Combination Immune Checkpoint Blockade Enhances IL-2 and CD107a Production from HIV-Specific T Cells Ex Vivo in People Living with HIV on Antiretroviral Therapy.

In people with HIV (PWH) on antiretroviral therapy (ART), immune dysfunction persists, including elevated expression of immune checkpoint (IC) proteins on total and HIV-specific T cells. Reversing immune exhaustion is one strategy to enhance the elimination of HIV-infected cells that persist in PWH on ART. We aimed to evaluate whether blocking CTL-associated protein 4 (CTLA-4), programmed cell death protein 1 (PD-1), T cell Ig domain and mucin domain 3 (TIM-3), T cell Ig and ITIM domain (TIGIT) and lymphocyte activation gene-3 (LAG-3) alone or in combination would enhance HIV-specific CD4+ and CD8+ T cell function ex vivo. Intracellular cytokine staining was performed using human PBMCs from PWH on ART (n = 11) and expression of CD107a, IFN-γ, TNF-α, and IL-2 was quantified with HIV peptides and Abs to IC. We found the following: 1) IC blockade enhanced the induction of CD107a and IL-2 but not IFN-γ and TNF-α in response to Gag and Nef peptides; 2) the induction of CD107a and IL-2 was greatest with multiple combinations of two Abs; and 3) Abs to LAG-3, CTLA-4, and TIGIT in combinations showed synergistic induction of IL-2 in HIV-specific CD8+ and CD107a and IL-2 production in HIV-specific CD4+ and CD8+ T cells. These results demonstrate that the combination of Abs to LAG-3, CTLA-4, or TIGIT can increase the frequency of cells expressing CD107a and IL-2 that associated with cytotoxicity and survival of HIV-specific CD4+ and CD8+ T cells in PWH on ART. These combinations should be further explored for an HIV cure.

Cutting Edge: SARS-CoV-2 Infection Induces Robust Germinal Center Activity in the Human Tonsil.

Understanding the generation of immunity to SARS-CoV-2 in lymphoid tissues draining the site of infection has implications for immunity to SARS-CoV-2. We performed tonsil biopsies under local anesthesia in 19 subjects who had recovered from SARS-CoV-2 infection 24-225 d previously. The biopsies yielded >3 million cells for flow cytometric analysis in 17 subjects. Total and SARS-CoV-2 spike-specific germinal center B cells, and T follicular helper cells, were readily detectable in human tonsils early after SARS-CoV-2 infection, as assessed by flow cytometry. Responses were higher in samples within 2 mo of infection but still detectable in some subjects out to 7 mo following infection. We conclude the tonsils are a secondary lymphoid organ that develop germinal center responses to SARS-CoV-2 infection and could play a role in the long-term development of immunity.

Balancing Statistical Power and Risk in HIV Cure Clinical Trial Design.

BACKGROUND: Analytical treatment interruptions (ATI) are pauses of antiretroviral therapy (ART) in the context of human immunodeficiency virus (HIV) cure trials. They are the gold standard in determining if interventions being tested can achieve sustained virological control in the absence of ART. However, withholding ART comes with risks and discomforts to trial participant. We used mathematical models to explore how ATI study design can be improved to maximize statistical power, while minimizing risks to participants. METHODS: Using previously observed dynamics of time to viral rebound (TVR) post-ATI, we modelled estimates for optimal sample size, frequency, and ATI duration required to detect a significant difference in the TVR between control and intervention groups. Groups were compared using a log-rank test, and analytical and stochastic techniques. RESULTS: In placebo-controlled TVR studies, 120 participants are required in each arm to detect 30% difference in frequency of viral reactivation at 80% power. There was little statistical advantage to measuring viral load more frequently than weekly, or interrupting ART beyond 5 weeks in a TVR study. CONCLUSIONS: Current TVR HIV cure studies are underpowered to detect statistically significant changes in frequency of viral reactivation. Alternate study designs can improve the statistical power of ATI trials.

Successful expanded clinic network collaboration and patient tracing for retention in HIV care.

