Transcriptome sequencing has become common in cancer research, resulting in the generation of a substantial volume of RNA sequencing (RNA-Seq) data. The ability to extract immune repertoires from these data is crucial for obtaining information on infiltrating T- and B-lymphocyte clones when dedicated amplicon T-cell/B-cell receptors sequencing (TCR-Seq/BCR-Seq) methods are unavailable. In response to this demand, several dedicated computational methods have been developed, including MiXCR, TRUST and ImRep. In the recent publication in Briefings in Bioinformatics, Peng et al. have conducted an intensive, systematic comparison of the three previously mentioned tools. Although their effort is commendable, we do have a few constructive critiques regarding technical elements of their analysis.

• Keywords
• RNA sequencing, T-cell receptor, TCR sequencing, benchmarking, cancer immunology, computational methods, immunogenomics,
• MeSH
• B-Lymphocytes MeSH
• Benchmarking * MeSH
• Humans MeSH
• Neoplasms * genetics   MeSH
• Receptors, Antigen, T-Cell genetics   MeSH
• Sequence Analysis, RNA MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Receptors, Antigen, T-Cell MeSH

A balanced immune response is a cornerstone of healthy aging. Here, we uncover distinctive features of the long-lived blind mole-rat (Spalax spp.) adaptive immune system, relative to humans and mice. The T-cell repertoire remains diverse throughout the Spalax lifespan, suggesting a paucity of large long-lived clones of effector-memory T cells. Expression of master transcription factors of T-cell differentiation, as well as checkpoint and cytotoxicity genes, remains low as Spalax ages. The thymus shrinks as in mice and humans, while interleukin-7 and interleukin-7 receptor expression remains high, potentially reflecting the sustained homeostasis of naive T cells. With aging, immunoglobulin hypermutation level does not increase and the immunoglobulin-M repertoire remains diverse, suggesting shorter B-cell memory and sustained homeostasis of innate-like B cells. The Spalax adaptive immune system thus appears biased towards sustained functional and receptor diversity over specialized, long-lived effector-memory clones-a unique organizational strategy that potentially underlies this animal's extraordinary longevity and healthy aging.

• MeSH
• Adaptive Immunity MeSH
• Immunoglobulins metabolism   MeSH
• Interleukin-7 metabolism   MeSH
• Humans MeSH
• Mole Rats MeSH
• Mice MeSH
• Spalax * genetics   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Immunoglobulins MeSH
• Interleukin-7 MeSH

The organizational integrity of the adaptive immune system is determined by functionally discrete subsets of CD4+ T cells, but it has remained unclear to what extent lineage choice is influenced by clonotypically expressed T-cell receptors (TCRs). To address this issue, we used a high-throughput approach to profile the αβ TCR repertoires of human naive and effector/memory CD4+ T-cell subsets, irrespective of antigen specificity. Highly conserved physicochemical and recombinatorial features were encoded on a subset-specific basis in the effector/memory compartment. Clonal tracking further identified forbidden and permitted transition pathways, mapping effector/memory subsets related by interconversion or ontogeny. Public sequences were largely confined to particular effector/memory subsets, including regulatory T cells (Tregs), which also displayed hardwired repertoire features in the naive compartment. Accordingly, these cumulative repertoire portraits establish a link between clonotype fate decisions in the complex world of CD4+ T cells and the intrinsic properties of somatically rearranged TCRs.

• Keywords
• CDR3 properties, TCR repertoire, helper CD4+ subsets, human, immunology, inflammation, plasticity of CD4+ subsets,
• MeSH
• Cell Lineage immunology   MeSH
• CD4-Positive T-Lymphocytes immunology   MeSH
• Humans MeSH
• Receptors, Antigen, T-Cell, alpha-beta immunology   MeSH
• T-Lymphocyte Subsets immunology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Receptors, Antigen, T-Cell, alpha-beta MeSH

