Interleukin 10 (IL-10)-producing regulatory T-cells (Tregs) play a central role in the maintenance of normal immune homeostasis. Tregs include the well-characterized FOXP3+ subset as well as a FOXP3-negative CD4+ T-cell type that secretes IL-10 and low levels or no IL-4 and co-expresses CD49b, LAG-3, inducible T-cell costimulator (ICOS), and/or CCR5 and PD-1 among others (Roncarolo et al., 2018).

We have shown that systemic delivery of nanoparticles (NPs) coated with mono-specific peptide-major histocompatibility complex class II (pMHCII) molecules (Singha et al., 2017) triggers the expansion and re-programming of cognate splenic T-follicular helper (TFH) CD4+ T-cells into expanded pools of transitional (referred to as TR1.1 or TR1-like) and terminally differentiated TR1 cells (referred to as TR1.2 or TR1) that resolve inflammation in various organ-specific autoimmune disease models in a disease-specific manner without impairing normal immunity (Clemente-Casares et al., 2016; Umeshappa et al., 2020; Umeshappa et al., 2019). These events result from the sustained assembly of large TCR microclusters, and rapid, robust, and prolonged TCR signaling on TFH cells (Singha et al., 2017; Solé et al., 2023b; Solé et al., 2023a). The antigen-specific TFH and TR1 sub-pools arising in response to pMHCII-NP therapy have nearly identical clonotypic composition but alternative functional properties and transcription factor (TF) expression profiles (Solé et al., 2023b). In addition, pMHCII-NPs trigger cognate TR1 cell formation in TFH cell-transfused immunodeficient hosts, and T-cell-specific deletion of two master regulators of TFH cell genesis (Bcl6 or Irf4) blunt both pMHCII-NP-induced TFH expansion and TR1 formation. In contrast, deletion of Prdm1 selectively abrogates the TFH-to-TR1 conversion. Together, these data indicated that TFH cells can differentiate into TR1 cells in vivo and that Blimp-1 is a gatekeeper of this cell re-programming event (Solé et al., 2023b). The work described herein was initiated to test the hypothesis that pMHCII-NP-induced conversion of TFH cells into TR1 cells involves epigenetic re-programming of the TFH precursors.

Chromatin can be modified at various levels, including alterations in chromatin accessibility to expose or shield specific genes from the gene expression machinery, through histone modifications that either promote or repress gene expression, or via changes in DNA methylation, typically hypomethylation. Although such processes have been shown to contribute to T-cell differentiation, such as during Th1/Th2 cell specification (O’Garra and Arai, 2000), they can also occur upon T-cell activation (Avni et al., 2002) or in response to environmental cues, such as cytokine stimulation (Ostuni et al., 2013). Likewise, it has been established that DNA methylation, histone modifications, and chromatin accessibility regulate T-cell activation and effector and memory responses during immune responses to infection. Effector T-cell-specific genes are demethylated and gain chromatin accessibility and naïve T-cell-associated genes are repressed. When these cells need to become memory, the naïve genes required for survival are demethylated (Belk et al., 2022). Another notable example of chromatin remodeling is in exhausted T-cells, where numerous effector genes, such as Ifng, become closed, and acquire open chromatin at genes that are upregulated in these cells, such as Pdcd1 (Belk et al., 2022).

Several notable examples of redistribution of histone marks during T-cell differentiation have been described. For example, Th1 differentiation results in H3K4 di-methylation and H3 and H4 acetylation and in the creation of chromatin accessible regions at regulatory elements within the Ifng locus, as well as in the loss of H3K27me3 through the locus (in addition to DNA demethylation of the locus) (Hatton et al., 2006; Schoenborn et al., 2007). In contrast, activation of the Th2 program leads to the loss of permissive histone modifications in Ifng, addition of repressive H3K27me3 marks along the locus and DNA methylation (Schoenborn et al., 2007; Jones and Chen, 2006; Chang and Aune, 2007). Th2 differentiation from naïve precursors also involves the acquisition of permissive histone modifications in Il4, Il5, Il13 and the locus control region (LCR), and the loss of H3K27me3 marks (Avni et al., 2002; Koyanagi et al., 2005). Likewise, the promoters and eight gene regulatory elements (GREs) of Il17a and Il17f genes in naïve CD4+ T-cells acquire H3K27ac marks upon culture in Th17-polarizing conditions (Akimzhanov et al., 2007). In memory T-cells, which display faster and greater levels of gene transcription than their naïve and effector counterparts, lineage-specific cytokine genes retain the positive histone modifications on their proximal and distal GREs that were acquired during differentiation of their precursor T-cells (e.g. H3K27ac and H3K9ac marks), even when expression of the genes ceases upon memory cell conversion, allowing rapid reactivation of these loci upon antigen re-encounter (Fann et al., 2006). Likewise, naive CD8+ T cells lose repressive H3K27me3 marks during the primary immune response against infections, allowing rapid upregulation of Ifng and Gzmb expression by their memory CD8+ T-cell counterparts (Araki et al., 2009). Overall, memory T-cells appear to retain the epigenetic signatures of their effector progenitors, thus allowing a quicker and more efficient response in subsequent antigen encounters.

