ESTRO 2020 Abstract Book

S37 ESTRO 2020

Conclusion NK cells may play a critical role in supporting the therapeutic capacity of RT and immunotherapy combinations, particularly in the context of high BED RT. Understanding the regulation and kinetics of radiation- induced NK cell responses reveals opportunities for optimising RT and immunotherapy strategies. OC-0081 Immuno-regulatory properties of irradiated tumor fibroblasts T. Hellevik 1 , R. Berzaghi 2 , A. Islam 2 , L. Gorchs 2 , I. Pettersen 2 , T. Stuge 3 , M.A. Akhtar 2 , I. Martinez- Zubiaurre 2 1 University Hospital of Northern Norway, Departmetn of Radiation Oncology, Tromsø, Norway ; 2 University of Tromsø, Clinical Medicine, Tromsø, Norway ; 3 University of Tromsø, Medical Biology, Tromsø, Norway Purpose or Objective Cancer-associated fibroblasts are key modulators of the tumor microenvironment, facilitating tumor initiation, tumor growth, invasion and metastasis, in addition to function as sentinel cells of the immune system. Nevertheless, the impact of the abundant and heterogeneous CAF-community towards treatment resistance is only starting to emerge. With the ultimate goal of enhancing therapeutic outcome, our laboratory is exploring how CAFs are influencing responses to radiotherapy and immunomodulation. Material and Methods Experiments were carried out with human CAFs freshly isolated from (NSCLC) lung tumor specimens. Primary CAF cultures were exposed to ionizing radiation delivered as single-high or fractionated medium-high doses. The secretory profiles of irradiated and control CAFs were compared by multiplex protein arrays and high-throughput proteomics. Immuno-regulatory properties of irradiated CAFs (iCAFs) were explored in functional in vitro assays, where different types of immune cells, such as macrophages, dendritic cells, natural killer cells or T- lymphocytes, were individually co-cultured with iCAFs or exposed to iCAF-conditioned medium or iCAF-derived exosomes. Radiation-induced events that could potentially interfere with anti-tumor immunity, including immunogenic cell death (ICD) and type-I interferon expression (IFN-I), was also explored in iCAFs. At the in vivo level, tumor development and inflammatory responses were studied upon transplantation of tumor cells admixed with irradiated or control CAFs. Results Cultured CAFs display a noticeable radio-resistant phenotype. However, exposure to high radiation doses provoke induction of cellular senescence and important phenotypic changes including persistent DDR-responses, elevated surface levels of integrins and significant changes in the secretory profile. Importantly, radiation exposure does not elicit ICD or IFN-I release in iCAFs. CAF-derived soluble factors, but not exosomes, show powerful immunosuppressive effects over activated lymphocytes, macrophages and dendritic cells, however, most of the observed effects seem to be unchanged upon radiation exposure. In vivo studies in xenografts propose that the net effect of IR on CAFs is advantageous, since the natural pro-tumorigenic effects of CAFs become abrogated. Conclusion Irradiated CAFs change substantially their phenotype upon exposure to single-high radiation doses, which could impact tumor responses to therapy. Of note, most of the studied CAF-mediated immunomodulatory functions are maintained following irradiation. However, the heterotypic nature of CAFs in true in vivo circumstances, and the existence of CAF subsets possibly exerting opposing effects in the context of radiotherapy and immunotherapy needs to be carefully considered for optimized treatment strategies.

SABR is highly unlikely to be cost-effective at currently commissioned costs and if all patients currently undergoing palliative radiotherapy for bone metastases were considered eligible. This latter limitation results from the limited survival of many patients with bone metastases. Recognition of the learning curve seen in the cost of SABR delivery and careful selection of patients could, however, allow for cost-effective treatment delivery. The efficacy of SABR, demonstrated in early phase trials, must now be replicated in larger, randomised controlled studies if these treatments are to be cost- effective. OC-0080 Radiation therapy increases tumor vulnerability to natural killer cell-mediated control J.I. Sia 1,2 , I. Chindris 1 , J. Hagekyriakou 1 , R. Johnstone 1,2 , N. Haynes 1,2 1 Peter MacCallum Cancer Centre, Cancer Research, Melbourne, Australia ; 2 University of Melbourne, Sir Peter MacCallum Department of Oncology, Melbourne, Australia Purpose or Objective The immune modulatory effects of radiation therapy (RT) on the innate and adaptive arms of the immune system are increasingly recognised to be important. Utilising RT in a manner that supports host anti-cancer immune responses is a paradigm gaining interest, but how best to employ this remains unclear. Here we examine the impact of radiation dose-fractionation on tumor-associated natural killer (NK) cells, which form an important line of host innate immune defense against cancer. From this, we identify ways of better leveraging the therapeutic potential of NK cells with RT and cancer immunotherapy. Material and Methods Established AT3-OVA intra-mammary and MC-38 subcutaneous tumors growing in wildtype C57BL/6 mice were locally irradiated with different radiation dose- fractionations to dissect the effects of radiation dose per fraction (DPF) from total radiation dose (biological effective dose, BED). Depleting antibodies were used to assess the impact of immune subsets on tumor growth post irradiation. Effects on the tumor microenvironment were examined with flow cytometry and RNA sequencing. Such information guided the selection and application of antibody-based checkpoint blockade therapy to promote host innate and adaptive anti-cancer immune defenses . Results Radiation dose-fractionations are not equivalent in their capacity to engage CD8+ T cell and NK cell responses. Treatment with the 3x4Gy, 9x4Gy and 3x8Gy regimens elicited effective CD8+ T cell responses that supported the anti-cancer activity of anti-PD-1 therapy. The high DPF regimen of 1x20Gy, despite having a similar BED to 9x4Gy, was unable to engage effective CD8+ T cell responses. In contrast, only higher BED regimens (36-45Gy) evoked NK cell responses that significantly slowed tumor growth. While all radiation regimens tested promoted an acute increase in tumor-associated NK cell numbers, only the higher BED regimen supported a sustained elevation in tumor-associated NK cell numbers with higher activation markers and corresponding enrichment of the NK cell- mediated cytotoxicity gene signature. The anti-cancer activity of these NK cells is highly sensitive to control by a concurrent but brief wave of radiation-induced regulatory T cells. These early and late phase effects of RT on NK cell responses could be leveraged with checkpoint blockade therapy to achieve prolonged tumor control and, in some instances, complete regression. Proffered Papers: Proffered papers 2: Interactions between radiation and the immune response