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Abstract Major histocompatibility complex class I (MHC I) molecules present peptides to CD8+ T‐cells for immunosurveillance of infection and cancer. Recent studies indicate lineage‐specific heterogeneity in MHC I expression. While respiratory diseases rank among the leading causes of mortality, studies in mice have shown that lung epithelial cells (LECs) express the lowest levels of MHC I in the lung. This study aims to answer three questions: (i) Do human LECs express low levels of MHC I? (ii) Is LEC MHC I expression modulated in chronic respiratory diseases? (iii) Which factors regulate MHC I levels in human LECs? We analyzed human LECs from parenchymal explants using single‐cell RNA sequencing and immunostaining. We confirmed low constitutive MHC I expression in human LECs, with significant upregulation in chronic respiratory diseases. We observed a sexual dimorphism, with males having higher MHC I levels under steady‐state conditions, likely due to differential redox balance. Our study unveils the complex interplay between MHC I expression, sex, and respiratory disease. Since MHC I upregulation contributes to the development of immunopathologies in other models, we propose that it may have a similar impact on chronic lung disease.

Epithelial ovarian cancer (EOC) has not significantly benefited from advances in immunotherapy, mainly because of the lack of well-defined actionable antigen targets. Using proteogenomic analyses of primary EOC tumors, we previously identified 91 aberrantly expressed tumor-specific antigens (TSAs) originating from unmutated genomic sequences. Most of these TSAs derive from non-exonic regions, and their expression results from cancer-specific epigenetic changes. The present study aimed to evaluate the immunogenicity of 48 TSAs selected according to two criteria: presentation by highly prevalent HLA allotypes and expression in a significant fraction of EOC tumors. Using targeted mass spectrometry analyses, we found that pulsing with synthetic TSA peptides leads to a high-level presentation on dendritic cells. TSA abundance correlated with the predicted binding affinity to the HLA allotype. We stimulated naïve CD8 T cells from healthy blood donors with TSA-pulsed dendritic cells and assessed their expansion with two assays: MHC-peptide tetramer staining and TCR Vβ CDR3 sequencing. We report that these TSAs can expand sizeable populations of CD8 T cells and, therefore, represent attractive targets for EOC immunotherapy.

Transposable elements (TEs) are repetitive sequences representing ~45% of the human and mouse genomes and are highly expressed by medullary thymic epithelial cells (mTECs). In this study, we investigated the role of TEs on T-cell development in the thymus. We performed multiomic analyses of TEs in human and mouse thymic cells to elucidate their role in T-cell development. We report that TE expression in the human thymus is high and shows extensive age- and cell lineage-related variations. TE expression correlates with multiple transcription factors in all cell types of the human thymus. Two cell types express particularly broad TE repertoires: mTECs and plasmacytoid dendritic cells (pDCs). In mTECs, transcriptomic data suggest that TEs interact with transcription factors essential for mTEC development and function (e.g., PAX1 and REL), and immunopeptidomic data showed that TEs generate MHC-I-associated peptides implicated in thymocyte education. Notably, AIRE, FEZF2, and CHD4 regulate small yet non-redundant sets of TEs in murine mTECs. Human thymic pDCs homogenously express large numbers of TEs that likely form dsRNA, which can activate innate immune receptors, potentially explaining why thymic pDCs constitutively secrete IFN ɑ/β. This study highlights the diversity of interactions between TEs and the adaptive immune system. TEs are genetic parasites, and the two thymic cell types most affected by TEs (mTEcs and pDCs) are essential to establishing central T-cell tolerance. Therefore, we propose that orchestrating TE expression in thymic cells is critical to prevent autoimmunity in vertebrates.

Abstract MHC-I-associated peptides deriving from non-coding genomic regions and mutations can generate tumor-specific antigens, including neoantigens. Quantifying tumor-specific antigens’ RNA expression in malignant and benign tissues is critical for discriminating actionable targets. We present BamQuery, a tool attributing an exhaustive RNA expression to MHC-I-associated peptides of any origin from bulk and single-cell RNA-sequencing data. We show that many cryptic and mutated tumor-specific antigens can derive from multiple discrete genomic regions, abundantly expressed in normal tissues. BamQuery can also be used to predict MHC-I-associated peptides immunogenicity and identify actionable tumor-specific antigens de novo.

