Cynthia Hawkins – Hospital for Sick Children

Project Title: “Investigating treatment targets in the DIPG immune landscape using single cell technology”

Description of Project:

Diffuse intrinsic pontine gliomas (DIPG) are incurable tumours that account for nearly 10% of all pediatric brain cancers. Current therapies are ineffective, and the median survival remains under a year with near 100% fatality. DIPG tumour cells evade selective pressures of the immune environment which is key to it becoming an incurable entity. Modulating the evasive process has been a game-changer for skin, lung and breast cancer; transposing the existing drugs to improve outcomes in DIPG is promising. However we do not know yet if the molecular targets of these successful drugs are present in DIPG, in fact we know very little about the immune presence in DIPG. Further limitations are that current studies use bulk tumor tissue where highly heterogenous cell compositions of DIPG are masked by summative effects. Single-cell technologies have enabled us to identify cellular origins, genomics drivers of treatment resilience and highlighted treatment vulnerabilities in other cancers. Here we propose for the first time to leverage single-cell RNA sequencing to investigate the landscape of immune cells in DIPG tumor and interrogate for existing drug targets. We expected our results to allow the repurposing of successful drugs to improve outcomes in this incurable disease.

What receiving this award means:

I am very grateful to the Brain Tumour Foundation of Canada for supporting our DIPG research. These funds will allow us to use cutting edge technologies to investigate how the immune system interacts with brain tumour cells and brain tissue to help or hinder the growth and invasion of the cancer cells. This funding is critical to allow us to pursue novel ideas and generate breakthroughs for this devastating cancer.

Midpoint Review- September 2023

Aim 1: Investigate immune cell composition of DIPG by anatomical location

In our research project, we aimed to investigate the different types of immune cells in DIPG (Diffuse Intrinsic Pontine Glioma) based on where they are found in the body. We looked at samples from DIPG patients before and after treatment, including biopsies and tissues from autopsies taken from different parts of the body.

We managed to study around 20,000 immune cells from these DIPG samples, and we took a really close look at each individual cell. What we found were distinct programs or patterns of activity in these immune cells which are dependent on the location of the tumor and the treatment. This helped us understand more about how the immune system works in DIPG. One important discovery was that we could categorize immune cells into different groups. Some of these immune cells, called macrophages, seemed to be causing inflammation (M1-type) or promoting the growth of cancer (M2-type). Others, like T-regulatory cells, were responsible for suppressing the immune system. These distinct sub-populations of immune cells seem to be involved in how DIPG progresses and why it’s so hard to treat. Right now, we’re working on confirming these findings using additional analyses and in Aim 2.

Aim 2: Investigate the expression of immune targets in DIPG for drugs currently in clinical trials

Building on what we learned in Aim 1, we wanted to see if there are any targets in DIPG that could be utilized for potential drug treatments. We used all the data we collected to explore how we could use this information to make better cancer treatments. In our initial analysis, we found something called CSFR1, which could be a good target for immunotherapy, just like in other types of cancer.

We also checked for the presence of other markers, like CCL2, CCR2, CD40, CD47, SIRPa, PI3Kg, and TREM2. These markers are either being tested in clinical trials or are part of wider research/clinical studies. We are looking at how much of these markers are expressed in these distinct and well characterized immune cell populations. To further validate these findings, we’re going to test using more archival tissue samples housed at the Hospital for Sick Children and make sure that these markers are actually active and are therapeutically viable targets.

Disease impact and significance

Our work is groundbreaking because it’s the first time anyone has studied the immune system in DIPG at such a detailed level, looking at individual cells. The results we’re getting could help us plan new treatments that change the way we fight DIPG. Several of these treatment methods are currently undergoing clinical trials or have gained FDA approval, although they may have been tested in other types of cancer rather than specifically in DIPG. Finding these new treatments could be a game-changer for DIPG, a disease that’s really tough to treat using currently available options. We expect to finish our study by the end of 2023 and have the results ready in early 2024.

Final Report – March 2025

Diffuse Intrinsic Pontine Glioma (DIPG) is a deadly childhood brain cancer with limited treatment options and poor survival rates. This study explored the immune environment within DIPG tumours using advanced single-cell analysis of over 17,000 immune cells to identify potential new treatment targets. The research revealed that DIPG tumours, contrary to previous beliefs, contain a complex mix of immune cells, including microglia (the most abundant cell type), monocytes and T cells. A significant portion of these microglia showed signs of activation and inflammation, expressing pro-inflammatory markers and suggesting they may contribute to the tumour’s growth. The study also identified specific proteins, such as Ferritin Light Chain (FTL) and Solute Carrier Family 16 Member 10 (SLC16A10), which were highly expressed in tumour-associated macrophages after treatment, potentially promoting tumour progression and resistance. Additionally, the FOXP1 gene was found to be elevated in DIPG samples compared to normal brain tissue, indicating its potential involvement in tumour development and immunosuppression. These findings offer valuable insights into the DIPG immune landscape and highlight potential targets for the development of new immunotherapies to combat this devastating disease.