Abatacept increases T cell exhaustion in early RA individuals who carry HLA risk alleles.

Exhausted CD8 T cells (TEX) are associated with worse outcome in cancer yet better outcome in autoimmunity. Building on our past findings of increased TIGIT+KLRG1+ TEX with teplizumab therapy in type 1 diabetes (T1D), in the absence of treatment we found that the frequency of TIGIT+KLRG1+ TEX is stable within an individual but differs across individuals in both T1D and healthy control (HC) cohorts. This TIGIT+KLRG1+ CD8 TEX population shares an exhaustion-associated EOMES gene signature in HC, T1D, rheumatoid arthritis (RA), and cancer subjects, expresses multiple inhibitory receptors, and is hyporesponsive in vitro, together suggesting co-expression of TIGIT and KLRG1 may broadly define human peripheral exhausted cells. In HC and RA subjects, lower levels of EOMES transcriptional modules and frequency of TIGIT+KLRG1+ TEX were associated with RA HLA risk alleles (DR0401, 0404, 0405, 0408, 1001) even when considering disease status and cytomegalovirus (CMV) seropositivity. Moreover, the frequency of TIGIT+KLRG1+ TEX was significantly increased in RA HLA risk but not non-risk subjects treated with abatacept (CTLA4Ig). The DR4 association and selective modulation with abatacept suggests that therapeutic modulation of TEX may be more effective in DR4 subjects and TEX may be indirectly influenced by cellular interactions that are blocked by abatacept.

Inhibitory effect of selected Indian honey on colon cancer cell growth by inducing apoptosis and targeting the ß-catenin/Wnt pathway.

Colon cancer is the most prevalent cause of death from cancer across the globe. Although chemotherapy drugs are predominantly used, their toxicity always remains a cause of concern. As an alternative to synthetic drugs, natural compounds or nutraceuticals are comparatively less toxic. Honey is widely used across different cultures as an alternative form of medicine. It represents a prominent source of plant-phenolic compounds and there is demonstrable evidence of its anti-oxidant and anti-microbial activities. The aim of the present work was to investigate the anti-proliferative effect of some Indian honeys and analyze their mechanism of action in colon cancer. In order to establish the composition-activity relationship, we evaluated the bioactive components present in selected honey samples by GC-MS and HPLC analysis. Indian honey samples showed a significant inhibitory impact on cell growth by restricting cell proliferation, causing apoptosis, and restricting the cell cycle in the G2/M phase specifically for colon cancer cells. The apoptotic activities, as imparted by the honey samples, were established by Annexin V/PI staining, real-time PCR, and immunoblot analyses. The treated cells showed increased expressions of p53 and caspases 3, 8, and 9, thus indicating the involvement of both extrinsic and intrinsic apoptotic pathways. The honey samples were also found to inhibit the ß-catenin/Wnt pathway. In the next phase of the study, the efficacy of these honey samples was evaluated in colon carcinoma induced SD-rats. Overall, these findings demonstrated that selected Indian honeys could be established as effective nutraceuticals for the prevention as well as cure of colon cancer.

HLA-DRB1 risk alleles for RA are associated with differential clinical responsiveness to abatacept and adalimumab: data from a head-to-head, randomized, single-blind study in autoantibody-positive early RA.

BACKGROUND: Certain risk alleles associated with autoantibody-positive rheumatoid arthritis (RA) have been linked to poorer prognoses. In patients with autoantibody-positive RA, abatacept shows differential efficacy to tumor necrosis factor inhibitors. Our aim was to investigate the relationship between clinical response to abatacept and to adalimumab and presence of risk alleles encoding human leukocyte antigen (HLA)-DRB1 shared epitope (SE) in RA. METHODS: In this head-to-head study, biologic-naïve adults with early (≤ 12 months), moderate-to-severe RA and inadequate response to methotrexate (MTX-IR), autoantibody-positive for both anti-cyclic citrullinated peptide 2 and rheumatoid factor, were randomized 1:1 to receive subcutaneous abatacept 125 mg weekly or subcutaneous adalimumab 40 mg every 2 weeks for 24 weeks with stable, weekly oral MTX. An open-label period to 48 weeks followed, during which adalimumab-treated patients were switched to abatacept. Patients were genotyped for HLA-DRB1 alleles and classified as SE-positive (≥ 1 SE allele) or SE-negative (no SE alleles). Efficacy was assessed at weeks 24 and 48. RESULTS: Forty patients each received abatacept (9 SE-negative, 30 SE-positive, one unknown) or adalimumab (9 SE-negative, 31 SE-positive). Mean age and disease duration were 46.0 years and 5.5 months, respectively. At week 24, a greater percentage of abatacept patients achieved 50% improvement in ACR criteria (ACR50) compared with adalimumab patients (73% vs 45%, respectively) and estimate of difference (95% confidence interval [CI]), 28 (5, 48). In SE-positive patients, ACR50 estimate of difference (95% CI) was 32 (7, 55). During the open-label period, responses were sustained in the abatacept non-switch group and showed trends toward further improvement in the adalimumab-to-abatacept switch group at week 48, in both the overall and the SE-positive subpopulation. No new safety signals were identified. CONCLUSIONS: In MTX-IR patients with early, autoantibody-positive RA, abatacept resulted in numerically higher efficacy responses versus adalimumab after 24 weeks, with more pronounced treatment differences in SE-positive patients. After 48 weeks, responses were sustained in patients who continued abatacept while those who switched to abatacept showed further clinical improvement, overall, and in SE-positive patients. This supports co-stimulation blockade as an effective treatment strategy for patients with early, autoantibody-positive RA, particularly among SE-positive patients. TRIAL REGISTRATION: NIH US National Library of Medicine, NCT02557100 . Registered on September 23, 2015.

