Post-translational modifications such as citrullination are excellent targets for cancer therapy

V.A. Brentville, M. Vankemmelbeke, R.L. Metheringham, L.G. Durrant

ABSTRACT: Under conditions of cellular stress, proteins can be post-translationally modified causing them to be recognized by the immune system. One such stress-induced post-translational modification (siPTM) is citrullination, the conversion of arginine residues to citrulline by peptidylarginine deiminase (PAD) enzymes. PAD enzymes are activated by millimolar concentrations of calcium which can occur during apoptosis, leading to precipitation of proteins, their subsequent uptake by B cells and stimulation of antibody responses. Detection of anti-citrullinated protein antibodies (ACPAs) is a diagnostic of rheumatoid arthritis (RA), where immune complexes stimulate inflammation around the joints. More recently, autophagy has been shown to play a role in the presentation of citrullinated peptides on MHC class II molecules to CD4+ helper T cells, suggesting that citrullination may be a way of alerting immune cells to cellular stress.  Additionally, inflammation-induced IFNγ and concomitant MHC class II expression on target cells contributes to immune activation. Stressful conditions in the tumour microenvironment induce autophagy in cancer cells as a pro-survival mechanism. Cancer cells also over express PAD enzymes and in light of this the hypothesis that citrullinated peptides stimulate CD4+ T cell responses that would recognize these siPTM’s produced during autophagy has been investigated. The induction of potent citrullinated peptide-specific CD4 responses has been shown in both humans and HLA transgenic mouse models. Responses in mouse models resulted in potent anti-tumour responses against tumours expressing either constitutive or IFNγ-inducible MHC class II. The anti-tumour effect relied upon direct recognition of tumours by specific CD4 T cells suggesting that citrullinated peptides are attractive targets for cancer vaccines.

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

Victoria A Brentville, Peter Symonds, Katherine  W Cook, Ian Daniels, Tracy Pitt, Mohamed Gijon, Poonam Vaghela, Wei Xue, Sabaria Shah, Rachael E Metheringham, and Lindy Durrant

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 enhanced 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 target 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 d 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.

Post-translationally modified antigens are good targets for cancer immunotherapy but some patients have antigen specific T-regs that may need to be neutralized

Suha Atabani, Victoria Brentville, Ian Daniels, Ruhul Choudhury, Katherine Cook, Poulam Patel and Lindy Durrant

Improving selection criteria for post translationally modified CD4 epitopes using computer algorithms.

K Cook, P Symonds, A Skinner, S Shah, R Metheringham, S Paston, V Brentville and L Durrant

Carbamylation of lysine residues mediated by MDSCs in the tumour environment make excellent targets for CD4 T cell mediated cancer immunotherapy

K Cook, W Xue, I Daniels, P Symonds, M Gijon, D Boocock, C Coveney, A Miles, P Vaghela, R Choudhury, S Shah, S Atabani, R Metheringham, V Brentville and L Durrant

Targeting citrullinated vimentin and enolase with cytotoxic CD4 T cells, relies upon MHC-II expression by tumors, reduces myeloid suppressor cells and directly kills tumor cells

V Brentville, R Metheringham, I Daniels, S Atabani, P Symonds, K Cook, R Choudhury, P Vaghela, M Gijon, G Meiners, W-J Krebber, CJM Melief and L Durrant

Citrullinated glucose-regulated protein 78 is a candidate target for cancer immunotherapy

V Brentville, J Chua, S Atabani, P Symonds, K Cook, R Choudhury, I Daniels, S Shah and L Durrant

An ultraspecificmonoclonal antibody recognises a novel marker on stem memory T cells and induce cell proliferation and differentiation in vitro and in vivo

J Chua, E Cid, M Vankemmelbeke, R McIntosh, R Metheringham, I Daniels, V Brentville and L Durrant

Cancer vaccine:how far are we from curing cancer?

Ruhul Choudhury and Lindy Durrant

Healthy individuals have a repertoire of CD4 cells that recognize and respond to citrullinated peptides - a possible new target for cancer immunotherapy?

Ruhul Hasan Choudhury, Ian Daniels, Vicky Brentville, Katherine Cook, Peter Symonds, Poonam Bilimoria, Suha Atabani, Rachael Metheringham, Lindy Durrant 

CRI 2018 : Post translationally modified homocitrulline induced by MDSCs can be an effective anti-tumour target for CD4 T cells

K. Cook, W. Xue, P. Symonds, M. Gijon, P. Vaghela, R. Choudhury, S. Shah, S. Atabani, R. Metheringham, V. Brentville, L. Durrant 

CRI 2018 : Pre-existing citrulline specific CD4 T cells can be efficiently harnessed for tumour therapy

V. Brentville, P. Symonds, K. Cook, I. Daniels, S. Atabani, R. Choudhury, P. Vaghela, R. Metheringham, M. Gijon, W. Xue, L. Durrant

CRI 2018 : Targeting gastrointestinal tumors with constant region-engineered anti-glycan antibodies

M. Vankemmelbeke, T. Kirk, C. Papagregoriou, S. Aziz, J.X. Chua, R.S. McIntosh, L.G. Durrant

Current Strategies to Enhance Anti-Tumour Immunity (2018)

Katherine W. Cook, Lindy G. Durrant and Victoria A. Brentville

ABSTRACT: The interaction of the immune system with cancer is complex, but new approaches are resulting in exciting therapeutic benefits. In order to enhance the immune response to cancer, immune therapies seek to either induce high avidity immune responses to tumour specific antigens or to convert the tumour to a more pro-inflammatory microenvironment. Strategies, including vaccination, oncolytic viruses, and adoptive cell transfer all seek to induce anti-tumour immunity. To overcome the suppressive tumour microenvironment checkpoint inhibitors and modulators of regulatory cell populations have been investigated. This review summarizes the recent advances in immune therapies and discusses the importance of combination therapies in the treatment of cancers.

