Uterine Epithelial Cell Regulation of DC-SIGN Expression Inhibits Transmitted/Founder HIV-1 Trans Infection by Immature Dendritic Cells

Sexual transmission accounts for the majority of HIV-1 infections. In over 75% of cases, infection is initiated by a single variant (transmitted/founder virus). However, the determinants of virus selection during transmission are unknown. Host cell-cell interactions in the mucosa may be critical in regulating susceptibility to infection. We hypothesized in this study that specific immune modulators secreted by uterine epithelial cells modulate susceptibility of dendritic cells (DC) to infection with HIV-1.Here we report that uterine epithelial cell secretions (i.e. conditioned medium, CM) decreased DC-SIGN expression on immature dendritic cells via a transforming growth factor beta (TGF-β) mechanism. Further, CM inhibited dendritic cell-mediated trans infection of HIV-1 expressing envelope proteins of prototypic reference. Similarly, CM inhibited trans infection of HIV-1 constructs expressing envelopes of transmitted/founder viruses, variants that are selected during sexual transmission. In contrast, whereas recombinant TGF- β1 inhibited trans infection of prototypic reference HIV-1 by dendritic cells, TGF-β1 had a minimal effect on trans infection of transmitted/founder variants irrespective of the reporter system used to measure trans infection.Our results provide the first direct evidence for uterine epithelial cell regulation of dendritic cell transmission of infection with reference and transmitted/founder HIV-1 variants. These findings have immediate implications for designing strategies to prevent sexual transmission of HIV-1

Where it all begins: exploring Dendritic cell control of viral infection and cell development in the bone marrow

The bone marrow (BM) is considered the birthplace of haematopoiesis. It consists of specialised microenvironments involved in the survival and development of precursor cells and the guidance of haematopoiesis. More recently the BM has been implicated as a potential site for the initiation of primary immune responses against pathogens. Such immune responses within the BM have the potential to affect the development of hematopoietic lineages therein. However, the accessory cells potentially capable of promoting immune responses within the BM, such as dendritic cells (DC), have been only crudely characterised. Here we identify all DC populations of mouse BM and discover DC subsets that express comparable levels of MHCII and co-stimulatory markers to mature DC of the spleen. These mature, BM resident DC produce pro-inflammatory cytokines and chemokines but in the steady-state appear potentially tolerogenic with relatively poor ability to stimulate naïve T cells. While plasmacytoid (p)DC are known to mature in the BM, conventional (c)DC are thought to migrate from the BM as pre-cDC to mature in peripheral organs. As well as DC of mature phenotype, the BM also contains immature DC that are subset committed and express MHCII, but are phenotypically and functionally underdeveloped compared to DC found in peripheral lymphoid organs. Our identification of several stages of cDC development within the BM implicates it as an organ where DC can mature in entirety without the need to access peripheral organs for final stages of maturation. Lymphocytic choriomeningitis virus (LCMV) is a BM tropic virus. We investigated the behaviour of BM DC during LCMV infection. DC numbers in the BM are increased during the acute and then chronic phases of infection. Acute DC increases are attributable to enhanced development of immature and pre-DC within the BM in response to endogenous, inflammation induced fms-related tyrosine kinase 3 ligand (FL). Interestingly, BM DC developing to maturity under conditions of increased FL show enhanced antigen presenting cell functions. Furthermore, during chronic stage LCMV infection the most mature DC subset of the BM, present in very low numbers during steady-state, is remarkably increased within the BM. It thus appears the BM plays varying roles supporting these novel DC subsets during acute and chronic phases of infection. Our findings implicate BM DC as regulators of cellular homeostasis during steady-state conditions that can rapidly respond to viral infection by increasing in number and antigen presentation potency. Such responses reveal the BM microenvironment to be a site for the rapid development of “emergency”, inflammation induced DC that likely re-seed DC populations of peripheral organs during infection, but also as a site for the accumulation of mature DC potentially capable of initiating immune responses therein. Our elucidation of these mature and precursor DC within the BM identifies novel cellular targets for clinical manipulation of immune responses

Abstract 2793: Minimal residual disease (MRD) as a predictive biomarker/endpoint for novel drug development