There are also several noteworthy examples of changes in DNA methylation in T-cells. For example, the Cd4 locus is hypermethylated in CD4–CD8–, CD4+CD8+ thymocytes and mature CD8+ T cells, but is demethylated in CD4+CD8–cells (Teghanemt et al., 2022). Likewise, in naïve T-cells, Il4, Il5, Il13, and the corresponding LCR are hypermethylated and Ifng is hypomethylated (Fields et al., 2004; Lee et al., 2002). Whereas this methylation pattern is maintained during the Th0-Th1 differentiation process, it undergoes dramatic changes during Th2 formation, such that Ifng becomes hypermethylated, and Il4, Il5, Il13, and the LCR demethylate key GREs (Fields et al., 2004; Lee et al., 2002; Kim et al., 2007), in addition to acquiring the type of permissive histone modifications discussed above (Avni et al., 2002; Koyanagi et al., 2005). Likewise, TFH formation from naïve T-cell precursors involves demethylation of BCL-6 binding sites (Liu et al., 2016). In Foxp3+ Tregs, the expression of Foxp3 and genes coding for some Treg-function-associated molecules, such as Ctla4 and Il2ra, but not genes coding for TFs controlling other cell fates or cytokine genes that are repressed in Foxp3+ Tregs, are also associated with DNA demethylation (Liu et al., 2010; Ohkura et al., 2012). The development of memory T-cells also involves progressive demethylation of promoter-distal GREs of key genes (Durek et al., 2016).

Collectively, the above observations indicate that, in most cases, acquisition of new functional states by peripheral T-cells, including their differentiation into different T-cell subsets, involves extensive and diverse modifications of the chromatin around specific loci, although global (i.e. genome-wide) epigenetic changes during these processes were not always explored. Here, we show, through a broad combination of transcriptomic and epigenetic studies, that the TFH-to-TR1 transdifferentiation process does not follow this pattern. Specifically, we find that conventional TFH cells are epigenetically poised to differentiate into TR1 cells, and that the TFH-to-TR1 transdifferentiation process is associated with extensive contraction of the chromatin and the upregulation of genes that are already epigenetically poised for expression, yet are silent, at the TFH cell stage, such as Il10. These genes are closed and hypermethylated in Tconv cells, but are accessible, hypomethylated, and enriched for H3K27ac-marked and hypomethylated active enhancers in TFH cells. These data suggest that genomic imprinting is a key enabler of the TFH-TR1 cell transdifferentiation process, as documented in non-immune cell types (Holliday and Pugh, 1975; Muramoto et al., 2010; Raas et al., 2022).

Having established that TFH cells can transdifferentiate into TR1-like and terminally differentiated TR1 cells in vivo in response to certain stimuli (Solé et al., 2023b; Solé et al., 2023a), we sought to explore the epigenetic events underpinning this process. Our comprehensive transcriptional and epigenetic studies at the bulk and/or single-cell levels indicate that conventional antigen experienced TFH cells are epigenetically poised to become TR1 cells. One of our main findings is that the TFH-to-TR1 differentiation process is associated with massive closure of OCRs, and that the vast majority of the OCRs found in TR1-like and TR1 cells are also found in KLH-DNP-induced TFH cells. Furthermore, most of the genes harbored in these shared OCRs, such as Cxcr5, Tox2, Il21, Il10, and Ctla4, to just name a few, contain nearly identical patterns of histone deposition marks, including H3K4me3, H3K27me3, and H3K27Ac, equally similar DNA methylation patterns in gene bodies, proximal promoters, and gene-distal regulatory elements and a similar distribution of active enhancers across the genome, even when not expressed in either cell type. Altogether, these data indicate that the TR1-poised epigenome of TFH cells is a key enabler of this transdifferentiation process, and that transdifferentiation of TFH cells into TR1 cells is likely driven by changes in the expression of TFH-stabilizing and TR1-promoting TFs, possibly in response to sustained ligation of TCRs.

