Physical and in silico immunopeptidomic profiling of a cancer antigen prostatic acid phosphatase reveals targets enabling TCR isolation.

Tissue-specific antigens can serve as targets for adoptive T cell transfer-based cancer immunotherapy. Recognition of tumor by T cells is mediated by interaction between peptide-major histocompatibility complexes (pMHCs) and T cell receptors (TCRs). Revealing the identity of peptides bound to MHC is critical in discovering cognate TCRs and predicting potential toxicity. We performed multimodal immunopeptidomic analyses for human prostatic acid phosphatase (PAP), a well-recognized tissue antigen. Three physical methods, including mild acid elution, coimmunoprecipitation, and secreted MHC precipitation, were used to capture a thorough signature of PAP on HLA-A*02:01. Eleven PAP peptides that are potentially A*02:01-restricted were identified, including five predicted strong binders by NetMHCpan 4.0. Peripheral blood mononuclear cells (PBMCs) from more than 20 healthy donors were screened with the PAP peptides. Seven cognate TCRs were isolated which can recognize three distinct epitopes when expressed in PBMCs. One TCR shows reactivity toward cell lines expressing both full-length PAP and HLA-A*02:01. Our results show that a combined multimodal immunopeptidomic approach is productive in revealing target peptides and defining the cloned TCR sequences reactive with prostatic acid phosphatase epitopes.

Peptide Presentation to T Cells: Solving the Immunogenic Puzzle: Systems Immunology Profiling of Antigen Presentation for Prediction of CD8+ T Cell Immunogenicity.

The vertebrate immune system uses an impressive arsenal of mechanisms to combat harmful cellular states such as infection. One way is via cells delivering real-time snapshots of their protein content to the cell surface in the form of short peptides. Specialized immune cells (T cells) sample these peptides and assess whether they are foreign, warranting an action such as destruction of the infected cell. The delivery of peptides to the cell surface is termed antigen processing and presentation, and decades of research have provided unprecedented understanding of this process. However, predicting the capacity for a given peptide to be immunogenic-to elicit a T cell response-has remained both enigmatic and a long sought-after goal. In the era of big data, a point is being approached where the steps of antigen processing and presentation can be quantified and assessed against peptide immunogenicity in order to build predictive models. This review presents new findings in this area and contemplates challenges ahead.

Treg-derived TGF-ß1 dampens cGAS-STING signaling to downregulate the expression of class I MHC complex in multiple myeloma.

Multiple myeloma (MM) progression involves diminished tumor antigen presentation and an immunosuppressive microenvironment, characterized by diminished expression of major histocompatibility complexes (MHC) class I molecule and elevated programmed death ligand 1 (PDL1) in MM cells, along with an enriched population of regulatory T cells (Tregs). To investigate Treg's influence on MM cells, we established a co-culture system using Tregs from MM patients and the MM cell lines (MM.1S and SK-MM-1) in vitro and assessed the effects of intervening in the relevant pathways connecting Tregs and MM cells in vivo. In vitro, Tregs induced transforming growth factor beta-1 (TGF-ß1) production, downregulated MHC I members, and increased PDL1 expression in MM cells. Treg-derived TGF-ß1 suppressed the cGAS-STING pathway, contributing to the loss of MHC I molecule expression and PDL1 upregulation. Correspondingly, neutralizing TGF-ß1 or activating the cGAS-STING pathway restored MHC I and PDL1 expression, effectively countering the pro-tumorigenic effect of Tregs on MM cells in vivo. These data elucidated how Tregs influence tumor antigen presentation and immunosuppressive signal in MM cells, potentially providing therapeutic strategies, such as neutralizing TGF-ß1 or activating the cGAS-STING pathway, to address the immune escape and immunosuppressive dynamics in MM.

The electrostatic landscape of MHC-peptide binding revealed using inception networks.

Predictive modeling of macromolecular recognition and protein-protein complementarity represents one of the cornerstones of biophysical sciences. However, such models are often hindered by the combinatorial complexity of interactions at the molecular interfaces. Exemplary of this problem is peptide presentation by the highly polymorphic major histocompatibility complex class I (MHC-I) molecule, a principal component of immune recognition. We developed human leukocyte antigen (HLA)-Inception, a deep biophysical convolutional neural network, which integrates molecular electrostatics to capture non-bonded interactions for predicting peptide binding motifs across 5,821 MHC-I alleles. These predictions of generated motifs correlate strongly with experimental peptide binding and presentation data. Beyond molecular interactions, the study demonstrates the application of predicted motifs in analyzing MHC-I allele associations with HIV disease progression and patient response to immune checkpoint inhibitors. A record of this paper's transparent peer review process is included in the supplemental information.

Self-Induced Back-Action Actuated Nanopore Electrophoresis (SANE) Sensor for Label-Free Detection of Cancer Immunotherapy-Relevant Antibody-Ligand Interactions.

