David Glass

All Publications

• Multi-omic profiling reveals the endogenous and neoplastic responses to immunotherapies in cutaneous T cell lymphoma. Cell reports. Medicine Glass, D. R., Mayer-Blackwell, K., Ramchurren, N., Parks, K. R., Duran, G. E., Wright, A. K., Bastidas Torres, A. N., Islas, L., Kim, Y. H., Fling, S. P., Khodadoust, M. S., Newell, E. W. 2024: 101527

  Abstract

  Cutaneous T cell lymphomas (CTCLs) are skin cancers with poor survival rates and limited treatments. While immunotherapies have shown some efficacy, the immunological consequences of administering immune-activating agents to CTCL patients have not been systematically characterized. We apply a suite of high-dimensional technologies to investigate the local, cellular, and systemic responses in CTCL patients receiving either mono- or combination anti-PD-1 plus interferon-gamma (IFN-γ) therapy. Neoplastic T cells display no evidence of activation after immunotherapy. IFN-γ induces muted endogenous immunological responses, while anti-PD-1 elicits broader changes, including increased abundance of CLA+CD39+ T cells. We develop an unbiased multi-omic profiling approach enabling discovery of immune modules stratifying patients. We identify an enrichment of activated regulatory CLA+CD39+ T cells in non-responders and activated cytotoxic CLA+CD39+ T cells in leukemic patients. Our results provide insights into the effects of immunotherapy in CTCL patients and a generalizable framework for multi-omic analysis of clinical trials.

  View details for DOI 10.1016/j.xcrm.2024.101527

  View details for PubMedID 38670099

• Type 2-polarized memory B cells hold allergen-specific IgE memory. Science translational medicine Koenig, J. F., Knudsen, N. P., Phelps, A., Bruton, K., Hoof, I., Lund, G., Libera, D. D., Lund, A., Christensen, L. H., Glass, D. R., Walker, T. D., Fang, A., Waserman, S., Jordana, M., Andersen, P. S. 2024; 16 (733): eadi0944

  Abstract

  Allergen-specific immunoglobulin E (IgE) antibodies mediate pathology in diseases such as allergic rhinitis and food allergy. Memory B cells (MBCs) contribute to circulating IgE by regenerating IgE-producing plasma cells upon allergen encounter. Here, we report a population of type 2-polarized MBCs defined as CD23hi, IL-4Rαhi, and CD32low at both the transcriptional and surface protein levels. These MBC2s are enriched in IgG1- and IgG4-expressing cells while constitutively expressing germline transcripts for IgE. Allergen-specific B cells from patients with allergic rhinitis and food allergy were enriched in MBC2s. Furthermore, MBC2s generated allergen-specific IgE during sublingual immunotherapy, thereby identifying these cells as a major reservoir for IgE. The identification of MBC2s provides insights into the maintenance of IgE memory, which is detrimental in allergic diseases but could be beneficial in protection against venoms and helminths.

  View details for DOI 10.1126/scitranslmed.adi0944

  View details for PubMedID 38324637

• Unravelling human hematopoietic progenitor cell diversity through association with intrinsic regulatory factors. bioRxiv : the preprint server for biology Favaro, P., Glass, D. R., Borges, L., Baskar, R., Reynolds, W., Ho, D., Bruce, T., Tebaykin, D., Scanlon, V. M., Shestopalov, I., Bendall, S. C. 2023

  Abstract

  Hematopoietic stem and progenitor cell (HSPC) transplantation is an essential therapy for hematological conditions, but finer definitions of human HSPC subsets with associated function could enable better tuning of grafts and more routine, lower-risk application. To deeply phenotype HSPCs, following a screen of 328 antigens, we quantified 41 surface proteins and functional regulators on millions of CD34+ and CD34- cells, spanning four primary human hematopoietic tissues: bone marrow, mobilized peripheral blood, cord blood, and fetal liver. We propose more granular definitions of HSPC subsets and provide new, detailed differentiation trajectories of erythroid and myeloid lineages. These aspects of our revised human hematopoietic model were validated with corresponding epigenetic analysis and in vitro clonal differentiation assays. Overall, we demonstrate the utility of using molecular regulators as surrogates for cellular identity and functional potential, providing a framework for description, prospective isolation, and cross-tissue comparison of HSPCs in humans.