There is considerable clinical and fundamental value in measuring the clonal heterogeneity of T and B cell expansions in tumors and tumor-associated lymphoid structures-along with the associated heterogeneity of the tumor neoantigen landscape-but such analyses remain challenging to perform. Here, we propose a straightforward approach to analyze the heterogeneity of immune repertoires between different tissue sections in a quantitative and controlled way, based on a beta-binomial noise model trained on control replicates obtained at the level of single-cell suspensions. This approach allows to identify local clonal expansions with high accuracy. We reveal in situ proliferation of clonal T cells in a mouse model of melanoma, and analyze heterogeneity of immunoglobulin repertoires between sections of a metastatically-infiltrated lymph node in human melanoma and primary human colon tumor. On the latter example, we demonstrate the importance of training the noise model on datasets with depth and content that is comparable to the samples being studied. Altogether, we describe here the crucial basic instrumentarium needed to facilitate proper experimental setup planning in the rapidly evolving field of intratumoral immune repertoires, from the wet lab to bioinformatics analysis.

• Keywords
• TCR repertoire, clonal expansions, immunoglobulin repertoire, tumour clonality, tumour heterogeneity,
• Publication type
• Journal Article MeSH

In mice, the ability of naive T (TN) cells to mount an effector response correlates with TCR sensitivity for self-derived Ags, which can be quantified indirectly by measuring surface expression levels of CD5. Equivalent findings have not been reported previously in humans. We identified two discrete subsets of human CD8+ TN cells, defined by the absence or presence of the chemokine receptor CXCR3. The more abundant CXCR3+ TN cell subset displayed an effector-like transcriptional profile and expressed TCRs with physicochemical characteristics indicative of enhanced interactions with peptide-HLA class I Ags. Moreover, CXCR3+ TN cells frequently produced IL-2 and TNF in response to nonspecific activation directly ex vivo and differentiated readily into Ag-specific effector cells in vitro. Comparative analyses further revealed that human CXCR3+ TN cells were transcriptionally equivalent to murine CXCR3+ TN cells, which expressed high levels of CD5. These findings provide support for the notion that effector differentiation is shaped by heterogeneity in the preimmune repertoire of human CD8+ T cells.

• MeSH
• Lymphocyte Activation immunology   MeSH
• Biomarkers MeSH
• Cell Differentiation immunology   MeSH
• CD8-Positive T-Lymphocytes immunology   metabolism   MeSH
• Adult MeSH
• Immunophenotyping MeSH
• Immunologic Memory MeSH
• Cells, Cultured MeSH
• Middle Aged MeSH
• Humans MeSH
• Young Adult MeSH
• Mice MeSH
• Receptors, CXCR3 metabolism   MeSH
• Aged MeSH
• T-Lymphocyte Subsets immunology   metabolism   MeSH
• Age Factors MeSH
• Animals MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Young Adult MeSH
• Male MeSH
• Mice MeSH
• Aged MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Intramural MeSH
• Names of Substances
• Biomarkers MeSH
• CXCR3 protein, human MeSH Browser
• Receptors, CXCR3 MeSH

Increasing evidence suggests that antibody-drug conjugates (ADCs) can enhance anti-tumor immunity and improve clinical outcome. Here, we elucidate the therapeutic efficacy and immune-mediated mechanisms of a novel HER2-targeting ADC bearing a potent anthracycline derivate as payload (T-PNU) in a human HER2-expressing syngeneic breast cancer model resistant to trastuzumab and ado-trastuzumab emtansine. Mechanistically, the anthracycline component of the novel ADC induced immunogenic cell death leading to exposure and secretion of danger-associated molecular signals. RNA sequencing derived immunogenomic signatures and TCRβ clonotype analysis of tumor-infiltrating lymphocytes revealed a prominent role of the adaptive immune system in the regulation of T-PNU mediated anti-cancer activity. Depletion of CD8 T cells severely reduced T-PNU efficacy, thus confirming the role of cytotoxic T cells as drivers of the T-PNU mediated anti-tumor immune response. Furthermore, T-PNU therapy promoted immunological memory formation in tumor-bearing animals protecting those from tumor rechallenge. Finally, the combination of T-PNU and checkpoint inhibition, such as α-PD1, significantly enhanced tumor eradication following the treatment. In summary, a novel PNU-armed, HER2-targeting ADC elicited long-lasting immune protection in a murine orthotopic breast cancer model resistant to other HER2-directed therapies. Our findings delineate the therapeutic potential of this novel ADC payload and support its clinical development for breast cancer patients and potentially other HER2 expressing malignancies.

