Abstract Disclosure: K. Sauter: None. D.L. Takahashi: None. G. Webb: None. H. Hofmeister: None. O. Varlamov: None. J. Sacha: None. P. Kievit: None. Current antiretroviral therapy (ART) regimens have rendered HIV a manageable chronic condition rather than a fatal disease, with people who initiate ART soon after infection achieving nearly normal life expectancy. As a consequence of increased survival, more women living with HIV (WLWH) will now undergo menopause and be subject to a disease burden that reflects age along with adverse skeletal consequences of postmenopausal estrogen deficiency. In addition to known bone loss and increased fracture risk in postmenopausal women, ART also causes decreased bone mineral density. Therefore ART-suppressed WLWH may have a compounded detrimental effect on bone due to long-term ART treatment combined with menopause that may be attenuated with estrogen replacement. Eleven rhesus macaques were infected intravenously with SIVmac239M and received ART two weeks post-infection (PI). All animals were ovariectomized (OVX) at 35 weeks PI and received estrogen implants (n=6) or cholesterol implants (n=5). Weekly blood samples were collected and animals were monitored longitudinally for body composition including bone mineral content (BMC) via dual x-ray absorptiometry (DEXA). Infection with SIV caused a significant decrease, -8.2%, in pelvis BMC after 14 days of peak viremia with SIV. These changes were not observed in the BMC of spine or extremities. BMC remained suppressed by ∼7% for an additional 25 weeks PI during ART treatment. Measurement of circulating bone turnover markers (BTMs), osteocalcin and C-terminal telopeptide of type 1 collagen (CTX), decreased post infection without significant improvement after ART suppression. Induction of menopause via OVX and loss of estrogen induced a drastic increase in BTMs with a much larger increase in CTX, indicating potential global bone loss due to OVX. Post-implant, the control animals’ pelvis BMC continued to decrease to -20.9% and pelvis BMD to -9.3% indicating that OVX further exacerbated the effect of SIV and ART. However, animals that received estrogen implants demonstrated marked improvement in both measures to greater than baseline. SIV and subsequent long-term ART negatively impact pelvic BMC and BMD and this effect is compounded by menopause in our model of post-menopausal WLWH. However, estrogen replacement post-menopause may attenuate these detrimental bone effects despite long-term ART treatment. Presentation: 6/1/2024

Adoptive T-cell therapy using natural T-cell receptor (TCR) redirection is a promising approach to fight solid cancers and viral infections in liver and other organs. However, clinical efficacy of such TCR+-T cells has been limited so far. One reason is that syngeneic preclinical models to evaluate safety and efficacy of TCR+-T cells are missing. We therefore developed an efficient viral vector strategy mediating expression of human MHC-I in hepatocytes, which allows evaluation of TCR-T cell therapies targeting diseased liver cells. We designed adeno-associated virus (AAV) and adenoviral vectors encoding either the human-mouse chimeric HLA-A*02-like molecule HHD, or fully human HLA-A*02 and human 2 microglobulin (h2m). Upon transduction of murine hepatocytes, the HLA-A*02 construct proved superior in terms of expression levels, presentation of endogenously processed peptides and activation of murine TCR+-T cells grafted with HLA-A*02-restricted, hepatitis B virus (HBV)-specific TCRs. In vivo, these T cells elicited effector function, controlled HBV replication, reduced HBV viral load and antigen expression specifically in livers of mice that had received AAV-HBV and AAV-HLA-A*02. We then demonstrated the utility of this approach by expressing the HBV-specific TCRs on macaque primary T cells enabling them to recognize HBV-infected macaque hepatocytes expressing HLA-A*02 upon adenoviral transduction. In conclusion, AAV and adenovirus vectors are suitable for delivery of HLA-A*02 and h2m into mouse and macaque hepatocytes. When recognizing HBV in the HLA-A*02-transduced mouse livers or on macaque hepatocytes, HLA-A*02-restricted, HBV-specific TCR+-T cells become activated and exert antiviral effector functions. This approach is applicable to other MHC restrictions and target diseases, paving the way for safety and efficacy studies of human TCR-based therapies in physiologically relevant preclinical animal models.

Few countermeasures to treat Yellow Fever virus (YFV) infection are under development, because vaccines have helped to limit new infections. Unfortunately, vaccine hesitancy, supply deficits, and a paucity of therapeutic options have left individuals at risk. Here, we tested potent YFV-specific neutralizing monoclonal antibodies in rodents and non-human primates. We administered antibodies during acute pathogenic YFV infection and demonstrate that we can prevent severe disease and death. Given the severity of YFV-induced disease, our results show that these antibodies could be effective in saving lives and fill a much-needed void in managing Yellow Fever cases during outbreaks around the world. One Sentence Summary Therapeutic monoclonal antibodies prevent death from YFV infection.