Efficacy and safety of abatacept in active primary Sjögren's syndrome: results of a phase III, randomised, placebo-controlled trial.

OBJECTIVES: To evaluate efficacy and safety of abatacept in adults with active primary Sjögren's syndrome (pSS) in a phase III, randomised, double-blind, placebo-controlled trial. METHODS: Eligible patients (moderate-to-severe pSS [2016 ACR/European League Against Rheumatism (EULAR) criteria], EULAR Sjögren's Syndrome Disease Activity Index [ESSDAI] ≥5, anti-SS-related antigen A/anti-Ro antibody positive) received weekly subcutaneous abatacept 125 mg or placebo for 169 days followed by an open-label extension to day 365. Primary endpoint was mean change from baseline in ESSDAI at day 169. Key secondary endpoints were mean change from baseline in EULAR Sjögren's Syndrome Patient Reported Index (ESSPRI) and stimulated whole salivary flow (SWSF) at day 169. Other secondary clinical endpoints included glandular functions and patient-reported outcomes. Selected biomarkers and immune cell phenotypes were examined. Safety was monitored. RESULTS: Of 187 patients randomised, 168 completed double-blind period and 165 continued into open-label period. Mean (SD) baseline ESSDAI and ESSPRI total scores were 9.4 (4.3) and 6.5 (2.0), respectively. Statistical significance was not reached for primary (ESSDAI -3.2 abatacept vs -3.7 placebo, p=0.442) or key secondary endpoints (ESSPRI, p=0.337; SWSF, p=0.584). No clinical benefit of abatacept over placebo at day 169 was seen with other clinical and PRO endpoints. Relative to baseline, abatacept was associated with significant differences vs placebo in some disease-relevant biomarkers (including IgG, IgA, IgM-rheumatoid factor) and pathogenic cell subpopulations (post hoc analyses). No new safety signals were identified. CONCLUSIONS: Abatacept treatment did not result in significant clinical efficacy compared with placebo in patients with moderate-to-severe pSS, despite evidence of biological activity.

Composite of Relevant Endpoints for Sjögren's Syndrome (CRESS): development and validation of a novel outcome measure.