Pregnancy causes abrupt thymic atrophy. This atrophy is characterized by a severe decrease in the number of all thymocyte subsets and qualitative (but not quantitative) changes in thymic epithelial cells (TECs). Pregnancy-related thymic involution is triggered by progesterone-induced functional changes affecting mainly cortical TECs (cTECs). Remarkably, this severe involution is rapidly corrected following parturition. We postulated that understanding the mechanisms of pregnancy-related thymic changes could provide novel insights into signaling pathways regulating TEC function. When we analyzed genes whose expression in TECs was modified during late pregnancy, we found a strong enrichment in genes bearing KLF4 transcription factor binding motifs. We, therefore, engineered a Psmb11-iCre : Klf4lox/lox mouse model to study the impact of TEC-specific Klf4 deletion in steady-state conditions and during late pregnancy. Under steady-state conditions, Klf4 deletion had a minimal effect on TEC subsets and did not affect thymic architecture. However, pregnancy-induced thymic involution was much more pronounced in pregnant females lacking Klf4 expression in TECs. These mice displayed a substantial ablation of TECs with a more pronounced loss of thymocytes. Transcriptomic and phenotypic analyses of Klf4-/- TECs revealed that Klf4 maintains cTEC numbers by supporting cell survival and preventing epithelial-to-mesenchymal plasticity during late pregnancy. We conclude that Klf4 is essential for preserving TEC’s integrity and mitigating thymic involution during late pregnancy.

Summary: Based on analyses of TCR sequences from over 1,000 individuals, we report that the TCR repertoire is composed of two ontogenically and functionally distinct types of TCRs. Their production is regulated by variations in thymic output and terminal deoxynucleotidyl transferase (TDT) activity. Neonatal TCRs derived from TDT-negative progenitors persist throughout life, are highly shared among subjects, and are reported as disease-associated. Thus, 10%–30% of most frequent cord blood TCRs are associated with common pathogens and autoantigens. TDT-dependent TCRs present distinct structural features and are less shared among subjects. TDT-dependent TCRs are produced in maximal numbers during infancy when thymic output and TDT activity reach a summit, are more abundant in subjects with AIRE mutations, and seem to play a dominant role in graft-versus-host disease. Factors decreasing thymic output (age, male sex) negatively impact TCR diversity. Males compensate for their lower repertoire diversity via hyperexpansion of selected TCR clonotypes.

Abstract Background Endogenous retroelements (EREs) constitute about 42% of the human genome and have been implicated in common human diseases such as autoimmunity and cancer. The dominant paradigm holds that EREs are expressed in embryonic stem cells (ESCs) and germline cells but are repressed in differentiated somatic cells. Despite evidence that some EREs can be expressed at the RNA and protein levels in specific contexts, a system-level evaluation of their expression in human tissues is lacking. Methods Using RNA sequencing data, we analyzed ERE expression in 32 human tissues and cell types, including medullary thymic epithelial cells (mTECs). A tissue specificity index was computed to identify tissue-restricted ERE families. We also analyzed the transcriptome of mTECs in wild-type and autoimmune regulator (AIRE)-deficient mice. Finally, we developed a proteogenomic workflow combining RNA sequencing and mass spectrometry (MS) in order to evaluate whether EREs might be translated and generate MHC I-associated peptides (MAP) in B-lymphoblastoid cell lines (B-LCL) from 16 individuals. Results We report that all human tissues express EREs, but the breadth and magnitude of ERE expression are very heterogeneous from one tissue to another. ERE expression was particularly high in two MHC I-deficient tissues (ESCs and testis) and one MHC I-expressing tissue, mTECs. In mutant mice, we report that the exceptional expression of EREs in mTECs was AIRE-independent. MS analyses identified 103 non-redundant ERE-derived MAPs (ereMAPs) in B-LCLs. These ereMAPs preferentially derived from sense translation of intronic EREs. Notably, detailed analyses of their amino acid composition revealed that ERE-derived MAPs presented homology to viral MAPs. Conclusions This study shows that ERE expression in somatic tissues is more pervasive and heterogeneous than anticipated. The high and diversified expression of EREs in mTECs and their ability to generate MAPs suggest that EREs may play an important role in the establishment of self-tolerance. The viral-like properties of ERE-derived MAPs suggest that those not expressed in mTECs can be highly immunogenic.

