HU CFAR Annual Scientific Symposium & Poster Session: Immunodysregulation in Human Diseases

Dr. Engelman is a Professor of Medicine at HMS with an affiliated appointment in the Department of Microbiology. Dr. Engelman’s research focuses on virus-host interactions during the early phase of HIV replication, including integration. His research helped to explain the mechanism of action of integrase strand transfer inhibitors, which are widely used in the clinic, as well as mechanisms of drug resistance. Dr Engelman’s R01 grant on HIV integration has twice been awarded R37 MERIT status. Dr Engelman co-directs a T32 Training Program on HIV Related Research at DFCI, and has served on over 50 NIH Study Sections, including 17 as Chair.

Dr. Juelg is an Infectious Diseases physician at MGH where he directs the Infectious Disease Clinical Research Unit and serves as the associate director of the HU CFAR clinical core. He is also a member of the Ragon Institute where he maintains his lab. Dr Juelg’ s work focuses on identifying, developing, and evaluating immunotherapeutic interventions including therapeutic vaccination and monoclonal antibodies with an emphasis on translating discoveries from bench to bedside.

Dr. Tsibris is an Assistant Professor of Medicine at HMS and the co-director of the HU CFAR Developmental and Mentoring Core. His research focusses on the biology and dynamics of the HIV reservoir in blood and tissues.

Dr. Gaiha is an Assistant Professor of Medicine at HMS who is developing novel T cell vaccine candidates against HIV-1 and other viral pathogens that target epitopes derived from structurally-constrained regions of viral proteins.

Dr. Lauer is an Associate Professor of Medicine at HMS who studies the T cell response against hepatitis B and C viruses, with or without HIV co-infection. His focus is on supporting the development of HBV immunotherapies and prophylactic vaccines for HCV, but also on using these infections as model diseases to understand basic mechanisms of human immunology.

Dr. Reuben Harris is an investigator of the Howard Hughes Medical Institute (HHMI) and chair of the Biochemistry and Structural Biology department at University of Texas Health San Antonio. He received his B.S. (1993) and Ph.D. (1997) degrees from the University of Alberta and performed postdoctoral work at Baylor College of Medicine (1997-1998), Yale University (1998), and Cambridge University (1998-2003). He joined the University of Minnesota as an Assistant Professor in 2003 and was promoted to Associate Professor with Tenure in 2008 and to Full Professor in 2013.

In 2022, Dr. Harris moved his laboratory to University of Texas Health San Antonio. Dr. Harris has received numerous grants and awards, including a Searle Scholarship, membership to the American Academy of Microbiology, NIH Merit Award, a Distinguished McKnight University Professorship, and the KT Jeang Prize. In 2015, he was also appointed as a Howard Hughes Medical Institute Investigator. Dr. Harris is an Associate Editor for Science Advances and an Editorial Board Member for Journal of Biological Chemistry, Journal of Virology, and Cancer Research. He has published over 200 manuscripts, contributed to 13 patent applications, and co-founded a cancer therapeutics company. Dr. Harris’s scientific passion is elucidating mechanisms of mutation and establishing relevance to human biology and disease. As a doctoral student, he discovered a novel recombination-dependent mutation process operative in stationary-phase bacteria with implications for antibiotic resistance and microbial evolution. As a postdoctoral fellow, he helped solve an immunology Rosetta stone by discovering the DNA cytosine deaminase activity of AID and proposing a DNA deamination model for antibody gene diversification. Also as a postdoctoral fellow, he discovered the DNA cytosine deaminase activity of several APOBEC family members and, during the transition to faculty, elucidated a new mechanism of antiviral immunity by demonstrating APOBEC3G-catalyzed retroviral cDNA hypermutation. As a Principal Investigator, Dr. Harris has become known for his work on APOBEC enzymes in antiviral immunity. This body of work has shed light on fundamental mechanisms of antiviral immunity and yielded new strategies for drug development. In recent years, Dr. Harris’s virology studies have also enabled a major breakthrough in cancer research. His group found that APOBEC3 enzymes are responsible for a large proportion of mutations in breast, head/neck, lung, bladder, cervical, and other cancers. Independent work has confirmed these results and indicated that “APOBEC mutagenesis” far exceeds most other sources of mutations in cancer, including those attributable to smoking and UV rays. These breakthroughs have created new opportunities for cancer diagnosis, prognosis, and treatment by targeting tumor evolvability.

One of his other passions is training and mentoring the next-generation of scientists, and he has trained a multitude of students and postdoctoral fellows over his career. Importantly, all of these trainees have succeeded in obtaining independent positions in academia or industry.

Related Links: NIH PubMed- https://pubmed.ncbi.nlm.nih.gov/?term=Reuben%20S.%20Harris%20&sort=date

Dr. Susan Moir obtained her Ph.D. in Immunology and Microbiology from the Université Laval, Québec, Canada in 1996. Shortly after, she began a postdoctoral fellowship on HIV immunopathogenesis with Dr. Anthony Fauci in the Laboratory of Immunoregulation (LIR) of the National Institute of Allergy and Infectious Diseases at NIH. In 2015, Dr. Moir was selected as a Stadtman Tenure Track Investigator in the LIR and became a tenured Senior Investigator and Chief of the B-Cell Immunology Section in 2020. The primary focus of her research program is to study the role of B cells in HIV pathogenesis and apply this knowledge to the study of other infectious and non-infectious diseases that affect B cells.

Dr. Gordon Freeman is in the Department of Medical Oncology at Dana-Farber Cancer Institute and is Professor of Medicine at Harvard Medical School.  Dr. Freeman earned his BA in Biochemistry and Molecular Biology, and PhD in Microbiology and Molecular Genetics from Harvard University. 

Dr. Freeman’s research identified the major pathways that control the immune response by inhibiting T cell activation (PD-L1/PD-1 and B7-2/CTLA-4) or stimulating T cell activation (B7-2/CD28). In 2000, Dr. Freeman discovered PD-L1 and PD-L2, and showed they were ligands for PD-1, thus defining the PD-1 pathway and the drug target: block the interaction.  He showed the function of PD-1 was to inhibit immune responses and that blockade enhanced immune responses. He showed that PD-L1 is highly expressed on many solid tumors such as breast and lung, as well as some hematologic malignancies and allows these tumors to inhibit immune attack.  This work provided the critical translational insights for development of a successful strategy for cancer immunotherapy – blocking the PD-1-ligand interaction.

Dr. Freeman has published over 400 scientific papers and holds over 90 US patents on immunotherapies. He is a member of the National Academy of Sciences, the National Academy of Inventors, the American Academy of Arts and Sciences and is a Fellow of the AACR and SITC Academies. He has received numerous awards including the 2014 William B. Coley Award for Distinguished Research in Tumor Immunology, 2017 Warren Alpert Foundation award, and the 2020 Richard Smalley, MD, memorial award for his work that led to development of PD-L1/PD-1 pathway blockade for cancer immunotherapy.

Dr. Cigall Kadoch is an academic investigator as well as entrepreneur in the biomedical sciences. She is currently an Associate Professor of Pediatric Oncology at the Dana-Farber Cancer Institute and Harvard Medical School, Institute Member and Epigenomics Program Co-Director at the Broad Institute, and an Investigator of the Howard Hughes Medical Institute (HHMI). Dr. Kadoch established her independent laboratory in 2014, at age 28, immediately following completion of her Ph.D. studies at Stanford University. She is a leading expert in chromatin and gene regulation and is internationally recognized for her groundbreaking studies in these areas leading to recent clinical translation. Specifically, her laboratory studies the structure and function of nuclear protein complexes called chromatin remodeling complexes that govern DNA architecture and gene expression, perturbations in which are implicated in over 50% of human cancers, including their roles as oncogenic drivers of several aggressive pediatric cancers. Based on her seminal work, Dr. Kadoch is also the Scientific Founder of Foghorn Therapeutics (NASDAQ: FHTX), a company advancing a new class of medicines in oncology and beyond.

Dr. E. John Wherry is the Barbara and Richard Schiffrin President’s Distinguished Professor, Chair of the Department of Systems Pharmacology and Translational Therapeutics in the Perelman School of Medicine and Director of the UPenn Institute for Immunology.  Dr. Wherry received his Ph.D. at Thomas Jefferson University in 2000 and performed postdoctoral research at Emory University from 2000-2004. Dr. Wherry has received numerous honors including the Distinguished Alumni award from the Thomas Jefferson University, the Cancer Research Institute’s Frederick W. Alt Award for New Discoveries in Immunology, the Stanley N. Cohen Biomedical Research Award from the University of Pennsylvania Perelman School of Medicine and was inducted as an AAAS Fellow in 2021.  As of November 2022, Dr. Wherry has over 300 publications, an H-Index of 126, and his publications have been cited over 86,000 times. 

Dr. Wherry helped pioneered the field of T cell exhaustion, the mechanisms by which T cell responses are attenuated during chronic infections and cancer. He helped identify the role of the “checkpoint” molecule PD-1 and others for reinvigoration of exhausted T cells in cancer. Dr. Wherry’s work has defined the underlying molecular and epigenetic mechanisms of exhausted T cells. His laboratory has also recently focused on applying systems immunology approaches to define Immune Health patients across a spectrum of diseases. In 2020-2021, Dr. Wherry’s laboratory focused considerable efforts on the immunology of COVID-19 and SARS-CoV-2 vaccination including establishing a new Immune Health Project to interrogate and use immune features to identify novel treatment opportunities. 

Dr. Shiv Pillai is a Professor of Medicine and Health Sciences and Technology at Harvard Medical School.  He studied medicine at Christian Medical College in Vellore, completed his PhD with Bimal Bacchawat, and was a Postdoctoral Fellow with David Baltimore at the Whitehead institute and MIT. He is the Program Director of an NIH-funded Basic Autoimmune Center of Excellence at Massachusetts General Hospital, the Director of the Harvard Immunology PhD and Masters’ in Medical Sciences Programs and director of the HMS-HST MD student research program. He was elected as a Distinguished Fellow of the American Association of Immunology in 2023.   He has been the recipient of a number of teaching awards at Harvard including the Irving M. London Award for Teaching, the Thomas McMahon Mentoring Award, and has been listed on Harvard Crimson’s Professors of the Year. 

Dr. Pillai coined the term “surrogate light chains” for proteins that he identified as part of the pre-B receptor, that drives early B cell development. His laboratory postulated and provided evidence for the first ligand-independent signaling model during lymphocyte development and showed that BTK, the product of the gene mutated in X-linked agammaglobulinemia, is functionally linked to the pre-B receptor and the B cell receptor. Btk inhibitors are now widely used in lymphoid malignancies and autoimmunity. Apart from the pro-B to pre-B cell transition, his group developed the concept of the follicular versus marginal zone B lymphoid cell-fate decision, has identified a metabolic transitional to follicular B cell switch that is blocked in human common variable immunodeficiency, and identified a block in T follicular helper cell development in COVID-19 that prevents the formation of germinal centers. His laboratory actively studies the dysfunctional extra-follicular B cell response and its links to cytotoxic CD4+T cells in autoimmune and inflammatory diseases that include IgG4-related disease, systemic sclerosis, fibrosing mediastinitis and COVID-19. The group also studies B cell tolerance defects in patients with single gene mutations in CTLA4, NFKB1, PI3KCD and more and explores the biology of B cell memory.

Dr. Pillai is the author of a monograph “Lymphocyte Development” and co-author with Abul Abbas and Andrew Lichtman of two widely used textbooks of immunology.

Upasana completed her doctoral studies in Cancer signaling and Immunology from a collaborative international PhD program between Max Delbruck Centre of Molecular Medicine, Germany and Hebrew University of Jerusalem, Israel securing the Marie Curie Doctoral Fellowship. She joined the Kwon Lab to study Immunity at the epithelial barriers in the context of HIV. Here, she established patient derived ex vivo 3-dimensional mini gut organoids complexed with matched patient autologous immune cells to study HIV-associated intestinal barrier integrity in people with HIV. Recently she was awarded the K99/R00 Pathway to Independence award from NIDDK/NIH.

Kathryn is an Infectious Diseases physician at Brigham and Women’s Hospital and Massachusetts General Hospital, and a postdoctoral fellow at the Ragon Institute and Harvard School of Public Health, mentored by Sarah Fortune, MD.  Her research is focused on the humoral immune response to Tuberculosis and HIV/TB co-infection, and understanding how antibody function can be leveraged both for biomarker development and in the rational design of vaccines and therapeutics in tuberculosis and more broadly across other infectious diseases.

