Session: 622. Lymphomas: Translational–Non-Genetic: Poster III
Hematology Disease Topics & Pathways:
Poster-only abstracts
Methods: We used a tissue microarray (TMA) containing formalin-fixed paraffin-embedded samples from 115 diagnostic cHL tumors. The TMA was stained with a 7-plex immunohistochemistry panel to characterize CAFs [platelet-derived growth factor (PDGFR) -alpha and -beta, fibroblast-activating protein (FAP), secreted protein acidic and rich in cysteine (SPARC)], macrophages (CD68), Hodgkin-Reed Sternberg (HRS) cells (CD30), and leukocytes (CD45). Image processing and quality control were performed by combining Ilastik and CellProfiler softwares, and nuclei segmented by a pretrained deep learning segmentation model. Single cell features from the images were extracted using HistoCAT software. We utilized the Phenograph clustering algorithm for cell phenotyping and a permutation test by HistoCAT for neighborhood analysis.
Results: We identified a total of 952,099 single cells, which were split into 25 distinct phenotype subsets by the Phenograph clustering spanning CAFs, macrophages, leukocytes, and HRS cells. CAFs were classified into seven distinct subsets based on the status of the different CAF markers. Median proportion of all CAFs was 28% (range 2-80%), the proportion being higher in the nodular sclerosis subtype compared to the others. In general, higher proportions of CAFs associated with favorable freedom from treatment failure (FFTF) independently of the subtype, age, and stage (P<0.01). On the contrary, a subcluster of CD45+ immune cells with strong FAP-positivity, which were characterized as macrophages, was enriched in other than nodular sclerosis subtype (P<0.001) and associated with worse FFTF independently of age, stage, and subtype (P=0.01). The neighborhood analysis allowed identification of colocalization and statistically significant interaction or avoidance between pairs of cell phenotypes. For instance, higher proportions of FAP+PDGFRalpha+ CAFs interacting with other cells predicted better FFTF (P<0.01), whereas interactions of FAP+ macrophages with other cells were associated with worse FFTF. Despite the positive impact of CAF proportions on cHL outcome, patients with CD30+ HRS cells surrounded by PDGFRbeta+ CAFs had worse FFTF (P<0.001).
Conclusions: We have characterized distinct CAF subsets in cHL and demonstrate their favorable clinical impact on cHL outcome. We also identified a novel subset of FAP-positive macrophages, the proportions and interactions of which translated to poor outcome. Our data highlight that not only the cell proportions, but the cell-cell interactions and spatial context play a crucial role. Further validation of the findings is ongoing.
Disclosures: Leppä: Genmab: Consultancy; Beigene: Consultancy; Hutchmed: Research Funding; Gilead: Consultancy, Honoraria; Bayer AG: Research Funding; Nordic Nanovector: Research Funding; Incyte: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene/BMS: Research Funding; Abbvie: Consultancy; Novartis: Honoraria, Research Funding; Roche: Consultancy, Research Funding; Sobi: Consultancy; Orion: Consultancy.
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