Our earlier scRNAseq and mass cytometry studies of cognate (antigen-specific, tetramer+) pMHCII-NP-induced CD4+ T cells demonstrated the presence of a significant Tet+ TFH-like cluster that separated away from its TR1 counterpart (Solé et al., 2023b; Solé et al., 2023a). When compared to each other, these two major clusters of tetramer+ cells were remarkably similar, but the TR1 sub-pool had significantly downregulated key TFH-specific genes, including S1pr2, Cxcr4, Cxcr5, Pdcd1, Il4, Ascl2, Bcl6, Cba2t3, Cebpa, Id3, Nfia, Pou2af1, Tox2, while upregulating TR1-associated genes, such as Ccr5, Havcr2, Il10, Ahr, Myc, and Prdm1. Importantly, these two clusters were developmentally related because they harbored identical clonotypes (i.e. identical TCRαβ sequences at different transcriptional states). Additional studies indicated that the TFH-to-TR1 conversion proceeds through a transitional TR1-like subset, whereby the progenitors undergo progressive downregulation of TFH-associated transcripts (i.e. Bcl6, Cxcr5, among others) and progressive upregulation of TR1-associated transcripts (i.e. il10, Ccr5, and Prdm1, among others). The suspected TFH origin of pMHCII-NP-induced TR1 cells was further supported by two additional lines of evidence. First, treatment of NOD.Scid mice engrafted with total CXCR5^highPD-1^high CD4+ T-cells (containing pMHCII-NP-expanded TFH-like cells but devoid of terminally differentiated Tet+ TR1 cells) with pMHCII-NPs led to the formation of cognate TR1-like and terminally differentiated TR1 cell pools in the hosts. Second, the two-dimensional scMultiome profile of Tet+ TFH-like cells arising in response to BDC2.5mi/IA^g7-NP therapy was essentially identical to that corresponding to an effector TFH-like sub-pool of TFH-like cells (CD4⁺CD44^hiCXCR5^hiPD1^hi) induced by immunization with the KLH-DNP conjugate (Bcl6^hiTox2^hiIl21⁺Pdcd1⁺; referred to as TFH.1).

Abrogation of Prdm1 (encoding BLIMP-1) expression enabled the conversion of TFH.1 cells into TR1-like progeny, but completely blunted the TR1-like–>TR1 conversion, indicating that this process requires the expression of BLIMP-1, a transcriptional repressor that antagonizes BCL-6 expression and function in both B- and T-cells, including TFH cells (Crotty et al., 2010). Since expression of BLIMP-1 in T-cells is restricted to activated T-cells and is induced by TCR ligation, and since pMHCII-NP therapy triggers the formation and expansion of cognate, antigen-specific TR1-like cell pools via sustained TCR signaling (Singha et al., 2017), we suspect that BLIMP-1 expression in TFH cells is induced by repetitive encounters of cognate TFH cells with these compounds. Progressive downregulation of Bcl6 and upregulation of Prdm1 at the TR1-like cell stage, immediately preceding TR1 development, might be facilitated by the loss of Lef1 and significant downregulation of Tcf7 (encoding TCF-1) expression (positive regulators of Bcl6 expression and negative regulators of Prdm1 expression) in pMHCII-NP-challenged vs. vaccine-induced TFH cells. In fact, the Tet+ TR1-like and terminally differentiated TR1 cells (BLIMP-1-independent or -dependent, respectively) differ in the expression levels of a significant number of genes whose expression has been previously associated with this TF, including Il10, Ctla4, Lag3, Icos, Havcr2, Tnfrsf4, and Tnfrsf18, among others (Chihara et al., 2018).