Charles is a research technician in the lab of Dr. Gaurav Gaiha at the Ragon Institute of Massachusetts General Hospital, MIT, and Harvard. There he studies CD8+ T cell-mediated immune mechanisms in HIV controllers. Prior to joining the Gaiha lab, he graduated summa cum laude from Northeastern University with his B.S. in Cell and Molecular Biology and minors in Mathematics and Data Science, where he received the President’s Award and was inducted as a member of the Tri Beta Biological Honors Society. As an undergraduate, he conducted research in the lab of Dr. Jonathan Li at Brigham and Women’s Hospital, where he studied SARS-CoV-2 mutational patterns in cases of persistent SARS-CoV-2 infection and in response to monoclonal antibody therapy.

Malika holds an engineering degree in computer science and a Ph.D. in Computational Biology and Bioinformatics. She is a principal investigator and the head of the Computational Genomics and Epigenetics research group at the Center for Virology and Vaccine Research. Her research focus includes implementing machine learning and AI-based tools for multi-omics data analysis and integration to investigate the host-virus interface using transcriptomics, proteomics, and metabolomics profiling; Investigating the host early innate responses to vaccines to identify host early factors that impede vaccine response efficacy and durability; and leveraging AI for the development of the next generation of mRNA vaccines. She is also exploring disease pathogenesis through epigenetic profiling, with a keen interest in decoding the epigenetic determinants of HIV integration and persistence as well as investigating potential correlates of HIV rebound after ATI.

TITLE: Molecular mechanisms underlying the persistent viral RNA in lymphoid tissues in SIV-infected ART-suppressed
rhesus macaques.

ABSTRACT: Tissue viral reservoir is rapidly and broadly seeded early during acute SIV infection in rhesus macaques [1, 2]. In a recent
report [3], a comprehensive necroscopic analysis of the SIV viral reservoir in multiple lymphoid and non-lymphoid tissues from SIV-infected rhesus macaques suppressed with ART for one year. ART was initiated during both acute (Early ART group) and chronic (Late ART group) infection for a comprehensive evaluation of the viral reservoir in both settings. In this study [3], we observed that animals were fully suppressed on ART at the time of necropsy. We quantitated cell-associated viral DNA and viral RNA from 24 different tissue sites using cell-associated gag qPCR and qRT-PCR, respectively and we showed that viral DNA is observed broadly in multiple tissues and is comparable in animals that had initiated ART at week 1 or week 52 of infection. In contrast, we observed that viral RNA persists in lymph nodes and other secondary lymphoid tissues despite 1–3 years of suppressive ART [3]. Therefore, our study suggested viral latency does not appear to be universally transcriptionally silent across all analyzed tissues, and that the reservoir may include a spectrum of latency depths as previously suggested by several studies [4-6]. Additionally, our study suggested the presence of a small fraction of cells that remain transcriptionally active in LN despite ART suppression. Herein, we investigated the molecular mechanisms underlying the persistent viral RNA in lymph nodes compared to other anatomical compartments. We performed bulk transcriptomic profiling of lymphoid and non-lymphoid tissues at the time of necropsy from 19 macaques, including (Axillary LN, Inguinal LN, Mesenteric LN, Ileocecal LN, Pararectal LN, Sacral LN, Pelvic LN), Spleen, Bonne Marrow, Mid-Vagina, Cervix, Brain tissues (FC and BG), Duodenum, Distal Jejunum, Ileum, Cecum, Colon, and Rectum. Plasma was collected for proteomics profiling before ART initiation and at three-time intervals on ART: early time points on ART (weeks 0,1 and 2), intermediate time points on ART (weeks 12, 16, and 20), and late time points on ART (weeks 32, 36, and 40) before necropsy. We identified key cytokines and immune cell states that correlate with persistent viral RNA in LN tissues suggesting potential host mechanisms associated with persistent viral transcription and immune activation despite years of suppressive ART.

References:

1. Whitney, J.B., et al., Rapid seeding of the viral reservoir prior to SIV viraemia in rhesus monkeys. Nature, 2014.
   512(7512): p. 74-7.
2. Whitney, J.B., et al., Prevention of SIVmac251 reservoir seeding in rhesus monkeys by early antiretroviral
   therapy. Nat Commun, 2018. 9(1): p. 5429.
3. Cadena, A.M., et al., Persistence of viral RNA in lymph nodes in ART-suppressed SIV/SHIV-infected Rhesus
   Macaques. Nat Commun, 2021. 12(1): p. 1474.
4. Bradley, T., et al., Single-Cell Analysis of Quiescent HIV Infection Reveals Host Transcriptional Profiles that
   Regulate Proviral Latency. Cell Rep, 2018. 25(1): p. 107-117 e3.
5. Golumbeanu, M., et al., Single-Cell RNA-Seq Reveals Transcriptional Heterogeneity in Latent and Reactivated
   HIV-Infected Cells. Cell Rep, 2018. 23(4): p. 942-950.
6. Spina, C.A., et al., An in-depth comparison of latent HIV-1 reactivation in multiple cell model systems and
   resting CD4+ T cells from aviremic patients. PLoS Pathog, 2013. 9(12): p. e1003834.

Kathryn is an Infectious Diseases physician at Brigham and Women’s Hospital and Massachusetts General Hospital, and a postdoctoral fellow at the Ragon Institute and Harvard School of Public Health, mentored by Sarah Fortune, MD.  Her research is focused on the humoral immune response to Tuberculosis and HIV/TB co-infection, and understanding how antibody function can be leveraged both for biomarker development and in the rational design of vaccines and therapeutics in tuberculosis and more broadly across other infectious diseases.

Title: Serological profiling identifies humoral shifts in immune tone in HIV/TB co-infection

ABSTRACT: HIV-1 co-infection is the most important risk factor for development of active tuberculosis, but the mechanistic basis of this remains unclear. Recent work has demonstrated that Mtb-specific antibodies can define states of Mtb exposure and disease and serve as an independent measure of exposure from CD4 dependent interferon-gamma release assays (IGRA) for detection of latent infection in HIV-infected individuals (Lu et al., 2016; Fischinger et al., 2021).  Additionally, emerging data point to a role for antibodies in the control of Mtb (Achkar et al., 2015; Irvine et al., 2021). To evaluate and prior work has demonstrated TB-specific antibody profile differences in HIV-positive individuals with active TB (Bell and Noursadeghi, 2018; van Woudenbergh et al., 2020).  To evaluate the in-depth humoral signatures associated with Mtb disease in immunosuppressed and endemically TB-exposed populations, . However, the performance of these Mtb-specific antibody signatures in individuals with HIV remains unclear. Prior work has demonstrated TB-specific antibody profile differences in HIV-positive individuals with active TB (Bell and Noursadeghi, 2018; van Woudenbergh et al., 2020). Here We performed humoral immune profiling of Mtb antibody responses from a Cambodian cohort of 67 ART-naïve HIV-infected individuals (CD4+ T cell counts ≤200/mm³) with (n=40) or without (n=27) newly diagnosed smear-positive active tuberculosis, and 40 HIV-uninfected individuals, including individuals with active TB (n=20), latent TB (n=10), or without evidence of TB infection (n=10)(Blanc et al., 2011).  Within this especially immunosuppressed and endemically TB-exposed population, an in-depth understanding of humoral signatures of HIV+ individuals without TB may be of particular importance. We utilized a multiplexed Luminex-based platform targeting a broad range of Mtb antigens to profile antibody changes in Mtb-specific IgM, IgA₁, and IgG isotype and subclasses, Fc gamma receptor binding, and Fc glycosylation state. We then analyzed Mtb-specific Fc-mediated effector functions including antibody-dependent monocyte phagocytosis and neutrophil phagocytosis, and antibody-dependent complement activation. Univariate and multivariate analyses revealed significant Mtb-specific isotype and subclass differences and antigen-specific enrichment in antibody glycosylation and Fc receptor binding in HIV-positive individuals. HIV-infected individuals also exhibited a shift in MTb-specific Fc-dependent effector function towards enhance neutrophil phagocytosis. Among HIV-infected individuals, Mtb-specific antibodies differentially correlated with viral load and CD4 count in those with active TB compared with latent TB. Taken together, these data reveal that HIV infection is associated with a broad shift in TB-specific humoral immune tone, which may be useful as clinically relevant biomarkers to detect Mtb exposure in this highly vulnerable population.

AUTHORS: Kathryn A Bowman (1,2), Leela RL Davies (1,2), Galit Alter (1,3), Anne Goldfeld (4), Sarah Fortune (5)

¹Ragon Institute of MGH, MIT and Harvard, Boston, MA, USA; ²Brigham and Women’s Hospital, Boston, MA, USA; ³Harvard Medical School, Boston, MA; ⁴Program in Cellular and Molecular Medicine, Boston Children’s Hospital, Harvard Medical School, Boston, Massachusetts, USA; ⁵Department of Immunology and Infectious Diseases, Harvard T. H. Chan School of Public Health, Boston, MA, USA.

REFERENCES:

1. Lu LL, Chung AW, Rosebrock TR, Ghebremichael M, Yu WH, Grace PS, Schoen MK, Tafesse F, Martin C, Leung V, Mahan AE, Sips M, Kumar MP, Tedesco J, Robinson H, Tkachenko E, Draghi M, Freedberg KJ, Streeck H, Suscovich TJ, Lauffenburger DA, Restrepo BI, Day C, Fortune SM, Alter G. A Functional Role for Antibodies in Tuberculosis. Cell. 2016 Oct 6;167(2):433-443.e14. doi: 10.1016/j.cell.2016.08.072. Epub 2016 Sep 22. PMID: 27667685; PMCID: PMC5526202.
2. Fischinger S, Cizmeci D, Shin S, Davies L, Grace PS, Sivro A, Yende-Zuma N, Streeck H, Fortune SM, Lauffenburger DA, Naidoo K, Alter G. A Mycobacterium tuberculosis Specific IgG3 Signature of Recurrent Tuberculosis. Front Immunol. 2021 Sep 22;12:729186. doi: 10.3389/fimmu.2021.729186. PMID: 34630406; PMCID: PMC8493041.
3. Achkar JM, Chan J, Casadevall A. B cells and antibodies in the defense against Mycobacterium tuberculosis infection. Immunol Rev. 2015 Mar;264(1):167-81. doi: 10.1111/imr.12276. PMID: 25703559; PMCID: PMC4629253.
4. Irvine EB, O’Neil A, Darrah PA, Shin S, Choudhary A, Li W, Honnen W, Mehra S, Kaushal D, Gideon HP, Flynn JL, Roederer M, Seder RA, Pinter A, Fortune S, Alter G. Robust IgM responses following intravenous vaccination with Bacille Calmette-Guérin associate with prevention of Mycobacterium tuberculosis infection in macaques. Nat Immunol. 2021 Dec;22(12):1515-1523. doi: 10.1038/s41590-021-01066-1. Epub 2021 Nov 22. PMID: 34811542; PMCID: PMC8642241.
5. Bell LCK, Noursadeghi M. Pathogenesis of HIV-1 and Mycobacterium tuberculosis co-infection. Nat Rev Microbiol. 2018 Feb;16(2):80-90. doi: 10.1038/nrmicro.2017.128. Epub 2017 Nov 7. PMID: 29109555.
6. van Woudenbergh E, Irvine EB, Davies L, de Kock M, Hanekom WA, Day CL, Fortune S, Alter G. HIV Is Associated with Modified Humoral Immune Responses in the Setting of HIV/TB Coinfection. mSphere. 2020 May 20;5(3):e00104-20. doi: 10.1128/mSphere.00104-20. PMID: 32434838; PMCID: PMC7380575.
7. Blanc FX, Sok T, Laureillard D, Borand L, Rekacewicz C, Nerrienet E, Madec Y, Marcy O, Chan S, Prak N, Kim C, Lak KK, Hak C, Dim B, Sin CI, Sun S, Guillard B, Sar B, Vong S, Fernandez M, Fox L, Delfraissy JF, Goldfeld AE; CAMELIA (ANRS 1295–CIPRA KH001) Study Team. Earlier versus later start of antiretroviral therapy in HIV-infected adults with tuberculosis. N Engl J Med. 2011 Oct 20;365(16):1471-81. doi: 10.1056/NEJMoa1013911. PMID: 22010913; PMCID: PMC4879711.