  View details for DOI 10.1101/2023.08.30.555623

  View details for PubMedID 37693547

  View details for PubMedCentralID PMC10491219

• Magnitude and kinetics of the human immune cell response associated with severe dengue progression by single-cell proteomics. Science advances Robinson, M. L., Glass, D. R., Duran, V., Agudelo Rojas, O. L., Sanz, A. M., Consuegra, M., Sahoo, M. K., Hartmann, F. J., Bosse, M., Gelvez, R. M., Bueno, N., Pinsky, B. A., Montoya, J. G., Maecker, H., Estupiñan Cardenas, M. I., Villar Centeno, L. A., Garrido, E. M., Rosso, F., Bendall, S. C., Einav, S. 2023; 9 (12): eade7702

  Abstract

  Approximately 5 million dengue virus-infected patients progress to a potentially life-threatening severe dengue (SD) infection annually. To identify the immune features and temporal dynamics underlying SD progression, we performed deep immune profiling by mass cytometry of PBMCs collected longitudinally from SD progressors (SDp) and uncomplicated dengue (D) patients. While D is characterized by early activation of innate immune responses, in SDp there is rapid expansion and activation of IgG-secreting plasma cells and memory and regulatory T cells. Concurrently, SDp, particularly children, demonstrate increased proinflammatory NK cells, inadequate expansion of CD16+ monocytes, and high expression of the FcγR CD64 on myeloid cells, yet a signature of diminished antigen presentation. Syndrome-specific determinants include suppressed dendritic cell abundance in shock/hemorrhage versus enriched plasma cell expansion in organ impairment. This study reveals uncoordinated immune responses in SDp and provides insights into SD pathogenesis in humans with potential implications for prediction and treatment.

  View details for DOI 10.1126/sciadv.ade7702

  View details for PubMedID 36961888

• Supervised dimensionality reduction for exploration of single-cell data by HSS-LDA. Patterns (New York, N.Y.) Amouzgar, M., Glass, D. R., Baskar, R., Averbukh, I., Kimmey, S. C., Tsai, A. G., Hartmann, F. J., Bendall, S. C. 2022; 3 (8): 100536

  Abstract

  Single-cell technologies generate large, high-dimensional datasets encompassing a diversity of omics. Dimensionality reduction captures the structure and heterogeneity of the original dataset, creating low-dimensional visualizations that contribute to the human understanding of data. Existing algorithms are typically unsupervised, using measured features to generate manifolds, disregarding known biological labels such as cell type or experimental time point. We repurpose the classification algorithm, linear discriminant analysis (LDA), for supervised dimensionality reduction of single-cell data. LDA identifies linear combinations of predictors that optimally separate a priori classes, enabling the study of specific aspects of cellular heterogeneity. We implement feature selection by hybrid subset selection (HSS) and demonstrate that this computationally efficient approach generates non-stochastic, interpretable axesamenable to diverse biological processes such as differentiation over time and cell cycle. We benchmark HSS-LDA against several popular dimensionality-reduction algorithms and illustrate its utility and versatility for the exploration of single-cell mass cytometry, transcriptomics, and chromatin accessibility data.

  View details for DOI 10.1016/j.patter.2022.100536

  View details for PubMedID 36033591

• Data science through the lens of systems immunology. Patterns (New York, N.Y.) Glass, D. R., Amouzgar, M. 2022; 3 (8): 100574

  Abstract

  Glass, a post-doctoral researcher, and Amouzgar, a PhD student, in Bendall lab proposed a supervised dimensionality reduction method to explore and analyze single-cell data. Their Patterns paper highlights the advantages of supervised learning in single-cell datasets with class labels. They talk about the essential role of data science in this project and in their lives.