BACKGROUND: Recent randomised controlled trials (RCTs) in primary Sjögren's syndrome used the European League Against Rheumatism (EULAR) Sjögren's Syndrome Disease Activity Index (ESSDAI) as their primary endpoint. Given the heterogeneous and complex nature of primary Sjögren's syndrome, it might be more appropriate to also assess other clinically relevant disease features. We aimed to develop a novel composite endpoint for assessing treatment efficacy in patients with primary Sjögren's syndrome: the Composite of Relevant Endpoints for Sjögren's Syndrome (CRESS). METHODS: A multidisciplinary expert team selected clinically relevant items and candidate measurements for inclusion in the composite score. For each measurement, cutoff points for response to treatment were chosen based on expert opinion, previously published data on minimal clinically important improvements, and trial data, primarily the week-24 data of the single-centre ASAP-III trial of abatacept versus placebo. CRESS was validated using data from three independent RCTs: one trial of rituximab (TRACTISS), one of abatacept (multinational trial), and one of tocilizumab (ETAP). We calculated the number and percentage of patients who were responders in the separate CRESS items, and the percentage of responders based on the total CRESS at the primary endpoint visits (week 48 for TRACTISS, week 24 for the other two trials). Patients with fewer than three items available for evaluating CRESS response were imputed as non-responders. FINDINGS: Based on expert opinion, five complementary items were selected to assess response: (1) systemic disease activity by Clinical ESSDAI (less than 5 points); (2) patient-reported symptoms by EULAR Sjögren's Syndrome Patient Reported Index, assessed by a decrease of at least 1 point or at least 15% from baseline; (3) tear gland item by Schirmer's test and ocular staining score, assessed by an increase of at least 5 mm or decrease of at least 2 points, respectively, in patients with abnormal Schirmer's test or ocular staining score findings at baseline, or, in patients with normal baseline values, assessed by no change to abnormal for both; (4) salivary gland item, assessed by unstimulated whole saliva secretion (increase of at least 25%) and salivary gland ultrasonography (decrease of at least 25%); and (5) serology, assessed by rheumatoid factor (decrease of at least 25%) and IgG (decrease of at least 10%). Total CRESS response is defined as response on at least three of five items. Post-hoc assessment of phase 3 trial data showed that CRESS response rates at the primary endpoint visits were 60% (24 of 40) for abatacept versus 18% (seven of 39) for placebo (p<0·0001) in ASAP-III, 49% (33 of 67) for rituximab versus 30% (20 of 66) for placebo (p=0·026) in the TRACTISS trial, 45% (41 of 92) for abatacept versus 32% (30 of 95) for placebo (p=0·067) in the multinational abatacept trial, and 18% (10 of 55) for tocilizumab versus 24% (13 of 55) for placebo (p=0·48) in the ETAP trial. INTERPRETATION: The CRESS is a feasible, well-balanced, composite endpoint for use in trials of primary Sjögren's syndrome. As a next step, the CRESS will require validation in a prospective RCT. FUNDING: None. TRANSLATION: For the Dutch translation of the abstract see Supplementary Materials section.

Resistance profile of the HIV-1 maturation inhibitor GSK3532795 in vitro and in a clinical study.

GSK3532795 (formerly BMS955176) is a second-generation maturation inhibitor (MI) that progressed through a Phase 2b study for treatment of HIV-1 infection. Resistance development to GSK3532795 was evaluated through in vitro methods and was correlated with information obtained in a Phase 2a proof-of-concept study in HIV-1 infected participants. Both low and high concentrations of GSK3532795 were used for selections in vitro, and reduced susceptibility to GSK3532795 mapped specifically to amino acids near the capsid/ spacer peptide 1 (SP1) junction, the cleavage of which is blocked by MIs. Two key substitutions, A364V or V362I, were selected, the latter requiring secondary substitutions to reduce susceptibility to GSK3532795. Three main types of secondary substitutions were observed, none of which reduced GSK3532795 susceptibility in isolation. The first type was in the capsid C-terminal domain and downstream SP1 region (including (Gag numbering) R286K, A326T, T332S/N, I333V and V370A/M). The second, was an R41G substitution in viral protease that occurred with V362I. The third was seen in the capsid N-terminal domain, within the cyclophilin A binding domain (V218A/M, H219Q and G221E). H219Q increased viral replication capacity and reduced susceptibility of poorly growing viruses. In the Phase 2a study, a subset of these substitutions was also observed at baseline and some were selected following GSK35323795 treatment in HIV-1-infected participants.

Immunohistochemical evaluation of stress-responsive protein sestrin2 and its correlation with p53 mutational status in eyelid sebaceous gland carcinoma.

BACKGROUND: p53 is a stress-activated tumour suppressor gene, and its mutation has been associated with solid tumours including non-melanoma skin cancers. Sestrin2 expression is associated with DNA damage and oxidative stress and has been described as a downstream target of p53 network. However, its role in sebaceous gland carcinoma (SGC) remains unexplored. OBJECTIVES: To determine the role of p53 and its downstream target gene sestrin2 expression and p53 gene mutation status in SGC. METHODS: Twenty cases of eyelid SGC tumour and circulating cell-free DNA (ccfDNA) were subjected to mutational analysis of p53 gene. p53 and sesrin2 expression was evaluated by immunohistochemistry. Results were correlated with the clinicopathological features of eyelid SGC. RESULTS: p53 gene mutations was detected in 25% of the SGC cases. A C>T transition was identified in exon 6 in a single patient in both tumour and ccfDNA. A G>T transversion leading to amino acid change D259Y was seen in four patients. A splice site mutation affected a single case in exon 6. p53 expression was observed in 55% SGC. Loss of sestrin2 in 55% SGC cases correlated with poor tumour differentiation (P=0.0001), upper eyelid involvement (P=0.004), p53 mutation (P=0.039) and with mutant p53 expression (P=0.0001). CONCLUSION: Sestrin2 expression was found to be significantly reduced in p53 mutated SGC cases and in cases with strong p53 nuclear immunopositivity, suggesting that loss of sestrin2 may be of biological significance in the development of SGC and as a key downstream component of p53 tumour suppression network in eyelid SGC.