MHC-I associated peptides (MAPs) play a central role in the elimination of virus-infected and neoplastic cells by CD8 T cells. However, accurately predicting the MAP repertoire remains difficult, because only a fraction of the transcriptome generates MAPs. In this study, we investigated whether codon arrangement (usage and placement) regulates MAP biogenesis. We developed an artificial neural network called Codon Arrangement MAP Predictor (CAMAP), predicting MAP presentation solely from mRNA sequences flanking the MAP-coding codons (MCCs), while excluding the MCC per se. CAMAP predictions were significantly more accurate when using original codon sequences than shuffled codon sequences which reflect amino acid usage. Furthermore, predictions were independent of mRNA expression and MAP binding affinity to MHC-I molecules and applied to several cell types and species. Combining MAP ligand scores, transcript expression level and CAMAP scores was particularly useful to increase MAP prediction accuracy. Using an in vitro assay, we showed that varying the synonymous codons in the regions flanking the MCCs (without changing the amino acid sequence) resulted in significant modulation of MAP presentation at the cell surface. Taken together, our results demonstrate the role of codon arrangement in the regulation of MAP presentation and support integration of both translational and post-translational events in predictive algorithms to ameliorate modeling of the immunopeptidome. Author summary MHC-I associated peptides (MAPs) are small fragments of intracellular proteins presented at the surface of cells and used by the immune system to detect and eliminate cancerous or virus-infected cells. While it is theoretically possible to predict which portions of the intracellular proteins will be naturally processed by the cells to ultimately reach the surface, current methodologies have prohibitively high false discovery rates. Here we introduce an artificial neural network called Codon Arrangement MAP Predictor (CAMAP) which integrates information from mRNA-to-protein translation to other factors regulating MAP biogenesis (e.g. MAP ligand score and transcript expression levels) to improve MAP prediction accuracy. While most MAP predictive approaches focus on MAP sequences per se, CAMAP’s novelty is to analyze the MAP-flanking mRNA sequences, thereby providing completely independent information for MAP prediction. We show on several datasets that the integration of CAMAP scores with other known factors involved in MAP presentation (i.e. MAP ligand score and mRNA expression) significantly improves MAP prediction accuracy, and further validate CAMAP learned features using an in-vitro assay. These findings may have major implications for the design of vaccines against cancers and viruses, and in times of pandemics could accelerate the identification of relevant MAPs of viral origins.

Summary: Combining RNA sequencing, ribosome profiling, and mass spectrometry, we elucidate the contribution of non-canonical translation to the proteome and major histocompatibility complex (MHC) class I immunopeptidome. Remarkably, of 14,498 proteins identified in three human B cell lymphomas, 2,503 are non-canonical proteins. Of these, 28% are novel isoforms and 72% are cryptic proteins encoded by ostensibly non-coding regions (60%) or frameshifted canonical genes (12%). Cryptic proteins are translated as efficiently as canonical proteins, have more predicted disordered residues and lower stability, and critically generate MHC-I peptides 5-fold more efficiently per translation event. Translating 5′ “untranslated” regions hinders downstream translation of genes involved in transcription, translation, and antiviral responses. Novel protein isoforms show strong enrichment for signaling pathways deregulated in cancer. Only a small fraction of cryptic proteins detected in the proteome contribute to the MHC-I immunopeptidome, demonstrating the high preferential access of cryptic defective ribosomal products to the class I pathway.

The search for tumor-specific antigens (TSAs) has considerably accelerated during the past decade due to the improvement of proteogenomic detection methods. This provides new opportunities for the development of novel antitumoral immunotherapies to mount an efficient T cell response against one or multiple types of tumors. While the identification of mutated antigens originating from coding exons has provided relatively few TSA candidates, the possibility of enlarging the repertoire of targetable TSAs by looking at antigens arising from non-canonical open reading frames opens up interesting avenues for cancer immunotherapy. In this review, we outline the potential sources of TSAs and the mechanisms responsible for their expression strictly in cancer cells. In line with the heterogeneity of cancer, we propose that discrete families of TSAs may be enriched in specific cancer types.