Elise is from Saco, Maine and earned her Bachelor’s degree in Microbiology from the University of Vermont. She has been working in the Tsibris lab since 2022, investigating LTR sequence diversity in the context of HIV latency reversal. She aspires to study bioinformatics and the gut microbiome in graduate school. In her free time, Elise loves to rock climb, read, and bake sourdough.

Title: The within-individual proviral HIV-2 long terminal repeat reservoir is genetically diverse

Background: The long terminal repeat (LTR) is a retroviral genetic element that coordinates transcription factor binding and virus transcription. In HIV infection, the LTR sequence may be highly conserved. Recently, the HIV-2 LTR was found to have greater inter-participant genetic variability than was previously appreciated. The within-participant diversity of the HIV-2 LTR, and its effect on predicted transcription factor (TF) binding, is unknown.

Methods: We performed single genome amplifications (SGA) of proviral HIV-2 5’ and 3’ LTRs from cryopreserved peripheral blood mononuclear cells (PBMC) isolated from ten aviremic participants with HIV-2; consensus primers were designed to minimize LTR sequence bias. To determine the proportion of APOBEC-induced hypermutated sequences, we used the LANL hypermut tool. To assess sequence variability, mean pairwise distances (MPD) for each participant were calculated, relative to the HIV-2 ROD reference standard and to a consensus HIV-2 LTR sequence for each participant; significance was assessed by t-test. Shannon entropies were calculated across the LTR length. LTR SGAs were aligned to a reference HIV-2 5’ LTR and phylogenetic analyses were performed. We used a core set of TF profiles to predict the number and types of TF binding motifs present in the HIV-2 LTR SGAs we generated.

Results: A total of 427 HIV-2 LTR SGA amplicons (SGAs) were sequenced and analyzed. APOBEC-induced hypermutated sequences comprised 5.8% (95% CI [1.9, 9.78]) and 10.6% (95% CI [4.9, 16.2]) of 5’ and 3’ LTRs sequences, respectively. An unrooted consensus
neighbor-joining tree using a Tamura-Nei distance model and 1,000 bootstrapped replicates identified nine HIV-2 group A and one HIV-2 group B infections that clustered distinctly. The MPD was similar between a participant-specific consensus 5’ and 3’ LTR sequence, 10.9% vs 14.6% (p=0.70), when compared to the HIV-2 ROD reference. The MPD between participant SGAs and the corresponding consensus LTR were also similar, 1.9% vs 1.5% (p=0.23), and were significantly lower than the MPD to the ROD reference (p<0.002). Shannon entropy quantifications identified the greatest variability in the U3 LTR region, proximal to the transactivation response (TAR) element, when compared to the R and U5 regions. The greatest absolute number of predicted TF binding sites, and their variance, across HIV-2 LTR SGAs was greatest for zinc finger, NF-κB REL, and ETS TF motifs.

Conclusions: Contrary to conventional wisdom, the proviral HIV-2 LTR reservoir is genetically variable at the inter- and intra-individual levels. This variability was greatest proximal to the TAR region and impacted the number and types of predicted TF motifs. Future work will compare proviral HIV-2 and HIV-1 LTR diversity and the impact this diversity may have on transcription factor binding and LTR activity in response to latency reversal.

AUTHORS: Elise Courtney (1), Jennifer Bouzy (1), Tyler Lilie (1), Stacey M. Gaudreau (2), Susan Cu-Uvin (2,3), Trevor Siggers (4), Athe Tsibris (1,5).

(1) Brigham and Women’s Hospital, Boston, MA, USA, (2) The Miriam Hospital, Providence, RI, USA, (3) Brown University, Providence, RI, USA, (4) Bioinformatics Program, Department of Biology, and Biological Design Center, Boston University, Boston, MA, USA, (5) Harvard Medical School, Boston, MA, USA.

Charles is a research technician in the lab of Dr. Gaurav Gaiha at the Ragon Institute of Massachusetts General Hospital, MIT, and Harvard. There he studies CD8+ T cell-mediated immune mechanisms in HIV controllers. Prior to joining the Gaiha lab, he graduated summa cum laude from Northeastern University with his B.S. in Cell and Molecular Biology and minors in Mathematics and Data Science, where he received the President’s Award and was inducted as a member of the Tri Beta Biological Honors Society. As an undergraduate, he conducted research in the lab of Dr. Jonathan Li at Brigham and Women’s Hospital, where he studied SARS-CoV-2 mutational patterns in cases of persistent SARS-CoV-2 infection and in response to monoclonal antibody therapy.

Title: HIV Post-treatment Controllers Show Enhanced CD8+ T Cell Proliferative Capacity After ART Cessation

Background: HIV post-treatment controllers (PTCs) are individuals who control viremia to low or undetectable levels following ART discontinuation. While functional CD8+ T cell responses have been shown to be a defining feature of spontaneous (“elite”) control of
HIV, characterization of features of CD8+ T cell responses in PTCs has been limited. Here, we aimed to characterize the frequency, functionality, and specificity of CD8+ T cells in PTCs following ART cessation.

Methods: We evaluated HIV-specific CD8+ T cell responses of 7 PTCs alongside a group of 8 spontaneous controllers who had received and later discontinued ART (treated controllers, TC) and 11 non-controllers (NC) during the post-treatment interruption (TI)
period. Both PTCs and TCs were defined as having plasma viral loads <10,000 copies/mL for at least 52 weeks post-TI; individuals meeting these criteria were defined as TC if their viral load immediately preceding ART initiation was <10,000 copies/mL, and PTC if
otherwise. Ex-vivo CD8+ T cell reactivity was evaluated by IFN-γ ELISpot and functional proliferative capacity through a six-day CFSE-based proliferation assay with individual optimal clade B epitopes matched to each participant’s HLA haplotype.

Results: HIV-specific CD8+ T cell reactivity post-TI as measured by ELISpot was significantly lower in PTCs compared to TCs (P<0.01), with no significant differences observed between TCs and NCs. Of note, median plasma viral loads at time of analysis
were 50, 638, and 36200 copies/mL in the PTCs, TCs, and NCs respectively. In contrast, both PTCs and TCs had significantly stronger proliferative responses to HIV epitopes than NCs (P<0.01 and P<0.05, respectively). Additionally, we calculated the capacity for
antigen-specific CD8+ T cells to expand after antigen exposure by normalizing the magnitude of proliferation to direct ex vivo ELISpot magnitude and observed an even substantially higher proliferative capacity in both PTCs and TCs compared to NCs (P<0.001 and P<0.05, respectively).

Conclusions: Compared to NCs, PTCs exhibited a distinct HIV-specific CD8+ T cell profile, with less direct ex vivo reactivity but greater proliferative capacity following antigen exposure. These findings warrant further investigation into the mechanisms by which
highly expandible antigen-specific CD8+ T cell populations are formed and maintained, and lend further support to therapeutic vaccine strategies aimed at inducing T cell responses that retain high proliferative capacity.

Authors: Charles R. Crain (1*), Anusha Nathan (1,2), Behzad Etemad (3), Prerana Shrestha (3), Rachel L. Rutishauser (4), Steven G. Deeks (4), Jonathan Z. Li (3), Gaurav D. Gaiha (1,5).

1. Ragon Institute of Mass General, MIT, and Harvard
2. Program in Health Sciences and Technology, Harvard Medical School and MIT
3. Brigham and Women’s Hospital, Harvard Medical School
4. University of California, San Francisco
5. Massachusetts General Hospital, Harvard Medical School *Presenting author

Upasana completed her doctoral studies in Cancer signaling and Immunology from a collaborative international PhD program between Max Delbruck Centre of Molecular Medicine, Germany and Hebrew University of Jerusalem, Israel securing the Marie Curie Doctoral Fellowship. She joined the Kwon Lab to study Immunity at the epithelial barriers in the context of HIV. Here, she established patient derived ex vivo 3-dimensional mini gut organoids complexed with matched patient autologous immune cells to study HIV-associated intestinal barrier integrity in people with HIV. Recently she was awarded the K99/R00 Pathway to Independence award from NIDDK/NIH.

TITLE: Role of Immunometabolic Dysregulation of Intestinal CD8+ T Cells in the Disruption of Intestinal Barrier Integrity in People with HIV

Background: The disruption of intestinal barrier integrity is a persistent issue in people with HIV (PWH) even after long-term treatment with suppressive antiretroviral therapy (ART). Loss of barrier integrity leads to the translocation of microbial products into the bloodstream, resulting in chronic systemic immune activation. This immune activation plays a critical role in the development of non-communicable diseases (NCDs), such as diabetes, cardiovascular diseases, obesity, and liver disease. While ART has improved the lives of PWH, the burden of NCDs has significantly increased morbidity and mortality in this population. However, the underlying mechanisms of intestinal barrier disruption in PWH on ART remain poorly understood.

Results: We report a novel mechanism by which tissue resident CD8+ T cells scavenge lipids from intestinal epithelium in PWH while on ART, leading to epithelial apoptosis. Our findings reveal that colonic CD8+ T cells rely on fatty acid oxidation (FAO) to generate energy for their maintenance. These cells utilize fatty acids (FAs) released from their intrinsic lipid reserves to fuel mitochondrial FAO. However, colonic CD8+ T cells from PWH on ART exhibit reduced lipid reserves and impaired FAO compared to HIV-uninfected individuals. We show that these metabolic alterations are associated with the downregulation of peroxisome proliferator-activated
receptors (PPARs), a family of lipid-sensing receptors known to enhance CD8+ T cell memory and effector immune function. This reduction of PPAR activity in colonic CD8+ T cells results in their failure to utilize intrinsic lipid stores for fueling mitochondrial FAO. To compensate for their lipid homeostasis, these CD8+ T cells then acquire lipids from nearby epithelial cells, which contributes to epithelial cell death. Using ex vivo 3D mini-gut organoids cocultured with autologous colonic CD8+ T cells, we demonstrate that treatment with PPAR agonists increases intrinsic lipid stores and decreases epithelial apoptosis in samples derived from PWH.

Additionally, in a genetically engineered murine model, we show that the deletion of PPARs in colonic CD8+ T cells leads to epithelial apoptosis and the loss of barrier integrity. These findings indicate that metabolic dysregulation of intestinal CD8+ T cells contributes to impairment of intestinal barrier function.

Conclusion: The immunometabolic dysregulation of colonic CD8+ T cells in PWH has a detrimental effect on their surveillance capacity to maintain intestinal barrier homeostasis. Understanding the mechanisms behind immunometabolic dysregulation and its consequences on heterotypic cellular interaction is critical to developing strategies to preserve the integrity of the intestinal barrier. Therapies developed to mitigate these disruptions for PWH could then be applied to a broader set of similar NCDs, including colitis and other situations where the stability of the intestinal mucosa is compromised.

Wilfredo is originally from Puerto Rico, where he obtained a bachelor’s in Chemistry from the University of Puerto Rico-Rio Piedras in 2010. He subsequently entered into a cross-institutional MD/PhD program where he received his PhD degree in Immunology from Harvard University and his medical degree from Harvard Medical School and Massachusetts Institute of Technology (MIT) in 2018. He pursued clinical specialty training in Clinical Pathology with sub-specialty training in Transfusion Medicine at Massachusetts General Hospital (MGH), and carried out post-doctoral research in NK-cell biology and cellular therapies as well as immune responses to SARS-CoV-2 infection and vaccines. He is currently leading his own laboratory as a Clinician-Scientist Fellow at Ragon Institute of Mass General, MIT and Harvard studying the fundamental biology and how to harness them for immune and cellular therapies against cancer and chronic viral infections.

TITLE: Deciphering the role of natural killer cells in mpox virus infection

BACKGROUND: Mpox virus (MPXV), a poxvirus of the orthopoxvirus genus endemic to Central and West Africa, gained global attention following its 2022 outbreak and worldwide spread, disproportionately affecting individuals living with HIV. This has spurred interest in understanding MPXV susceptibility and immune control. Despite limited characterization of host-pathogen interactions in human MPXV infection, the significance of natural killer (NK) cells as a first-line defense to achieve poxvirus control is recognized in small-animal and non-human primate models. Our study aims to elucidate how MPXV infection alters infected cells and impacts NK-cell recognition and killing.

METHODS: We established in-vitro infection models with flow cytometry detection of infection using A549 cells and human peripheral blood mononuclear cells (PBMCs) and performed flow cytometry and bulk RNA-sequencing on MPXV-infected A549 cells to assess phenotypic and transcriptional alterations.

