Integrative genomic analysis identifies unique immune environments associated with immunotherapy response in diffuse large B cell lymphoma.

Most diffuse large B-cell lymphoma (DLBCL) patients treated with bispecific antibodies (BsAb) or chimeric antigen receptor (CAR) T cells fail to achieve durable treatment responses, underscoring the need for a deeper understanding of mechanisms that regulate the immune environment and response to treatment. Here, an integrative, multi-omic approach was employed to characterize DLBCL immune environments, which effectively segregated DLBCLs into four quadrants - termed DLBCL-immune quadrants (IQ) - defined by cell-of-origin and immune-related gene set expression scores. Recurrent genomic alterations were enriched in each IQ, suggesting that lymphoma cell-intrinsic alterations contribute to orchestrating unique DLBCL immune environments. In relapsed/refractory DLBCL patients, DLBCL-IQ assignment correlated significantly with clinical benefit with the CD20 x CD3 BsAb, mosunetuzumab, but not with CD19-directed CAR T cells. DLBCL-IQ provides a new framework to conceptualize the DLBCL immune landscape and uncovers the differential impact of the endogenous immune environment on outcomes to BsAb and CAR T cell treatment.

MHC cross-dressing in antigen presentation.

Dendritic cells (DCs) orchestrate T cell responses by presenting antigenic peptides on major histocompatibility complex (MHC) and providing costimulation and other instructive signals. Professional antigen presenting cells (APCs), including DCs, are uniquely capable of generating and presenting peptide antigens derived from exogenous proteins. In addition to these canonical cross-presentation and MHC-II presentation pathways, APCs can also display exogenous peptide/MHC (p/MHC) acquired from neighboring cells and extracellular vesicles (EVs). This process, known as MHC cross-dressing, has been implicated in the regulation of T cell responses in a variety of in vivo contexts, including allogeneic solid organ transplantation, tumors, and viral infection. Although the occurrence of MHC cross-dressing has been clearly demonstrated, the importance of this antigen presentation mechanism continues to be elucidated. The contribution of MHC cross-dressing to overall antigen presentation has been obfuscated by the fact that DCs express the same MHC alleles as all other cells in the host, making it difficult to distinguish p/MHC generated within the DC from p/MHC acquired from another cell. As a result, much of what is known about MHC cross-dressing comes from studies using allogeneic organ transplantation and bone marrow chimeric mice, though recent development of mice bearing conditional knockout MHC and ß2-microglobulin alleles should facilitate substantial progress in the coming years. In this review, we highlight recent advances in our understanding of MHC cross-dressing and its role in activating T cell responses in various contexts, as well as the experimental insights into the mechanism by which it occurs.

Speed and navigation control of thymocyte development by the fetal T-cell gene regulatory network.

T-cell differentiation is a tightly regulated developmental program governed by interactions between transcription factors (TFs) and chromatin landscapes and affected by signals received from the thymic stroma. This process is marked by a series of checkpoints: T-lineage commitment, T-cell receptor (TCR)ß selection, and positive and negative selection. Dynamically changing combinations of TFs drive differentiation along the T-lineage trajectory, through mechanisms that have been most extensively dissected in adult mouse T-lineage cells. However, fetal T-cell development differs from adult in ways that suggest that these TF mechanisms are not fully deterministic. The first wave of fetal T-cell differentiation occurs during a unique developmental window during thymic morphogenesis, shows more rapid kinetics of differentiation with fewer rounds of cell division, and gives rise to unique populations of innate lymphoid cells (ILCs) and invariant γδT cells that are not generated in the adult thymus. As the characteristic kinetics and progeny biases are cell-intrinsic properties of thymic progenitors, the differences could be based on distinct TF network circuitry within the progenitors themselves. Here, we review recent single-cell transcriptome data that illuminate the TF networks involved in T-cell differentiation in the fetal and adult mouse thymus.

Dendritic cells can prime anti-tumor CD8+ T cell responses through major histocompatibility complex cross-dressing.

Antigen cross-presentation, wherein dendritic cells (DCs) present exogenous antigen on major histocompatibility class I (MHC-I) molecules, is considered the primary mechanism by which DCs initiate tumor-specific CD8+ T cell responses. Here, we demonstrate that MHC-I cross-dressing, an antigen presentation pathway in which DCs acquire and display intact tumor-derived peptide:MHC-I molecules, is also important in orchestrating anti-tumor immunity. Cancer cell MHC-I expression was required for optimal CD8+ T cell activation in two subcutaneous tumor models. In vivo acquisition of tumor-derived peptide:MHC-I molecules by DCs was sufficient to induce antigen-specific CD8+ T cell priming. Transfer of tumor-derived human leukocyte antigen (HLA) molecules to myeloid cells was detected in vitro and in human tumor xenografts. In conclusion, MHC-I cross-dressing is crucial for anti-tumor CD8+ T cell priming by DCs. In addition to quantitatively enhancing tumor antigen presentation, MHC cross-dressing might also enable DCs to more faithfully and efficiently mirror the cancer cell peptidome.

Divergent fates of antigen-specific CD8+ T cell clones in mice with acute leukemia.

