation, M50I, S119P, and E157Q led to decreased susceptibility to elvitegravir but remained sensitive to dolutegravir and bictegravir.

C-Reactive Protein Values After Colorectal Resection: Can We Discharge a Patient With a C-Reactive Protein Value >100? A Retrospective Cohort Study.

BACKGROUND: C-reactive protein is a useful negative predictive test for the development of anastomotic leakage following colorectal surgery. Evolution of procedures (laparoscopy, enhanced recovery program, early discharge, complex redo surgery) may influence C-reactive protein values; however, this is poorly studied to date. OBJECTIVE: The aim of this study is to evaluate C-reactive protein as an indicator of postoperative complication and as a predictor for discharge. DESIGN: This is retrospective study of a consecutive monocentric cohort. SETTINGS: All patients undergoing a colorectal resection with anastomosis (2014-2015) were included. MAIN OUTCOMES MEASURES: C-reactive protein, leukocytosis, type of resection, and postoperative course were the primary outcomes measured. RESULTS: A total of 522 patients were included. The majority had either a colorectal (n = 159, 31%) or coloanal anastomosis (n = 150, 29%). Overall morbidity was 29.3%. C-reactive protein was significantly higher among patient having intra-abdominal complications at an early stage (day 1-2) (164.6 vs 136.2; p = 0.0028) and late stage (day 3-4) (209.4 vs 132.1; p < 0.0001). In multivariate analysis, early C-reactive protein was associated with BMI (coefficient, 4.9; 95% CI, 3.2-6.5; p < 0.0001) and open surgical procedures (coefficient, 43.1; 95% CI, 27-59.1; p < 0.0001), while late C-reactive protein value was influenced by BMI (coefficient, 4.8; 95% CI, 2.5-7.0; p = 0.0024) and associated extracolonic procedures (coefficient, 34.2; 95% CI, 2.7-65.6; p = 0.033). Sensitivity, specificity, negative predictive values, and positive predictive values for intra-abdominal complication were 85.9%, 33.6%, 89.3%, and 27.1% for an early C-reactive protein <100 mg/L and 72.7%, 75.4%, 89.4%, and 49.2% for a late C-reactive protein <100 mg/L. Four hundred seven patients with an uneventful postoperative course were discharged at day 8 ± 6.4 with a mean discharge C-reactive protein of 83.5 ± 67.4. Thirty-eight patients (9.3%) were readmitted and had a significantly higher discharge C-reactive protein (138.6 ± 94.1 vs 77.8 ± 61.2, p = 0.0004). Readmission rate was 16.5% for patients with a discharge C-reactive protein >100 mg/L vs 6% with C-reactive protein <100 mg/L (p = 0.0008). For patients included in an enhanced recovery program (discharge at day 4 ± 2.4), the threshold should be higher because discharge is around day 3 or 4. With a C-reactive protein <140, readmission rate was 2% vs 19%, (p = 0.056). LIMITATIONS: This study includes retrospective data. CONCLUSION: C-reactive protein <100 mg/L is associated with a lower risk of intra-abdominal complication and readmission rates. See Video Abstract at http://links.lww.com/DCR/A749.

Commonly Transmitted HIV-1 Drug Resistance Mutations in Reverse-Transcriptase and Protease in Antiretroviral Treatment-Naive Patients and Response to Regimens Containing Tenofovir Disoproxil Fumarate or Tenofovir Alafenamide.

Background: The presence of transmitted drug resistance mutations (TDRMs) in antiretroviral treatment (ART)-naive patients can adversely affect the outcome of ART. Methods: Resistance testing was conducted in 6704 ART-naive subjects predominantly from the United States and Europe in 9 clinical studies conducted by Gilead Sciences from 2000 to 2013. Results: The presence of TDRMs increased during this period (from 5.2% to 11.4%), primarily driven by an increase in nonnucleoside reverse-transcriptase (RT) inhibitor (NNRTI) resistance mutations (from 0.3% to 7.1%), particularly K103N/S (increase from 0.3% to 5.3%). Nucleoside/nucleotide RT inhibitor mutations were found in 3.1% of patients. Only 1 patient had K65R (0.01%) and 7 had M184V/I (0.1%), despite high use of tenofovir disoproxil fumarate (TDF), emtricitabine, and lamivudine and potential transmission of resistance to these drugs. At least 1 thymidine-analogue mutations was present in 2.7% of patients with 0.07% harboring T215Y/F and 2.7% harboring T215 revertant mutations (T215rev). Patients with the combination of M41L + L210W + T215rev showed full human immunodeficiency virus RNA suppression while receiving a TDF- or tenofovir alafenamide-containing regimen. Conclusions: There was an overall increase of TDRMs among patients enrolling in clinical trials from 2000 through 2013, driven primarily by an increase in NNRTI resistance. However, the presence of common TDRMs, including thymidine-analogue mutations/T215rev, showed no impact on response to TDF- or tenofovir alafenamide-containing regimens.