BACKGROUND: There are more than 7,800 people living with human immunodeficiency virus (HIV) in Victoria, Australia. Crucial in maximising the individual and population level benefits from antiretroviral therapy (ART) is understanding how to achieve patient retention in care and the factors that drive it. This study was an expansion of a 2015 assessment of HIV-care retention in Victoria, which sought out to determine whether the inclusion of a broader range of HIV-healthcare sites would yield more accurate estimates of retention in HIV-care. We aimed to improve our understanding of HIV-care retention in Victoria, Australia, identify people living with HIV (PLHIV) with unknown outcomes, and attempt to re-engage PLHIV in care. METHODS: A network of 15 HIV-care sites was established in Victoria, Australia across diverse care settings which ranged from low-caseload rural sites to high-caseload metropolitan GP clinics and hospitals. Individuals who had an HIV viral load (VL) performed in both calendar years of 2016 and 2017 were classified as retained in care. Individuals with a VL test in 2016 but not in 2017 were considered to potentially have unknown outcomes as they may have been receiving care elsewhere, have disengaged from care or died. For this group, an intervention of cross-referencing partially de-identified data between healthcare sites, and contact tracing individuals who still had unknown outcomes was performed. RESULTS: For 5223 individuals considered to be retained in care across 15 healthcare sites in the study period, 49 had unconfirmed transfers of care to an alternative provider and 79 had unknown outcomes. After the intervention, the number of unconfirmed care transfers was reduced to 17 and unknown outcomes reduced to 51. These changes were largely attributed to people being reclassified as confirmed transfers of care. Retention in care estimates that did not include the patient outcome of confirmed transfer of care ranged from 76.2 to 95.8% and did not alter with the intervention. However, retention in care estimates which considered confirmed transfers and those that re-entered care at a new site as retained in care significantly increased across five of the sites with estimates ranging from 80.9 to 98.3% pre-intervention to 83.3-100% post-intervention. Individuals whose outcomes remained unknown post-intervention were more often men who have sex with men (MSM) when compared to other categories (person who injects drugs (PWID), combined PWID/MSM, men who identify as heterosexual or unknown) (74.5% vs. 53.5%, [p = 0.06]) and receiving ART at their last HIV-care visit (84.3% vs. 67.8% [p = 0.09]). CONCLUSION: This study confirmed high retention in HIV-care and low numbers of people disengaged from HIV-care in Victoria. This was demonstrated across a larger number of sites with varying models of care than a prior assessment in 2015. These data align with national and state targets aiming for 95% of PLHIV retained in HIV-care.

Monkeypox infection presenting as genital rash, Australia, May 2022.

Rapid diagnosis and whole genome sequencing confirmed a case of monkeypox in an HIV-positive individual receiving antiretroviral therapy. The patient had a normal CD4+ T-cell count and suppressed HIV viral load and presented with a genital rash in Melbourne, Australia after return from Europe in May 2022. He subsequently developed systemic illness and disseminated rash and 11 days after symptom onset, he was hospitalised to manage painful bacterial cellulitis of the genital area.

Bringing evidence from press release to the clinic in the era of COVID-19.

The urgent need to develop effective therapeutics and disseminate information from clinical studies has led to data from clinical trials being made available by alternate methods prior to peer-reviewed publication, including press releases, social media and pre-print papers. While this allows clinicians more open access to these data, a trust has to be placed with the investigators releasing these data without the availability of scientifically rigorous peer review. The examples of results from trials studying dexamethasone and hydroxychloroquine for treatment of COVID-19 have had contrasting outcomes, including the potential for significant numbers of lives saved with the early release of results from the RECOVERY trial studying dexamethasone contrasting with unsubstantiated data being presented from trials studying hydroxychloroquine. Clinicians and researchers must maintain a healthy scepticism when reviewing results prior to peer-reviewed publication, but also consider when these opportunities may allow for early implementation of potentially lifesaving interventions for people infected with COVID-19.

Acceptability, motivation and the prospect of cure for people living with HIV and their healthcare providers in HIV cure-focused treatment interruption studies.

BACKGROUND: Analytical treatment interruptions (ATI) are commonly used clinical endpoints to assess interventions aimed at curing HIV or achieving antiretroviral therapy (ART)-free HIV remission. Understanding the acceptability of ATI amongst people living with HIV (PLHIV) and their HIV healthcare providers (HHP) is limited. METHODS: Two online surveys for PLHIV and HHP assessed awareness and acceptability of ATI, and understanding of the prospect for HIV cure in the future. Responses were collected from July 2017-January 2018. A descriptive analysis was performed and similar questions across the two surveys were compared using χ squared test. RESULTS: 442 PLHIV and 144 HHP completed the survey. 105/400 (26%) PLHIV had ever interrupted ART, 8% of which were in a clinical trial. Altruistic motivations were drivers of participation of PLHIV in cure related research. 81/135 (60%) HHP would support their patients wishing to enrol in an HIV cure-focused trial, but fewer would promote and allow such participation (25% and 31% respectively). Compared to HHP, PLHIV were more likely to believe that an HIV cure would be achievable within 10 years (55% vs. 19%, p < 0.001), had less awareness of ATI (46% vs. 62%, p < 0.001) and were less likely to have had experience of either participation or enrolment in an ATI study (5% vs. 18%, p < 0.001) CONCLUSION: PLHIV were more optimistic about the potential for HIV cure. HHP had more direct experience with HIV cure-focused studies. Educational strategies are required for both groups to increase understanding around ATIs in HIV cure research but should be tailored specifically to each group.