• Keywords
• Anthracycline, Antibody-drug conjugates, Checkpoint inhibitor combination therapy, HER2-positive breast cancer,
• MeSH
• Programmed Cell Death 1 Receptor antagonists & inhibitors   MeSH
• Anthracyclines therapeutic use   MeSH
• CD8-Positive T-Lymphocytes drug effects   immunology   MeSH
• Mammary Neoplasms, Experimental drug therapy   immunology   MeSH
• Immunoconjugates therapeutic use   MeSH
• Immunologic Memory drug effects   MeSH
• Humans MeSH
• Mice, Inbred BALB C MeSH
• Cell Line, Tumor MeSH
• Antineoplastic Agents, Immunological therapeutic use   MeSH
• Receptor, ErbB-2 antagonists & inhibitors   genetics   MeSH
• Trastuzumab therapeutic use   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Programmed Cell Death 1 Receptor MeSH
• Anthracyclines MeSH
• ERBB2 protein, human MeSH Browser
• Immunoconjugates MeSH
• Pdcd1 protein, mouse MeSH Browser
• Antineoplastic Agents, Immunological MeSH
• Receptor, ErbB-2 MeSH
• Trastuzumab MeSH

• MeSH
• Gene Regulatory Networks * MeSH
• RNA * MeSH
• Sequence Analysis, RNA MeSH
• Publication type
• Journal Article MeSH
• Comment MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• RNA * MeSH

Diverse repertoires of hypervariable immunoglobulin receptors (TCR and BCR) recognize antigens in the adaptive immune system. The development of immunoglobulin receptor repertoire sequencing methods makes it possible to perform repertoire-wide disease association studies of antigen receptor sequences. We developed a statistical framework for associating receptors to disease from only a small cohort of patients, with no need for a control cohort. Our method successfully identifies previously validated Cytomegalovirus and type one diabetes responsive TCR[Formula: see text] sequences .

• Keywords
• computational biology, condition associated receptors, human, immunology, inflammation, repertoire sequencing, statistical analysis, systems biology,
• MeSH
• Adaptive Immunity genetics   MeSH
• Cytomegalovirus immunology   MeSH
• Diabetes Mellitus genetics   immunology   MeSH
• Genetic Variation immunology   MeSH
• Complementarity Determining Regions genetics   MeSH
• Humans MeSH
• Receptors, Antigen, B-Cell genetics   MeSH
• Receptors, Antigen, T-Cell genetics   immunology   MeSH
• Receptors, Antigen genetics   immunology   MeSH
• Receptors, Immunologic genetics   immunology   MeSH
• High-Throughput Nucleotide Sequencing MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Complementarity Determining Regions MeSH
• Receptors, Antigen, B-Cell MeSH
• Receptors, Antigen, T-Cell MeSH
• Receptors, Antigen MeSH
• Receptors, Immunologic MeSH

For understanding the rules and laws of adaptive immunity, high-throughput profiling of T-cell receptor (TCR) repertoires becomes a powerful tool. The structure of TCR repertoires is instructive even before the antigen specificity of each particular receptor becomes available. It embodies information about the thymic and peripheral selection of T cells; the readiness of an adaptive immunity to withstand new challenges; the character, magnitude and memory of immune responses; and the aetiological and functional proximity of T-cell subsets. Here, we describe our current analytical approaches for the comparative analysis of murine TCR repertoires, and show several examples of how these approaches can be applied for particular experimental settings. We analyse the efficiency of different metrics used for estimation of repertoire diversity, repertoire overlap, V-gene and J-gene segments usage similarity, and amino acid composition of CDR3. We discuss basic differences of these metrics and their advantages and limitations in different experimental models, and we provide guidelines for choosing an efficient way to lead a comparative analysis of TCR repertoires. Applied to the various known and newly developed mouse models, such analysis should allow us to disentangle multiple sophisticated puzzles in adaptive immunity.

• Keywords
• T cell, T-cell receptor repertoires, aging, diversity, functional T-cell subsets,
• MeSH
• Immunity, Cellular physiology   MeSH
• Complementarity Determining Regions genetics   immunology   MeSH
• Mice MeSH
• T-Lymphocyte Subsets cytology   immunology   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Complementarity Determining Regions MeSH