Rare emergence of drug resistance in HIV-1 treatment-naïve patients after 48 weeks of treatment with elvitegravir/cobicistat/emtricitabine/tenofovir alafenamide.

Tenofovir alafenamide (TAF), a novel prodrug of the NtRTI tenofovir (TFV), delivers TFV-diphosphate (TFV-DP) to target cells more efficiently than the current prodrug, tenofovir disoproxil fumarate (TDF), with a 90% reduction in TFV plasma exposure. TAF, within the fixed dose combination of elvitegravir /cobicistat / emtricitabine (FTC)/TAF (E/C/F/TAF), has been evaluated in one Phase 2 and two Phase 3 randomized, double-blinded studies in HIV-infected treatment-naive patients, comparing E/C/F/TAF to E/C/F/TDF. In these studies, the TAF-containing group demonstrated non-inferior efficacy to the TDF-containing comparator group with 91.9% of E/C/F/TAF patients having <50 copies/mL of HIV-1 RNA at week 48. An integrated resistance analysis across these three studies was conducted, including HIV-1 genotypic analysis at screening, and genotypic/phenotypic analysis for patients with HIV-1 RNA>400 copies/mL at virologic failure. Pre-existing primary resistance-associated mutations (RAMs) were observed at screening among the 1903 randomized and treated patients: 7.5% had NRTI-RAMs, 18.2% had NNRTI-RAMs, and 3.4% had primary PI-RAMs. Pre-treatment RAMs did not influence treatment response at Week 48. In the E/C/F/TAF group, resistance development was rare; seven patients (0.7%, 7/978) developed NRTI-RAMs, five of whom (0.5%, 5/978) also developed primary INSTI-RAMs. In the E/C/F/TDF group, resistance development was also rare; seven patients (0.8%, 7/925) developed NRTI-RAMs, four of whom (0.4%, 4/925) also developed primary INSTI-RAMs. An additional analysis by deep sequencing in virologic failures revealed minimal differences compared to population sequencing. Overall, resistance development was rare in E/C/F/TAF-treated patients, and the pattern of emergent mutations was similar to E/C/F/TDF.

Characterization of HIV-1 Resistance to Tenofovir Alafenamide In Vitro.

Tenofovir alafenamide (TAF) is an investigational prodrug of the HIV-1 nucleotide reverse transcriptase (RT) inhibitor (NtRTI) tenofovir (TFV), with improved potency and drug delivery properties over the current prodrug, tenofovir disoproxil fumarate (TDF). TAF is currently in phase 3 clinical studies for the treatment of HIV-1 infection, in combination with other antiretroviral agents. Phase 1 and 2 studies have shown that TAF was associated with increased peripheral blood mononuclear cell (PBMC) drug loading and increased suppression of HIV-1 replication compared to treatment with TDF. In this study, selection of in vitro resistance to both TAF and the parent compound, TFV, led to the emergence of HIV-1 with the K65R amino acid substitution in RT with 6.5-fold-reduced susceptibility to TAF. Although TAF is more potent than TFV in vitro, the antiviral susceptibilities to TAF and TFV of a large panel of nucleoside/nucleotide RT inhibitor (NRTI)-resistant mutants were highly correlated (R(2) = 0.97), indicating that the two compounds have virtually the same resistance profile when assessed as fold change from the wild type. TAF showed full antiviral activity in PBMCs against primary HIV-1 isolates with protease inhibitor, nonnucleoside RT inhibitor (NNRTI), or integrase strand transfer inhibitor resistance but reduced activity against isolates with extensive NRTI resistance amino acid substitutions. However, the increased cell loading of TFV with TAF versus TDF observed in vivo suggests that TAF may retain activity against TDF-resistant mutant viruses.

Cobicistat-boosted darunavir in HIV-1-infected adults: week 48 results of a Phase IIIb, open-label single-arm trial.