Clinical Trial of the Anti-PD-L1 Antibody BMS-936559 in HIV-1 Infected Participants on Suppressive Antiretroviral Therapy.

Background: Reversing immune exhaustion with an anti-PD-L1 antibody may improve human immunodeficiency virus type 1 (HIV-1)-specific immunity and increase clearance of HIV-1-expressing cells. Methods: We conducted a phase I, randomized, double-blind, placebo-controlled, dose-escalating study of BMS-936559, including HIV-1-infected adults aged >18 to <70 years on suppressive antiretroviral therapy with CD4+ counts >350 cells/µL and detectable plasma HIV-1 RNA by single-copy assay. Data on single infusions of BMS-936559 (0.3 mg/kg) versus placebo are described. The primary outcomes were safety defined as any grade 3 or greater or immune-related adverse event (AE) and the change in HIV-1 Gag-specific CD8+ T cell responses from baseline to day 28 after infusion. Results: Eight men enrolled: 6 received 0.3 mg/kg of BMS-936559, and 2 received placebo infusions. There were no BMS-936559-related grade 3 or greater AEs. In 1 participant, asymptomatic hypophysitis (a protocol-defined immune-related AE) was identified 266 days after BMS-936559 infusion; it resolved over time. The mean percentage of HIV-1 Gag-specific CD8+ T cells expressing interferon γ increased from baseline (0.09%) through day 28 (0.20%; P = .14), driven by substantial increases in 2 participants who received BMS-936559. Conclusions: In this first evaluation of an immunologic checkpoint inhibitor in healthy HIV-1-infected persons, single low-dose BMS-936559 infusions appeared to enhance HIV-1-specific immunity in a subset of participants. Clinical Trials Registration: NCT02028403.

Antiviral Activity, Safety, and Exposure-Response Relationships of GSK3532795, a Second-Generation Human Immunodeficiency Virus Type 1 Maturation Inhibitor, Administered as Monotherapy or in Combination With Atazanavir With or Without Ritonavir in a Phase 2a Randomized, Dose-Ranging, Controlled Trial (AI468002).

BACKGROUND: GSK3532795 is a second-generation human immunodeficiency virus type 1 (HIV-1) maturation inhibitor that targets HIV-1 Gag, inhibiting the final protease cleavage between capsid protein p24 and spacer protein-1, producing immature, noninfectious virions. METHODS: This was a phase 2a, randomized, dose-ranging multipart trial. In part A, subtype B-infected subjects received 5-120 mg GSK3532795 (or placebo) once daily for 10 days. In part B, subtype B-infected subjects received 40 mg or 80 mg GSK3532795 once daily with atazanavir (ATV) with or without (±) ritonavir (RTV) or standard of care (SOC) (tenofovir disoproxil fumarate 300 mg, emtricitabine 200 mg, and ATV/RTV 300 mg/100 mg) for 28 days. In part C, subtype C-infected subjects received 40 mg or 120 mg GSK3532795 once daily (or placebo) for 10 days. Endpoints included change in HIV-1 RNA from baseline on day 11 (parts A/C) or day 29 (part B). RESULTS: A >1 log10 median decline in HIV-1 RNA was achieved by day 11 in parts A and C and day 29 in part B at GSK3532795 doses ≥40 mg; part B subjects receiving GSK3532795 and ATV ± RTV achieved similar declines to those receiving SOC. Median of the maximum declines in HIV-1 RNA were similar for the 40-120 mg once-daily dose groups regardless of baseline Gag polymorphisms. There were no deaths, adverse events leading to discontinuation, or serious adverse events. CONCLUSIONS: GSK3532795 demonstrated potent antiviral activity against subtype B (monotherapy or with ATV ± RTV) and subtype C, and was generally well tolerated, which supported continued development of GSK3532795 in subjects with HIV-1 subtype B or subtype C. CLINICAL TRIALS REGISTRATION: NCT01803074.

The Second-Generation Maturation Inhibitor GSK3532795 Maintains Potent Activity Toward HIV Protease Inhibitor-Resistant Clinical Isolates.

