951 Spatial atlases of immunological development within the lymph node

• DOI: 10.1136/jitc-2022-sitc2022.0951
• Publication Date: 2022-11-08T01:13:46Z
• Authors: Tyler Hether; Stephanie Zimmerman; Zachary Lewis; Shanshan He; Liang Zhang; Kathy Ton; Liuliu Pan; Wei Yang; Stefan Phelan; Charles Glaser; Andy Nam; Emily Brown; Michael Patrick; Gary Geiss; Sarah Church; Michael Rhodes; Yan Liang; Jason Reeves; Joseph Beechem

<h3>Background</h3> Immunological mechanisms regulating detection and clearance of cancer, including CTLA-4 PD-L1, were discovered by studying the natural physiological processes immune cell maturation, attenuation, dissemination throughout body. Single-cell atlases mapping cells provide hints to these aspects immunology but lack essential spatio-temporal relationships between cells. With advent spatial 'omics we can resolve thousands RNA or more than 60 protein molecules simultaneously<i> in situ</i>, enabling direct insight into dynamics occurring as mature migrate through tissue. <h3>Methods</h3> We profiled lymph node samples using complementary platforms: GeoMx^® Digital Spatial Profiler CosMx™ Molecular Imager. GeoMx, whole transcriptomes from 5 patients focusing on key structures within germinal center, mantle zones, medulla, paracortex. CosMx, examine one FFPE with clear center zonation, analyzing 1000 genes 55 proteins across serial sections covering &gt;100mm² &gt;1.4 million cells/section. used a novel, semi-supervised clustering algorithm map CosMx scRNAseq. <h3>Results</h3> Across GeoMx observed 11,316 above background &gt;10% tissue regions profiled. identified 2,618 (FDR&lt;0.05) associated distinct functional regions, 928 significant pathways (FDR&lt;0.05). Profiling 27 types, 6 which not captured dissociated reference. Integrating results 643 enriched dark (n=283) light (n=206) zones at their interface (n=154), well 139 ligand-receptor interactions driving such pathways. For example, found co-stimulation CD28 was zone (FDR&lt;7.5e-06, GeoMx). confirmed that CD86 ligands B significantly colocalized receptors T_fH (FDR&lt;1.5e-05). also IL18 signaling macrophages (FDR&lt;5.6e-09), profiling CosMx. <h3>Conclusions</h3> The ecosystem provided platforms captures an unprecedented view architecture, hard-to-profile cells, immunological happening These findings shed novel interfaces, be compared infiltrate tumors identify perturbed active during tumor escape.