Novel tumour antigens and the development of optimal vaccine design (2018)

Victoria A Brentville, Suha Atabani, Katherine Cook and Lindy G Durrant

ABSTRACT: The interplay between tumours and the immune system has long been known to involve complex interactions between tumour cells, immune cells and the tumour microenvironment. The progress of checkpoint inhibitors in the clinic in the last decade has highlighted again the role of the immune system in the fight against cancer. Numerous efforts have been undertaken to develop ways of stimulating the cellular immune response to eradicate tumours. These interventions include the identification of appropriate tumour antigens as targets for therapy. In this review, we summarize progress in selection of target tumour antigen. Targeting self antigens has the problem of thymic deletion of high-affinity T-cell responses leaving a diminished repertoire of low-affinity T cells that fail to kill tumour cells. Thymic regulation appears to be less stringent for differentiation of cancer–testis antigens, as many tumour rejection antigens fall into this category. More recently, targeting neo-epitopes or post-translational modifications such as a phosphorylation or stress-induced citrullination has shown great promise in preclinical studies. Of particular interest is that the responses can be mediated by both CD4 and CD8 T cells. Previous vaccines have targeted CD8 T-cell responses but more recently, the central role of CD4 T cells in orchestrating inflammation within tumours and also differentiating into potent killer cells has been recognized. The design of vaccines to induce such immune responses is discussed herein. Liposomally encoded ribonucleic acid (RNA), targeted deoxyribonucleic acid (DNA) or long peptides linked to toll-like receptor (TLR) adjuvants are the most promising new vaccine approaches. These exciting new approaches suggest that the ‘Holy Grail’ of a simple nontoxic cancer vaccine may be on the horizon. A major hurdle in tumour therapy is also to overcome the suppressive tumour environment. We address current progress in combination therapies and suggest that these are likely to show the most promise for the future.

Targeting gp100 and TRP-2 with a DNA vaccine: Incorporating T cell epitopes with a human IgG1 antibody induces potent T cell responses that are associated with favourable clinical outcome in a phase I/II trial

Poulam M. Patel, Christian H. Ottensmeier, Clive Mulatero, Paul Lorigan, Ruth Plummer, Hardev Pandha, Somaia Elsheikh, Efthymios Hadjimichael, Naty Villasanti, Sally E. Adams, Michelle Cunnell, Rachael L. Metheringham, Victoria A. Brentville, Lee Machado, Ian Daniels, Mohamed Gijon, Drew Hannaman and Lindy G. Durrant

ABSTRACT: A DNA vaccine, SCIB1, incorporating two CD8 and two CD4 epitopes from TRP-2/gp100 was evaluated in patients with metastatic melanoma. Each patient received SCIB1 via intramuscular injection with electroporation. The trial was designed to find the safest dose of SCIB1 which induced immune/clinical responses in patients with or without tumour. Fifteen patients with tumor received SCIB1 doses of 0.4-8 mg whilst 20 fully-resected patients received 2–8 mg doses. Twelve patients elected to continue immunization every 3 months for up to 39 months. SCIB1 induced dose-dependent T cell responses in 88% of patients with no serious adverse effects or dose limiting toxicities. The intensity of the T cell responses was significantly higher in patients receiving 4 mg doses without tumor when compared to those with tumor (ρ < 0.01). In contrast, patients with tumor showed a significantly higher response to the 8 mg dose than the 4 mg dose (ρ < 0.03) but there was no significant difference in the patients without tumor. One of 15 patients with measurable disease showed an objective tumor response and 7/15 showed stable disease. 5/20 fully-resected patients have experienced disease recurrence but all remained alive at the cut-off date with a median observation time of 37 months. A positive clinical outcome was associated with MHC-I and MHC-II expression on tumors prior to therapy (ρ = 0.027).

We conclude that SCIB1 is well tolerated and stimulates potent T cell responses in melanoma patients. It deserves further evaluation as a single agent adjuvant therapy or in combination with checkpoint inhibitors in advanced disease.

CRI 2017 Peptide/TLR agonist conjugates poster

V. Brentville, K. Cook, P. Symonds, G.G. Zom, W. Xue, R. Metheringham, W-J. Krebber, C.J.M. Melief and L.G. Durrant

CRI 2017 Modi-1, a novel cancer vaccine

L. G. Durrant, R. Metheringham, I. Daniels, K. Cook, P. Symonds, T. Pitt, W. Xue, M. Gijon and V. Brentville