Tumor draining lymph nodes (TDLNs) function as an anatomic niche which primes immune cells to generate anti-cancer responses. While tumors are frequently staged based on the presence or absence of TDLN metastasis, mechanisms for how tumors impact the TDLN microenvironment and immunologic responses are incompletely understood. Dogs with spontaneously arising tumors provide an opportune model to study immuno-oncology, sharing many similarities to human patients with their environment, clinical care, and pathobiology. To evaluate how tumors impact TDLN responses, we conducted single cell RNA sequencing (scSeq) on cryopreserved TDLNs of dogs with osteosarcoma (n=4) and healthy controls (n=2). We obtained an average of 3,984 cells per sample (range 1,603-5,259) with a total of 4,780 cells from healthy samples and 19,121 cells from TDLN samples. All samples were integrated into one dataset, then unsupervised clustering was completed. Each unique cell cluster was assigned an identity using canonical markers and reference mapping to human datasets. Following cell classification, we observed notable changes in gene expression across all cells isolated from the LNs, with most differences arising from changes to the transcriptome of myeloid cells. Although still preliminary, the data also demonstrate differences in the abundance of T, B, and myeloid cells between healthy and tumor draining LNs. Taken together, this dataset provides indications of the changes TDLNs undergo in dogs with osteosarcoma.

Citation Format: Samuel A. Brill, Dylan T. Ammons, Anne C. Avery, Douglas H. Thamm. Evaluation of tumor draining lymph nodes in dogs with spontaneously arising osteosarcoma using single-cell sequencing. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4465.