BACKGROUND: Protease inhibitor (PI)-resistant HIV-1 isolates with primary substitutions in protease (PR) and secondary substitutions in Gag could potentially exhibit cross-resistance to maturation inhibitors. We evaluated the second-generation maturation inhibitor, GSK3532795, for activity toward clinical isolates with genotypic and phenotypic characteristics associated with PI resistance (longitudinal). METHODS: Longitudinal clinical isolates from 15 PI-treated patients and 7 highly PI-resistant (nonlongitudinal) viruses containing major and minor PI resistance-associated mutations were evaluated for GSK3532795 sensitivity. Phenotypic sensitivity was determined using the PhenoSense Gag/PR assay (Monogram Biosciences) or in-house single- and multiple-cycle assays. Changes from baseline [CFB; ratio of post- to pre-treatment FC-IC50 (fold-change in IC50 versus wild-type virus)] <3 were considered to be within the no-effect level. RESULTS: All nonlongitudinal viruses tested were sensitive to GSK3532795 (FC-IC50 range 0.16-0.68). Among longitudinal isolates, all post-PI treatment samples had major PI resistance-associated mutations in PR and 17/21 had PI resistance-associated changes in Gag. Nineteen of the 21 post-PI treatment samples had GSK3532795 CFB <3. Median (range) CFB was 0.83 (0.05-27.4) [Monogram (11 patients)] and 1.5 (1.0-2.2) [single-cycle (4 patients)]. The 2 post-PI treatment samples showing GSK3532795 CFB >3 (Monogram) were retested using single- and multiple-cycle assays. Neither sample had meaningful sensitivity changes in the multiple-cycle assay. Gag changes were not associated with an increased GSK3532795 CFB. CONCLUSIONS: GSK3532795 maintained antiviral activity against PI-resistant isolates with emergent PR and/or Gag mutations. This finding supports continued development of GSK3532795 in treatment-experienced patients with or without previous PI therapy.

Efficacy, safety, bone and metabolic effects of HIV nucleoside reverse transcriptase inhibitor BMS-986001 (AI467003): a phase 2b randomised, controlled, partly blinded trial.

BACKGROUND: BMS-986001 is a thymidine analogue nucleoside reverse transcriptase inhibitor (NRTI) designed to maintain in-vitro antiviral activity while minimising off-target effects. We assessed the efficacy and safety of BMS-986001 versus tenofovir disoproxil fumarate in treatment-naive patients with HIV-1. METHODS: In this phase 2b, randomised, active-controlled trial (AI467003), we recruited treatment-naive (no current or previous exposure to an antiretroviral drug for >1 week) adults (aged at least 18 years) with HIV-1 from 47 sites across Asia, Australia, Europe, North America, South Africa, and South America. Patients with plasma HIV-1 RNA greater than 5000 copies per mL and CD4 counts greater than 200 cells per µL were randomly assigned (2:2:2:3) to receive BMS-986001 100 mg, 200 mg, or 400 mg once a day or to receive tenofovir disoproxil fumarate 300 mg once a day; each allocation was given with efavirenz 600 mg once a day and lamivudine 300 mg once a day. Both patients and investigators were masked to BMS-986001 dose (achieved with similar looking placebo tablets), but not allocation up to and including week 48. The primary endpoints were the proportion of patients with plasma HIV-1 RNA less than 50 copies per mL and safety events (serious adverse events and adverse events leading to discontinuation) through week 24; the main analysis was with a modified intention-to-treat population. Resistance analysis was a secondary endpoint, and additional safety parameters were exploratory endpoints. This trial is registered with ClinicalTrials.gov, number NCT01489046, and the European Clinical Trials Database, number EudraCT 2011-003329-89. FINDINGS: Patients were recruited between Jan 25, 2012, and Oct 3, 2012; 757 patients were assessed for eligibility and 301 were randomly assigned to receive either BMS-986001 once a day (67 patients to 100 mg, 67 to 200 mg, and 66 to 400 mg) or tenofovir disoproxil fumarate (n=101). 297 patients received at least one dose of study drug. At week 24, 57 (88%) of 65 patients for whom there were data in the 100 mg group, 54 (81%) of 67 in the 200 mg group, 62 (94%) of 66 in the 400 mg group achieved HIV-1 RNA less than 50 copies per mL, compared with 88 (89%) of 99 in the tenofovir disoproxil fumarate group (modified intention-to-treat population). BMS-986001 was generally well tolerated through week 48. Two patients had BMS-986001-related serious adverse events (atypical drug eruption and thrombocytopenia) and two in the tenofovir disoproxil fumarate group had study drug-related serious adverse events (potential drug-induced liver injury and depression or lipodystrophy) that led to discontinuation. NRTI resistance-associated mutations were reported in four (2%) of 198 patients, and non-NRTI mutations in 17 (9%) of 198 patients receiving BMS-986001 versus none of 99 and one (1%) of 99 patients receiving tenofovir disoproxil fumarate, respectively. Compared with tenofovir disoproxil fumarate, individuals in the BMS-986001 groups showed a smaller decrease in lumbar spine and hip bone mineral density but greater accumulation of limb and trunk fat, subcutaneous and visceral adipose tissue, and increased total cholesterol. INTERPRETATION: BMS-986001 had similar efficacy to that of tenofovir disoproxil fumarate and was associated with a smaller decrease in bone mineral density; however, greater resistance and gains in both peripheral and central fat accumulation were recorded for the investigational drug. Bristol-Myers Squibb has discontinued its involvement in the development of BMS-986001, and future decisions on development will be made by Oncolys BioPharma. FUNDING: Bristol-Myers Squibb.