Here, we have shown that pMHCII-NP-driven transdifferentiation of cognate TFH cells into TR1 progeny is driven by both massive closure of OCRs and major changes in the transcriptional factor make-up of the cells, rather than by TCR signaling-induced changes in the epigenetic status of the genes contained within the shared OCRs. There are some noteworthy differences between the evolution of the epigenetic landscape in effector, memory, and exhausted T-cells as compared to what we see in the TFH-TR1 pathway. In T-cells, acquisition of the epigenetic programs of effector, memory, and exhaustion states are stepwise processes. In exhausted T-cells, for example, ATACseq and H3K27AC ChIPseq analyses of different exhausted states are associated with distinct epigenetic states, including differential chromatin accessibility and active enhancer landscapes, yet shared TF profiles (Belk et al., 2022). In contrast, transdifferentiation of TFH cells into TR1 cells appears to be driven by changes in the expression of TFH-stabilizing and TR1-promoting TFs, in the face of remarkable epigenetic similarity, including a shared methylome, consistent with the idea that TFH cells are poised to rapidly become TR1 when appropriately activated. Interestingly, the TF-coding genes that are significantly downregulated during the TFH-to-TR1 conversion, unlike those that are upregulated, experience a significant reduction in chromatin accessibility. In contrast, as expected based on the epigenetic similarity of TFH vs. TR1 cells, upregulation of TF-coding genes important for TR1 cell genesis is largely dissociated from the different epigenetic marks studied here, including changes in chromatin accessibility. This suggests that changes in TF expression, particularly the loss of TFH-associated TFs, are driven, in part, via chromatin remodeling of the coding loci.

The combination of TFs that become available (i.e. Prdm1) or unavailable (i.e. Bcl6) during the TFH-TR1 conversion would be responsible for enabling/driving the formation of the active enhancer-promoter contacts required for gene expression and establishing TR1 transcriptional identity. This would explain why TR1-associated genes that are already epigenetically marked for expression in TFH cells (i.e. Il10) remain silent at the TFH stage. This is not a unique feature of the TFH-to-TR1 transdifferentiation process. For example, although the Il10 locus is closed and transcriptionally silent in naive CD4+ T-cells, it is exposed (Miraldi et al., 2019) and incorporates permissive H3K4me3 marks but not repressive H3K27me3 marks in differentiated T helper subsets (Ciofani et al., 2012; Wei et al., 2010), promoting a transcriptional competent yet silent state. Since TFs regulating Th1, Th2, and Th17 subsets, including T-bet, GATA-3, or RORγt, do no inhibit IL-10 expression, but rather enhance it, the above observations imply that different effector cell programs can co-exist with much broader gene expression-competent states. Our data indicate that this is also true for TFH cells. In fact, the expression of Il10 in Th cell subsets can only occur until the TFs required for gene expression become available. For example, BATF (an AP1 family member), IRF4, NFAT, c-MAF, AHR, and BLIMP-1 are known to collectively promote Il10 expression by binding to the promoter and/or cis-regulatory elements of Il10 (Pot et al., 2011). Likewise, although Th lineage-specific cytokines present active histone marks in the corresponding lineage and repressive marks in the others, TF-coding genes are not always so strictly marked and display expression-permissive epigenetic patterns. For example, Tbx21, encoding T-bet, harbors activating H3K4me3 marks in the promoter of Th1 cells, but bivalent modifications in other Th subsets. This is also true for Gata3 and Rorc or Bcl6 in Th2 vs. non-Th2, Th17 vs. non-Th17, and TFH vs. non-TFH cells, respectively (Lu et al., 2011; Wei et al., 2009). Thus, whereas Bcl6 is also marked with expression-promoting H3K4me3 marks in other Th cell subsets, loci encoding Th subset-specifying TFs (Tbx21, Gata3, and Rorc) also appear to be poised for expression in TFH cells (Lu et al., 2011). Because BCL-6 can regulate the expression other TFs, it is likely the collective TF make-up of the cell that determines its phenotype at a given point in time. It is thus reasonable to suspect that the epigenomes of polarized T-cell subsets, including TFH cells, are programmed to enable their conversion into alternative transcriptional states when required (i.e. in response to excessive antigenic stimulation).

