Abstract 4635: Single cell definition of the immunogenic synapse between the tumorigenic schwann cells and tumor associated macrophages in Plexiform Neurofibroma
Authors: On Thomas J. , Zhang Xiyuan , Abbas Shahroze , Shern John 
Published: April 2023

Abstract
Background: Patients with Neurofibromatosis Type 1 (NF1) frequently develop benign tumors called Plexiform neurofibromas (PN). Macrophages compose 30-50% of the total cellular composition of PNs and have previously been shown to play a critical role in the PN development and growth. This study defines components of the immunogenic synapse between the neoplastic Schwann cell and the tumor associated myeloid compartment and investigates alterations of the synapse during treatment with MEK inhibition (MEKi). The IL34-CSF1R axis is nominated as a key node in the maintenance of the tumor associated macrophages in PN.
Methods: In this work, we applied single cell RNA sequencing (scRNAseq) to human primary tumor PN samples to comprehensively evaluate tumor cellular populations and their interactions. Resulting cellular populations were compared to a single cell atlas of tumor associated immune cells taken from a panel of tumor types. Ligand and receptor interactions were informatically extracted from scRNAseq data and immunohistochemistry was used to validate scRNAseq observations. scRNAseq was also used on pre and post treatment samples taken from three patients receiving the MEKi Selumetinib. Human primary macrophage cell cultures and qPCR were used to interrogate the roles of the IL34-CSF1R signaling and MEKi in PN.
Results: scRNAseq generated data on 45101 PN myeloid cells. Integration of these cells with a publicly available tumor atlas of 11 different tumors containing a total of 52195 cells discovered correlation of cell populations between the PN as compared to other tumor types. Tumor-promoting (M2) macrophages represented the largest portion (48%) of the PN myeloid population. Receptor-ligand analysis using cellphoneDB highlighted IL34-CSF1R (p&lt;.0001), TNF-TNFRSF1A (p&lt;.0001) and SIRPA-CD47 (p&lt;.0001) as significant interactions between the neoplastic Schwann cell and macrophages. IL34 and CSF1R protein expression was confirmed in human PN tumors with immunohistochemistry. scRNAseq data from NF1 patients PN tumors taken pre and post MEK inhibitor treatment, demonstrated decreased M2 macrophage gene expression of CD163(p&lt;.001), MRC1(p&lt;.001), TREM2(p&lt;.001) after treatment. Finally, IL34 stimulated primary human monocytes produced macrophages with an M2-like phenotype. MEKi treatment of these macrophages resulted in increased expression of inflammatory macrophage genes CXCL9 (p&lt;.05), CXCL10(p&lt;.001), IDO1(p&lt;.001).
Conclusions: PN myeloid populations contain large amounts of M2 macrophages with expression profiles consistent with TAM seen in a variety of malignant tumors. We postulate that IL34 expression by the neoplastic Schwann cell leads to expansion and maintenance of this TAM population. MEKi treatment on human PNs reduces number of macrophages in PNs and shifts macrophage gene expression towards inflammatory genes.
Citation Format: Thomas J. On, Xiyuan Zhang, Shahroze Abbas, John Shern. Single cell definition of the immunogenic synapse between the tumorigenic schwann cells and tumor associated macrophages in Plexiform Neurofibroma. [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 4635.