Changes in biomarkers in HIV-1-infected treatment-naive patients treated with tenofovir DF/emtricitabine plus atazanavir/ritonavir or lopinavir/ritonavir for 96 weeks: the CASTLE biomarker substudy.

BACKGROUND: The impact of boosted protease inhibitor therapy on inflammatory and cardiovascular biomarker levels in treatment-naive HIV-infected patients remains unclear and may differ between agents. Unconjugated bilirubin elevation, which favourably affects vascular biomarkers and cardiovascular disease risk in Gilbert's syndrome, occurs with atazanavir. METHODS: CASTLE was a 96-week study comparing efficacy and safety in treatment-naive HIV-1-infected patients randomized to atazanavir/ritonavir (ATV/r) versus lopinavir/ritonavir (LPV/r), each in combination with tenofovir disoproxil fumarate/emtricitabine. In this substudy, fasting plasma tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6), high sensitivity C-reactive protein (hs-CRP), plasminogen activator inhibitor-1 (PAI-1) and fibrinogen were assessed at baseline, week 12, 24, 48 and 96. Impact of grade 3-4 hyperbilirubinaemia on biomarkers was examined. RESULTS: CASTLE demonstrated similar efficacy in both treatment arms with higher rates of hyperbilirubinaemia on ATV/r and elevated lipids on LPV/r. In this substudy (n=224), patterns of biomarker expression were similar between the ATV/r and LPV/r groups and between-group differences in biomarker percentage change from baseline were not significant at 48 and/or 96 weeks. Hyperbilirubinaemia did not influence fasting biomarker expression. CONCLUSIONS: No significant differences were noted between ATV/r and LPV/r for biomarker percentage changes from baseline. Furthermore, no association was found between total bilirubin levels and biomarker expression.

Genotypic correlates of susceptibility to HIV-1 attachment inhibitor BMS-626529, the active agent of the prodrug BMS-663068.

OBJECTIVES: In an 8 day monotherapy study of subjects infected with HIV-1 (subtype B) (NCT01009814), BMS-626529 (an attachment inhibitor that binds to HIV-1 envelope glycoprotein gp120), administered as the prodrug BMS-663068, produced substantial declines in plasma HIV-1 RNA. However, large variability in susceptibility to BMS-626529 was noted and virus with low susceptibility was less likely to be suppressed by BMS-663068 administration. The current analysis sought to investigate the genotypic correlates of susceptibility to BMS-626529. METHODS: In vitro selection experiments, evaluation of clinical samples of subtype B from the monotherapy study and evaluation of intrinsically resistant subtype AE viruses were conducted. Reverse genetics was used to identify key substitutions in envelope clones responsible for reduced susceptibility. RESULTS: An M426L or S375M change were the major substitutions associated with reductions in susceptibility to BMS-626529 in baseline samples of subtype B viruses from the monotherapy study, with M434I and M475I contributing to a lesser extent. Class resistance in subtype AE viruses was mapped to 375H and 475I substitutions, found in the vast majority of these viruses. Analysis of multiple envelope clones from infected subjects showed higher intrasubject variability in susceptibility to BMS-626529 compared with other classes of entry inhibitors. CONCLUSIONS: These data define key genotypic substitutions in HIV-1 gp120 that could confer phenotypic resistance to BMS-626529.

Activity of the HIV-1 attachment inhibitor BMS-626529, the active component of the prodrug BMS-663068, against CD4-independent viruses and HIV-1 envelopes resistant to other entry inhibitors.

