Next, we have Derek Lee, who's going to talk about tumor-induced immunosuppression and how that promotes brain metastases in patients with non-small cell lung cancer. Thank you to the AANS Annual Scientific Meeting Committee for the invitation to present my research looking at tumor-induced immunosuppression in patients with brain metastases from non-small cell lung cancer. So, I have no disclosures. Lung cancer is one of the most common cancers and is the leading cause of cancer death in both men and women. Up to one in every three advanced-age patients develops brain metastases, which are a significant source of morbidity and mortality for these patients. Brain mets are actually the most common type of brain tumor. And as you can see from here, non-small cell lung cancer is the leading primary source of brain metastases. So as been said before, one therapeutic approach that's been gaining traction or that has gained traction in cancer treatment is immunotherapy. The immune system has the potential to target and destroy tumor cells through interactions between cytotoxic CD8 T-cells and the tumor cells. The overall aim of immunotherapy is to stimulate and support the immune system as it fights cancer. However, cancer cells have evolved mechanisms to evade and counteract the immune surveillance. As an activated immune cell becomes exhausted in the inflammatory microenvironment of the tumor, these immune checkpoints are upregulated. Our study focuses on the PD-L1 checkpoint, which has been shown to induce T-cell energy and apoptosis through binding with PD-1. And so consequently, drugs have been developed to inhibit these immune checkpoints, and multiple clinical trials have demonstrated favorable results, including these two on the right, looking at patients with non-small cell lung cancer. We also know that these immune checkpoints are not only expressed in the tumor microenvironment. Various cell types throughout the systemic immune compartment also express PD-L1. Prior work from our lab has demonstrated an upregulation of peripheral myeloid PD-L1 in patients with glioblastoma, and in patients who were treated with autologous heat shock protein vaccination, patients who had higher levels of peripheral myeloid PD-L1 were found to have worse overall survival. So given these findings, we wanted to further examine the systemic immune phenotype of patients with brain metastatic lung cancer, as well as investigate the mechanisms that induce immunosuppression in these patients. So here we present our cohort of patients. We were able to obtain peripheral blood samples and tumor sections from 34 patients. The median age was 66. About 60% were females. Most of the patients had ECOG performance scores of 0 or 1. 20% had EGFR mutation, 30% had KRAS mutation, and 3% had Alkali arrangement. Now on the right, we just show our survival data by gender and ECOG performance score. So first we quantified peripheral MDSCs and Tregs using flow cytometry with our gating scheme shown on the left, and we found that in patients who had brain metastatic lung cancer, they had increased MDSCs and Tregs in their blood. We also looked at peripheral myeloid PD-L1 and found that peripheral myeloid PD-L1 was also increased in patients who had brain metastases. And we divided these patients up into PD-L1 high and low groups around the median as shown on the right to look at survival. And we found that peripheral myeloid PD-L1 was significantly associated with worse survival in terms of progression-free and overall survival. We also looked at the impact of MDSCs and Tregs on survival, but neither of which were significant. Therefore we've decided to focus our studies on peripheral myeloid PD-L1. And from prior work in our lab with glioblastoma, we found that glioma-derived IL-6 induces myeloid PD-L1 expression. We showed in GBM that tumor cells secrete soluble factors, including IL-6, and that resulted in an upregulation of myeloid cell PD-L1. That resulted in T cell energy and apoptosis. And so we hypothesized that this chain of events was also occurring in patients with brain metastases from non-small cell lung cancer. We analyzed plasma IL-6 levels using ELISA and found that plasma IL-6 was significantly correlated with peripheral myeloid PD-L1 levels. So next we stimulated naive monocytes with tumor condition media from X-plant cell lines that we were able to generate from six patients who were undergoing resection of their brain metastasis. And we found a significant relationship between tumor condition media IL-6 levels and monocyte PD-L1. So that is to say the condition medias with higher levels of IL-6 induced a greater upregulation of myeloid PD-L1 expression. In order to determine that IL-6 was the predominant driver of myeloid PD-L1, we repeated the tumor condition media stimulation experiment with the addition of pharmacologic inhibitors. So we used sultuximab, which is an anti-IL-6 antibody, and tocilizumab, which is an anti-IL-6 receptor antibody. And at clinically relevant doses, both sultuximab and tocilizumab were able to return myeloid PD-L1 levels back down to baseline. So lastly, we wanted to confirm that IL-6 is primarily secreted from tumor cells in the brain metastasis microenvironment. To do this, we did immunoflush and staining on our tumor sections using IL-6, CD68 for infiltrating macrophages, and TTF1 for tumor cells. And as shown here, IL-6 predominantly co-localized with the TTF1-positive tumor cells. And so our model is that the primary lung cancer secretes these factors, including IL-6, and that these factors enter into systemic circulation, which then induces an upregulation of peripheral myeloid PD-L1 expression. This results in T cell energy and apoptosis systemically, which allows the cancer cells to enter into systemic circulation without being targeted by these circulating lymphocytes, resulting in late-stage events such as brain metastases. So in summary, we demonstrated that patients with brain metastatic lung cancer have systemic signs of immunosuppression, and that increased peripheral myeloid PD-L1 was associated with worse progression-free and overall survival. This upregulation in peripheral myeloid PD-L1 was primarily driven by a tumor-derived IL-6, and that pharmacologic inhibition of IL-6 could then return the myeloid PD-L1 levels back to baseline, and thus could provide a novel immunotherapeutic approach for brain metastatic non-small cell lung cancer patients. I'd like to thank Dr. Block and the rest of the lab, as well as Dr. Horbinski, who runs the Brain Tumor Bank at Northwestern. I'd also like to thank the Howard Hughes Medical Institute for funding this research through the MedFellows program. Thank you for your attention.

tumor-induced immunosuppression

brain metastases

non-small cell lung cancer

peripheral myeloid PD-L1 expression

IL-6 inhibition