T cell repertoire to citrullinated self-peptides in healthy humans is not confined to the HLA-DR SE alleles; Targeting of citrullinated self-peptides presented by HLA-DP4 for tumour therapy

Abstract

Post-translational modifications are induced in stressed cells which cause them to be recognised by the system. One such modification is citrullination where the positive charged arginine is modified to a neutral citrulline. We demonstrate most healthy donors show an oligoclonal CD4 response in vitro to at least one citrullinated vimentin or enolase peptide. Unlike rheumatoid arthritis patients, these T cell responses were not restricted by HLA-DRB1 shared epitope (SE) alleles, suggesting they could be presented by other MHC class II alleles. As HLA-DP is less polymorphic than HLA-DR, we investigated whether the common allele, HLA-DP4 could present citrullinated epitopes. The modification of arginine to citrulline enhanced binding of the peptides to HLA-DP4 and induced high-frequency CD4 responses in HLA-DP4 transgenic mouse models. Our previous studies have shown that tumours present citrullinated peptides restricted through HLA-DR4 which are good targets for anti-tumour immunity. In this study, we show that citrullinated vimentin and enolase peptides also induced strong anti-tumour immunity (100% survival, p < 0.0001) against established B16 tumours and against the LLC/2 lung cancer model (p = 0.034) both expressing HLA-DP4. Since most tumours do not constitutively express MHC class II molecules, models were engineered that expressed MHC class II under the control of an IFNγ inducible promoter. Immunisation with citrullinated peptides resulted in 90% survival (p < 0.001) against established B16 HHD tumour expressing IFNγ inducible DP4. These studies show that citrullinated peptides can be presented by a range of MHC class II molecules, including for the first time HLA-DP4, and are strong targets for anti-tumour immunity.

Homocitrullination of lysine residues mediated by myeloid-derived suppressor cells in the tumor environment is a target for cancer immunotherapy

“…In contrast, immunization with Aldo238-256 Hcit peptide showed no significant survival advantage despite inducing an immune response and also providing evidence that CpG/MPLA adjuvant has no antitumor effect in this model which is consistent with previous data. 5 These results suggest that Aldo74-93 and Aldo140-157 peptides are naturally homocitrullinated and presented on MHC-II, whereas Aldo238-256 is either not naturally homocitrullinated or not presented in vivo.…”

“…Previous studies have shown that citrullinated peptides are presented on HLA-DP4, an HLA allele that is frequently expressed in the Caucasian population. 5 We aimed to determine if homocitrullinated peptides could be presented on HLA-DP4 molecules on the cell surface. IEDB algorithm was used to predict potential peptides from aldolase, Bip, Cyk8 and enolase that have a strong or moderate predicted binding affinity to HLA-DP4 (DPA1*01/DPB1*04:01) and contain lysine residues in the predicted core binding regions.…”

“…RNA was purified from sorted cells, reverse transcription polymerase chain reaction (RT-PCR) performed, the cDNA was subjected to amplicon rescued multiplex PCR using human TCR α and β 250 PER primers (iRepertoire, Huntsville, Alabama, USA). 5 Ten assessed sample libraries were pooled and sequenced using the Illumina MiSeq platform (Illumina, USA). Tree maps and D50 diversity values were generated using iRweb software.…”

Cancer Vaccines, Adjuvants, and Delivery Systems

“…Due to its ubiquitous expression, ENO1 is often degraded during autophagy; previous studies have also shown that ENO1 can be citrullinated ( 65 , 83 ). We have shown that these citrullinated peptides are recognized and presented to CD4+ T cells by both MHC class II HLA-DR4 and HLA-DP4 molecules ( 49 , 84 , 85 ). HLA transgenic mice vaccinated with citrullinated vimentin and α-enolase peptides linked to an adjuvant (Modi-1 vaccine) can stimulate CD4+ T cells ( 49 , 64 , 86 ) and generate potent anti-tumor responses resulting in tumor regression and eradication with no associated toxicity ( 49 , 87 ).…”

Insights of rheumatoid arthritis risk factors and associations

“…The high diagnostic sensitivity and specificity of anti-MCV as well as its predictive value for erosive disease provide indirect evidence for citrullinated vimentin as an important molecule for driving the pathological immune response in RA. These earlier reports of RA demonstrate that at least one citrullinated peptide, but not its arginine-containing counterpart, can drive a T-cell response in the context of the HLA-DRB1*0401 molecule [ 47 ]. However, unlike citrullinated fibrinogen, citrullinated vimentin has yet to be demonstrated to be arthritogenic in an in vivo animal model.…”