S2614
Radiobiology - Translational radiobiology
ESTRO 2026
Conclusion: We developed and validated a 20-gene pan-cancer hypoxia signature with reproducible prognostic associations across LUAD/LUSC, HNSC, and PAAD. Although the incremental gain in discrimination was modest, it consistently improved prognostic accuracy beyond standard clinical variables. This cross-tumour signature supports further evaluation of hypoxia- informed biomarkers as complementary tools for risk stratification in solid tumours. References: 1. Harris, A. L. Hypoxia — a key regulatory factor in tumour growth. Nature Reviews Cancer 2002 2:12, 38–47 (2002).2. Buffa, F. M., Harris, A. L., West, C. M. & Miller, C. J. Large meta-analysis of multiple cancers reveals a common, compact and highly prognostic hypoxia metagene. British Journal of Cancer 2010 102:2102, 428–435 (2010).3. Toustrup, K. et al. Development of a hypoxia gene expression classifier with predictive impact for hypoxic modification of radiotherapy in head and neck cancer. Cancer Res71, 5923–5931 (2011).4. Abou Khouzam, R. et al. An Eight-Gene Hypoxia Signature Predicts Survival in Pancreatic Cancer and Is Associated With an Immunosuppressed Tumor Microenvironment. Front Immunol12, 680435 (2021). Keywords: Hypoxia Signature, Prognostic Pan-Cancer Analysis Digital Poster Highlight 687 Fractionation-dependent modulation of intratumoral NK and CD8 ⁺ T-cell infiltration drives synergy between radiotherapy and dual PD- L1/NKG2A blockade. Jérémy Baude 1,2 , Alexia Tavares 2 , Lisa Froidurot 2 , Adeline Delfour 2 , Riad Ladjohounlou 2 , Emeric Limagne 3,4 , Eric Vivier 5 , Fabien Milliat 6 , Céline Mirjolet 2 1 Radiation Oncology, Georges-François Leclerc Cancer Centre (CGFL), Dijon, France. 2 XRain team, INSERM UMR1231, Dijon, France. 3 CTM Trecs team, INSERM UMR1231, Dijon, France. 4 PTBC, Georges-François Leclerc Cancer Centre (CGFL), Dijon, France. 5 CIML, Centre d'immunologie de Marseille Luminy, Marseille, France. 6 LRMed, Autorité de sureté nucléaire et de radioprotection (ASNR), Fontenay-aux-Roses, France Purpose/Objective: Beyond its cytotoxic effect, radiotherapy (RT) acts as a potent immunomodulatory agent. The radiation fractionation schedule can markedly influence the intratumoral immune landscape, particularly CD8 ⁺ T cell infiltration and the PD-1/PD-L1 axis, both of which are critical determinants of RT - immunotherapy interactions1,2. Natural killer (NK) cells, key effectors of the innate immune response, exert antitumor
activity through a balance of activating (e.g., NKG2D) and inhibitory (e.g., NKG2A-HLA-E) signaling pathways. However, the impact of RT on NK-cell-mediated immune modulation remains poorly understood3.This study aimed to characterize the fractionation- dependent modulation of CD8 ⁺ T cells and NK cells following RT and to identify the optimal combination strategy with dual PD-L1 and NKG2A blockade. Material/Methods: We used an in vivo heterotopic murine model of colorectal cancer (CT26) to evaluate two RT regimens with equivalent EQD2: normofractionated (18 × 2 Gy) and hypofractionated (3 × 8 Gy) schedules. Irradiations were performed using the SARRP system, those protocol is described elsewhere4. Immune modulation was assessed by flow cytometry at days 3, 7, and 10 after the first irradiation, focusing on CD8 ⁺ T cells, NK cells, and the NKG2A-Qa1 axis (Qa1 being the murine homolog of HLA-E). Single cell RNA sequencing (scRNAseq) was performed at day 7 to further characterize immune changes. To explore the functional relevance of NKG2A, RT response was compared between CT26 wild type and CT26 Qa1 ⁻ / ⁻ models. Finally, therapeutic efficacy was assessed in vivo by combining RT with anti-PD-L1 and anti-NKG2A monoclonal antibodies administered intraperitoneally. Results: RT induced a time-dependent increase in intratumoral NK and CD8 ⁺ T cell infiltration, more pronounced with the hypofractionated regimen. Both RT schemes modulated immune cell activation and phenotype, but in distinct ways. Normofractionated RT led to a stronger upregulation of NKG2A expression on both NK and CD8 ⁺ T cells compared with hypofractionation. Moreover, the antitumor efficacy of normofractionated RT was dependent on the Qa1- NKG2A axis, whereas hypofractionated RT was not. Combining RT with dual anti-PD-L1 and anti-NKG2A blockade significantly improved survival, with the best outcomes observed in the normofractionated group (70% complete responses, figure 1).
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