During gestation, sex hormones cause a significant thymic involution which enhances fertility. This thymic involution is rapidly corrected following parturition. As thymic epithelial cells (TECs) are responsible for the regulation of thymopoiesis, we analyzed the sequential phenotypic and transcriptomic changes in TECs during the postpartum period in order to identify mechanisms triggering postpartum thymic regeneration. In particular, we performed flow cytometry analyses and deep RNA-sequencing on purified TEC subsets at several time points before and after parturition. We report that pregnancy-induced involution is not caused by loss of TECs since their number does not change during or after pregnancy. However, during pregnancy, we observed a significant depletion of all thymocyte subsets downstream of the double-negative 1 (DN1) differentiation stage. Variations in thymocyte numbers correlated with conspicuous changes in the transcriptome of cortical TECs (cTECs). The transcriptomic changes affected predominantly cTEC expression of Foxn1, its targets and several genes that are essential for thymopoiesis. By contrast, medullary TECs (mTECs) showed very little transcriptomic changes in the early postpartum regenerative phase, but seemed to respond to the expansion of single-positive (SP) thymocytes in the late phase of regeneration. Together, these results show that postpartum thymic regeneration is orchestrated by variations in expression of a well-defined subset of cTEC genes, that occur very early after parturition.

Intensive fundamental and clinical research in cancer immunotherapy has led to the emergence and evolution of two parallel universes with surprisingly little interactions: the realm of hematologic malignancies and that of solid tumors. Treatment of hematologic cancers using allogeneic hematopoietic cell transplantation (AHCT) serendipitously led to the discovery that T cells specific for minor histocompatibility antigens (MiHAs) could cure hematopoietic cancers. Besides, studies based on treatment of solid tumor with ex vivo-expanded tumor infiltrating lymphocytes or immune checkpoint therapy demonstrated that anti-tumor responses could be achieved by targeting tumor-specific antigens (TSAs). It is our contention that much insight can be gained by sharing the tremendous amount of data generated in the two-abovementioned universes. Our perspective article has two specific goals. First, to discuss the value of methods currently used for MiHA and TSA discovery and to explain the key role of mass spectrometry analyses in this process. Second, to demonstrate the importance of broadening the scope of TSA discovery efforts beyond classic annotated protein-coding genomic sequences.

Editorial summary The nature of the tumor antigens that are detectable by T cells remains unclear. In melanoma, T cells were shown to react against major histocompatibility complex (MHC)-associated peptides (MAPs) that are derived from exonic mutations. A recent multi-omic study of hepatocellular carcinomas suggests, however, that mutated exonic MAPs were exceedingly rare, bringing the accuracy of the current methods for antigen identification into question and demonstrating the importance of broadening tumor-antigen discovery efforts.

The sole nonredundant role of the thymic medulla is to induce central tolerance, a vital process that depends on promiscuous gene expression (pGE), a unique feature of medullary thymic epithelial cells (mTECs). Although pGE enhances transcription of >3,000 genes in mTECs, its impact on the regulation of protein homeostasis remains unexplored. Here, we report that, because of pGE, mature mTECs synthesize substantially more proteins than other cell types and are exquisitely sensitive to loss of immunoproteasomes (IPs). Indeed, IP deficiency causes proteotoxic stress in mTECs and leads to exhaustion of postnatal mTEC progenitors. Moreover, IP-deficient mice show accelerated thymic involution, which is characterized by a selective loss of mTECs and multiorgan autoimmune manifestations. We conclude that pGE, the quintessential feature of mTECs, is a major burden for the maintenance of proteostasis, which is alleviated by the constitutive expression of IPs in mTECs.

Cryptic translation of the 'non-coding' genome is increasingly recognised, however its biological significance remains unclear. Laumont et al.employ proteogenomic techniques to map the human immunoproteome, and find that approximately 10% of MHC class I-associated peptides are cryptic.

