Gobbini, E.; Hubert, M.; Doffin, A.-C.; Eberhardt, A.; Hermet, L.; Li, D.; Duplouye, P.; Barrin, S.; Berthet, J.; Benboubker, V.; Grimont, M.; Sakref, C.; Perrot, J.; Tondeur, G.; Harou, O.; Lopez, J.; Dubois, B.; Dalle, S.; Caux, C.; Caramel, J.; Valladeau-Guilemond, J.

Background: Dendritic cells (DCs) are promising targets for cancer immunotherapies owing to their central role in the initiation and the control of immune responses. Their functions encompass a wide range of mechanisms mediated by different DC subsets. Several studies have identified human tumor-associated DC (TA-DC) populations through limited marker-based technologies, such as immunostaining or flow cytometry. However, tumor infiltration, spatial organization and specific functions in response to immunotherapy of each DC subset remain to be defined. Methods: Here, we implemented a multiplexed immunofluorescence analysis pipeline coupled with bio-informatic analyses to decipher the tumor DC landscape and its spatial organization within melanoma patients lesions, and its association with patients response to immune checkpoint inhibitors (ICI). For this aim, we analyze a cohort of 41 advanced melanoma patients treated with anti-PD1 alone or associated with anti-CTLA4. Distance and cell network analyses were performed to gain further insight into the spatial organization of tumor-associated DCs. A Digital Spatial Profiling analysis further characterized ecosystem of tumor-infiltrating DCs. Results: Plasmacytoid DCs (pDCs) were the most abundant DC population, followed by conventional cDC1 and mature DCs, present in equal proportions. In contrast to CD8+ T cell frequency, and despite varying densities, all DC subsets were associated with a favorable response to ICI. Distance and cell network analyses demonstrated that tumor-infiltrating DCs were largely organized in dense areas with high homotypic connections, except for cDC1 that exhibited a more scattered distribution. We identified four patterns of ecosystems with distinct preferential interactions between DC subsets. Significantly, the proximity and interactions between CD8+ T cells and cDC1 were positively associated with patients response to ICI. Conclusions: Our study unravels the complex spatial organization of DC subsets and their interactions in melanoma patient lesions, shedding light on their pivotal role in shaping the response to ICI. Our discoveries regarding the spatial arrangement of cDC1, especially with CD8+ T cells, provide valuable clues for improving immunotherapeutic strategies in melanoma patients.