Abstract MRD is defined as the continual presence of cancer cells in patients post therapy that is below the threshold of detection by conventional morphologic or radiologic assessments. Monitoring patient status clinically as MRD negative or MRD positive has become the basis for predicting outcomes, remission or reoccurrence, as well as determining therapeutic options. The three primary methods to assess MRD status in patients include Flow Cytometry (FC), Polymerase Chain Reaction (PCR) and Next-Generation Sequencing (NGS). All three are highly quantitative measurements with a high degree of sensitivity to detect “new” circulating tumor cells or ctDNA that can potentially indicate disease recurrence before it happens. The choice of MRD methodology usually depends on the tumor type, liquid or solid. Flow Cytometry has been widely used to monitor MRD status in patients with hematologic malignancies. Specifically designed MRD panels have been used clinically in CLL, MM, CML and AML patients. In addition to its prognostic/diagnostic relevance, recent guidance by the FDA is now driving the use of MRD testing as a predictive biomarker/endpoint for novel cancer drug development. NeoGenomics Pharma Services has provided flow cytometry MRD expertise to multiple sponsors conducting clinical trials to evaluate the effectiveness of novel cancer therapy. In a CLL study, NeoGenomics employed the ERIC MRD assessment to monitor patient status during therapeutic intervention at various points throughout treatment. Representative patient data show progression from full, active disease prior to therapy to non-detectable disease levels below 0.01%MRD. These examples show patient progression through multiple cycles of treatment to full remission. Clearly, this is an example of how MRD can serve as the primary “biomarker” to evaluate a patient’s response to novel drug therapy in clinical trials. This presentation will further discuss the data and the relevance of MRD testing to support new drug development. Citation Format: Nicholas Jones, Brian Ngo, Floyd Davis, Benjamin Fancke. Minimal residual disease (MRD) as a predictive biomarker/endpoint for novel drug development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2793.</jats:p

Monitoring dendritic cell activation and maturation

Since the 1997 discovery that the first identified human homolog of Drosophila Toll could activate the innate immune system, the innate arm of immunity has rapidly taken on a new light as an important player in the recognition of pathogens and damaged self. The recognition of danger by dendritic cells (DC) is a crucial step in activating the adaptive immune system. Different DC express varied subsets of pattern recognition receptors (PRR), enabling both overlap and exclusivity in the recognition of danger signals by DC. PRR-mediated DC maturation and activation can be measured by changes in the surface expression of costimulatory as well as coinhibitory molecules, changes in size and shape of the DC and by their production of different cytokines

Use of Receptor Occupancy Assays on Cells from Peripheral Blood As a Model to Determine Therapeutic Efficacy

Abstract In the development of monoclonal antibodies for the treatment of liquid or solid tumors, monitoring receptor occupancy (RO) on peripheral blood lymphocytes can help illustrate potential therapeutic activity occurring systemically, or in the tumor environment. There is increased interest in using these assessments on cells from the periphery to assess the efficacy of investigational therapies. In this area of therapeutic development, much focus has been placed on checkpoint inhibition proteins/receptors such as PD-1, PD-L1, TIM3, CTLA-4, TIGIT, and multiple others. These immune checkpoint inhibitors are suitable targets since they can be highly modulated on exhausted T cells in cancer patients. In the case of solid tumors, assessing the expression of these markers in the tumor itself via biopsy would be an invasive process that could not be performed sequentially and, therefore, would provide little information to correlate with pharmacokinetic results throughout the course of a clinical trial to determine timing and optimal dose selection. However, assessment of the receptor expression and occupancy in combination with pharmacokinetics can lead to a better understanding of drug levels required to achieve optimal therapeutic performance. Monitoring the expression and binding of these molecules by a drug can determine specific treatment based on the ability of the patient's own immune system to act, while providing information to identify therapeutic responses against cancer. This poster will present an example of receptor occupancy (RO) assessment in its development and implementation. Disclosures No relevant conflicts of interest to declare. </jats:sec

Abstract 4041: Use of Receptor Occupancy Assays on cells from Peripheral Blood as a Surrogate Model of the Tumor Microenvironment

Abstract In the development of monoclonal antibodies for the treatment of solid tumors, monitoring receptor occupancy (RO) on peripheral blood lymphocytes can help illustrate potential therapeutic activity occurring in the tumor environment as an assessment of pharmacodynamics (PD). There is increased interest in using these PD assessments on cells from the periphery to assess the effectiveness and efficacy of investigational therapies. In this area of therapeutic development, much focus has been placed on checkpoint inhibitors proteins/receptors such as PD-1, PD-L1, TIM3, CTLA-4, TIGIT, and multiple others. These immune checkpoint inhibitors are suitable targets since they can be upregulated and/or modulated on exhausted T cells in cancer patients. Assessing the expression of these markers in the tumor itself would be an invasive process and provide little information to correlate with pharmacokinetic results throughout the course of a clinical trial to determine optimal dose selection. However, assessment of the receptor expression and occupancy in combination with pharmacokinetics can lead to a better understanding of drug levels required to achieve optimal therapeutic performance. Monitoring the expression and binding of these molecules by drug can determine specific treatment based on the ability of the patient’s own immune system to act, while providing information to identify therapeutic responses against cancer. This poster will present an example of one such RO assessment in its development and implementation. Citation Format: Nicholas Jones, Brian Ngo, Benjamin Fancke, Jessica Limson, Floyd Davis. Use of Receptor Occupancy Assays on cells from Peripheral Blood as a Surrogate Model of the Tumor Microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4041.</jats:p