Re-irradiation plus pembrolizumab: Phase II study for recurrent glioblastoma patients.

Purpose: Radiation therapy may enhance anti-tumor immune responses by several mechanisms including induction of immunogenic cell death. We performed a phase 2 study of pembrolizumab with re-irradiation in patients with recurrent glioblastoma.

Methods: Sixty recurrent glioblastoma patients received pembrolizumab with re-irradiation alone (cohort A, bevacizumab-naïve; n=30) or with bevacizumab continuation (cohort B, n=30).

Regulatory T cells use heparanase to access IL-2 bound to extracellular matrix in inflamed tissue.

Although FOXP3 regulatory T cells (Treg) depend on IL-2 produced by other cells for their survival and function, the levels of IL-2 in inflamed tissue are low, making it unclear how Treg access this critical resource. Here, we show that Treg use heparanase (HPSE) to access IL-2 sequestered by heparan sulfate (HS) within the extracellular matrix (ECM) of inflamed central nervous system tissue. HPSE expression distinguishes human and murine Treg from conventional T cells and is regulated by the availability of IL-2.

T cell-mediated curation and restructuring of tumor tissue coordinates an effective immune response.

Antigen-specific T cells traffic to, are influenced by, and create unique cellular microenvironments. Here we characterize these microenvironments over time with multiplexed imaging in a melanoma model of adoptive T cell therapy and human patients with melanoma treated with checkpoint inhibitor therapy. Multicellular neighborhood analysis reveals dynamic immune cell infiltration and inflamed tumor cell neighborhoods associated with CD8 T cells.

FOXP3 regulatory T cells use heparanase to access IL-2 bound to ECM in inflamed tissues.

FOXP3 regulatory T cells (Treg) depend on exogenous IL-2 for their survival and function, but circulating levels of IL-2 are low, making it unclear how Treg access this critical resource . Here, we show that Treg use heparanase (HPSE) to access IL-2 sequestered by heparan sulfate (HS) within the extracellular matrix (ECM) of inflamed central nervous system tissue. HPSE expression distinguishes human and murine Treg from conventional T cells and is regulated by the availability of IL-2.

A tissue atlas of ulcerative colitis revealing evidence of sex-dependent differences in disease-driving inflammatory cell types and resistance to TNF inhibitor therapy.

Although literature suggests that resistance to TNF inhibitor (TNFi) therapy in patients with ulcerative colitis (UC) is partially linked to immune cell populations in the inflamed region, there is still substantial uncertainty underlying the relevant spatial context. Here, we used the highly multiplexed immunofluorescence imaging technology CODEX to create a publicly browsable tissue atlas of inflammation in 42 tissue regions from 29 patients with UC and 5 healthy individuals. We analyzed 52 biomarkers on 1,710,973 spatially resolved single cells to determine cell types, cell-cell contacts, and cellular neighborhoods.

Immunotherapy of glioblastoma explants induces interferon-γ responses and spatial immune cell rearrangements in tumor center, but not periphery.

A patient-tailored, ex vivo drug response platform for glioblastoma (GBM) would facilitate therapy planning, provide insights into treatment-induced mechanisms in the immune tumor microenvironment (iTME), and enable the discovery of biomarkers of response. We cultured regionally annotated GBM explants in perfusion bioreactors to assess iTME responses to immunotherapy. Explants were treated with anti-CD47, anti-PD-1, or their combination, and analyzed by multiplexed microscopy [CO-Detection by indEXing (CODEX)], enabling the spatially resolved identification of >850,000 single cells, accompanied by explant secretome interrogation.

Biochemical, biophysical, and immunological characterization of respiratory secretions in severe SARS-CoV-2 infections.

Thick, viscous respiratory secretions are a major pathogenic feature of COVID-19, but the composition and physical properties of these secretions are poorly understood. We characterized the composition and rheological properties (i.e.

Combined protein and nucleic acid imaging reveals virus-dependent B cell and macrophage immunosuppression of tissue microenvironments.

Understanding the mechanisms of HIV tissue persistence necessitates the ability to visualize tissue microenvironments where infected cells reside; however, technological barriers limit our ability to dissect the cellular components of these HIV reservoirs. Here, we developed protein and nucleic acid in situ imaging (PANINI) to simultaneously quantify DNA, RNA, and protein levels within these tissue compartments. By coupling PANINI with multiplexed ion beam imaging (MIBI), we measured over 30 parameters simultaneously across archival lymphoid tissues from healthy or simian immunodeficiency virus (SIV)-infected nonhuman primates.