The TR1-poised state of TFH cells is also reflected on the global active enhancer landscapes of both cell types. For example, we have shown that most of the genes that remain open as BDC2.5mi/I-A^g7-NP-induced TFH.1 cells transition into TR1-like/TR1 cells also share active enhancers in both subsets. In addition, the demethylated status of most active enhancers associated with genes specifically upregulated in TR1 cells, such as Il10, were already so (hence imprinted) at the TFH stage. Furthermore, we find that many of the active enhancers linked to genes specifically upregulated at the TR1 cell stage, which, in turn, are highly hypomethylated and accessible at the TFH cell stage, are enriched for binding sites for at least three TFH-associated TFs, including TOX2, IRF4, and c-MAF. This suggests that these sites are likely already occupied by these TFs at the TFH stage. The remarkable similarity of the DNA methylome of the TFH and TR1 subsets further suggests that both TFH and TR1 cells share a stable epigenetic program.

The factors responsible for chromatin closure during the TFH-to-TR1 conversion remain to be determined, but changes in the expression of Tox2 (and the related TF Tox) may contribute to this event. TOX-2 expression in CD4+ T-cells is upregulated by BCL-6, and TOX-2 binds to loci associated with TFH generation, including Bcl6, Cxcr5, Pdcd1 (also upregulated in TR1-like cells) along with BATF and IRF4 or STAT-3, increasing their chromatin accessibility and promoting their expression (Xu et al., 2019). Importantly, whereas KLH-DNP- and pMHCII-NP-induced TFH cells express high levels of Tox2, their TR1-like and TR1 counterparts express substantially lower levels; suboptimal occupation of TOX-2 binding sites by TOX-2 in pMHCII-NP-challenged TFH cells may result in the closure of these sites, cessation of the expression of the corresponding genes and progressive differentiation into TR1 cells. Downregulation of TCF-1 (encoded by Tcf7) and LEF-1 expression as TFH cells become TR1 cells may be additional contributing factors to loss of chromatin accessibility (Gounari and Khazaie, 2022). We note that loss of TCF-1 in T-cell subsets is usually paralleled by loss of LEF-1, which belongs to the same TF family and recognizes a similar motif (Gounari and Khazaie, 2022). In developing thymocytes, upregulation of TCF-1 expression is associated with an increase in chromatin accessibility, suggesting that it may act as a pioneer TF. Furthermore, loss of TCF-1 in DP thymocytes or CD8+ T-cells, T-exhausted stem cells or activated CD4+ or CD8+ T-cells is associated with loss of chromatin accessibility at sites that had bound TCF-1 (Gounari and Khazaie, 2022). In addition, TCF-1, along with CTCF, promotes deposition of H3K27ac onto insulated enhancers and the recruitment of cohesin-loading factor NIPBL at active enhancers in developing thymocytes (Wang et al., 2022). In another recent study, TCF-1 was identified as a ‘placeholder’ TF, responsible for maintaining chromatin accessibility in naïve T-cells, and allowing activation-induced TFs to displace it (Zhong et al., 2022). Hence, downregulation of TCF-1 and LEF-1 may contribute to the loss of the TFH-specific gene expression program as these cells transdifferentiate into TR1, close previously open chromatin sites, and acquire new TFs orchestrating the TR1-specific gene expression program.

The current study provides a foundational understanding of how the epigenetic landscape of TFH cells evolves as they transdifferentiate into TR1 progeny in response to chronic ligation of cognate TCRs using pMHCII-NPs. Our current studies focus on functional validation of these observations, by carrying out extensive perturbation studies of the TFH-TR1 transdifferentiation pathway in conditional TF gene knockout mice. In these ongoing studies, genes coding for a series of TFs expressed along the TFH-TR1 pathway are selectively knocked out in T cells, to ascertain (1) the specific roles of key TFs in the various cell conversion events and transcriptional changes that take place along the TFH-TR1 cell axis; (2) the roles that such TFs play in the chromatin re-modeling events that underpin the TFH-TR1 transdifferentiation process; and (3) the effects of TF gene deletion on phenotypic and functional readouts of TFH and Treg function. Although the TFH-TR1 transdifferentiation was discovered in mice treated with pMHCII-NPs, we now have evidence that this is a naturally occurring pathway that also develops in other contexts (i.e. in mice that have not been treated with pMHCII-NPs). Importantly, the discovery of this transdifferentiation process affords a unique opportunity to further understand the transcriptional and epigenetic mechanisms underpinning T-cell plasticity; the findings reported here can help guide/inform not only upcoming translational studies of pMHCII-NP therapy in humans, but also other research in this area.