BMS-626529 is a novel small-molecule HIV-1 attachment inhibitor active against both CCR5- and CXCR4-tropic viruses. BMS-626529 functions by preventing gp120 from binding to CD4. A prodrug of this compound, BMS-663068, is currently in clinical development. As a theoretical resistance pathway to BMS-663068 could be the development of a CD4-independent phenotype, we examined the activity of BMS-626529 against CD4-independent viruses and investigated whether resistance to BMS-626529 could be associated with a CD4-independent phenotype. Finally, we evaluated whether cross-resistance exists between BMS-626529 and other HIV-1 entry inhibitors. Two laboratory-derived envelopes with a CD4-independent phenotype (one CXCR4 tropic and one CCR5 tropic), five envelopes from clinical isolates with preexisting BMS-626529 resistance, and several site-specific mutant BMS-626529-resistant envelopes were examined for their dependence on CD4 for infectivity or susceptibility to BMS-626529. Viruses resistant to other entry inhibitors (enfuvirtide, maraviroc, and ibalizumab) were also examined for susceptibility to BMS-626529. Both CD4-independent laboratory isolates retained sensitivity to BMS-626529 in CD4(-) cells, while HIV-1 envelopes from viruses resistant to BMS-626529 exhibited no evidence of a CD4-independent phenotype. BMS-626529 also exhibited inhibitory activity against ibalizumab- and enfuvirtide-resistant envelopes. While there appeared to be some association between maraviroc resistance and reduced susceptibility to BMS-626529, an absolute correlation cannot be presumed, since some CCR5-tropic maraviroc-resistant envelopes remained sensitive to BMS-626529. Clinical use of the prodrug BMS-663068 is unlikely to promote resistance via generation of CD4-independent virus. No cross-resistance between BMS-626529 and other HIV entry inhibitors was observed, which could allow for sequential or concurrent use with different classes of entry inhibitors.

Prediction of virological response and assessment of resistance emergence to the HIV-1 attachment inhibitor BMS-626529 during 8-day monotherapy with its prodrug BMS-663068.

BACKGROUND: BMS-663068 is the phosphonooxymethyl prodrug of BMS-626529, a small-molecule attachment inhibitor that targets the HIV-1 envelope glycoprotein gp120 preventing it from binding to CD4 T cells. In vitro investigations have demonstrated considerable variation in susceptibility of different HIV-1 isolates to BMS-626529. BMS-663068 monotherapy in HIV-1-infected subjects produced a mean maximum change from baseline of -1.64 log10 copies per milliliter, but the response was variable. METHODS: In this analysis, baseline and day 8 samples were analyzed for susceptibility to BMS-626529 and the presence of known HIV-1 attachment inhibitor resistance mutations. In addition, predictors of virological response (maximal HIV-1 RNA decline ≥1 log10 copies per milliliter) and resistance selection were investigated. RESULTS: The only factor associated with reduced virological response was low baseline susceptibility to BMS-626529. There was no apparent relationship between virological response and baseline treatment experience, coreceptor tropism, plasma HIV-1 RNA level, or CD4 T-cell count. Examination of all positions with known BMS-626529 resistance mutations based on in vitro selection studies showed that gp120 M426L was the primary substitution most clearly associated with nonresponse to BMS-663068. There was minimal change in susceptibility to BMS-626529 over the course of the study and no clear evidence of emergence of a known HIV-1 attachment inhibitor resistance mutation in the majority of subjects as measured by standard population-based phenotypic and genotypic approaches. CONCLUSIONS: Nonresponse to BMS-663068 was associated with low baseline susceptibility to BMS-626529 and the presence of M426L. In this short-term trial, there was minimal evidence of selection for BMS-626529 high-level resistance over 8 days of monotherapy with BMS-663068 by population-based approaches.

Pharmacodynamics, safety, and pharmacokinetics of BMS-663068, an oral HIV-1 attachment inhibitor in HIV-1-infected subjects.

BACKGROUND: BMS-663068 is a prodrug of the small-molecule inhibitor BMS-626529, which inhibits human immunodeficiency virus type 1 (HIV-1) infection by binding to gp120 and interfering with the attachment of virus to CD4+ T-cells. METHODS: Fifty HIV-1-infected subjects were randomized to 1 of 5 regimen groups (600 mg BMS-663068 plus 100 mg ritonavir every 12 hours [Q12H], 1200 mg BMS-663068 plus 100 mg ritonavir every bedtime, 1200 mg BMS-663068 plus 100 mg ritonavir Q12H, 1200 mg BMS-663068 Q12H plus 100 mg ritonavir every morning, or 1200 mg BMS-663068 Q12H) for 8 days in this open-label, multiple-dose, parallel study. The study assessed the pharmacodynamics, pharmacokinetics, and safety of BMS-663068. RESULTS: The maximum median decrease in plasma HIV-1 RNA load from baseline ranged from 1.21 to 1.73 log(10) copies/mL. Plasma concentrations of BMS-626529 were not associated with an antiviral response, while low baseline inhibitory concentrations and the minimum and average steady-state BMS-626529 plasma concentrations, when adjusted by the baseline protein binding-adjusted 90% inhibitory concentration (inhibitory quotient), were linked with antiviral response. BMS-663068 was generally well tolerated. CONCLUSIONS: Administration of BMS-663068 for 8 days with or without ritonavir resulted in substantial declines in plasma HIV-1 RNA levels and was generally well tolerated. Longer-term clinical trials of BMS-663068 as part of combination antiretroviral therapy are warranted. Clinical Trials Registration.NCT01009814.