We fabricated a novel single molecule nanosensor by integrating a solid-state nanopore and a double nanohole nanoaperture. The nanosensor employs Self-Induced Back-Action (SIBA) for optical trapping and enables SIBA-Actuated Nanopore Electrophoresis (SANE) for concurrent acquisition of bimodal optical and electrical signatures of molecular interactions. This work describes how to fabricate and use the SANE sensor to quantify antibody-ligand interactions. We describe how to analyze the bimodal optical-electrical data to improve upon the discrimination of antibody and ligand versus bound complex compared to electrical measurements alone. Example results for specific interaction detection are described for T-cell receptor-like antibodies (TCRmAbs) engineered to target peptide-presenting Major Histocompatibility Complex (pMHC) ligands, representing a model of target ligands presented on the surface of cancer cells. We also describe how to analyze the bimodal optical-electrical data to discriminate between specific and non-specific interactions between antibodies and ligands. Example results for non-specific interactions are shown for cancer-irrelevant TCRmAbs targeting the same pMHCs, as a control. These example results demonstrate the utility of the SANE sensor as a potential screening tool for ligand targets in cancer immunotherapy, though we believe that its potential uses are much broader.

Biological drug and drug delivery-mediated immunotherapy.

The initiation and development of major inflammatory diseases, i.e., cancer, vascular inflammation, and some autoimmune diseases are closely linked to the immune system. Biologics-based immunotherapy is exerting a critical role against these diseases, whereas the usage of the immunomodulators is always limited by various factors such as susceptibility to digestion by enzymes in vivo, poor penetration across biological barriers, and rapid clearance by the reticuloendothelial system. Drug delivery strategies are potent to promote their delivery. Herein, we reviewed the potential targets for immunotherapy against the major inflammatory diseases, discussed the biologics and drug delivery systems involved in the immunotherapy, particularly highlighted the approved therapy tactics, and finally offer perspectives in this field.

Effect of bacillus bacterial extracts on the expression of major histocompatibility complex Class I and Class II receptors on MDA-MB 231 breast cancer cells.

OBJECTIVE: The expression of human leukocyte antigen (HLA)-ABC and HLA-DR is linked to the development of breast cancer. This study was aimed to determine the effect of bacillus bacterial extracts on the expression of the major histocompatibility complexes (MHC) Class I and Class II receptors on breast cancer cells. METHODS: The expression of HLA-ABC and HLA-DR was assessed by flow cytometry and confocal microscope on the human breast tumor cell lines MDA-MB-231, which used a readily accessible model system. It is postulated that the HLA-ABC and HLA-DR receptors might be regulated by bacillus bacterial extracts designated (RA10, RA4, RA7, and RA16). RESULTS: It was observed that the treatment of cell line MDA-MB-231 with the RA10 resulted in an upregulation of the cell surface expression of the HLA-A, B, C receptors. It was observed that RA10 has the capacity to reduce the expression of HLA-A, B, C, it also shows a detectable degree of cytotoxicity when used at high concentrations. The data show that the cell surface expression of HLA-ABC is higher than HLA-DR. No significant changes of HLA-DR expression were observed on MDA-MB-231 cell lines. CONCLUSIONS: Improved understanding of the connection between HLA-ABC, HLA-DR, and bacterial extracts such as RA10 may lead to the development of drug design and therapies related to breast cancer condition in which these receptors are involved.

Empirical and Rational Design of T Cell Receptor-Based Immunotherapies.

The use of T cells reactive with intracellular tumor-associated or tumor-specific antigens has been a promising strategy for cancer immunotherapies in the past three decades, but the approach has been constrained by a limited understanding of the T cell receptor's (TCR) complex functions and specificities. Newer TCR and T cell-based approaches are in development, including engineered adoptive T cells with enhanced TCR affinities, TCR mimic antibodies, and T cell-redirecting bispecific agents. These new therapeutic modalities are exciting opportunities by which TCR recognition can be further exploited for therapeutic benefit. In this review we summarize the development of TCR-based therapeutic strategies and focus on balancing efficacy and potency versus specificity, and hence, possible toxicity, of these powerful therapeutic modalities.

A role for the thermal environment in defining co-stimulation requirements for CD4(+) T cell activation.

Maintenance of normal core body temperature is vigorously defended by long conserved, neurovascular homeostatic mechanisms that assist in heat dissipation during prolonged, heat generating exercise or exposure to warm environments. Moreover, during febrile episodes, body temperature can be significantly elevated for at least several hours at a time. Thus, as blood cells circulate throughout the body, physiologically relevant variations in surrounding tissue temperature can occur; moreover, shifts in core temperature occur during daily circadian cycles. This study has addressed the fundamental question of whether the threshold of stimulation needed to activate lymphocytes is influenced by temperature increases associated with physiologically relevant increases in temperature. We report that the need for co-stimulation of CD4+ T cells via CD28 ligation for the production of IL-2 is significantly reduced when cells are exposed to fever-range temperature. Moreover, even in the presence of sufficient CD28 ligation, provision of extra heat further increases IL-2 production. Additional in vivo and in vitro data (using both thermal and chemical modulation of membrane fluidity) support the hypothesis that the mechanism by which temperature modulates co-stimulation is linked to increases in membrane fluidity and membrane macromolecular clustering in the plasma membrane. Thermally-regulated changes in plasma membrane organization in response to physiological increases in temperature may assist in the geographical control of lymphocyte activation, i.e., stimulating activation in lymph nodes rather than in cooler surface regions, and further, may temporarily and reversibly enable CD4+ T cells to become more quickly and easily activated during times of infection during fever.