Background/Purpose:

We have previously reported in patients with RA abnormal inflammatory T cell responses to an E. coli peptide (dnaJP1) which shares homology with the HLA “shared epitope” as well as with human heat shock protein dnaJ. We hypothesised that these abnormal immune responses are part of pathogenesis, as they may lead to an imbalance in the microbiome and a consequent loss of systemic immune tolerance, through a multistep molecular mimicry mechanism. In an ongoing clinical development program, in which we have completed Phase I and IIa clinical trials and are approaching Phase IIb, we have shown that induction of immune tolerance to dnaJp1 results in clinical improvement in RA and thus represents a promising therapeutic intervention, conceptually similar to the induction of tolerance to an antigen in allergy.

Here, we employed a combination of high dimensional cytometry and ngRNA sequencing approaches to dissect the mechanisms associated with clinically relevant induction of immune tolerance.

Methods:

Peripheral Blood Mononuclear Cells (PMBCs) were obtained at the end of the Phase II trial (Day168), from clinical responders treated with dnaJP1 (n=6) and clinical non-responders treated with placebo (n=10). The T cell compartment was studied by flow cytometry using specifically designed antibody panels. Flow cytometry results were then analysed by clustering with Multi-Dimensional Automated Reduction and Visualization (MARVis). PD-1 positive and negative Tregs, and Teff were sorted viable and subject to NGRNA seq, which was analysed by gene ontology.

Results:

Analysis of the T cell immunomes of dnaJP1 responders and placebo non-responders revealed a subset of CD4+FoxP3+ regulatory T (Treg) cells exclusively in dnaJP1 responders that displayed a higher expression of the inhibitory immune checkpoint receptor, PD-1. The expression of PD-1 contributes to an enhancement of the tolerogencity of this Treg cell subset by upregulating the production of signature anti-inflammatory cytokines such as TGFβ. In addition, we observed a corresponding reshaping of the effector T (Teff) cell compartment in which the expression of pro-inflammatory cytokines such as IL-17A and IFNγ was downregulated. Importantly, epitope-specific immunotherapy also induced a subset of active antigen-experienced memory T cells (CD4+CD45RO+CD69+) which sustains the tolerogenic immune response by secreting TGFβ.

Strikingly, ngRNAseq data confirmed a dicothomous effect of therapy on Treg and Teff subsets, with a prevalence of inflammatory pathways in PD1- Treg, including TNFa production, while, conversely, PD1+ Treg were bona-fide tolerogenic. Teff in RA patients who responded to therapy were characterised by a prevalence of memory and pro-apoptotic pathways.

Conclusion:

Our data suggest the presence of a “molecular rheostat” between inflammation and tolerance, which pivots on an unique subset of Treg cells in which the immune checkpoint protein, PD-1 is switched on by the immune therapy.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/immune-therapy-of-rheumatoid-arthritis-patients-with-a-microbiome-derived-self-non-self-peptide-induces-clinically-relevant-immune-tolerance-pivoting-on-immune-checkpoint-expressing-therapy-induced/