Steric Shielding of Surface Epitopes and Impaired Immune Recognition Induced by the Ebola Virus Glycoprotein

Many viruses alter expression of proteins on the surface of infected cells including molecules important for immune recognition, such as the major histocompatibility complex (MHC) class I and II molecules. Virus-induced downregulation of surface proteins has been observed to occur by a variety of mechanisms including impaired transcription, blocks to synthesis, and increased turnover. Viral infection or transient expression of the Ebola virus (EBOV) glycoprotein (GP) was previously shown to result in loss of staining of various host cell surface proteins including MHC1 and β1 integrin; however, the mechanism responsible for this effect has not been delineated. In the present study we demonstrate that EBOV GP does not decrease surface levels of β1 integrin or MHC1, but rather impedes recognition by steric occlusion of these proteins on the cell surface. Furthermore, steric occlusion also occurs for epitopes on the EBOV glycoprotein itself. The occluded epitopes in host proteins and EBOV GP can be revealed by removal of the surface subunit of GP or by removal of surface N- and O- linked glycans, resulting in increased surface staining by flow cytometry. Importantly, expression of EBOV GP impairs CD8 T-cell recognition of MHC1 on antigen presenting cells. Glycan-mediated steric shielding of host cell surface proteins by EBOV GP represents a novel mechanism for a virus to affect host cell function, thereby escaping immune detection

Redirection of T cell antigen specificity through genetic transfer of supraphysiologic T cell receptors

Gene transfer of T cell receptor (TCR) αβ-chains into T cells is a promising strategy for providing a polyclonal population of T cells with redirected reactivity against tumor-associated and viral antigens. One potential problem with this approach is that TCRs, particularly those that react to tumor-associated antigens, bind to their cognate antigen with very low affinities. There is experimental evidence suggesting that TCR affinity is correlated to functional avidity, and that high-avidity T cells are more effective at clearing viral infections and tumors in murine models. In the setting of TCR gene transfer there are no studies analyzing TCR affinity and its relation to T cell effector function. In order to address this question, we studied an HLA-A*02 restricted, HIV-1 p17Gag77-85 (SL9; SLYNTVATL)-specific TCR and three derived mutants with increased affinities. We found that while we could detect differences in TCR-pMHC off-rates on a cell-based assay, CD8+ T cells transduced with all TCR constructs were able to proliferate to similar levels in response to HIVgag. Strikingly, when stimulated with targets loaded with SL9 the high-affinity TCR-transduced CD8 T cells show enhanced production of cytokines. Finally, when co-cultured with HIV-infected targets, the high affinity TCR-transduced cells are more effective at controlling HIV spread than parental TCR-transduced effectors. These data show that TCRs engineered with antibody-like affinities for antigen can induce a polyfunctional phenotype in CD8+ T cells. These receptors can be introduced into CD8- T cells to generate novel responses. The findings obtained from this work could lay the grounds to improved adoptive T cell transfer therapies for HIV and cancer

Improved Expansion and In Vivo Function of Patient T Cells by a Serum-free Medium

Improvements to T cell culture systems that promote long-term engraftment and function of adoptively transferred T cells will likely result in superior clinical benefit to more individuals. To this end, we recently developed a chemically defined cell culture medium that robustly expands all T cell subsets in the absence of human serum. Using a humanized mouse model, we observed that T cells expanded in the absence of human serum provided durable control of tumors, whereas T cells expanded in medium supplemented with human serum only mediated transient control of tumor growth. Importantly, our new medium effectively expanded more differentiated T cells from multiple myeloma patients in the absence of serum. These patient-derived T cells were also able to provide durable control of B cell tumors in vivo, and this long-term control of cancer was lost when T cells were expanded in the presence of serum. Thus, engineered T cells expanded in an optimized medium in the absence of serum may have improved therapeutic potential

Differential Reliance on Lipid Metabolism as a Salvage Pathway Underlies Functional Differences of T Cell Subsets in Poor Nutrient Environments

Summary: T cells compete with malignant cells for limited nutrients within the solid tumor microenvironment. We found that effector memory CD4 T cells respond distinctly from other T cell subsets to limiting glucose and can maintain high levels of interferon-γ (IFN-γ) production in a nutrient-poor environment. Unlike naive (TN) or central memory T (TCM) cells, effector memory T (TEM) cells fail to upregulate fatty acid synthesis, oxidative phosphorylation, and reductive glutaminolysis in limiting glucose. Interference of fatty acid synthesis in naive T cells dramatically upregulates IFN-γ, while increasing exogenous lipids in media inhibits production of IFN-γ by all subsets, suggesting that relative ratio of fatty acid metabolism to glycolysis is a direct predictor of T cell effector activity. Together, these data suggest that effector memory T cells are programmed to have limited ability to synthesize and metabolize fatty acids, which allows them to maintain T cell function in nutrient-depleted microenvironments. : Ecker et al. distinguish unique metabolic and functional properties of naive and memory T cell subsets during glucose limitation. During glucose starvation, T cells begin to differentially rely on fatty acid synthesis and glutamine utilization to survive. Unexpectedly, reliance on fatty acid synthesis alters the ability to produce IFN-γ. Keywords: lipid droplets, IFN-γ, oxidative phosphorylation, reductive glutaminolysis, serum-free media, naive T cell, glycolysis, effector memory T cell, fatty acid synthesi

Control of HIV-1 immune escape by CD8 T cells expressing enhanced T-cell receptor.

HIV's considerable capacity to vary its HLA-I-restricted peptide antigens allows it to escape from host cytotoxic T lymphocytes (CTLs). Nevertheless, therapeutics able to target HLA-I-associated antigens, with specificity for the spectrum of preferred CTL escape mutants, could prove effective. Here we use phage display to isolate and enhance a T-cell antigen receptor (TCR) originating from a CTL line derived from an infected person and specific for the immunodominant HLA-A(*)02-restricted, HIVgag-specific peptide SLYNTVATL (SL9). High-affinity (K(D) < 400 pM) TCRs were produced that bound with a half-life in excess of 2.5 h, retained specificity, targeted HIV-infected cells and recognized all common escape variants of this epitope. CD8 T cells transduced with this supraphysiologic TCR produced a greater range of soluble factors and more interleukin-2 than those transduced with natural SL9-specific TCR, and they effectively controlled wild-type and mutant strains of HIV at effector-to-target ratios that could be achieved by T-cell therapy