Cancer Inflammation and Immunity - Santiago Zelenay

Dr Santiago Zelenay obtained his undergraduate degree in biology from the University of Buenos Aires in 2002. As a student he worked on DNA vaccines in the laboratory of Juan Fló. He then undertook his PhD in Immunology at the Institute Gulbenkian of Science in Portugal where he studied regulatory T cells, working with Jocelyne Demengeot and Antonio Coutinho. In 2008, he joined the group of Caetano Reis e Sousa at the Cancer Research UK (CRUK) London Research Institute and later at The Francis Crick Institute, where he was awarded Marie Curie and EMBO long-term postdoctoral fellowships to investigate innate immune receptors and signalling pathways that trigger dendritic cell activation and drive T cell responses against viruses or tumours.

Santiago joined the CRUK Manchester Institute in 2015, as a Junior Group Leader to form the Cancer Inflammation and Immunity group. His group focuses on understanding the underlying mechanisms that mediate cancer-inhibitory versus tumour-promoting inflammation in order to design new therapies for cancer patients.

In 2017, he received the CRUK Future Leaders in Cancer Research Prize. More recently in 2019, Santiago and colleagues were awarded the inaugral BIAL Award in Biomedicine (worth €300,000) in recognition of their 'international high-quality scientific publication' Zelenay et al. Cell 2015.

Introduction

The extent to which the immune system acts as a natural barrier to cancer has been a subject of debate. This notion has recently gained great support from the clinical success of therapies aimed at exploiting cells from the immune system. Yet whether and how tumours trigger and simultaneously evade the immune system are longstanding questions in cancer biology. The Cancer Inflammation and Immunity group investigates the mechanisms underlying natural and therapy-induced tumour immunity, with a particular emphasis on identifying the cellular and molecular mediators that regulate the balance between tumour-protective versus tumour-promoting inflammation.

Inflammatory cells present at the tumour site are known for promoting several key aspects of carcinogenesis. At the same time, effective anti-tumour immunity depends on inflammatory mediators that contribute to the activation of innate immune cells such as dendritic cells. Combining the use of genetically engineered cancer models with the analysis of samples from cancer patients, our group aims to identify the underlying mechanisms that allow evasion of immune control and enable progressive tumour growth. In this context, the main objectives of the group are to elucidate the signals that trigger innate immune cell activation, initiating tumour immunity, and to   distinguish mediators that favour tumour elimination from those that support cancer progression. This distinction should allow stratification of subgroups of cancer patients with an immune promoting tumour environment likely to benefit from existing immunotherapy from those with active pathways resulting in local immune inhibition. The ultimate goal of the group is, therefore, to develop novel targeted interventions to disrupt immune suppression, promote tumour immunity and enhance the efficacy of cancer therapy.

Role of the inflammatory response in cancer

 

The inflammatory response that takes place at the tumour bed has a dual role in cancer. The most common type of inflammatory profile measured in solid tumours is long known for fostering aggressive tumour growth and for its strong association with dismal prognosis. In contrast, the remarkable clinical responses that some patients experienced following treatment with so-called immune checkpoint inhibitors (ICIs), have highlighted a type of inflammation with profound cancer inhibitory features. The latter inflammatory ‘flavour’ is less often present in clinically apparent tumours and is linked to increased infiltration by select innate and adaptive immune cells with key functions in anti-tumour immunity. Our group investigates the principles and rules that control the establishment of tumour inflammatory environments that either promote or restrain cancer growth spontaneously or following treatment. We argue that improved understanding of the determinants of the intratumoral inflammatory profile will help design better cancer treatments, especially those that exploit the anti-cancer function of the immune system. In this context, we combine fundamental and translational research to inform the design of novel interventions that boost tumour immunity and improve the efficacy of cancer therapy.

 

Cyclooxygenase (COX)-2/prostaglandin E2

 

Much of our group current efforts are centred on investigating the role of the cyclooxygenase (COX)-2/prostaglandin E2 (PGE2) pathway in tumour inflammation and immunity. This pathway is very often upregulated across many tumour types, including in lung, colorectal, breast, and pancreatic cancers and has been implicated in promoting several aspects of malignant tumour growth.

Importantly, our past work has identified PGE2 as a powerful instructive signal that stimulates a type of inflammatory flavour that fuels tumour growth and therapy resistance through enabling immune escape. Combining the use of genetically engineered in vitro and in vivo pre-clinical cancer models with the analysis of patient samples, our work specifically pointed to the inhibitory effects of PGE2 on the immune system as the basis for its protumourigenic role. Accordingly, genetic ablation of PGE2 synthesis on cancer cells, or of its receptors EP2 and EP4, on select immune cell subsets led to spontaneous immune-dependent control of tumours that otherwise grow aggressively in wild-type hosts (Zelenay et al. Cell 2015; Bonavita et al. Immunity 2020). Searching for the primary regulators of cancer inhibitory inflammation in these experimental systems, we showed that natural killer (NK) cells are direct targets of PGE2 activity in vivo. When NK cells were rendered insensitive to PGE2 through genetic ablation of EP2 and EP4, NK cells drove an intratumoral inflammatory remodelling characteristic of the so called ‘hot’ inflamed tumours that set the stage for T cell-mediated tumour control.