  View details for DOI 10.1016/j.patter.2022.100574

  View details for PubMedID 36033601

• Human IL-10-producing B cells have diverse states that are induced from multiple B cell subsets. Cell reports Glass, M. C., Glass, D. R., Oliveria, J. P., Mbiribindi, B., Esquivel, C. O., Krams, S. M., Bendall, S. C., Martinez, O. M. 2022; 39 (3): 110728

  Abstract

  Regulatory B cells (Bregs) suppress immune responses through the secretion of interleukin-10 (IL-10). This immunomodulatory capacity holds therapeutic potential, yet a definitional immunophenotype for enumeration and prospective isolation of B cells capable of IL-10 production remains elusive. Here, we simultaneously quantify cytokine production and immunophenotype in human peripheral B cells across a range of stimulatory conditions and time points using mass cytometry. Our analysis shows that multiple functional B cell subsets produce IL-10 and that no phenotype uniquely identifies IL-10+ B cells. Further, a significant portion of IL-10+ B cells co-express the pro-inflammatory cytokines IL-6 and tumor necrosis factor alpha (TNFα). Despite this heterogeneity, operationally tolerant liver transplant recipients have a unique enrichment of IL-10+, but not TNFα+ or IL-6+, B cells compared with transplant recipients receiving immunosuppression. Thus, human IL-10-producing B cells constitute an induced, transient state arising from a diversity of B cell subsets that may contribute to maintenance of immune homeostasis.

  View details for DOI 10.1016/j.celrep.2022.110728

  View details for PubMedID 35443184

• Transition to invasive breast cancer is associated with progressive changes in the structure and composition of tumor stroma. Cell Risom, T., Glass, D. R., Averbukh, I., Liu, C. C., Baranski, A., Kagel, A., McCaffrey, E. F., Greenwald, N. F., Rivero-Gutiérrez, B., Strand, S. H., Varma, S., Kong, A., Keren, L., Srivastava, S., Zhu, C., Khair, Z., Veis, D. J., Deschryver, K., Vennam, S., Maley, C., Hwang, E. S., Marks, J. R., Bendall, S. C., Colditz, G. A., West, R. B., Angelo, M. 2022; 185 (2): 299-310.e18

  Abstract

  Ductal carcinoma in situ (DCIS) is a pre-invasive lesion that is thought to be a precursor to invasive breast cancer (IBC). To understand the changes in the tumor microenvironment (TME) accompanying transition to IBC, we used multiplexed ion beam imaging by time of flight (MIBI-TOF) and a 37-plex antibody staining panel to interrogate 79 clinically annotated surgical resections using machine learning tools for cell segmentation, pixel-based clustering, and object morphometrics. Comparison of normal breast with patient-matched DCIS and IBC revealed coordinated transitions between four TME states that were delineated based on the location and function of myoepithelium, fibroblasts, and immune cells. Surprisingly, myoepithelial disruption was more advanced in DCIS patients that did not develop IBC, suggesting this process could be protective against recurrence. Taken together, this HTAN Breast PreCancer Atlas study offers insight into drivers of IBC relapse and emphasizes the importance of the TME in regulating these processes.

  View details for DOI 10.1016/j.cell.2021.12.023

  View details for PubMedID 35063072

• An Integrated Multi-omic Single-Cell Atlas of Human B Cell Identity. Immunity Glass, D. R., Tsai, A. G., Oliveria, J. P., Hartmann, F. J., Kimmey, S. C., Calderon, A. A., Borges, L. n., Glass, M. C., Wagar, L. E., Davis, M. M., Bendall, S. C. 2020; 53 (1): 217–32.e5

