Search

Tumor-specific antigens (TSAs) represent ideal targets for cancer immunotherapy, but few have been identified thus far. We therefore developed a proteogenomic approach to enable the high-throughput discovery of TSAs coded by potentially all genomic regions. In two murine cancer cell lines and seven human primary tumors, we identified a total of 40 TSAs, about 90% of which derived from allegedly noncoding regions and would have been missed by standard exome-based approaches. Moreover, most of these TSAs derived from nonmutated yet aberrantly expressed transcripts (such as endogenous retroelements) that could be shared by multiple tumor types. Last, we demonstrated that, in mice, the strength of antitumor responses after TSA vaccination was influenced by two parameters that can be estimated in humans and could serve for TSA prioritization in clinical studies: TSA expression and the frequency of TSA-responsive T cells in the preimmune repertoire. In conclusion, the strategy reported herein could considerably facilitate the identification and prioritization of actionable human TSAs.

Summary 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.

Progress in our understanding of thymic epithelial cell (TEC) renewal and homeostasis is hindered by the lack of markers for TEC progenitors. Stem and progenitor cell populations display remarkable diversity in their proliferative behavior. In some but not all tissues, stemness is associated with quiescence. The primary goal of our study was to discover whether quiescent cells were present in neonatal and adult TECs. To this end, we used a transgenic label-retaining cell (LRC) assay in which a histone H2B-GFP fusion protein is expressed under the control of the reverse tetracycline-controlled transactivator and the tetracycline operator minimal promoter. In adult mice, we found that both cortical and medullary TECs (cTECs and mTECs) proliferated more actively in females than males. Moreover, we observed three main differences between neonatal and adult TECs: 1) neonatal TECs proliferated more actively than adult TECs; 2) whereas cTECs and mTECs had similar turnover rates in young mice, the turnover of mTECs was more rapid than that of cTECs in adults; and 3) although no LRCs could be detected in young mice, LRCs were detectable after a 16-wk chase in adults. In female mice, LRCs were found almost exclusively among cTECs and expressed relatively low levels of p16INK4a, p19ARF, and Serpine1, and high levels of Bmi1, Foxn1, Trp63, and Wnt4. We conclude that LRCs in adult TECs are not senescent postmitotic cells and may represent the elusive progenitors responsible for TEC maintenance in the adult thymus.

Natural killer cells constitute potent innate lymphoid cells that play a major role in both tumor immunosurveillance and viral clearance via their effector functions. A four-stage model of NK cell functional maturation has been established according to the expression of CD11b and CD27, separating mature NK (mNK) cells into distinct populations that exhibit specific phenotypic and functional properties. To identify genetic factors involved in the regulation of NK cell functional maturation, we performed a linkage analysis on F2 (B6.Rag1−/− × NOD.Rag1−/− intercross) mice. We identified six loci on chromosomes 2, 4, 7, 10, 11, and 18 that were linked to one or more mNK cell subsets. Subsequently, we performed an in silico analysis exploiting mNK cell subset microarray data, highlighting various genes and microRNAs as potential regulators of the functional maturation of NK cells. Together, the combination of our unbiased genetic linkage study and the in silico analysis positions genes known to affect NK cell biology along the specific stages of NK cell functional maturation. Moreover, this approach allowed us to uncover a novel candidate gene in the regulation of NK cell maturation, namely Trp53. Using mice deficient for Trp53, we confirm that this tumor suppressor regulates NK cell functional maturation. Additional candidate genes revealed in this study may eventually serve as targets for the modulation of NK cell functional maturation to potentiate both tumor immunosurveillance and viral clearance.

Establishment of self-tolerance in the thymus depends on promiscuous expression of tissue-restricted Ags (TRA) by thymic epithelial cells (TEC). This promiscuous gene expression (pGE) is regulated in part by the autoimmune regulator (AIRE). To evaluate the commonalities and discrepancies between AIRE-dependent and -independent pGE, we analyzed the transcriptome of the three main TEC subsets in wild-type and Aire knockout mice. We found that the impact of AIRE-dependent pGE is not limited to generation of TRA. AIRE decreases, via non–cell autonomous mechanisms, the expression of genes coding for positive regulators of cell proliferation, and it thereby reduces the number of cortical TEC. In mature medullary TEC, AIRE-driven pGE upregulates non-TRA coding genes that enhance cell–cell interactions (e.g., claudins, integrins, and selectins) and are probably of prime relevance to tolerance induction. We also found that AIRE-dependent and -independent TRA present several distinctive features. In particular, relative to AIRE-induced TRA, AIRE-independent TRA are more numerous and show greater splicing complexity. Furthermore, we report that AIRE-dependent versus -independent TRA project nonredundant representations of peripheral tissues in the thymus.

Patients with chronic low back pain exhibit characteristics such as clinical pain, psychological symptoms and neuromuscular adaptations. The purpose of this study was to determine the independent contribution of clinical pain, psychological factors and neuromuscular adaptations to disability in patients with chronic low back pain. Clinical pain intensity, pain catastrophizing, fear-avoidance beliefs, anxiety, neuromuscular adaptations to chronic pain and neuromuscular responses to experimental pain were assessed in 52 patients with chronic low back pain. Lumbar muscle electromyographic activity was assessed during a flexion-extension task (flexion relaxation phenomenon) to assess both chronic neuromuscular adaptations and neuromuscular responses to experimental pain during the task. Multiple regressions showed that independent predictors of disability included neuromuscular adaptations to chronic pain (β=0.25, p=0.006, sr(2)=0.06), neuromuscular responses to experimental pain (β=-0.24, p=0.011, sr(2)=0.05), clinical pain intensity (β=0.28, p=0.002, sr(2)=0.08) and psychological factors (β=0.58, p0.001, sr(2)=0.32). Together, these predictors accounted for 65% of variance in disability (R(2)=0.65 p0.001). The current investigation revealed that neuromuscular adaptations are independent from clinical pain intensity and psychological factors, and contribute to inter-individual differences in patients' disability. This suggests that disability, in chronic low back pain patients, is determined by a combination of factors, including clinical pain, psychological factors and neuromuscular adaptations.

In order to gain novel insights into thymus biology, we analysed the whole transcriptome of cortical and medullary thymic epithelial cells (cTECs and mTECs) and of skin epithelial cells (ECs). Consistent with their ability to express ectopic genes, mTECs expressed more genes than other cell populations. Out of a total of 15,069 genes expressed in TECs, 25% were differentially expressed by at least 5-fold in cTECs vs. mTECs. Genes expressed at higher levels in cTECs than mTECs regulate numerous cell functions including cell differentiation, cell movement and microtubule dynamics. Many positive regulators of the cell cycle were overexpressed in skin ECs relative to TECs. Our RNA-seq data provide novel systems-level insights into the transcriptional landscape of TECs, highlight substantial divergences in the transcriptome of TEC subsets and suggest that cell cycle progression is differentially regulated in TECs and skin ECs.