 

Pharmacological inhibition of COX-2

 

Based on this genetic evidence highlighting PGE2 as a primary immunosuppressive factor in the tumour environment, we have explored if and how pharmacological inhibition of COX-2 could equally modulate the inflammatory landscape at the tumour bed and enhance the efficacy of ICIs (Pelly and Moeini et al. Cancer Discovery 2021). To test the value of therapeutically inhibiting the pathway in clinically relevant settings, we first used celecoxib, a selective COX-2 inhibitor widely prescribed for the management of conditions like arthritis. Daily oral administration of celecoxib, at a dose considered safe for human use, alongside treatment with ICI led to greater tumour control compared with the single treatments. Across multiple experiments in independent cancer models, we found that ICIs and selective COX-2 inhibition synergised. About two-thirds of animals fully eradicated their tumours after receiving combination therapy, while around only 25% eradicated their tumours when monotherapy was followed with ICIs, and <5% achieved full tumour eradication following monotherapy with celecoxib. We obtained similar results using forefront antagonist of the PGE2 receptors, EP2 and EP4, establishing a prevalent role of PGE2, among other prostaglandins and COX-2 products, and a potentially safer more targeted approach to limit the immunosuppressive effects of PGE2.

 

Synergy between immune checkpoint inhibitors and selective COX-2 inhibition

 

Most surprisingly, the synergy with ICIs was also observed when co-administering corticosteroids, widely considered to be potent immunosuppressants and frequently prescribed to cancer patients for the management of the adverse effects that often patients face following ICI therapy. These findings are of particular clinical relevance as they suggest that corticosteroids, while limiting the toxicities of ICIs, might concomitantly improve the anti-tumour response. Although it remains to be established in which exact settings corticosteroids might boost immune-dependent tumour control, our current work supports the notion that broad non-steroidal and steroidal anti-inflammatory drugs can somewhat paradoxically stimulate a cancer-restraining inflammatory response.

 

Figure 1: Anti-inflammatory Drugs to Turn Up the Heat of Intratumoural Inflammation. By performing in-depth inflammatory profiling of mice and human tumours, we have identified mechanisms by which anti-inflammatory drugs rapidly alter the tumour inflammatory profile, tilting the balance towards cancer inhibitory inflammation and enhancing the response to immunotherapy based on immune checkpoint inhibitors.

 

To investigate the underlying mechanisms by which pharmacological inhibition of PGE2 synthesis or signalling improved the efficacy of ICI therapy, we carried out deep cellular and molecular profiling of tumours early following treatment. In doing so, we found that both celecoxib and the EP2/EP4 antagonists can very rapidly stimulate a shift in the inflammatory profile, tilting the balance towards a type of inflammation that is also observed enriched in patients that respond to ICIs. Importantly, these rapid changes were also transient and waned over time unless we re-dosed the tumour-bearing animals with the COX-2/PGE2 inhibitors. This data exposed a notable plasticity in the intratumoural inflammatory landscape and implied that sustained inhibition of the COX-2/PGE2 axis might be required to maximise the efficacy of ICIs.

We demonstrated the relevance of our findings for human cancers by showing that addition of celecoxib also leads to a similar, rapid shift in the inflammatory profile of patient tumours using a recently established experimental platform developed by our collaborators Daniela Thommen and Ton Schumacher from the Netherlands Cancer Institute (NKI). Thanks to a CRUK travel award, Victoria Pelly, a former postdoc in the group spent four months at the NKI where she tested the effect of adding celecoxib to short-term cultures of freshly explanted surgical specimens from a variety of cancer types. She found, similarly as in the mouse cancer models, that celecoxib did not generally dampen inflammation but rather promoted an intratumoural inflammatory switch that was accompanied by enhanced T cell effector function.

 

Summary

 

Altogether, our findings establish PGE2 signalling as an independent immune checkpoint and support the rational for using readily available anti-inflammatory drugs to tilt the balance toward cancer-inhibitory inflammation and improve the efficacy of current immune-targeting drugs (Figure). Certainly, these findings have significantly contributed to the design of two investigator-led clinical trials sponsored by The Christie NHS Foundation Trust. The first one, IMpALA (IMmunological effects of AveLumab and Aspirin) is funded by a Breast Cancer Now Catalyst Programme and will test in a neoadjuvant setting the combination of high-dose aspirin, a widely used anti-inflammatory drug with the ICI avelumab. The primary objective is to determine whether co-administering aspirin, a COX-inhibitor with ICI treatment switches the intratumoural inflammatory profile towards one that is more favourable to T cell-mediated tumor control as observed in our preclinical models. The second trial, LION (Lifting Immune CheckpOints with NSAIDs), is a pan-tumour basket trial funded by the J P Moulton Charitable Foundation and The Christie NHS Foundation Trust, in which we will test if addition of a selective COX-2 inhibitor enhances the efficacy of the standard of care immunotherapy in non-small cell lung cancer, triple-negative breast cancer and clear cell renal cell cancer.