Thymic aging precedes that of other organs and is initiated by the gradual loss of thymic epithelial cells (TECs). Based on in vitro culture and transplantation assays, recent studies have reported on the presence of thymic epithelial progenitor cells (TEPCs) in young adult mice. However, the physiological role and properties of TEPC populations reported to date remain unclear. Using an in vivo label-retention assay, we previously identified a population of quiescent but non-senescent TECs. The goals of this study were therefore (i) to evaluate the contribution of these quiescent TECs to thymic regeneration following irradiation-induced acute thymic injury and (ii) to characterize their phenotypic and molecular profiles using flow cytometry, immunohistology, and transcriptome sequencing. We report that while UEA1+ cells cycle the most in steady state, they are greatly affected by irradiation, leading to cell loss and proliferative arrest following acute thymic involution. On the opposite, the UEA1– subset of quiescent TECs is radioresistant and proliferate in situ following acute thymic involution, thereby contributing to thymic regeneration in 28- to 30-week-old mice. UEA1– quiescent TECs display an undifferentiated phenotype (co-expression of K8 and K5 cytokeratins) and express high levels of genes that regulate stem cell activity in different tissues (e.g., Podxl and Ptprz1). In addition, two features suggest that UEA1– quiescent TECs occupy discrete stromal niches: (i) their preferential location in clusters adjacent to the cortico-medullary junction and (ii) their high expression of genes involved in cross talk with mesenchymal cells. The ability of UEA1– quiescent TECs to participate to TEC regeneration qualifies them as in vivo progenitor cells particularly relevant in the context of regeneration following acute thymic injury.

pyGeno is a Python package mainly intended for precision medicine applications that revolve around genomics and proteomics. It integrates reference sequences and annotations from Ensembl, genomic polymorphisms from the dbSNP database and data from next-gen sequencing into an easy to use, memory-efficient and fast framework, therefore allowing the user to easily explore subject-specific genomes and proteomes. Compared to a standalone program, pyGeno gives the user access to the complete expressivity of Python, a general programming language. Its range of application therefore encompasses both short scripts and large scale genome-wide studies.

Abstract Self/non‐self discrimination is a fundamental requirement of life. Endogenous peptides presented by major histocompatibility complex class I (MHC I) molecules represent the essence of self for CD8 T lymphocytes. These MHC I peptides (MIPs) are collectively referred to as the immunopeptidome. From a systems‐level perspective, very little is known about the origin, composition and plasticity of the immunopeptidome. Here, we show that the immunopeptidome, and therefore the nature of the immune self, is plastic and moulded by cellular metabolic activity. By using a quantitative high‐throughput mass spectrometry‐based approach, we found that altering cellular metabolism via the inhibition of the mammalian target of rapamycin results in dynamic changes in the cell surface MIPs landscape. Moreover, we provide systems‐level evidence that the immunopeptidome projects at the cell surface a representation of biochemical networks and metabolic events regulated at multiple levels inside the cell. Our findings open up new perspectives in systems immunology and predictive biology. Indeed, predicting variations in the immunopeptidome in response to cell‐intrinsic and ‐extrinsic factors could be relevant to the rational design of immunotherapeutic interventions.

Abstract The mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation. mTOR signaling is frequently dysregulated in oncogenic cells, and thus an attractive target for anticancer therapy. Using CellDesigner, a modeling support software for graphical notation, we present herein a comprehensive map of the mTOR signaling network, which includes 964 species connected by 777 reactions. The map complies with both the systems biology markup language (SBML) and graphical notation (SBGN) for computational analysis and graphical representation, respectively. As captured in the mTOR map, we review and discuss our current understanding of the mTOR signaling network and highlight the impact of mTOR feedback and crosstalk regulations on drug‐based cancer therapy. This map is available on the Payao platform, a Web 2.0 based community‐wide interactive process for creating more accurate and information‐rich databases. Thus, this comprehensive map of the mTOR network will serve as a tool to facilitate systems‐level study of up‐to‐date mTOR network components and signaling events toward the discovery of novel regulatory processes and therapeutic strategies for cancer.