BACKGROUND: Cobicistat is an alternative pharmacoenhancer to ritonavir. In healthy volunteers, darunavir exposure was comparable when darunavir 800 mg once daily was co-administered with cobicistat 150 mg once daily (as single agents or a fixed-dose combination) vs. with ritonavir 100 mg once daily. METHODS: This 48-week, Phase IIIb, single-arm, US multicenter study (NCT01440569) evaluated safety, efficacy and pharmacokinetics of darunavir/cobicistat 800/150 mg once daily (as single agents) plus two investigator-selected nucleoside/tide reverse transcriptase inhibitors (N[t]RTIs) in HIV-1-infected adults. Patients had no darunavir resistance-associated mutations (RAMs), plasma viral load (VL) ≥1000 HIV-1 RNA copies/ml, eGFR ≥80 ml/min and genotypic sensitivity to the two N[t]RTIs. The primary endpoint was any treatment-emergent grade 3 or 4 adverse events (AEs) through Week 24. RESULTS: The majority of the 313 intent-to-treat patients were treatment-naïve (295/313; 94%), male (89%), White (60%) and received a tenofovir-based regimen (99%). Median baseline VL and CD4(+) count overall were 4.8 log10 HIV-1 RNA copies/ml and 361 cells/mm(3), respectively. Overall, 86% of patients (268/313) completed the study. The majority of discontinuations were for AEs (15/313; 5%). The incidence of treatment-emergent grade 3 or 4 AEs regardless of causality was 6% through Week 24 and 8% through Week 48. Most common AEs through Week 48 were diarrhea (27%) and nausea (23%), which were grade 1 or 2 in severity. Week 48 virologic response rates (% with VL <50 HIV-1 RNA copies/ml; Snapshot analysis) were 81% overall and 83% in treatment-naïve patients; median increases in CD4(+) count at 48 weeks were 167 and 169 cells/mm(3), respectively. Of 15/313 patients who met the criteria for resistance analysis, one developed a darunavir RAM as a mixture with wild-type (I84I/V), without phenotypic resistance to darunavir. The mean population pharmacokinetic-derived darunavir areas under the plasma concentration-time curve were 102,000 overall and 100,620 ng•h/ml in treatment-naïve patients. No clinically relevant relationships were seen between darunavir exposure and virologic response, AEs or laboratory parameters. CONCLUSION: Darunavir/cobicistat 800/150 mg once daily was generally well tolerated through Week 48, with no new safety concerns. Pharmacokinetics, virologic and immunologic responses for darunavir/cobicistat were similar to previous data for darunavir/ritonavir 800/100 mg once daily.

Nucleoside reverse transcriptase inhibitor resistance mutations associated with first-line stavudine-containing antiretroviral therapy: programmatic implications for countries phasing out stavudine.

BACKGROUND: The World Health Organization Antiretroviral Treatment Guidelines recommend phasing-out stavudine because of its risk of long-term toxicity. There are two mutational pathways of stavudine resistance with different implications for zidovudine and tenofovir cross-resistance, the primary candidates for replacing stavudine. However, because resistance testing is rarely available in resource-limited settings, it is critical to identify the cross-resistance patterns associated with first-line stavudine failure. METHODS: We analyzed HIV-1 resistance mutations following first-line stavudine failure from 35 publications comprising 1,825 individuals. We also assessed the influence of concomitant nevirapine vs. efavirenz, therapy duration, and HIV-1 subtype on the proportions of mutations associated with zidovudine vs. tenofovir cross-resistance. RESULTS: Mutations with preferential zidovudine activity, K65R or K70E, occurred in 5.3% of individuals. Mutations with preferential tenofovir activity, ≥ two thymidine analog mutations (TAMs) or Q151M, occurred in 22% of individuals. Nevirapine increased the risk of TAMs, K65R, and Q151M. Longer therapy increased the risk of TAMs and Q151M but not K65R. Subtype C and CRF01_AE increased the risk of K65R, but only CRF01_AE increased the risk of K65R without Q151M. CONCLUSIONS: Regardless of concomitant nevirapine vs. efavirenz, therapy duration, or subtype, tenofovir was more likely than zidovudine to retain antiviral activity following first-line d4T therapy.

Impact of primary elvitegravir resistance-associated mutations in HIV-1 integrase on drug susceptibility and viral replication fitness.

Elvitegravir (EVG) is an effective HIV-1 integrase (IN) strand transfer inhibitor (INSTI) in advanced clinical development. Primary INSTI resistance-associated mutations (RAMs) at six IN positions have been identified in HIV-1-infected patients failing EVG-containing regimens in clinical studies: T66I/A/K, E92Q/G, T97A, S147G, Q148R/H/K, and N155H. In this study, the effect of these primary IN mutations, alone and in combination, on susceptibility to the INSTIs EVG, raltegravir (RAL), and dolutegravir (DTG); IN enzyme activities; and viral replication fitness was characterized. Recombinant viruses containing the six most common mutations exhibited a range of reduced EVG susceptibility: 92-fold for Q148R, 30-fold for N155H, 26-fold for E92Q, 10-fold for T66I, 4-fold for S147G, and 2-fold for T97A. Less commonly observed primary IN mutations also showed a range of reduced EVG susceptibilities: 40- to 94-fold for T66K and Q148K and 5- to 10-fold for T66A, E92G, and Q148H. Some primary IN mutations exhibited broad cross-resistance between EVG and RAL (T66K, E92Q, Q148R/H/K, and N155H), while others retained susceptibility to RAL (T66I/A, E92G, T97A, and S147G). Dual combinations of primary IN mutations further reduced INSTI susceptibility, replication capacity, and viral fitness relative to either mutation alone. Susceptibility to DTG was retained by single primary IN mutations but reduced by dual mutation combinations with Q148R. Primary EVG RAMs also diminished IN enzymatic activities, concordant with their structural proximity to the active site. Greater reductions in viral fitness of dual mutation combinations may explain why some primary INSTI RAMs do not readily coexist on the same HIV-1 genome but rather establish independent pathways of resistance to EVG.