Risk factors for unstructured treatment interruptions and association with survival in low to middle income countries.

BACKGROUND: Antiretroviral therapy (ART) treatment interruptions lead to poor clinical outcomes with unplanned or unstructured TIs (uTIs) likely to be underreported. This study describes; uTIs, their risk factors and association with survival. METHODS: Analysis of ART programmatic data from 11 countries across Asia and Africa between 2003 and 2013 where an uTI was defined as a ≥90-day patient initiated break from ART calculated from the last day the previous ART prescription would have run out until the date of the next ART prescription. Factors predicting uTI were assessed with a conditional risk-set multiple failure time-to-event model to account for repeated events per subject. Association between uTI and mortality was assessed using Cox proportional hazards, with a competing risks extension to test for the influence of lost to follow-up (LTFU). RESULTS: 40,632 patients were included from 11 countries across 33 sites (17 Africa, 16 Asia). Median duration of follow-up was 1.61 years (IQR 0.54-3.31 years), 3386 (8.3 %) patients died, and 3453 (8.5 %) were LTFU. There were 14,817 uTIs, with 10,162 (25 %) patients having more than one uTI. In the adjusted model males were at lower risk of uTI (aHR 0.94, p < 0.01, and age 20-59 was protective compared to <20 years (20-39 years aHR 0.87, p < 0.01; 40-59 years aHR 0.86, p < 0.01). Preserved immune function, as measured by higher CD4 cell count, was associated with a reduced rate of uTI compared to CD4 <200 cells/µL (CD4 200-350 cells/µL aHR 0.89, p < 0.01; CD4 >350 cells/µL aHR 0.87, p < 0.01), whereas advanced clinical disease was associated with increased uTI rate (WHO stage 3 aHR 1.10, p < 0.01; WHO stage 4 aHR 1.21, p < 0.01). There was no relationship between uTI and mortality after adjusting for disease status and considering LTFU as a competing risk. CONCLUSIONS: uTIs were frequent in people in ART programs in low-middle income countries and associated with younger age, female gender and advanced HIV. uTI did not predict survival when loss to follow-up was considered a competing risk. Further evaluation of uTI predictors and interventions to reduce their occurrence is warranted.

Long-term Virological Outcomes of First-Line Antiretroviral Therapy for HIV-1 in Low- and Middle-Income Countries: A Systematic Review and Meta-analysis.

BACKGROUND: More than 11.7 million people are currently receiving antiretroviral therapy (ART) in low- and middle-income countries (LMICs), and focused efforts are needed to ensure high levels of adherence and to minimize treatment failure. Recently, international targets have emphasized the importance of long-term virological suppression as a key measure of program performance. METHODS: We systematically reviewed publications and conference abstracts published between January 2006 and May 2013 that reported virological outcomes among human immunodeficiency virus type 1 (HIV-1)-infected adults receiving first-line ART for up to 5 years in LMICs. Summary estimates of virological suppression after 6, 12, 24, 36, 48, and 60 months of ART were analyzed using random-effects meta-analysis. Intention-to-treat (ITT) analysis assumed all participants who were lost to follow-up, died, or stopped ART as having virological failure. RESULTS: Summary estimates of virological suppression remained >80% for up to 60 months of ART for all 184 included cohorts. ITT analysis yielded 74.7% (95% confidence interval [CI], 72.2-77.2) suppression after 6 months and 61.8% (95% CI, 44.0-79.7) suppression after 48 months on ART. Switches to second-line ART were reported scarcely. CONCLUSIONS: Among individuals retained on ART, virological suppression rates during the first 5 years of ART were high (>80%) and stable. Suppression rates in ITT analysis declined during 4 years.

Targets for intervention to improve virological outcomes for patients receiving free antiretroviral therapy in Tamil Nadu, India.