RESULTS: In-vitro infections identified A549 cells and monocytes – but no other PBMC subset – as permissive to MPXV. Flow cytometry revealed MPXV-induced surface upregulation of MHC ligands and other NK-cell receptor ligands, such as MICA/B (NKG2D ligand), HLA-E (NKG2A and NKG2C ligand), HLA-F (KIR3DS1 ligand), B7-H6/NCR3LG1 (NKp30 ligand), PVR (DNAM-1 and TIGIT ligand), and the integrin LFA-3 (CD2 ligand). Bulk mRNA sequencing of infected A549 cells highlighted transcriptional signatures of immune activation and viral infection and identified upregulated transcripts of NK-cell receptor ligands, including NCR3LG1 and MICB, in line with our flow cytometry data.

CONCLUSIONS: Overall, our findings demonstrate MPXV infection modulates NKcell receptor ligands, with particular roles for the NKp30:B7-H6 and MIC-B:NKG2D axes. Ongoing studies will directly assess the contribution of these axes to NK-cell degranulation and killing of MPXV-infected cells in healthy versus HIV-positive individuals. This is particularly relevant given prior studies describing NK-cell dysfunction and receptor downregulation in individuals living with HIV, which may provide an explanation for their increased susceptibility to MPXV.

AUTHORS & AFFILITATIONS: Dagny C. Reese (1,2), Madison R. Leone (1,*), Amy K. Barczak (1,2,3), Julie Boucau (1), Wilfredo F. Garcia-Beltran ( 1,4)

1. Ragon Institute of Mass General, MIT and Harvard, Cambridge, MA, USA
2. 2 Harvard Medical School, Boston, MA, USA
3. 3 Department of Medicine, Massachusetts General Hospital, Boston, MA, USA
4. 4 Department of Pathology, Massachusetts General Hospital, Boston, MA, USA

Tiffany is a licensed psychologist and NIAID T32 post-doctoral fellow at Harvard Medical School with a joint appointment in Behavioral Medicine/Psychiatry at MGH and Emergency Medicine at BWH. She has a PhD in clinical health psychology and MS in public health. Her clinical research program is based in community-informed methods and examines how the interaction of biopsychosocial factors create barriers to health equity and development/implementation of multilevel interventions to address such disparities. Within this area, a majority of her work has had a focus on HIV and substance use.

Title: Characterizing syndemic HIV risk profiles and mHealth intervention acceptability among patients in the emergency department

Background: Syndemic theory proposes that co-occurring, mutually reinforcing psychosocial challenges (mental health, substance use, minority stress [discrimination/stigma], abuse, unmet basic needs) drive HIV risk behavior and create barriers to care for marginalized populations. It is thus necessary to address this holistic, complex picture in HIV prevention. Emergency department (ED) visits are a prime opportunity to engage key risk groups, given their low engagement in regular clinic-based care and high utilization of drop-in care via EDs. Yet, EDs are overburdened and under-resourced; mHealth may be a vehicle for intervention delivery in this context. This study aimed to 1) characterize demographics, syndemic profiles, and HIV risk behavior among ED patients and 2) assess the acceptability of addressing syndemic issues, particularly via an mHealth approach, in the ED.

Methods: A sample of N=198 ED patients with an indication of HIV risk completed a crosssectional psychosocial assessment. Descriptive statistics and bivariate correlations between syndemic issues were examined.

Results: Patients presenting to the ED reported marginalized identities and complex syndemic profiles including mental health issues (77%), problematic substance use (30%), childhood abuse (35%), adult abuse (31%), minority stress (63%), and unmet basic needs (37%). Over half the sample reported at least three syndemic issues (54%). All syndemic issues were significantly correlated with each other, supporting their synergistic nature. The sample reported indicators of HIV risk including lack of PrEP awareness (33%)/uptake (94%), condomless sex (37%), and not testing for HIV (41%). Majority reported syndemic profiles have never been addressed in the ED
(71%), think it would be helpful (88%), and are willing to utilize mHealth during an ED visit (76%).

Conclusions: The current study provides information to guide next steps for ED-based point-of care HIV prevention, and more broadly, working towards equitable HIV prevention services reaching those missed by existing interventions.

Co-authors: Simran S. Khanna; Mohammad Adrian Hasdianda; Conall O’Cleirigh; Peter Chai

Julia is a 3^rd year PhD student in the group of Prof. Bruce Walker at the Ragon Institute of MGH, MIT and Harvard. Her research project focuses on understanding the mechanisms of spontaneous HIV control mediated by lymph node CD8 T cell responses and how they relate to responses defined in CD8 T cells from peripheral blood. Part of this research was published in Science Immunology in 2023. Julia holds a master’s degree in Immunology from FAU Erlangen in Germany where she is also pursuing her doctorate. When she is not busy attending to her beloved CD8 T cells, Julia enjoys cycling and spending time outdoors.

TITLE: Transcriptional profiling of HIV-specific CD8 T cell responses from lymph nodes and blood of natural controllers

ABSTRACT: HIV is a chronic disease requiring life-long antiretroviral treatment to prevent the progression to AIDS. A vaccine or cure for this disease remains elusive, yet a small subset of individuals living with HIV controls infection naturally in the absence of antiretroviral treatment. Studying their immune response to HIV has helped to shed light on mechanisms of successful viral
suppression. Natural control of HIV is associated with strong, proliferative, cytolytic CD8 T cell responses in the blood, yet differences in phenotype and cytolytic potential have been observed in lymph node (LN)-derived CD8 T cells, a major site of HIV persistence. To better understand the functional, phenotypic, and clonotypic relationship of circulating and LN tissue-derived CD8 T cells and their contribution to HIV control, we applied a novel single-cell RNAseq technique using barcoded peptide-MHC tetramers to fluorescently sort HIV- and CMV-specific CD8 T cells from people living with HIV in the presence or absence of natural control. We sequenced the
transcriptome, T cell receptor sequences and antibody/tetramer-derived tags for surface protein expression and T cell receptor specificity of over 41,000 cells enriched for HIV- and CMVspecific CD8 T cells from 11 individuals. Of these 11 individuals, five exhibited long-term natural HIV control, four individuals were on antiretroviral therapy and two individuals were negative for HIV.

While analysis of this rich dataset is still ongoing, we have already identified different clusters of CD8 T cells that show varying enrichment levels for cells isolated from PB or LN. We identified a large cytotoxic cluster with elevated expression of CX3CR1, GNLY, GZMB and GZMH, that is enriched for cells from peripheral blood but also contains lymph node CD8 T cells. We also identified clusters that are enriched with CD8 T cells from lymph nodes that have elevated expression of T follicular helper cell markers like TNFRSF4 (OX40) and CD40LG, as well as the expression of inflammation-associated genes like IL17RB, IFIT1 and IFIT3. With the help of this data, we plan to address the extent of compartmentalization or overlap between circulating and tissue-resident HIV-specific CD8 T cell clonotypes, the gene expression differences of lymphoid HIV- and CMV-specific CD8 T cells relating to their function, as well as the CD8 T cell subset composition in LN HIV-specific CD8 T cells of spontaneous controllers and people with HIV on antiretroviral therapy. Examining the similarities and differences between natural controllers and people on treatment, as well as their CTL immune responses in blood and lymph node could help to clarify better the mechanisms of viral control in the tissue setting and inform immune-based cure strategies.

AUTHORS: Julia Hitschfel (1,2), David R. Collins (1,3), Jonathan Urbach (1,3), Bruce D. Walker(1,3,4,*) (1) Ragon Institute of MGH, MIT and Harvard; (2) Friedrich-Alexander Universität Erlangen-Nürnberg; (3) Howard Hughes Medical Institute; (4) Institute for Medical Engineering and Sciences and Department of Biology, Massachusetts Institute of Technology *Corresponding Author (bwalker@mgh.harvard.edu)

Jana is a physician in Infectious Diseases at Massachusetts General Hospital and a clinical research fellow at the Medical Practice Evaluation Center at MGH. Dr. Jarolimova’s research uses implementation science methods to evaluate new approaches to testing and treatment for curable sexually transmitted infections (STIs) in the context of HIV prevention and other sexual and reproductive health services, with research studies based in South Africa and in Boston.

TITLE: Sexually transmitted infection testing integrated with HIV prevention and contraceptive services in hair salons in urban South Africa

Background: Curable sexually transmitted infections (STIs) increase HIV transmission and acquisition and cause morbidity for women, yet access to STI testing is limited for women at risk for HIV and STIs in sub-Saharan Africa. Offering STI care in novel community-based venues may address barriers to access. We are evaluating the implementation of STI testing integrated with HIV prevention and contraceptive services in hair salons in urban KwaZulu-Natal, South Africa.

Methods: Women accessing oral HIV pre-exposure prophylaxis (PrEP) or hormonal contraception in hair salons in an ongoing study are offered testing for four curable STIs. Self-collected vaginal swabs are tested by polymerase chain reaction for gonorrhea, chlamydia, and trichomoniasis. Fingerstick blood is tested by non-treponemal and treponemal assays for syphilis. Participants with positive results are offered treatment at the salon or local clinic. Demographics, STI history, symptoms, risk factors, and risk perception are collected using structured questionnaires.

Results: Currently, we have enrolled 112 women taking either PrEP or contraceptives (or both) in the hair salons, with median age 26 years [IQR 23-29]. Fifty of 102 participants (49%) reported a primary partner ≥5y older and 52/81 (64%) reported never using condoms in the preceding month. Nineteen participants (17%) reported current STI symptoms, and 21/104 (20%) reported being treated for an STI in the past year. Twenty-nine participants (26%) perceived having a ‘moderate’ or ‘great’ chance of acquiring an STI within the next year, and 21/89 (24%) felt they had a ‘moderate’ or ‘great’ chance of acquiring HIV within the next year. Among the 112 eligible participants, 108 (96%) accepted STI testing: 104 (93%) provided vaginal swabs and fingerstick blood and four (4%) provided blood only. Among 103 participants with complete results, 37 (36%) had at least one STI: 9/103 (9%) gonorrhea, 24/103 (23%) chlamydia, 2/103 (2%) trichomoniasis, and 9/103 (9%) syphilis. The majority (30/37, 81%) with an STI were asymptomatic. Of the 37 participants with STIs, 34 (92%) elected to receive treatment in the salons.

Conclusions: STI testing in hair salons in urban South Africa is acceptable, reaches women with risk factors for STIs
and HIV, and reveals a high prevalence of mostly asymptomatic STIs. Hair salons may serve as novel venues to increase the reach of STI testing to women at risk for HIV and STIs.

Minhae is a laboratory manager at the Pain and Neuroinflammation Imaging Laboratory at the A. A. Martinos Center for Biomedical Imaging, Massachusetts General Hospital, Harvard Medical School. She studied Psychology at Connecticut College and Premedical Sciences at Columbia University. She has extensive experience in clinical research and has been the primary study staff on the study of neuroinflammation in people living with HIV since 2021.

TITLE: Neuro-glial Dysfunction in HIV-Related Neuropathic Pain

Background: Approximately 38 million people worldwide live with human immunodeficiency virus (HIV). A common neurological complication of HIV and its treatment in people living with HIV (PWH) is HIV-associated sensory neuropathy, with neuropathic pain being its most highly prevalent, treatment-resistant, and debilitating symptom. Our group has previously observed elevated levels of the 18kDa translocator protein (TSPO), a marker of glial activation and neuroinflammation, in human chronic pain and HIV using [11C]PBR28 positron emission tomography (PET) [1-2]. In this study, we used [11C]PBR28 PET to evaluate the hypothesis that
HIV-related neuropathic pain is accompanied by elevated levels of TSPO expression in PWH.

Methods: Twenty-six PWH (11 with neuropathic pain, PWHpain; 15 without pain, PWHnopain) and 27 healthy controls (HC) completed [11C]PBR28 PET/MRI scans. All participants were genotyped for the Ala147Thr polymorphism in the TSPO gene, which predicts binding affinity to [11C]PBR28, and low affinity binders were considered ineligible. All PWH were on stable antiretroviral therapy and virally suppressed (plasma HIV RNA <200 copies/mL) at the time of screening. PWH completed the PainDETECT self-reported questionnaire, where higher scores (range: 0-38) indicate higher levels of neuropathic pain. Standardized uptake values (SUV) were computed from 60-90 min post-[11C]PBR28 injection data, spatially smoothed (FWHM=8mm), and normalized to the MNI template using FSL and FreeSurfer. A T1-weighted volume (MEMPRAGE; TR/TE1/TE2/TE3/TE4=2530/1.64/3.5/5.36/7.22ms, flip angle=7º, voxel size=1x1x1mm3) was used for attenuation correction of PET data and spatial normalization to the MNI space. Whole-brain voxel-wise analyses were run using whole-brain normalized PET data and analyzed using Randomize with a z=2.3 cluster-forming threshold.