Both physiological and psychological stress cause thymic atrophy via glucocorticoïd (GC)-dependent apoptosis of double-positive (DP) thymocytes. Given the pervasiveness of stress, GC-induced thymic atrophy is arguably the most common type of acquired immunodeficiency. We recently reported that interleukin-21 (IL-21) has a unique ability to expand the small subset of DP thymocytes (CD69(+)) which are ongoing positive selection, and that administration of IL-21 increases thymic output in aged mice. The goal of this study was to evaluate whether IL-21 could mitigate GC-induced thymic atrophy. In contrast to double-negative (DN) and single-positive (SP) thymocytes, most DP thymocytes (CD69(-)) do not constitutively express the IL-21 receptor (IL-21R). Accordingly, CD69(-) DP thymocytes from PBS-treated mice were unresponsive to IL-21 administration. However, following GC injection, surviving CD69(-) DP thymocytes up-regulated IL-21R and responded to IL-21 treatment as evidenced by enhancement of Bcl6 expression and phosphorylation of STAT1, STAT3 and STAT5. Consequently, IL-21 administration to GC-treated mice accelerated thymic recovery by expanding considerably DP thymocytes and, to a lesser extent, DN thymocytes. However, IL-21-induced expansion of DN/DP thymocytes did not alter the diversity of the intrathymic or peripheral T-cell receptor (TCR) repertoire. We conclude that IL-21 dramatically accelerates recovery from GC-induced thymic atrophy.

While the role of canonical (β-catenin-mediated) Wnt signaling in hematolymphopoiesis has been studied extensively, little is known of the potential importance of non-canonical Wnt signals in hematopoietic cells. Wnt4 is one of the Wnt proteins that can elicit non-canonical pathways. We have previously shown that retroviral overexpression of Wnt4 by hematopoietic cells increased thymic cellularity as well as the frequency of early thymic progenitors and bone marrow hematopoietic progenitor cells (HPCs). However, the molecular pathways responsible for its effect in HPCs are not known.Here we report that Wnt4 stimulation resulted in the activation of the small GTPase Rac1 as well as Jnk kinases in an HPC cell line. Jnk activity was necessary, while β-catenin was dispensable, for the Wnt4-mediated expansion of primary fetal liver HPCs in culture. Furthermore, Jnk2-deficient and Wnt4 hemizygous mice presented lower numbers of HPCs in their bone marrow, and Jnk2-deficient HPCs showed increased rates of apoptosis. Wnt4 also improved HPC activity in a competitive reconstitution model in a cell-autonomous, Jnk2-dependent manner. Lastly, we identified Fz6 as a receptor for Wnt4 in immature HPCs and showed that the absence of Wnt4 led to a decreased expression of four polarity complex genes.Our results establish a functional role for non-canonical Wnt signaling in hematopoiesis through a pathway involving Wnt4, Fz6, Rac1 and Jnk kinases.

The cystatin protein superfamily is characterized by the presence of conserved sequences that display cysteine protease inhibitory activity (e.g., towards cathepsins). Type 1 and 2 cystatins are encoded by 25 genes of which 23 are grouped in 2 clusters localized on mouse chromosomes 16 and 2. The expression and essential roles of most of these genes in mouse development and hematopoiesis remain poorly characterized. In this study, we describe a set of quantitative real-time PCR assays and a global expression profile of cystatin genes in normal mouse tissues. Benefiting from our collection of DelES embryonic stem cell clones harboring large chromosomal deletions (to be reported elsewhere), we selected a clone in which a 95-kb region of chromosome 16 is missing (Del(16qB3Delta/+)). In this particular clone, 2 cystatin genes, namely Csta and Stfa2l1 are absent along with 2 other genes (Fam162a, Ccdc58) and associated intergenic regions. From this line, we established a new homozygous mutant mouse model (Del(16qB3Delta/16qB3Delta)) to assess the in vivo biological functions of the 2 deleted cystatins. Stfa2l1 gene expression is high in wild-type fetal liver, bone marrow, and spleen, while Csta is ubiquitously expressed. Homozygous Del(16qB3Delta/16qB3Delta) animals are phenotypically normal, fertile, and not overtly susceptible to spontaneous or irradiation-induced tumor formation. The hematopoietic stem and progenitor cell activity in these mutant mice are also normal. Interestingly, quantitative real-time PCR expression profiling reveals a marked increase in the expression levels of Stfa2l1/Csta phylogenetically-related genes (Stfa1, Stfa2, and Stfa3) in Del(16qB3Delta/16qB3Delta) hematopoietic tissues, suggesting that these candidate genes might be contributing to compensatory mechanisms. Overall, this study presents an optimized approach to globally monitor cystatin gene expression as well as a new mouse model deficient in Stfa2l1/Csta genes, expanding the available tools to dissect cystatin roles under normal and pathological conditions.