Operational research to identify factors predicting poor clinical outcomes is critical to maximize patient care and prolong first-line regimens for those receiving free antiretroviral therapy (ART) in India. We sought to identify social or clinical factors amenable to intervention that predict virological outcomes after 12 months of ART. We examined a retrospective cohort of consecutive adults initiating free nonnucleoside reverse transcriptase inhibitor-based regimens. Individuals remaining in care 12 months post-ART initiation were tested for HIV viral load and surveyed to identify barriers and facilitators to adherence, and to determine clinic travel times and associated costs. Uni- and multivariate logistic regression identified factors predicting HIV viral load >200 copies/mL after 12 months of ART. Of 230 adults initiating ART, 10% of patients died, 8% transferred out, 5% were lost to follow-up, and 174/230 (76%) completed 12 months of ART, the questionnaire, and viral load testing. HIV viral load was <200 copies/mL in 140/174 (80%) patients. In multivariate models, being busy with work or caring for others (OR 2.9, p < 0.01), having clinic transport times ≥ 3 hours (OR 3.0, p = 0.02), and alcohol use (OR 4.8, p = 0.03) predicted viral load >200 copies/mL after 12 months of ART. Clinical outcomes following ART are related to programmatic factors such as prolonged travel time and individual factors such as being busy with family or using alcohol. Simple interventions that alter these factors should be evaluated to improve clinical outcomes for populations receiving free ART in similar settings.

Assessing understandings and perceptions of HIV cure among peer navigators and treatment officers in Australia.

HIV community peer navigators and treatment officers are important sources of information and guidance for people living with HIV. We conducted an anonymous online survey with members of the treatment outreach network of the Australian National Association of People Living with HIV. The survey explored understandings and acceptance of HIV cure research and was disseminated before and following an interactive workshop on HIV cure. We demonstrated an improvement in understandings of HIV cure science and increased willingness to participate in interventional clinical trials after the workshop. Concerns around interrupting HIV treatment and detrimental impacts on health were identified as barriers to participation. Altruism was a motivator to enrol in HIV cure trials, along with endorsement from a trusted member of community.

Viral suppression after 12 months of antiretroviral therapy in low- and middle-income countries: a systematic review.

OBJECTIVE: To establish estimates of viral suppression in low- and middle-income countries (LMICs) in patients who received antiretroviral therapy (ART) for human immunodeficiency virus (HIV) infection. METHODS: Data on viral suppression after 12 months of ART in LMICs were collected from articles published in 2003 to 2011 and from abstracts of conferences held between 2009 and 2011. Pooled proportions for on-treatment and intention-to-treat populations were used as summary estimates. Random-effects models were used for heterogeneous groups of studies (I (2) > 75%). FINDINGS: Overall, 49 studies covering 48 cohorts and 30 016 individuals met the inclusion criteria. With thresholds for suppression between 300 and 500 copies of viral ribonucleic acid (RNA) per ml of plasma, 84.3% (95% confidence interval, CI: 80.4-87.9) of the pooled on-treatment population and 70.5% (95% CI: 65.2-75.6) of the intention-to-treat population showed suppression. Use of different viral RNA thresholds changed the proportions showing suppression: to 84% and 76% of the on-treatment population with thresholds set above 300 and at or below 200 RNA copies per ml, respectively, and to 78%, 71% and 63% of the intention-to-treat population at thresholds set at 1000, 300 to 500, and 200 or fewer copies per ml, respectively. CONCLUSION: The pooled estimates of viral suppression recorded after 12 months of ART in LMICs provide benchmarks that other ART programmes can use to set realistic goals and perform predictive modelling. Evidence from this review suggests that the current international target - i.e. viral suppression in > 70% of the intention-to-treat population, with a threshold of 1000 copies per ml - should be revised upwards.

Pharmacy and self-report adherence measures to predict virological outcomes for patients on free antiretroviral therapy in Tamil Nadu, India.

Over 480,000 individuals receive free antiretroviral therapy (ART) in India yet data associating ART adherence with HIV viral load for populations exclusively receiving free ART are not available. Additionally estimates of adherence using pharmacy data on ART pick-up are not available for any population in India. After 12-months ART we found self-reported estimates of adherence were not associated with HIV viral load. Individuals with <100% adherence using pharmacy data predicted HIV viral load, and estimates combining pharmacy data and self-report were also predictive. Pharmacy adherence measures proved a feasible method to estimate adherence in India and appear more predictive of virological outcomes than self-report. Predictive adherence measures identified in this study warrant further investigation in populations receiving free ART in India to allow for identification of individuals at risk of virological failure and in need of adherence support.