The existence of a dysfunctional CD8+ T cell state in cancer is well established. However, the degree to which CD8+ T cell fates are influenced by the context in which they encounter cognate tumor antigen is less clear. We previously demonstrated that CD8+ T cells reactive to a model leukemia antigen were deleted by antigen cross-presenting type 1 conventional dendritic cells (cDC1s). Here, through a study of T cell receptor (TCR) transgenic CD8+ T cells (TCRTg101) reactive to a native C1498 leukemia cell antigen, we uncover a different mode of T cell tolerance in which TCRTg101 undergo progressive expansion and differentiation into an exhausted state. Antigen encounter by TCRTg101 requires leukemia cell major histocompatibility complex (MHC)-I expression and is independent of DCs, implying that leukemia cells directly mediate the exhausted TCRTg101 phenotype. Collectively, our data reveal that leukemia antigens are presented to CD8+ T cells via discrete pathways, leading to distinct tolerant states.

Negligible Role for Deletion Mediated by cDC1 in CD8+ T Cell Tolerance.

Deletion of CD8+ T cells by dendritic cells (DCs) is recognized as a critical mechanism of immune tolerance to self-antigens. Although DC-mediated peripheral deletion of autoreactive CD8+ T cells has been demonstrated using T cells reactive to model Ags, its role in shaping the naturally occurring polyclonal CD8+ T cell repertoire has not been defined. Using Batf3-/- mice lacking cross-presenting CD8α+ and CD103+ DCs (also known as type 1 conventional [cDC1]), we demonstrate that peripheral deletion of CD8+ T cells reactive to a model tissue Ag is dependent on cDC1. However, endogenous CD8+ T cells from the periphery of Batf3-/- mice do not exhibit heightened self-reactivity, and deep TCR sequencing of CD8+ T cells from Batf3-/- and Batf3+/+ mice reveals that cDC1 have a minimal impact on shaping the peripheral CD8+ T cell repertoire. Thus, although evident in reductionist systems, deletion of polyclonal self-specific CD8+ T cells by cDC1 plays a negligible role in enforcing tolerance to natural self-ligands.

PD-L1 gene alterations identify a subset of diffuse large B-cell lymphoma harboring a T-cell-inflamed phenotype.

Programmed death-ligand 1 (PD-L1) expression on malignant cells is a dominant immune escape mechanism across a variety of human cancers. A unique genetic mechanism underlying PD-L1 upregulation has been uncovered in classical Hodgkin lymphoma (cHL), in which copy gains of the chromosomal region (9p24.1) containing the programmed death-1 (PD-1) ligands PD-L1 and PD-L2 are recurrently observed. While chromosome 9p24.1 copy-number alterations are ubiquitous in cHL, they also occur in diffuse large B-cell lymphoma (DLBCL), albeit with a lower incidence. Here, fluorescence in situ hybridization was used to identify DLBCLs harboring PD-L1 gene alterations, thereby enabling a characterization of the immunogenomic landscape of these lymphomas. Among 105 DLBCL cases analyzed, PD-L1 alterations were identified in 27%. PD-L1 alterations were highly enriched among non-germinal center DLBCLs and exhibited robust PD-L1 protein expression. These lymphomas were heavily infiltrated by clonally restricted T cells and frequently downregulated human leukocyte antigen expression. RNA sequencing of PD-L1-altered DLBCLs revealed upregulation of genes involved in negative T-cell regulation and NF-κB pathway activation, while whole-exome sequencing identified frequent mutations in genes involved in antigen presentation and T-cell costimulation. Many of these findings were validated in a large external data set. Interestingly, DLBCL patients with PD-L1 alterations had inferior progression-free survival following front-line chemoimmunotherapy; however, in the relapsed/refractory setting, PD-L1 alterations were associated with response to anti-PD-1 therapy. Collectively, our results indicate that PD-L1 alterations identify a unique biological subset of DLBCL in which an endogenous antilymphoma immune response has been activated, and that is associated with responsiveness to PD-1 blockade therapy.

CD8α+ Dendritic Cells Dictate Leukemia-Specific CD8+ T Cell Fates.

APCs are essential for the orchestration of antitumor T cell responses. Batf3-lineage CD8α+ and CD103+ dendritic cells (DCs), in particular, are required for the spontaneous initiation of CD8+ T cell priming against solid tumors. In contrast, little is known about the APCs that regulate CD8+ T cell responses against hematological malignancies. Using an unbiased approach, we aimed to characterize the APCs responsible for regulating CD8+ T cell responses in a syngeneic murine leukemia model. We show with single-cell resolution that CD8α+ DCs alone acquire and cross-present leukemia Ags in vivo, culminating in the induction of leukemia-specific CD8+ T cell tolerance. Furthermore, we demonstrate that the mere acquisition of leukemia cell cargo is associated with a unique transcriptional program that may be important in regulating tolerogenic CD8α+ DC functions in mice with leukemia. Finally, we show that systemic CD8α+ DC activation with a TLR3 agonist completely prevents their ability to generate leukemia-specific CD8+ T cell tolerance in vivo, resulting instead in the induction of potent antileukemia T cell immunity and prolonged survival of leukemia-bearing mice. Together, our data reveal that Batf3-lineage DCs imprint disparate CD8+ T cell fates in hosts with solid tumors versus systemic leukemia.