The PLGG Program is investigating how the tumor microenvironment affects the growth and development of pediatric low-grade gliomas by asking questions such as why do tumors that express the KIAA1549-BRAF fusion genes only occur in the cerebellum? Tumors are mixtures of different cell types, including diverse types of cancer and non-cancer cells. This has implications for cancer treatment as distinct cancer cell types may respond differently to individual therapies. Moreover, immune cells present within a tumor can be harnessed to attack the cancer cells, an approach used in many other cancers such as melanoma. The Program’s researchers are taking two approaches to exploring the cellular diversity of tumors: developing culture systems for growing primary tumors in the laboratory and performing single cell RNA sequencing on individual tumor cells.
Defining cell types in PLGGs is important for the development of effective treatments of both primary and recurrent tumors. Single cell RNA sequencing (scRNAseq) is a powerful approach to characterizing individual cells in tumors. Pratiti Bandopadhayay, MD, MBBS, Rameen Beroukhim, MD, PhD, and Keith Ligon, MD, PhD developed technologies to 1) disaggregate PLGGs and sort cells according to expression of a glial progenitor marker, and 2) determine BRAF fusion status at the single cell level, among cells subject to single cell RNA sequencing. These technologies have allowed them to robustly distinguish between cancer cells and non-cancer cells, and to determine transcriptional profiles of each cell type at the single-cell level.
They found that the cancer cells formed two clusters, one containing radial glia/oligodendroglial precursor-like cells, and the other resembling more mature astrocytes. These clusters contrast with higher-grade tumors whose constituent cells are more similar immature neural progenitors, and include both astrocytic and oligodendroglial components. Consistent with the understanding of low-grade gliomas, the majority of the cancer cells were not in the cell cycle.
In addition to the BRAF-mutant cells, they identified multiple, non-cancer cells with the tumors. They found that non-cancer cells formed three major clusters of immune cells: microglia, T-cells and macrophages. Intriguingly, they found that the expression of immune checkpoint receptors and ligands varied among the tumor samples, a result that may be useful for developing immune therapies for pediatric tumors.