  Abstract

  B cells are capable of a wide range of effector functions including antibody secretion, antigen presentation, cytokine production, and generation of immunological memory. A consistent strategy for classifying human B cells by using surface molecules is essential to harness this functional diversity for clinical translation. We developed a highly multiplexed screen to quantify the co-expression of 351 surface molecules on millions of human B cells. We identified differentially expressed molecules and aligned their variance with isotype usage, VDJ sequence, metabolic profile, biosynthesis activity, and signaling response. Based on these analyses, we propose a classification scheme to segregate B cells from four lymphoid tissues into twelve unique subsets, including a CD45RB+CD27- early memory population, a class-switched CD39+ tonsil-resident population, and a CD19hiCD11c+ memory population that potently responds to immune activation. This classification framework and underlying datasets provide a resource for further investigations of human B cell identity and function.

  View details for DOI 10.1016/j.immuni.2020.06.013

  View details for PubMedID 32668225

• Single-cell metabolic profiling of human cytotoxic T cells. Nature biotechnology Hartmann, F. J., Mrdjen, D. n., McCaffrey, E. n., Glass, D. R., Greenwald, N. F., Bharadwaj, A. n., Khair, Z. n., Verberk, S. G., Baranski, A. n., Baskar, R. n., Graf, W. n., Van Valen, D. n., Van den Bossche, J. n., Angelo, M. n., Bendall, S. C. 2020

  Abstract

  Cellular metabolism regulates immune cell activation, differentiation and effector functions, but current metabolic approaches lack single-cell resolution and simultaneous characterization of cellular phenotype. In this study, we developed an approach to characterize the metabolic regulome of single cells together with their phenotypic identity. The method, termed single-cell metabolic regulome profiling (scMEP), quantifies proteins that regulate metabolic pathway activity using high-dimensional antibody-based technologies. We employed mass cytometry (cytometry by time of flight, CyTOF) to benchmark scMEP against bulk metabolic assays by reconstructing the metabolic remodeling of in vitro-activated naive and memory CD8+ T cells. We applied the approach to clinical samples and identified tissue-restricted, metabolically repressed cytotoxic T cells in human colorectal carcinoma. Combining our method with multiplexed ion beam imaging by time of flight (MIBI-TOF), we uncovered the spatial organization of metabolic programs in human tissues, which indicated exclusion of metabolically repressed immune cells from the tumor-immune boundary. Overall, our approach enables robust approximation of metabolic and functional states in individual cells.

  View details for DOI 10.1038/s41587-020-0651-8

  View details for PubMedID 32868913

• Multiplexed single-cell morphometry for hematopathology diagnostics. Nature medicine Tsai, A. G., Glass, D. R., Juntilla, M. n., Hartmann, F. J., Oak, J. S., Fernandez-Pol, S. n., Ohgami, R. S., Bendall, S. C. 2020; 26 (3): 408–17

  Abstract

  The diagnosis of lymphomas and leukemias requires hematopathologists to integrate microscopically visible cellular morphology with antibody-identified cell surface molecule expression. To merge these into one high-throughput, highly multiplexed, single-cell assay, we quantify cell morphological features by their underlying, antibody-measurable molecular components, which empowers mass cytometers to 'see' like pathologists. When applied to 71 diverse clinical samples, single-cell morphometric profiling reveals robust and distinct patterns of 'morphometric' markers for each major cell type. Individually, lamin B1 highlights acute leukemias, lamin A/C helps distinguish normal from neoplastic mature T cells, and VAMP-7 recapitulates light-cytometric side scatter. Combined with machine learning, morphometric markers form intuitive visualizations of normal and neoplastic cellular distribution and differentiation. When recalibrated for myelomonocytic blast enumeration, this approach is superior to flow cytometry and comparable to expert microscopy, bypassing years of specialized training. The contextualization of traditional surface markers on independent morphometric frameworks permits more sensitive and automated diagnosis of complex hematopoietic diseases.

  View details for DOI 10.1038/s41591-020-0783-x

  View details for PubMedID 32161403