Immunogenicity, effectiveness, and safety of SARS-CoV-2 vaccination in people with HIV.

OBJECTIVES: People with HIV (PWH) experience a greater risk of morbidity and mortality following COVID-19 infection, and poorer immunological responses to several vaccines. We explored existing evidence regarding the immunogenicity, effectiveness, and safety of SARS-CoV-2 vaccines in PWH compared with controls. METHODS: We conducted a systematic search of electronic databases from January 2020 until June 2022, in addition to conference databases, to identify studies comparing clinical, immunogenicity, and safety in PWH and controls. We compared results between those with low (<350 cells/µl) and high (>350 cells/µl) CD4 + T-cell counts where possible. We performed a meta-analysis of seroconversion and neutralization responses to calculate a pooled risk ratio as the measure of effect. RESULTS: We identified 30 studies, including four reporting clinical effectiveness, 27 immunogenicity, and 12 reporting safety outcomes. PWH were 3% [risk ratio 0.97, 95% confidence interval (95% CI) 0.95-0.99] less likely to seroconvert and 5% less likely to demonstrate neutralization responses (risk ratio 0.95, 95% CI 0.91-0.99) following a primary vaccine schedule. Having a CD4 + T-cell count less than 350 cells/µl (risk ratio 0.91, 95% CI 0.83-0.99) compared with a CD4 + T-cell count more than 350 cells/µl, and receipt of a non-mRNA vaccine in PWH compared with controls (risk ratio 0.86, 95% CI 0.77-0.96) were associated with reduced seroconversion. Two studies reported worse clinical outcomes in PWH. CONCLUSION: Although vaccines appear well tolerated in PWH, this group experience poorer immunological responses following vaccination than controls, particularly with non-mRNA vaccines and low CD4 + T-cell counts. PWH should be prioritized for mRNA COVID-19 vaccines, especially PWH with more advanced immunodeficiency.

Soluble immune checkpoints as correlates for HIV persistence and T cell function in people with HIV on antiretroviral therapy.

Introduction: In people with HIV (PWH) both off and on antiretroviral therapy (ART), the expression of immune checkpoint (IC) proteins is elevated on the surface of total and HIV-specific T-cells, indicating T-cell exhaustion. Soluble IC proteins and their ligands can also be detected in plasma, but have not been systematically examined in PWH. Since T-cell exhaustion is associated with HIV persistence on ART, we aimed to determine if soluble IC proteins and their ligands also correlated with the size of the HIV reservoir and HIV-specific T-cell function. Methods: We used multiplex bead-based immunoassay to quantify soluble programmed cell death protein 1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), lymphocyte activation gene-3 (LAG-3), T cell immunoglobulin domain and mucin domain 3 (TIM-3), PD-1 Ligand 1 (PD-L1) and PD-1 Ligand 2 (PD-L2) in plasma from PWH off ART (n=20), on suppressive ART (n=75) and uninfected controls (n=20). We also quantified expression of membrane-bound IC and frequencies of functional T-cells to Gag and Nef peptide stimulation on CD4+ and CD8+ T-cells using flow cytometry. The HIV reservoir was quantified in circulating CD4+ T-cells using qPCR for total and integrated HIV DNA, cell-associated unspliced HIV RNA and 2LTR circles. Results: Soluble (s) PD-L2 level was higher in PWH off and on ART compared to uninfected controls. Higher levels of sPD-L2 correlated with lower levels of HIV total DNA and higher frequencies of gag-specific CD8+ T-cells expressing CD107a, IFNγ or TNFα. In contrast, the concentration of sLAG-3 was similar in uninfected individuals and PWH on ART, but was significantly elevated in PWH off ART. Higher levels of sLAG-3 correlated with higher levels of HIV total and integrated DNA, and lower frequency of gag-specific CD4+ T cells expressing CD107a. Similar to sLAG-3, levels of sPD-1 were elevated in PWH off ART and normalized in PWH on ART. sPD-1 was positively correlated with the frequency of gag-specific CD4+ T cells expressing TNF-a and the expression of membrane-bound PD-1 on total CD8+ T-cells in PWH on ART. Discussion: Plasma soluble IC proteins and their ligands correlate with markers of the HIV reservoir and HIV-specific T-cell function and should be investigated further in in large population-based studies of the HIV reservoir or cure interventions in PWH on ART.