Biochemical, Biophysical, and Immunological Characterization of Respiratory Secretions in Severe SARS-CoV-2 (COVID-19) Infections.

Thick, viscous respiratory secretions are a major pathogenic feature of COVID-19 disease, but the composition and physical properties of these secretions are poorly understood. We characterized the composition and rheological properties (i.e.

CellSeg: a robust, pre-trained nucleus segmentation and pixel quantification software for highly multiplexed fluorescence images.

Background: Algorithmic cellular segmentation is an essential step for the quantitative analysis of highly multiplexed tissue images. Current segmentation pipelines often require manual dataset annotation and additional training, significant parameter tuning, or a sophisticated understanding of programming to adapt the software to the researcher's need. Here, we present CellSeg, an open-source, pre-trained nucleus segmentation and signal quantification software based on the Mask region-convolutional neural network (R-CNN) architecture.

Immune cell topography predicts response to PD-1 blockade in cutaneous T cell lymphoma.

Cutaneous T cell lymphomas (CTCL) are rare but aggressive cancers without effective treatments. While a subset of patients derive benefit from PD-1 blockade, there is a critically unmet need for predictive biomarkers of response. Herein, we perform CODEX multiplexed tissue imaging and RNA sequencing on 70 tumor regions from 14 advanced CTCL patients enrolled in a pembrolizumab clinical trial (NCT02243579).

Tissue schematics map the specialization of immune tissue motifs and their appropriation by tumors.

A schematic of a biological system, i.e., a representation of its pieces, how they are combined, and what they do, would facilitate understanding its essential organization and alteration in pathogenesis or evolution.

Subcellular localization of biomolecules and drug distribution by high-definition ion beam imaging.

Simultaneous visualization of the relationship between multiple biomolecules and their ligands or small molecules at the nanometer scale in cells will enable greater understanding of how biological processes operate. We present here high-definition multiplex ion beam imaging (HD-MIBI), a secondary ion mass spectrometry approach capable of high-parameter imaging in 3D of targeted biological entities and exogenously added structurally-unmodified small molecules. With this technology, the atomic constituents of the biomolecules themselves can be used in our system as the "tag" and we demonstrate measurements down to ~30 nm lateral resolution.

Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction.

Multiparameter tissue imaging enables analysis of cell-cell interactions in situ, the cellular basis for tissue structure, and novel cell types that are spatially restricted, giving clues to biological mechanisms behind tissue homeostasis and disease. Here, we streamlined and simplified the multiplexed imaging method CO-Detection by indEXing (CODEX) by validating 58 unique oligonucleotide barcodes that can be conjugated to antibodies. We showed that barcoded antibodies retained their specificity for staining cognate targets in human tissue.

Hyaluronan synthesis inhibition impairs antigen presentation and delays transplantation rejection.

A coat of pericellular hyaluronan surrounds mature dendritic cells (DC) and contributes to cell-cell interactions. We asked whether 4-methylumbelliferone (4MU), an oral inhibitor of HA synthesis, could inhibit antigen presentation. We find that 4MU treatment reduces pericellular hyaluronan, destabilizes interactions between DC and T-cells, and prevents T-cell proliferation in vitro and in vivo.

Biochemical and Biophysical Characterization of Respiratory Secretions in Severe SARS-CoV-2 (COVID-19) Infections.

Thick, viscous respiratory secretions are a major pathogenic feature of COVID-19 disease, but the composition and physical properties of these secretions are poorly understood. We characterized the composition and rheological properties (i.e.

Coordinated Cellular Neighborhoods Orchestrate Antitumoral Immunity at the Colorectal Cancer Invasive Front.

Antitumoral immunity requires organized, spatially nuanced interactions between components of the immune tumor microenvironment (iTME). Understanding this coordinated behavior in effective versus ineffective tumor control will advance immunotherapies. We re-engineered co-detection by indexing (CODEX) for paraffin-embedded tissue microarrays, enabling simultaneous profiling of 140 tissue regions from 35 advanced-stage colorectal cancer (CRC) patients with 56 protein markers.