In vitro antiviral characteristics of HIV-1 attachment inhibitor BMS-626529, the active component of the prodrug BMS-663068.

BMS-663068 is the phosphonooxymethyl prodrug of BMS-626529, a novel small-molecule attachment inhibitor that targets HIV-1 gp120 and prevents its binding to CD4(+) T cells. The activity of BMS-626529 is virus dependent, due to heterogeneity within gp120. In order to better understand the anti-HIV-1 spectrum of BMS-626529 against HIV-1, in vitro activities against a wide variety of laboratory strains and clinical isolates were determined. BMS-626529 had half-maximal effective concentration (EC(50)) values of <10 nM against the vast majority of viral isolates; however, susceptibility varied by >6 log(10), with half-maximal effective concentration values in the low pM range against the most susceptible viruses. The in vitro antiviral activity of BMS-626529 was generally not associated with either tropism or subtype, with few exceptions. Measurement of the binding affinity of BMS-626529 for purified gp120 suggests that a contributory factor to its inhibitory potency may be a relatively long dissociative half-life. Finally, in two-drug combination studies, BMS-626529 demonstrated additive or synergistic interactions with antiretroviral drugs of different mechanistic classes. These results suggest that BMS-626529 should be active against the majority of HIV-1 viruses and support the continued clinical development of the compound.

Enhanced phosphatase activity attenuates α-synucleinopathy in a mouse model.

α-Synuclein (α-Syn) is a key protein that accumulates as hyperphosphorylated aggregates in pathologic hallmark features of Parkinson's disease (PD) and other neurodegenerative disorders. Phosphorylation of this protein at serine 129 is believed to promote its aggregation and neurotoxicity, suggesting that this post-translational modification could be a therapeutic target. Here, we demonstrate that phosphoprotein phosphatase 2A (PP2A) dephosphorylates α-Syn at serine 129 and that this activity is greatly enhanced by carboxyl methylation of the catalytic C subunit of PP2A. α-Syn-transgenic mice raised on a diet supplemented with eicosanoyl-5-hydroxytryptamide, an agent that enhances PP2A methylation, dramatically reduced both α-Syn phosphorylation at Serine 129 and α-Syn aggregation in the brain. These biochemical changes were associated with enhanced neuronal activity, increased dendritic arborizations, and reduced astroglial and microglial activation, as well as improved motor performance. These findings support the notion that serine 129 phosphorylation of α-Syn is of pathogenetic significance and that promoting PP2A activity is a viable disease-modifying therapeutic strategy for α-synucleinopathies such as PD.

HIV type 1 from a patient with baseline resistance to CCR5 antagonists uses drug-bound receptor for entry.

CCR5 antagonists are a new class of antiretroviral drugs that block viral entry by disrupting interactions between the viral envelope (Env) glycoprotein and coreceptor. During the CCR100136 (EPIC) Phase IIb study of the CCR5 antagonist aplaviroc (APL) in treatment-naive individuals, a patient was identified who harbored virus strains that exhibited partial resistance to APL at the time of virologic failure. Retrospectively, it was found that APL resistance was present at baseline as well. To investigate the mechanism of APL resistance in this patient, we cloned HIV-1 env genes from plasma obtained at baseline and after virologic failure. Approximately 85% of cloned Envs were functional, and all exhibited partial resistance to APL. All Envs were R5-tropic, were partially resistant to other CCR5 antagonists including maraviroc on cells with high CCR5 expression, but remained sensitive to the fusion inhibitor enfuvirtide. Competition studies with natural CCR5 ligands revealed that the mechanism of drug resistance entailed the use of the drug-bound conformation of CCR5 by the Env proteins obtained from this individual. The degree of drug resistance varied between Env clones, and also varied depending on the cell line used or the donor from whom the primary T cells were obtained. Thus, both virus and host factors contribute to CCR5 antagonist resistance. This study shows that R5 HIV-1 strains resistant to CCR5 inhibitors can arise in patients, confirming a mechanism of resistance previously characterized in vitro. In addition, some patients can harbor CCR5 antagonist-resistant viruses prior to treatment, which may have implications for the clinical use of this new class of antiretrovirals.