Results: PWH showed elevated TSPO expression compared to HC in the frontal white and grey matter including the ventrolateral, ventromedial, and dorsolateral prefrontal cortices, frontal pole, and secondary somatosensory cortex. PWHpain showed higher TSPO expression compared to PWHnopain in the anterior and posterior midcingulate (aMCC and pMCC, respectively) and posterior cingulate cortices, dorsolateral and dorsomedial prefrontal cortices (dLPFC and dMPFC, respectively), precentral gyrus, superior parietal lobule (SPL), supplementary motor area (SMA), and primary somatosensory and motor cortices (S1 and M1, respectively). In PWH, TSPO expression was positively correlated with PainDETECT scores in the cuneus, aMCC, pMCC, SMA, dMPFC, dLPFC, SPL, S1 and M1.

Conclusion: Overall, PWH showed elevated TSPO expression compared to HC, and PWHpain showed elevated [11C]PBR28 signal in the cingulate and prefrontal cortices and somatosensory and sensorimotor areas compared to PWHnopain. In PWH, [11C]PBR28 signal was positively correlated with PainDETECT scores in the prefrontal, cingulate, and primary somatosensory and motor cortices. Our findings suggest an association between neuropathic pain and neuroinflammation, and that neuroinflammation could be a therapeutic target for neuropathic pain. Future research with a larger sample size is required to assess the generalizability and validity of these observations.

AUTHORS: Minhae Kim, Angelica Sandström, Aarushi Tandon, Akila Weerasekera, Kelly Castro-Blanco, Shibani S Mukerji, Vitaly Napadow, Robert R Edwards, Eva-Maria Ratai, Marco L Loggia

Maanasa is a senior at Harvard College studying Neuroscience and Global Health and Health Policy. She has worked at the Ryan White HIV/AIDS program, Alter Lab of the Ragon Institute, and the Botswana Harvard Partnership. She is an aspiring physician-scientist.

TITLE: Long-Term ART is not Associated with Reduced Anogenital Cancer Risk: A Case-Cohort Study

Background: People with HIV (PWH) are at increased risk of anogenital cancer. Malignancies of the cervix, vulva, anus, and penis have become leading causes of morbidity and death for PWH globally. HIV-related immune dysfunction may contribute to excess risk. We sought to assess whether sustained antiretroviral treatment (ART) reduces excess risk of anogenital cancer in PWH. Methods: We conducted a case-cohort study in Botswana involving citizens aged 20 to 65. Adults with anogenital cancer were prospectively enrolled in a cancer cohort (“Thabatse”) from 2012-2020 at the 4 principal cancer treatment centers in Botswana. The subcohort was drawn from the Ya Tsie trial, which included a random 20% household sample of 30 communities (2013-2015) and an 80% sample in 6 communities (2017) in Botswana. ART duration was divided into three categories: No ART, ART < 5 years, and ART ≥ 5 years. We estimated the marginal relative risk (compared to HIV-uninfected) of incident anogenital cancer by ART duration using Gcomputation with inverse probability of treatment weights (IPTW), accounting for common factors of HPV and HIV, ART duration, and access to cancer treatment: 5-year age strata, education, age at first intercourse, smoking status, geographic region, and time period.

Results: A total of 17,321 participants were enrolled, including 1,377 cancer cases. HIV prevalence was higher in individuals with cancer (80%) than persons without cancer (30%). PWH who received longer duration ART (≥ 5 years) were older than PWH who received shorter duration ART (< 5 years) (median age 44 and 38, respectively). Proportions of PWH on ART with CD4 nadir greater than 350 cells/µL were 55% and 58%, respectively. Greater than 95% of participants on ART had HIV virologic suppression. Following IPTW, the analytic cohort was balanced with standardized mean difference (SMD) <0.05 for included factors, and parameters of access to cancer treatment were similar across groups (SMD <0.15). In all categories of ART duration, HIV-infection was associated with a large increase in risk for each cancer studied (Figure). Compared with shorter duration, longer duration ART was associated with greater risk of anogenital cancers: anus RR 1.86 (95% CI 1.2, 2.8); vulva RR 1.83 (95% CI 1.4, 2.5); penis RR 2.58, (95% CI 1.8, 3.7); cervix RR 1.12, (95% CI 1.0, 1.3, non-significant).

Conclusion: Prolonged ART does not reduce age-stratified anogenital cancer risk. Further interventions are needed to address excess risk in PWH on ART.

Catherine is a T32 postdoctoral fellow in the Cancer Prevention Education Program at the Harvard School of Public Health in the Department of Social and Behavioral Sciences, with a joint appointment at Massachusetts General Hospital. Her current research focuses on improving tobacco quit rates in vulnerable groups using mixed methods. The research presented at the HU CFAR Annual Scientific Symposium will describe tobacco use patterns among patients with HIV living in Chennai, India.

Title: UNDERSTANDING TOBACCO USE & CESSATION AMONG PEOPLE IN HIV-CARE IN CHENNAI, INDIA

Background: Tobacco use is the leading preventable cause of death and disproportionately impacts people with HIV (PWH). Engagement in HIV care offers an opportunity to intervene. Understanding of diverse tobacco use patterns, health beliefs and attitudes towards cessation in low- and middle-income countries is needed to inform intervention strategies.

Methods: Prospective, longitudinal study of PWH initiating HIV care in Chennai, India between October 2019 to December 2021. Individual, in-person survey (n=154) in English, Tamil or Telugu measuring self-reported and biochemically confirmed (urine cotinine ≥50ng/ml) tobacco use among adults. A subset of participants completed a 3-month telephone follow-up (n=77) to assess longitudinal tobacco use.

Results: Of 154 adults, 37.7% (n=58) self-reported current tobacco use (13.6% [n=21] smoking, 17.5% [n=27] smokeless tobacco [SLT], 6.5% [n=10] dual smoking and SLT) and 7.8% (n=12) reported former tobacco use (5.8% [n=9] smoking, 1.3% [n=2] SLT, 0.6% [n=1] dual). Among those currently using tobacco, most were male (smoking: 95.2% male; SLT: 74.1%; dual: 90.0%), completed ≤ primary school (smoking: 52.3%; SLT: 85.2%; dual: 70.0%), and many reported daily tobacco use (smoking: 52.4%, SLT: 48.1%, dual: 70.0%). Cotinine levels varied by use (smoking: 66.7% had cotinine ≥50ng/ml, mean detectable level 472.7ng/ml [SD 67.8]; SLT: 70.4% ≥50ng/ml, mean 363.4ng/mL [SD 189.3]; dual: 80.0% ≥50ng/ml, mean 462.0ng/ml [SD 107.6], no current tobacco: 44.8% ≥50ng/ml, mean 221.3 ng/ml, SD 188.0], p< 0.0001). Adults using SLT reported less interest in quitting (plan to quit within 30 days: smoking 66.6% vs. SLT 18.5%, p=0.01; dual 50.0% vs. SLT, p=0.02), fewer prior quit attempts (smoking 57.1% vs. SLT 14.8%, p=0.03; dual 40.0% vs. SLT, p=0.06), and less belief that tobacco has harmed their health some/a lot (smoking 85.8% vs. SLT 48.1%, p=0.01; dual 100% vs. SLT, p=0.004). No one currently using tobacco reported use of cessation medications; n=2 reported counseling. In the 3-month follow-up sample (n=77 [smoking: n=8, SLT: n=15, dual: n=6, non-user: n=48]), 37.5% of those smoking at baseline reported quitting, 60.0% quit SLT use, and 50% quit dual use.

Conclusions: Tobacco use is common and underreported among PWH in Chennai and treatment is underused. Those using smokeless tobacco were less interested in quitting and perceived fewer harms compared to those who smoke and may have different needs for cessation support.

AUTHORS:  Poongulali Selvamuthu¹ , Nagalingeswaran Kumarasamy¹ , Nancy Rigotti² , Catherine S. Nagawa² , Beulah Faith¹ , Bianca Porneala² , Gina Kruse²

¹Voluntary Health Services Chennai Antiviral Research and Treatment Clinical Research Site, Chennai, India, ²Massachusetts General Hospital, Boston, MA, USA

Funsho is a trained immunologist. He received his doctoral training at Africa Health Research Institute, Durban, South Africa, with a portion of his PhD studies conducted at UPenn. His research focuses on elucidating host-pathogen interactions, aiming to understand how the immune system effectively controls various pathogens encountered in everyday life. Dr. Ogunshola is particularly interested in developing technologies to investigate immune responses against diverse pathogens, with the overarching goal of delineating immune parameters that can be bolstered in vaccine or immunotherapies targeting infectious diseases and cancers. Dr. Ogunshola is currently a research associate with the Howard Hughes Medical Institute, operating within the Ragon Institute of MGH, MIT and Harvard under the mentorship of Prof. Bruce D. Walker.

TITLE: Evolution of T Cells Following HIV-1 infection

Background: In the acute phase of the immune response to viral pathogens, there is a preferential expansion of T cells harboring high-avidity T cells receptors (TCRs). However, there is incomplete information on how the cumulative duration of antigen exposure influences the functional properties of memory TCRs selected during infection. In this study, we analyzed evolution of the repertoire of memory T cells following acute HIV-1 infection comparing the impact of early versus delayed antiretroviral therapy (ART).

Methods: Persons were identified with acute HIV infection (Fiebig stage I and II) and immediately started on antiretroviral therapy (n=6), limiting antigen exposure, and compared to historical controls in whom treatment was started in the chronic phase of infection (n=6). Samples were obtained after 2 years of ART.  In order to control for HLA diversity, all were selected to express the relatively protective HLA-B*58:01 allele, and CD8⁺ T cell recognition of dominant epitope-specific responses was measured by ELISpot. Proviral sequences were analyzed to identify major and minor epitope variants. Peptide-HLA stability of consensus and epitope variants was quantitated. We then used barcoded tetramer staining to profile anMgen specific clonotypes against consensus and variant immunodominant epitopes. 

Results: HLA-B*58:01 restricted consensus and variant immunodominant peptides all stabilized surface expression of the HLA-B*58:01 allele. We detected both dominant and subdominant clonotypes that recognized same epitopes using barcoded tetramers. Notably, individuals with antigen exposure that was limited by early initiation of ART showed an enrichment of high-avidity clonotypes compared to those with prolonged antigen exposure (p=0.007). We identified public clonotypes across HIV-1 infected persons, as defined by CDR3 beta usage. Interestingly, some shared public clonotypes had different CDR1 and CDR2 beta loops.  

Conclusion: Our preliminary findings indicate that the duration of HIV-antigen exposure before treatment initiation modulates important features of the memory T cell clonotypes that are established. Ongoing studies aim to clone and functionally characterize the native TCRs we have identified in these individuals and validate the avidity data using surface plasmon resonance (SPR).

AUTHORS: Funsho J. Ogunshola^1,4, Nishant K. Singh^1,2,4, Vincent Bu8y², Liza Vecchiarello¹, Webber Annabelle¹, Sophia Cheever^1,2, Anusha Nathan¹, Kavidha Reddy³, Kamini Gounder^3,5, Musie Ghebremichael¹, Gaurav Gaiha¹, Krista Dong¹, Thumbi Ndung’u^1,3,5, Michael Birnbaum² and Bruce D. Walker1,2,4,5

¹Ragon Institute of MGH, MIT and Harvard, Boston, ²Massachuse8s Institute of Technology, Boston, ³Africa Health Research Institute, Durban, South Africa, ⁴Howard Hughes Medical Institute, ⁵HIV Pathogenesis Programme, The Doris Duke Medical Research Institute, University of KwaZulu-Natal, Durban South Africa

Tianling is a postdoctoral fellow at the Division of Infectious Diseases and Microbiome at the Broad Institute, is devoted to advancing a functional cure and a novel class of vaccines for HIV-1. Tianling has pioneered an innovative humanized mouse model through engineered B cells, establishing the groundwork for her research. Her current project focuses on developing an mRNA vaccine based on HIV-1 envelope glycoproteins. By leveraging engineered B cells, she evaluates vaccine candidates in vivo, and identifies and enhances broadly neutralizing antibodies against HIV-1 through in vivo evolution. Tianling’s research interests span genome engineering, vaccine models, immune profiling, and cell therapies. She is committed to utilizing these tools to make contributions to the ongoing quest for effective HIV-1 interventions.

