Abstract 1502: Deciphering the spatial and temporal dynamics of premetastatic niche in lung mediated by Treg and myeloid cell interactions

Abstract Introduction: The lung is a frequent site for metastases, where pre-metastatic niches (PMN) form in anticipation of tumor cell arrival. A deeper understanding of the molecular and immune features underlying the PMN formation may facilitate developing innovative strategies to prevent and treat metastasis more effectively in its early stages. Methods: In this study, we developed mouse models by subcutaneously implanting the 344SQ cell line, derived from the metastasis of a genetically engineered mouse model (GEMM) harboring KrasG12D/+; p53R172H/+ mutations, which reliably metastasizes to the lungs. We collected paired samples of the subcutaneous tumors, peripheral blood mononuclear cells (PBMCs), and lung tissue at pre-metastatic, early-metastatic, and late-metastatic phases. These samples were subjected to single-cell RNA sequencing (scRNA-seq), single-cell TCR/BCR sequencing (scTCR/BCR-seq), and spatial gene expression profiling with 10X Genomics Visium technology. Results: ScRNA analysis revealed large-scale comprehensive longitudinal changes in cell components throughout the invasion and metastasis cascade, which were validated by spatial gene expression. Furthermore, analysis of Visium data deduces the spatial information of these dynamic changes. We identified a subset of tumor-associated Tregs (TATregs, enriched in the subcutaneous tumor tissues) that existed in small amounts in the lung tissue in the pre-metastatic phase when there are no tumor cells detectable in the lung. These cells were characterized by high expression of Treg-associated suppressive genes (Tnfrsf9, Tnfrsf4, Tnfrsf18, Traf1, Nfkbia), inhibitory molecules (Ikzf2, Il2ra, Tight, Pdcd1), cytokines and chemokines (Ccr5, Ccr8, Ccl5, Cxcr6) and other Treg effector molecules (Id2, Hif1a, Rora, Il1r2, Itgav, Stat1, Icos, Areg). The scTCR-seq results and trajectory analysis revealed the TATregs might derive from minor clones of lung resident Tregs (LRTregs) and play an active role in remodeling an immunosuppressive microenvironment by closely interacting with other cell components in the TME. We also observed a significant alteration in the ratio of macrophage subpopulations between MacroSpic and MacroVcan. This ratio exhibited a continuous decrease in lung tissues, which could be an early indication of the initiation of pre-metastatic niche (PMN) conditioning. Conclusion: By simultaneously characterizing cellular gene expression, immune repertoires, and their corresponding spatial information, we revealed the dynamic changes of the lung tissue microenvironment (LTME) during from a normal state to pro-metastasis state at single-cell resolution. The LTME of metastatic organs may be primed before the establishment of metastasis. Citation Format: Yanhua Tian, Jian-Rong Li, Bo Zhu, Jared Fradette, Hong Chen, Chenyang Li, Leticia Rodriguez, Daniel Kraushaar, Luisa Solis, Alexandre Reuben, Ignacio Wistuba, Don L. Gibbons, Chao Cheng, Jianjun Zhang. Deciphering the spatial and temporal dynamics of premetastatic niche in lung mediated by Treg and myeloid cell interactions abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 1502.