Selected Publications

 

Bell CR, Pelly VS, Moeini A, Chiang SC, Flanagan E, Bromley CP, Clark C, Earnshaw CH, Koufaki MA, Bonavita E, Zelenay S. (2022)
Chemotherapy-induced COX-2 upregulation by cancer cells defines their inflammatory properties and limits the efficacy of chemoimmunotherapy combinations.
Nature Communications 13(1):2063. PubMed abstract (PMID: 35440553)

Pelly VS, Moeini A, Roelofsen LM, Bonavita E, Bell CR, Hutton C, Gomez AB, Banyard A, Bromley CP, Flanagan E, Chiang SC, Jørgensen C, Schumacher TN, Thommen DS, Zelenay S. (2021)
Anti-inflammatory drugs remodel the tumor immune environment to enhance immune checkpoint blockade efficacy. 
Cancer Discovery 11(10):2602-2619. PubMed abstract (PMID: 34031121)

Bonavita E, Bromley CP, Jonsson G, Pelly VS, Sahoo S, Walwyn-Brown K, Mensurado S, Moeini A, Flanagan E, Bell CR, Chiang SC, Gowda CPC, Rogers N, Silva-Santos B, Jaillon S, Mantovani A, Reis e Sousa C, Guerra N, Davis DM and Zelenay S. (2020)
Antagonistic inflammatory phenotypes dictate tumor fate and response to immune checkpoint blockade.
Immunity 53, 1–15. PubMed abstract

Böttcher JP, Bonavita E, Chakravarty P, Blees H, Cabeza-Cabrerizo M, Sammicheli S, Rogers NC, Sahai E, Zelenay S, Reis e Sousa C. (2018)
NK cells stimulate recruitment of cDC1 into the tumor microenvironment promoting cancer immune control.
Cell 172(5):1022-1037. Article

Zelenay S*, van der Veen AG, Böttcher JP, Snelgrove KJ, Rogers N, Acton SP, Chakravarty P, Girotti MR, Marais R, Quezada SA, Sahai E, Reis e Sousa C*. (2015)
Cyclooxygenase-dependent tumor growth through evasion of immunity.
Cell 162(6):1257-70. PubMed abstract *corresponding author

Helft J, Böttcher J, Chakravarty P, Zelenay S, Huotari J, Schraml BU, Goubau D, Reis e Sousa C. (2015)
GM-CSF mouse bone marrow cultures comprise a heterogeneous population of CD11c(+)MHCII(+) macrophages and dendritic cells.
Immunity 42(6):1197-211. PubMed abstract

Murillo MM, Zelenay S, Nye E, Castellano E, Lassailly F, Stamp G, Downward J. (2014).
RAS interaction with PI3K p110α is required for tumor-induced angiogenesis.
Journal of Clinical Investigation 124(8):3601–11. PubMed abstract

Zelenay S, Keller AM, Whitney PG, Schraml BU, Deddouche S, Rogers NC, Schulz O, Sancho D, Reis e Sousa C. (2012)
The dendritic cell receptor DNGR-1 controls endocytic handling of necrotic cell antigens to favor cross-priming of CTLs in virus-infected mice.
Journal of Clinical Investigation 122(5):1615–1627. PubMed abstract

Zelenay S*, Bergman M-L, Paiva RS, Lino AC, Martins AC, Duarte JH, Moraes Fontes MF, Bilate AM, Lafaille JJ, Demengeot J. (2010)
Cutting edge: Intrathymic differentiation of adaptive Foxp3+ regulatory T cells upon peripheral proinflammatory immunization.
Journal of Immunology 185(7):3829–3833. PubMed abstract *Corresponding author

Zelenay S, Lopes-Carvalho T, Caramalho I, Moraes-Fontes MF, Rebelo M, and Demengeot J. (2005)
Foxp3+ CD25- CD4 T cells constitute a reservoir of committed regulatory cells that regain CD25 expression upon homeostatic expansion.
Proceedings of the National Academy of Sciences USA 102(11):4091–4096. PubMed abstract

 

Postdoctoral Fellows
Agrin Moeini

Scientific Officer
Shih-Chieh Chiang

Graduate Students
Charlotte Bell
Maria Koufaki

Clinical Fellow
Charles Earnshaw

 

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