TITLE: Development and evaluation of immunogens that target HIV-1 broadly neutralizing antibody precursors

An effective HIV-1 vaccine will likely need to elicit broadly neutralizing antibodies (bnAbs). However, eliciting bnAbs through conventional vaccination remains a challenge because they usually require high levels of somatic hypermutation and years of active infection to develop. Moreover, bnAb precursors are present at low frequencies in most human antibody repertoires, and their key features are human-specific and not well-modeled in wild-type mice, rabbits, or macaques. To address this, we developed a novel mouse B cell model that emulates human antibody responses, facilitating the evaluation of HIV-1 envelope glycoprotein (Env) antigens and allowing us to monitor the maturation of HIV-1 bnAb precursors. To generate antigens that drive bnAb maturation, we applied directed evolution in mammalian cells to select stabilized Env trimers that efficiently engage HIV-1 bnAb precursors. As proof of concept, we focus on a class of HIV-1 bnAbs targeting the V2-glycan or apex region of Env. Precursors of apex bnAbs are relatively abundant in human B cell repertoires and largely dependent on their HCDR3 regions for the initial engagement with HIV-1 Env. These studies will explore the hypothesis that the most direct way to generate bnAbs and potent neutralizing sera in vivo is to engage and affinity mature apex bnAb precursors.

AUTHORS: Tianling Ou, Jinge Xu, Xin Liu, Wenhui He, Michael Farzan
Division of Infectious Diseases, Boston Children’s Hospital, Boston, MA; Department of Pediatrics, Harvard Medical School, Boston, MA; Center for Integrated Solutions to Infectious Diseases, Broad Institute of MIT and Harvard, Cambridge, MA

Hemi is a Clinical Research Program/Project Manager for the Mukerji Lab for Neuroinfectious Diseases, Mass General Brigham Neurology. Her research interests include social epidemiology, immigrant health, HIV/AIDS, and community-based interventions. She graduated from Temple University with a master’s degree in public health with a focus on epidemiology.

TITLE: Plasma Neurofilament Light Chain Suggests Advanced Neuronal Age in Cognitively Unimpaired People with HIV on Antiretroviral Therapy in ACTG A5322 (HAILO).

Background: Neurofilament light chain (NfL) is a biomarker of neuronal damage and could become routine in cognitive impairment assessments. Age is strongly associated with plasma NfL values in people with HIV (PWH) and without (PWoH). We assessed plasma NfL values in cognitively unimpaired, older treated PWH and estimated the magnitude of accelerated aging so that NfL values from PWH would be comparable to NfL reference values from healthy PWoH.

Methods: This secondary analysis of a cohort study of treated PWH, focused on the subset of cognitively unimpaired PWH ≥45y, HIV RNA<200 copies/ml, and eGFR≥60 mL/min/1.73m², given the independent association of NfL with cognitive function and kidney disease. Cognitive status was provided by HAILO and based on four neuropsychological (NP) tests standardized as z-scores. Individuals with two NP scores below -1 SD or one score below -2 SD were considered impaired, others unimpaired. Plasma NfL were quantified using single-molecule array platform. Age-partitioned NfL reference values (upper 95^thpercentile per age category) were defined using published data from 1,724 “neurologically healthy” PWoH (ages, 5-90y) (Simrén et al., 2022). The NfL value from PWH was identified as above or below the reference value based on age at plasma sampling. We fit a logistic regression model to estimate the accelerated aging needed for only 5% of the PWH responses to be above the PWoH cut-off, analogous to a dose-response approach (dose: accelerating age; response: proportion above the cut-off). 

Results: The analysis included 340 PWH; 59 (17%) were female, and 100 (29%) were non-white, with a median age of 52y (range, 45-74y). The mean (SD) NfL concentration was 11.28 (5.56) pg/mL. Eighty-two (24%) PWH had NfL values above the 95% cut-off value based on reference values. To approximate when NfL values would be comparable to PWoH, an age adjustment was added to the actual age until the proportion of PWH above the reference limit was <5% (Figure). Based on the dose-response approach, participants would have needed to be 11.64y (95% confidence limit, 10.96-12.31y) older for results to be consistent with a 5% rate of elevated NfL.

Conclusion: Elevated NfL predicts cognitive decline in people with vascular and Alzheimer’s dementia.  Our analysis suggests that cognitively unimpaired treated PWH without kidney disease have plasma NfL values comparable to a healthy population of PWoH 11-12 years older. NfL may be a marker of neuronal aging in PWH and warrants further longitudinal investigation to clarify the role and relevance of plasma NfL in older PWH.

Figure. Plasma levels of NfL in cognitively unimpaired PWH are comparable to PWoH who are 11.64y older. The dots indicate the anticipated values (proportion of PWH above the 95% cut-off levels) and the dotted lines are the lower and upper 95% prediction bands from the logistic regression model. The 5% line shows expected proportion above the cut-off based on age (PMID: 35865350). Predicted value crosses the line between 11y and 12y, suggesting the accelerated aging needed to make the NfL levels among PWH consistent with the age-specified cut-off for PWoH.

Authors: Hemi Park, MPH, BS^1,2; Shibani S. Mukerji, MD, PhD^1,2,3,4; Petra Bachanová, BSc²; Linzy V. Rosen, BA⁵; Pia Kivisäkk, MD, PhD^1,3; Rommi Kashlan, BS^1,2; Felicia C. Chow, MD, MAS⁶; Kunling Wu, MS⁷; Raha M. Dastgheyb, PhD⁸; Leah H. Rubin, PhD, MA, MPH⁹; Katherine Tassiopoulos, DSc, MPH¹⁰; Emily P. Hyle, MD, MSc^3,4,5,12; Robert A. Parker, ScD, MSc^3,11,12

Affiliations: ¹ Department of Neurology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.² Vaccine and Immunotherapy Center, Massachusetts General Hospital, Boston, MA, USA. ³ Harvard Medical School, Boston, MA, USA. ⁴ Division of Infectious Diseases, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA. ⁵ Medical Practice Evaluation Center, Massachusetts General Hospital, Boston, MA, USA. ⁶ Departments of Neurology and Medicine (Infectious Diseases) and Weill Institute for Neurosciences, University of California, San Francisco, CA, USA. ⁷ Center for Biostatistics in AIDS Research, Harvard T.H. Chan School of Public Health, Boston, MA, USA. ⁸ Johns Hopkins Bloomberg School of Public Health, Department of Epidemiology, Baltimore, MD, USA. ⁹ Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, USA. ¹⁰ Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA, USA. ¹¹ Biostatistics Center, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA. ¹² Center for AIDS Research, Harvard University, Cambridge, MA, USA.

Lien is a research scientist/gerontologist, currently affiliated with the Yale School of Public Health and Medical Practice and Evaluation Center and Center for Aging Research and Serious Illness at Massachusetts General Hospital. Her academic journey commenced with an MD from Hanoi Medical School, followed by an MPH from Brown University and a Ph.D. in Gerontology from UMass Boston. She further pursued a Postdoctoral Fellowship at Harvard Medical School. Specializing in the social determinants of health, Dr. Quach’s research delves into the relationships between social patterns and cognitive/physical function, examining the overall quality of life in older adults globally, both with and without HIV. With over a decade of experience in VA data, she also focuses on the epidemiology of aging health outcomes among older veterans. Dr. Quach has contributed significantly to her field with a portfolio comprising over 41 peer-reviewed articles and numerous books and book chapters. Her dedication underscores her commitment to advancing public health interventions for older individuals, including people living with HIV in the US and globally.

TITLE: Social Determinants of Falls among Older Adults With and Without HIV in Rural Uganda

Background: In the global north, over one in three older adults experience falls every year. These events escalate the risk of disability, hospitalization, and premature death. Older people with HIV (PWH) may be at elevated risk of falls, due in part to diminished social networks and support systems, which have been linked to  potential protective factors of falls. Nonetheless, little data are available about the epidemiology of falls among PWH in sub-Saharan Africa, where 70% of PWH reside. To address this gap, we sought to estimate the incidence and risk factors for falls in a mixed cohort of older people with and without HIV in rural Uganda.

Method: We analyzed data from the Quality of Life and Ageing Study in Uganda, which follows two groups of people: PWH on antiretroviral therapy (ART) for at least three years and sex- and age-similar people without HIV (PWOH) recruited from the HIV clinic catchment areas. Our outcome of interest was the self-reported number of falls in the past year. Primary exposures of interest were HIV serostatus and covariates that were theorized to correlate with fall risk, including: household size, physical and financial support, household wealth, social integration, and loneliness. We fit generalized linear models assuming a negative binomial distribution to estimate the associations between the number of falls and our explanatory variables, adjusting for age, sex, education, alcohol consumption, and number of comorbidities.

Results: The mean age of participants (n=571) was 58 years (SD: 6.5). In this sample, 24% of people experienced at least one fall in the past year, and 10% experienced two or more falls. Fall incidence increased with age, with 50% of people over 70 reporting a fall. We found no difference in falls by sex, (IRR: 1.36, 95% CI: 0.95, 1.94) or HIV serostatus (IRR:0.82, 95% CI: 0.58, 1.17). In multivariable models, each additional point on the loneliness scale was associated with an increase in the rates of falls (IRR:1.15, 95% CI: 1.02-1.31) and reduced household size (IRR: 0.91 per household member, 95% CI: 0.84-0.99), but not with social integration, physical or financial support, or household wealth.

Conclusion: People with HIV (PWH) in Uganda experience falls at a similar rate as those without HIV (PWOH). Interventions to reduce loneliness and enhance social networks may reduce fall risk among older adults in general and could be integrated into screening in fall prevention programs.

AUTHORS: Lien T. Quach^1,2,3; Christine S. Ritchie^1,3;  Alexander C. Tsai^1,3; Zahra Reynolds¹; Robert Paul ⁴; Janet Seeley⁵; Yao Tong¹; Susanne S. Hoeppner^1,3; Flavia Atwiine ⁶; Edna Tindimwebwa⁷;Samson Okello^8,9; Noeline Nakasujja⁹; Jeremy Tanner¹⁰; Brianne Olivieri-Mui¹¹; Deanna Saylor¹²; Meredith Greene^l3; Stephen Asiimwe⁹; Mark J. Siedner^1,3,6 

Affiliations: ¹ Massachusetts General Hospital, USA , ²  University of Massachusetts Boston, USA, ³  Harvard Medical School. USA, ⁴  University of Missouri – St Louis, USA, ⁵  London School of Hygiene & Tropical Medicine, UK, ⁶  Mbarara University of Science and Technology, Uganda, ⁷ Kabwohe Clinical Research Centre, Uganda, ⁸  Department of Epidemiology, Gillings School of Global Public Health, University of North Carolina, Chapel Hill, NC, USA, ⁹  Makerere University, Uganda, ¹⁰ The Biggs Institute for Alzheimer’s and Neurodegenerative Diseases at UT Health San Antonio , ¹¹  Northeastern University, USA, ¹² Johns Hopkins University School of Medicine, USA, ¹³ Department of Medicine, Indiana University School of Medicine and Indiana University Center for Aging Research, Regenstreif Institute, Inc., Indianapolis, IN, USA.

Claire is a fourth-year medical student at Harvard Medical School, pursuing a specialty in Internal Medicine Primary Care. She is interested in infectious disease and geriatrics, and she is committed to using quantitative methods to fill existing gaps in evidence-based care for those with chronic infectious diseases. She has recently completed a rotation at the CDC in applied epidemiology and hopes to bridge clinical work and epidemiology research in her future work as a physician.

TITLE: Comparative effectiveness and safety of oral anticoagulants in patients with atrial fibrillation living with HIV

BACKGROUND: People living with HIV are rapidly aging into the geriatric demographic. They will develop multiple comorbidities as they age, such as atrial fibrillation (AF), requiring anticoagulation which may interact with their antiretroviral therapy (ART). How HIV and the use of ART impact the effects of oral anticoagulants (OACs) in patients with AF is unknown. We sought to determine the comparative safety and effectiveness of specific OACs in older patients with HIV and non-valvular AF taking ARTs.