BACKGROUND: Injection of CD8 T cells primed against immunodominant minor histocompatibility antigens (MiHA) such as H7(a) can eradicate leukemia and solid tumors. To understand why MiHA-targeted T cells have such a potent antitumor effect it is essential to evaluate their in vivo behavior. In the present work, we therefore addressed two specific questions: what is the proliferative dynamics of H7(a)-specifc T cells in tumors, and do H7(a)-specific T cells persist long-term after adoptive transfer? METHODOLOGY/PRINCIPAL FINDINGS: By day 3 after adoptive transfer, we observed a selective infiltration of melanomas by anti-H7(a) T cells. Over the next five days, anti-H7(a) T cells expanded massively in the tumor but not in the spleen. Thus, by day 8 after injection, anti-H7(a) T cells in the tumor had undergone more cell divisions than those in the spleen. These data strongly suggest that anti-H7(a) T cells proliferate preferentially and extensively in the tumors. We also found that two host factors regulated long-term persistence of anti-H7(a) memory T cells: thymic function and expression of H7(a) by host cells. On day 100, anti-H7(a) memory T cells were abundant in euthymic H7(a)-negative (B10.H7(b)) mice, present in low numbers in thymectomized H7(a)-positive (B10) hosts, and undetectable in euthymic H7(a)-positive recipients. CONCLUSIONS/SIGNIFICANCE: Although in general the tumor environment is not propitious to T-cell invasion and expansion, the present work shows that this limitation may be overcome by adoptive transfer of primed CD8 T cells targeted to an immunodominant MiHA (here H7(a)). At least in some cases, prolonged persistence of adoptively transferred T cells may be valuable for prevention of late cancer relapse in adoptive hosts. Our findings therefore suggest that it may be advantageous to target MiHAs with a restricted tissue distribution in order to promote persistence of memory T cells and thereby minimize the risk of cancer recurrence.

BackgroundGraft-versus-host disease (GVHD) results from recognition of host antigens by donor T cells following allogeneic hematopoietic cell transplantation (AHCT). Notably, histoincompatibility between donor and recipient is necessary but not sufficient to elicit GVHD. Therefore, we tested the hypothesis that some donors may be "stronger alloresponders" than others, and consequently more likely to elicit GVHD.Methods and findingsTo this end, we measured the gene-expression profiles of CD4(+) and CD8(+) T cells from 50 AHCT donors with microarrays. We report that pre-AHCT gene-expression profiling segregates donors whose recipient suffered from GVHD or not. Using quantitative PCR, established statistical tests, and analysis of multiple independent training-test datasets, we found that for chronic GVHD the "dangerous donor" trait (occurrence of GVHD in the recipient) is under polygenic control and is shaped by the activity of genes that regulate transforming growth factor-beta signaling and cell proliferation.ConclusionsThese findings strongly suggest that the donor gene-expression profile has a dominant influence on the occurrence of GVHD in the recipient. The ability to discriminate strong and weak alloresponders using gene-expression profiling could pave the way to personalized transplantation medicine.

pyGeno is a python package mainly intended for precision medicine applications that revolve around genomics and proteomics. It integrates reference sequences and annotations from Ensembl, genomic polymorphisms from the dbSNP database and data from next-gen sequencing into an easy to use, memory-efficient and fast framework, therefore allowing the user to easily explore subject-specific genomes and proteomes. Compared to a standalone program, pyGeno gives the user access to the complete expressivity of python, a general programming language. Its range of application therefore encompasses both short scripts and large scale genome-wide studies.