Human leukocyte antigen-E (HLA-E) normally presents a HLA class Ia signal peptide to the NKG2A/C-CD94 regulatory receptors on natural killer (NK) cells and T cell subsets. Rhesus macaques immunized with a cytomegalovirus vectored simian immunodeficiency virus (SIV) vaccine, generated Mamu-E (HLA-E homolog) restricted T cell responses that mediated post-challenge SIV replication arrest in >50% of animals. However, human immunodeficiency virus type 1 (HIV-1) specific HLA-E restricted T cells have not been observed in HIV-1-infected individuals. Here we primed HLA-E restricted HIV-1 specific CD8+ T cells in vitro. These T cell clones, and allogeneic CD8+ T cells transduced with their T cell receptors, suppressed HIV-1 replication in CD4+ T cells in vitro. Vaccine induction of efficacious HLA-E restricted HIV-1 specific T cells should therefore be possible. One Sentence Summary CD8+ T cells that recognize a Gag peptide presented by HLA-E suppress HIV-1 replication in vitro.

Methods: Cynomolgus and rhesus macaques were infected with simian immunodeficiency virus (SIV) and followed longitudinally. Plasma levels of SIV were monitored using qRT-PCR. Bacterial genomic DNA in plasma was characterized and quantified longitudinally using 16S ribosomal deep sequencing and qPCR. ELISA-based assays were used to monitor intestinal permeability (IFABP) and perturbation of bacteria-specific host factors (sCD14 and EndoCab). Flow cytometry was used to track peripheral blood lymphocyte populations. In vitro assays were performed by exposing freshly isolated peripheral blood mononuclear cells to bacterial lysate prepared from major translocators. Effects of bacterial lysate on CD4'T cell activation and CD8'T cell cytotoxicity were measured using flow cytometry. Statistical significance was calculated using ANOVA or Wilcoxon signed-rank testing. Results: Prior to the peak of viremia, we observed a transient highlevel influx of microbial genomic DNA into peripheral blood. This microbial translocation was accompanied by perturbation of bacteria-specific host factors in plasma, as well as expansion of the CD4'CCR5'T cell compartment. Exposure of freshly isolated peripheral blood mononuclear cells to lysate prepared from major translocating taxa revealed differential taxa-specific effects on the CD4'CCR5'T cell compartment and cytotoxic granule expression within CD8'T cells.Conclusions: Altogether, our data identify the influx of microbial products into blood during hyperacute SIV infection as a candidate modifier of early interactions between the antiviral host response and nascent HIV infection. Over the next …

Introduction: Within the first weeks of human immunodeficiency virus (HIV) infection, virus replication reaches systemic circulation.Despite the critical, causal role of virus replication in determining transmissibility and kinetics of disease progression, there is limited understanding of the conditions required to transform a small localized transmitted founder population into a large and heterogeneous systemic infection.Methods: Cynomolgus and rhesus macaques were infected with simian immunodeficiency virus (SIV) and followed longitudinally.Plasma levels of SIV were monitored using qRT-PCR.Bacterial genomic DNA in plasma was characterized and quantified longitudinally using 16S ribosomal deep sequencing and qPCR.ELISA-based assays were used to monitor intestinal permeability (IFABP) and perturbation of bacteria-specific host factors (sCD14 and EndoCab).Flow cytometry was used to track peripheral blood lymphocyte populations.In vitro assays were performed by exposing freshly isolated peripheral blood mononuclear cells to bacterial lysate prepared from major translocators.Effects of bacterial lysate on CD4' T cell activation and CD8' T cell cytotoxicity were measured using flow cytometry.Statistical significance was calculated using ANOVA or Wilcoxon signed-rank testing.Results: Prior to the peak of viremia, we observed a transient highlevel influx of microbial genomic DNA into peripheral blood.This microbial translocation was accompanied by perturbation of bacteria-specific host factors in plasma, as well as expansion of the CD4'CCR5' T cell compartment.Exposure of freshly isolated peripheral blood mononuclear cells to lysate prepared from major translocating taxa revealed differential taxa-specific effects on the CD4'CCR5' T cell compartment and cytotoxic granule expression within CD8' T cells.Conclusions: Altogether, our data identify the influx of microbial products into blood during hyperacute SIV infection as a candidate modifier of early interactions between the antiviral host response and nascent HIV infection.Over the next few months, we will explore the effect of inducing microbial translocation during SIV infection, with particular interest on microbial reactivity within the CD4'CCR5' target cell compartment.