METHODS: We used US Medicare claims data to establish two cohorts with nonvalvular AF and HIV aged ≥ 65 years, comparing new use of warfarin vs. apixaban, and new use of rivaroxaban vs. apixaban, from January 2013 to December 2020 (cohort entry date = first dispensing date; baseline assessment period 180 days before entry). The primary outcome was a composite endpoint of hospitalization for ischemic stroke or major bleeding, and we studied each specific outcome in secondary analyses. We used propensity score-weighting to adjust for 96 potential confounders. We stratified our analysis in two subgroups: (1) patients with an active ART prescription at the time of anticoagulation prescription, and (2) patients without an overlapping ART prescription.

RESULTS: The study cohort compared 1,126 warfarin users (weighted mean age 73.9 years, female 30.4%)  with 1,525 apixaban users (weighted mean age 74.0 years, female 29.5%).  The second cohort compared 667 rivaroxaban users (weighted mean age 73.9 years, female 32.4 %) with 1,545 apixaban users (weighted mean age 74.0 years, female 31.4 %). Among our subgroup of ART users taking warfarin vs. apixaban, the rate of the composite endpoint in those starting warfarin was higher than that of those starting apixaban (84.77 vs 24.56 per 1000 person-years; adjusted hazard ratio [aHR], 3.08; 95% confidence interval [CI] 1.35-7.00). We did not see this difference in risk in patients not taking ARTs (aHR 3.08 vs. 1.03 in ART users vs. non-users; p for heterogeneity by ART use=0.03). The treatment effect heterogeneity in those on ART in this cohort was largely driven by the higher risk of gastrointestinal (GI) bleeding for those on warfarin vs. apixaban (aHR 4.75 vs. 0.87 in ART users vs. non-users; p for heterogeneity by ART use <0.01). Comparing rivaroxaban with apixaban users, we also observed a trend towards a higher risk of GI bleeding in ART users (aHR 4.78 vs. 1.11 in ART users vs. non-users; p for heterogeneity by ART use=0.06). However, we did not observe treatment effect heterogeneity by ART use for other outcomes when comparing rivaroxaban with apixaban users.

CONCLUSIONS: In a national cohort of older adults with HIV and AF, apixaban was associated with a lower risk of major bleeding compared with warfarin or rivaroxaban in ART users but not in ART non-users. Our findings suggest concomitant use of ART in this underserved population may potentiate the bleeding risks of warfarin and rivaroxaban.

Authors: Claire Quinlan¹, BA, Jerry Avorn¹, MD, Aaron S. Kesselheim¹, MD, JD, MPH, Daniel E Singer², MD, Yichi Zhang¹, MPH, Alex Cervone¹, BA, Kueiyu Joshua Lin^1,2, MD, SCD

1 Division of Pharmacoepidemiology and Pharmacoeconomics, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School; 2 Department of General Internal Medicine, Massachusetts General Hospital, Harvard Medical School

Larance is research Scientist at the Lingwood lab from the Ragon Institute of Massachusetts General Hospital, MIT and Harvard University, Cambridge. He has been actively involved in understanding the role of non-permissive and permissive B cell selections for the discovery of viral epitopes especially HIV bnAb site, advancing viral immunogen design approaches such as epitope enrichment for B cell responses toward conserved viral epitopes like Influenza, enhancing scaffolds-independent antibody responses to SARS Cov2 RBD by DNA origami scaffolds, de-coding the functional consequences of IL6 buffering system in the mouse model. He has completed PhD in Microbiology from the University of Delhi and from the National Institute of Immunology, India. For his PhD thesis, he has characterized the genetic and functional role of natural mutations occurring in the HIV-1 genes such as Vpu, Tat, Vpr, Vif within the HIV-1 infected individuals and understanding their participation in viral genes-mediated cellular functions. Also, he has been involved in studying and identifying novel mutations in the human CCR5 gene, a co-receptor for HIV-1 entry into the cells and their genetic association to HIV-1 progression rate.

TITLE: Engaging an HIV vaccine target through the acquisition of low B cell affinity

Background: Low affinity is common for germline B cell receptors (BCR) seeding development of broadly neutralizing antibodies (bnAbs) that engage hypervariable viruses, including HIV. Antibody affinity selection is also non-homogenizing, insuring the survival of low affinity B cell clones. We aimed to explore whether this provides a natural window for expanding human B cell lineages against conserved HIV vaccine targets.

Methods: To test our hypothesis, we deploy transgenic mice mimicking human antibody diversity and somatic hypermutation (SHM) and immunized with monomeric HIV glycoprotein120 (gp120) envelope immunogens. After sequential immunizations with four different HIV gp120 strains, we quantified CD4 binding site (CD4bs) antibodies by ELISA and they were cell sorted and sequenced to explore the SHM and were subjected to clonal analysis. Later, the vaccine expanded BCR lineages were membrane presented on the ramos cell line to predict specificities by calcium flux assay and by antigen binding flow cytometry while the BCR affinities were determined by SPR analysis.

Results: We report an immunization regimen that focuses B cell memory upon the conserved CD4bs through both conventional affinity maturation and reproducible expansion of low affinity BCR clones with public patterns in SHM. In the latter instance, SHM facilitates target acquisition by decreasing binding strength. We demonstrate that across multiple vaccine recipients’ permissiveness in B cell affinity selection supports expansion of public B cell clones targeting the HIV CD4bs. Notably, these public permissive B cell clones share conserved patterns in SHM which are independently reproduced in the different mice.

Conclusion: We show that both conventional affinity maturation and permissiveness in B cell selection are concomitantly operable, and both used to engage HIV Env. These results suggest that permissive B cell selection enhances discovery of antibody epitopes, enabling engagement of a conserved HIV bnAb site.

AUTHORS: Larance Ronsard¹, Ashraf S Yousif¹, Faez Amokrane Nait Mohamed¹, Jared Feldman¹, Vintus Okonkwo¹, Caitlin McCarthy¹, Julia Schnabel¹, Timothy Caradonna¹, Ralston M Barnes², Daniel Rohrer², Nils Lonberg², Aaron Schmidt¹, Daniel Lingwood¹.

1. The Ragon Institute of Mass General, The Massachusetts Institute of Technology and Harvard University, 400 Technology Square, Cambridge, MA, 02139, USA.
2. Bristol-Myers Squibb, 700 Bay Rd, Redwood City, CA, 94063-2478, USA.

Keywords: HIV envelope, CD4 binding site antibody, B cell receptors, low affinity, Somatic hypermutation, HIV vaccine.

Lintang is a physician in Indonesia who provides primary palliative care. He is currently pursuing a Master of Medical Sciences in Global Health Delivery at Harvard Medical School. He has been serving underserved populations requiring palliative care in the country, including the emerging group of older people living with HIV who develop chronic illnesses whose needs were often underlooked.

TITLE: Community-based Palliative Care for Estranged Aging People Living with HIV  

ABSTRACT: In the era of antiretroviral treatment (ART), people living with HIV (PLHIV) worldwide, including in Indonesia, are increasingly able to live longer, healthier lives and age with the condition. While life expectancy has improved, aging with HIV presents its own challenges. PLHIV experience earlier onset of aging symptoms and comorbidities. This condition multiplied with the chronic stigma associated with HIV and aging itself, affect their quality life and survivorship. These issues frequently lead to extended periods of disability, dependency, and the need for long-term care. However, health services tailored to meet the needs of this population are currently limited. In Indonesia, where family members typically care for the chronically ill, many aging PLHIV find themselves disconnected from any familial support. Recognizing this gap, three HIV/AIDS non-profits in Jakarta, Yogyakarta, and Surabaya have initiated community-based shelters providing informal palliative care for estranged, aging PLHIV. Through an ethnographic lens, this paper aims to highlight the overlooked palliative care needs of this population and explore the role of community-based shelters in providing palliative care to estranged individuals living with HIV who lack familial support.

Parmit is a Scientist II in the Engelman laboratory at the Department of Cancer Immunology and Virology within the Dana-Farber Cancer Institute and Harvard Medical School. His interest lies in understanding the mechanism of retroviral integration site selection into the host genome and the cellular processes determining viral integration preferences. His published work shows that HIV-1 prefers to integrate into highly spliced genes, cancer genes, and DNA regions proximal to nuclear speckles. To understand the preference for pre-mRNA splicing, he is currently investigating roles for promoter-proximal Pol II pausing and alternative polyadenylation in HIV-1 integration.

TITLE: mRNA splicing targets HIV integration into PAF-1-regulated genes

ABSTRACT: Pre-mRNA co-transcriptional splicing is coupled with promoter-proximal Pol II pausing and alternative polyadenylation (APA). Splicing inhibitors increase pausing and the use of upstream or proximal polyadenylation sites (PASs) by impairing the recruitment of positive transcription elongation factor b (P-TEFb), which is a core component of the super elongation complex (SEC). The cleavage factor Im (CFIm) complex consisting of cleavage and polyadenylation specificity factor (CPSF) 6 and CPSF5 regulates APA by promoting the use of distal PASs. CPSF6 binds capsid (CA) to license HIV-1 intranuclear trafficking and integration targeting into highly spliced or intron rich genes. Based on the interconnections between splicing, pausing, and APA, we hypothesized that APA and Pol II elongation might play a role in HIV-1 integration targeting. Indeed, in Jurat T cells, APA-regulated genes dependent on U2 snRNP (4.6% of human genes) for the selection of distal PASs harbored 24% of HIV-1 integration sites (3x compared to RIC or random integration control; p<1E-5). In contrast, genes independent of U2 snRNP for APA regulation were targeted similarly to all human genes (p< 0.2). Apart from splicing and CFIm complex, Pol II associated factor-1 (PAF-1) regulates pausing and the selection of distal PASs. Additionally, PAF-1 regulates the expression of integrated proviral DNA. We observed that paused genes regulated by PAF-1 (14% of human genes) were preferentially targeted (3.5x RIC, containing 40% HIV-1 sites; p<1E-5), whereas the reciprocal gene set was preferentially avoided (p<1E-5). To test the role of splicing, we infected Jurkat T cells in the presence of the U2 snRNP inhibitor Pladienolide B (Plad B) or the SEC inhibitor KL-2 and determined sites of HIV-1 integration. Whereas Plad B significantly reduced integration into PAF-1 paused genes, it failed to significantly effect integration into the reciprocal set of human genes. We called chromosomes with reduced genic integrations (p<1E-04) as Plad B sensitive chromosomes (PBSC) and the remaining chromosomes with increased genic integrations as Plad B insensitive chromosomes (PBIC; p<0.02)). Although both PBSC and PBIC genes had the same average number of introns, PBSC were comparatively gene enriched (12 genes/Mb) whereas PBIC were gene-poor (7 genes/Mb; the average genes/Mb in human genome is 9). KL-2 reduced genic integration significantly for PBSC but not for PBIC, suggesting that both splicing and SEC inhibitors reduced HIV-1 integration into the same sets of genes. To test the roles of integration targeting cofactors, we mapped sites for CPSF6-defective CA mutant viruses or wild type (WT) HIV-1 in LEDGF/p75 knockout (LKO) cells. PBSC supported significantly less genic integration for CA mutants and for WT virus in LKO cells (p<1E-7). However, while PBIC were significantly less targeted by WT virus in LKO cells, these genes were significantly more targeted by CA mutants (p<1E-7 for both comparisons). Thus, the CPSF6-CA interaction is critical for preferential HIV-1 integration targeting of paused genes and APA genes regulated through P-TEFb/SEC and splicing. Currently, we are planning to differentiate the role of CPSF6-dependent APA from CPSF6-dependent trafficking of CA in the targeting of paused and APA-regulated genes.

AUTHORS: Parmit K. Singh¹, Arun S. Annamalai², Mamuka Kvaratkshelia², Alan N. Engelman¹

¹Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA; ²University of Colorado School of Medicine, Aurora, CO

Dessie graduated from The University of Texas at Austin with degrees in Neuroscience and Plan II Honors, Dessie became a Project Coordinator at the Medical Practice Evaluation Center at Massachusetts General Hospital. She works with Dr. Jennifer Manne-Goehler to investigate HIV and comorbid obesity, diabetes, and CVD in South Africa.