Although the snapshot provided by our single-cell studies reported herein documents the simultaneous presence of the different subsets composing the TFH-TR1 cell pathway upon the termination of treatment, the transdifferentiation process itself is extremely fast, such that proliferated TFH cells already transdifferentiate into TR1 cells after a single pMHCII-NP dose (Solé et al., 2023b). This makes it extremely challenging to pursue dynamic experiments. Notwithstanding this caveat, ongoing studies of cognate T-cells post treatment withdrawal, coupled to single-cell studies of the TFH-TR1 pathway in TF gene knockout mice exhibiting perturbed transdifferentiation processes, are likely to shed light into the progression and stability of the epigenetic changes reported herein.

We have recently shown that αGalCer/CD1d-NPs can trigger the differentiation of liver-resident invariant NKT cells (LiNKT) into a TR1-like immunoregulatory, IL-10+IL-21-producing Zbtb16^highMaf^highTbx21⁺Gata3⁺Rorc^– subset (LiNKTR1) that can suppress local inflammatory phenomena (Umeshappa et al., 2022). Interestingly, epigenetic studies of liver iNKT cells both before and after in vivo delivery of aGalCer/CD1d-coated NPs have shown that unlike the case for pMHCII-NP-induced TR1 transdifferentiation, aGalCer/CD1d-NP-induced LiNKTR1 transdifferentiation involves the acquisition of a novel epigenetic state. Specifically, whereas for most genes, gene upregulation during the LinKT-to-LiNKTR1 cell transition is largely associated with treatment-induced hypomethylation, the most upregulated genes (i.e. Il10 and Il21, among others) are also those that accumulate additional epigenetic modifications favoring gene expression, such as acquisition of new OCRs and H3K27ac and H3K4me3 marks. Thus, whereas TFH cells largely require chromatin closure and changes in TF expression to become TR1 cells, LiNKT cells do not undergo massive changes in chromatin exposure and involve extensive gene demethylation to do so. Although the mechanisms underlying these differences remain unclear, they indicate that the processes that regulate responses of different T-cell types to similar cues are context-dependent and dynamic.

The raw and processed data supporting the findings of this study are available at the Gene Expression Omnibus (GEO) database. Accession numbers are as follows: GSE173681 (bulk RNA-seq data); GSE182636 (scRNA-seq data); and GSE248152 (ATAC-seq, ChIP-seq, methylome, and single-cell multiome data).

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Author details

1. Josep Garnica

   Institut D’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain

   Contribution

   Data curation, Formal analysis, Investigation, Visualization, Methodology, Writing – original draft

   Competing interests

   No competing interests declared

2. Patricia Sole

   Institut D’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain

   Contribution

   Investigation, Methodology

   Competing interests

   No competing interests declared

3. Jun Yamanouchi

   Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada

   Contribution

   Investigation

   Competing interests

   No competing interests declared

4. Joel Moro

   Institut D’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain

   Contribution

   Methodology

   Competing interests

   No competing interests declared

5. Debajyoti Mondal

   Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada

   Contribution

   Methodology

   Competing interests

   No competing interests declared

6. Cesar Fandos

   Institut D’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain

   Contribution

   Methodology

   Competing interests

   No competing interests declared

7. Pau Serra

   Institut D’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain

   Contribution

   Supervision

   Competing interests

   No competing interests declared

8. Pere Santamaria

   1. Institut D’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain
   2. Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada

   Contribution

   Conceptualization, Resources, Supervision, Funding acquisition, Investigation, Methodology, Writing – original draft, Project administration, Writing – review and editing

   For correspondence

   psantama@ucalgary.ca

   Competing interests

   P. Santamaria is founder, scientific officer and stockholder of Parvus Therapeutics and receives funding from the company. He also has a consulting agreement with Sanofi

   "This ORCID iD identifies the author of this article:" 0000-0003-3469-1586

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