TITLE: Dolutegravir-Based Therapy, Diet, Activity & Weight Gain: 48-Week Prospective Cohort in South Africa

Background: Weight gain has been reported among people with HIV (PWH) after transitioning to tenofovir, lamivudine, and dolutegravir (TLD) antiretroviral therapy (ART). However, the contribution of changes in diet and physical activity to weight gain in this population remains poorly understood. We estimated relationships between diet, physical activity, and clinically significant weight gain over 48 weeks among PWH in South Africa after transitioning to TLD.

Methods: The DISCO cohort study followed 500 PWH at four public-sector clinics in KwaZulu-Natal, South Africa. Eligible participants were >18 years of age and on efavirenz-based, first-line ART for >6 months before transitioning to TLD by clinic staff. Anthropomorphic, diet, and physical activity data were measured at enrollment, 24-week, and 48-week study visits. Our primary exposures of interest were change in four dietary and exercise behaviors over 48 weeks: (1) fruit consumption, (2) fast-food consumption, (3) sugar-sweetened beverage (SSB) consumption, and (4) physical activity (in metabolic equivalent-minutes/week). Our primary outcome was clinically significant weight gain, defined as ≥10% increase in weight from baseline to 48 weeks. We estimated differences in each exposure by presence versus absence of weight gain using multivariable logistic regression models adjusted for age, sex, education, and ART duration.

Results: We analyzed data from 367 PWH having a mean age of 43 (standard deviation = 12) years and 78% women. People with clinically significant weight gain had an increase in fruit intake while those without clinically significant weight gain had decreased intake (0.21 servings/week [95% CI: -0.9 to 1.3] v. -1.07 servings/week [95% CI: -1.5 to -0.7], respectively) (p=0.015), but there were no significant differences in frequency of fast-food intake, frequency of SSB consumption or physical activity (Figure 1). In adjusted models, change in fruit intake remained significantly associated with clinically significant weight gain over 48 weeks ([95% CI: 1.0 to 1.1], p=0.033).

Conclusions: Clinically significant weight gain was associated with increased fruit intake but not with other dietary or physical activity changes over 48 weeks after transitioning ART. Interventions to modify behavior in this population may have a limited role in mitigating clinical obesity. Pharmacological interventions to mitigate clinical obesity in this population may be needed.

AUTHORS: Nomathemba Chandiwana^1*, Dessie Tien^2*, Gugulethu Shazi³, Geoffrey Chen², Smart Mabweazara³, Mahomed-Yunus S Moosa⁴, Ravindra K. Gupta^3,5, Deenan Pillay⁶, Vincent C. Marconi⁷, Bethany Hedt-Gauthier⁸, Willem D.F. Venter¹, Mark J. Siedner^2,3,8,9,10, Suzanne M. McCluskey^2,8,10**, Jennifer Manne-Goehler^2,10,11**

¹Wits Ezintsha, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; ²Medical Practice Evaluation Center, Massachusetts General Hospital, Boston, MA, USA; ³Africa Health Research Institute, Durban, South Africa; ⁴Department of Infectious Diseases, University of KwaZulu-Natal, Durban, South Africa; ⁵University of Cambridge, Cambridge, United Kingdom; ⁶University College London, London, United Kingdom; ⁷Emory University School of Medicine and Rollins School of Public Health, Emory University, Atlanta, GA, USA; ⁸Harvard Medical School, Boston, MA, USA; ⁹Mbarara University of Science and Technology, Mbarara, Uganda; ¹⁰Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA, USA; ¹¹Division of Infectious Diseases, Brigham and Women’s Hospital, Boston, MA, USA; *Delineates co-authorship. **Delineates senior co-authorship.

Jia Le is an artificial intelligence scientist in healthcare. She is the founder and principal investigator at Lizora Technologies, where she leads the development of an autonomous treatment platform targeting COVID-19, long COVID, and influenza with machine learning. Her work merges clinical medicine with machine learning, aiming to transform personalized digital health solutions and streamline disease management. Lizora is a member of NVIDIA’s Deep Learning and AI Incubator. Beyond her entrepreneurial endeavors, Xian is deeply involved in academic research as a visiting scholar at the Institute of Pure and Applied Mathematics at UCLA. There, she contributes to projects such as ‘Geometry and Learning from Data in 3D and Beyond’ and explores the mathematical challenges of decision-making in autonomous vehicles. Additionally, Xian has received four years of clinical training in traditional Chinese medicine.

Throughout her career, Xian has had the privilege of contributing to global health initiatives as a program manager at the Clinton Health Access Initiative. In this role, she supported the World Health Organization’s HIV/AIDS Prevention and Treatment Program and contributed to China’s first national pediatric AIDS treatment program, efforts she considers deeply meaningful.

Abstract Title: Using an Autonomous Treatment Platform for COVID-19, Long COVID, and Influenza with Machine Learning

Background: Despite progress in public health measures and vaccination efforts, COVID19 remains a formidable challenge to global health due to the mutable nature of the virus, the observed waning immunity post-vaccination and subsequent to natural infections, and a risk profile markedly graver than that of influenza. We hypothesize that treating multifaceted COVID infections—much like solving a Rubik’s Cube, which— despite trillions of combinations—has only three rotating layers on each axis (X, Y, Z)— requires a focus on three aspects: immune system strength, symptom alleviation, and viral load reduction. To this end, we propose precision medicine that tailors dosages based on individual characteristics such as age, pre-existing health conditions, and symptom severity to optimize personalized treatment plans. This method aims to improve antiviral efficacy and reduce adverse side effects. We also envision an autonomous machine learning platform, ACT-ML, informed by Traditional Chinese Medicine (TCM) principles, to offer scalable, personalized antiviral treatment in outpatient settings. This platform is designed to convert clinical triage into a classification task in machine learning.

Methods: Our approach involved the development of ACT-ML, leveraging 340 million synthetic patient records, which focused on 44 symptoms across three age groups: 1440, 41-60, and 60 and above. The validation included a 49-patient retrospective simulation study and a clinical feasibility trial among 27 patients (13 with mild and moderate COVID-19 cases, one severe outpatient COVID-19 case, one severe COVID-19 intensive care unit patient, 10 long COVID cases, and two influenza cases) from June to August 2023.

Results: The platform demonstrated 100% diagnostic alignment with TCM experts in both studies. In the clinical trial, there were no COVID-19 or influenza hospitalizations or deaths from any cause, and no cases of SARS-CoV-2 reinfection through day 28.

Conclusion: ACT-ML’s diagnostic alignment with experts, its scalability, and its ability to facilitate contactless treatment operations and swift medication delivery show promise in alleviating clinician shortages and administering personalized antiviral treatment at scale, especially during health crises.

Shijian is a Scientist II at Dana-Farber Cancer Institute, Research Associate in Microbiology at the Department of Microbiology Harvard Medical School. He has been working on HIV-1 Envelope (Env) membrane protein for almost ten years. HIV-1 Env is metastable and undergoes dramatic conformation changes under different physical and chemical agitations.  He has found that during HIV-1 Env production in cells, it goes through two pathways to present on cell surface: Golgi-bypass pathway, and conventional Golgi pathway. HIV-1 Env going through Golgi-bypass pathway can not be cleaved into mature functional HIV-1 Env. The uncleaved HIV-1 Env exposes poorly neutralizing antibody (pNAb) epitopes and loses some broadly neutralizing antibody (bNAb) epitopes. The epitope presentation shift of the uncleaved HIV-1 Env would skew host immune response against HIV-1 infection. His focus has been trying to establish good HIV-1 Env preparation approaches for single-particle Cryo-EM study to understand the molecular architecture of HIV-1 Env and guide the HIV-1 immunogen design. After several years of efforts, he has worked out several ways to prepare pure HIV-1 Env with the same antigenicity as that on cell surface. These novel HIV-1 Env preparation approaches will help us not only to look into HIV-1 Env structure but also investigate the immunogenicity of HIV-1 Env in right conformations.

Title: Conformations of HIV-1 Env in amphipol A18 lipid nanoparticles

Background: HIV-1 Env is the only membrane protein on HIV-1 viruses responsible for the virus entry into host cells, and the sole target for neutralizing antibodies to block virus entry. Metastable HIV-1 Env samples different conformations: state 1 (pretriggered closed conformation), state 2 (default intermediate conformation), state 3 (open conformation). Some broadly neutralizing antibodies preferentially bind to State-1 conformation and understanding State-1 conformation will guide HIV-1 immunogen design. However, State-1 conformation has not been solved yet, although several liganded or unliganded HIV-1 Env structures are available. One important reason is that it’s difficult to purify HIV-1 Env in State-1 conformation from cell membranes. A good approach to purify HIV-1 Env in State-1 conformation will be a prerequisite to solve the conformation. In this study, we compared different amphipathic copolymers to purify HIV-1 Env from cell membranes and evaluated the antigenicity of the purified HIV-1 Envs.

Methods: We screened several recently available amphipathic copolymers by purifying HIV-1 Env using these copolymers and evaluating HIV-1 Env antigenicity in the copolymer formed lipid nanoparticles.

Results: HIV-1 Env in amphipathic copolymer A-18 mostly resembles that on cell surface in terms of antigenicity, compared with other copolymers. Lipidomic analysis results demonstrated that HIV-1 Env in A-18 lipid nanoparticles rather than in other copolymer formed lipid particles contained cholesterol, signifying the importance of cholesterol to maintain HIV-1 Env pretriggered conformation.

Conclusion: HIV-1 Env in A-18 mostly resembles HIV-1 Env on cell surface in antigenicity, which might provide a new avenue to prepare HIV-1 Env for Cryo-EM and immunogenicity studies.

Zhiqing obtained her Ph.D from School of Public Health at Xiamen University in 2020. During her Ph.D study, she focused on investigating the HIV-1 capsid (CA) protein assembly and the role of CA as a scaffold to display HIV-1 Envelope (Env) glycoproteins. In 2021, she joined Joseph Sodroski’s lab at Dana-Farber Cancer Institute to carry out her postdoctoral research in the field of HIV-1 Env glycoproteins. Dr. Zhiqing Zhang has a long-standing interest in studying HIV-1 Env conformation and developing HIV-1 vaccine candidates.

Title: Membrane HIV-1 Envelope Glycoproteins Stabilized in a Pretriggered Conformation to a Degree Beyond That of Natural Virus Strains

Background: The pretriggered (State-1) conformation of the human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer ((gp120/gp41)3) is targeted by virus entry inhibitors and broadly neutralizing antibodies (bNAbs), which would be desirable to generate with a vaccine immunogen.  Differences in conformation and glycosylation have been observed between membrane State-1 Envs and sgp140 SOSIP.664 Env trimers, which have not elicited bNAbs to date. In natural HIV-1 Env trimers, the State 1 conformation is stabilized by furin cleavage of the Env precursor and is dependent on Env interactions with the membrane. These characteristics, along with the lability of the pretriggered Env conformation, have hindered characterization of State-1 Env structure and immunogenicity.

Methods: We screened natural polymorphisms in HIV-1 Envs for the ability to generate viral phenotypes associated with increased State-1 stability. To begin with wild-type HIV-1AD8 Env, Envs incorporating individual changes and multiple combinations of these changes were evaluated for processing, subunit association, gp120 association with detergent-solubilized trimers, ability to mediate cell-cell fusion, pseudovirus infectivity, CD4mcs inhibition and cold inactivation. Additionally, we examized the antigenicity and CD4mc/cold-induced shedding of gp120 from virus particles to distinguish Envs. We also compared the sensitivity of pseudoviruses with the wt and State-1-stabilized mutant Envs to neutralization by sCD4-Ig and a panel of bNAbs and pNAbs.

Results: Here, we produce membrane Env variants progressively stabilized in pretriggered conformations, in some cases to a degree beyond that found in natural HIV-1 strains.  State-1 stability correlated with stronger trimer subunit association, increased virus sensitivity to bNAb neutralization and decreased capacity to mediate cell-cell fusion and virus entry.  For some highly stabilized Env mutants, after virus-host cell engagement, the normally inaccessible gp120 V3 region on an Env intermediate became targetable by otherwise poorly neutralizing antibodies.

Conclusion: Thus, evolutionary pressure on HIV-1 Env to maintain trimer integrity, responsiveness to the CD4 receptor, and resistance to antibodies modulates State-1 stability.  The strongly State-1-stabilized membrane Envs reported here will facilitate further characterization of this functionally important conformation.

AUTHORS: Zhiqing Zhang, Saumya Anang, Hanh T. Nguyen, Christopher Fritschi, Amos B. Smith III, Joseph G. Sodroski