So, we're going to move on to our abstracts. The next is by Dr. Chen's group, where they're going to present their data on microglia, glioblastoma interactions, and how that influences survival. Thank you. Good afternoon, everyone. I'm Sanjay Dhagun from University of Minnesota. Many thanks to my mentor, Dr. Clark Chen, for giving me this opportunity to present this talk on his behalf. And I will be talking on the microglia-glioblastoma interaction influencing the clinical survival. And no disclosures as such. Glioblastoma is a deadly disease, and it needs no introduction to this audience. While we know that glioblastoma patients, patients who have mutations in isocitrate dehydrogenase, they have a relatively better survival prognosis as compared to those with wild-type IDH. But there still exists a small subset of those glioblastoma patients with wild-type IDH who exhibit an exceptional response. They survive beyond expectations. Maybe less than 10% of them. But then the molecular basis of this exceptional response still remains poorly understood. Talking about the cellular composition of gliomas, it has both the neoplastic tumor cells and the non-neoplastic cells like pericytes, endothelial cells, astrocytes, tumor-associated macrophages. These tumor-associated macrophages are recruited by the tumor bulk and by the glioma stem cells. These recruited and reprogrammed macrophages, they secrete soluble factors, which are responsible to expand the tumor bulk and the glioma stem cells as well. Survival analysis using three independent glioblastoma patient cohorts with wild-type IDH who underwent T-mozillavire treatment was performed. We tested 18 published glioblastoma survival gene signatures, including the Chinese genome atlas, the cancer genome atlas and the Rembrandt, and found that decreased expression of two inflammatory signatures, Kim and Lewis, was associated with an improved overall survival. The survival effect of these inflammatory signatures remain robust despite adjusting for variations including age, the Karnovsky-Promofan score, the extent of surgical resection and the concurrent T-mozillamide therapy and radiation. An integrated cell-based analysis was performed, and it told that the glioblastoma-associated microglia was a principal mediator in these inflammatory signatures. We isolated mouse microglia from a GL261 intracranial tumor model. Also, the microglia were isolated from a normal mouse brain. When these microglia from a tumor model were co-cultured with an independent cell line, we found they increased the colony formation by three-folds. Even co-implantation of these tumor microglia, the mouse microglia, with an independent cell line, they increased in vivo tumorigenicity and also enhanced the T-mozillamide resistance. These, it says that these glioblastomas, the microenvironment composed of microglia, they are inducing this glioblastoma to adopt a stem cell-like state, as evident by increase in tumorigenicity in vivo and enhanced T-mozillamide resistance. We tried to recapitulate these effects using a conditioned medium. These mice microglia were grown in a medium for three days, now called a conditioned medium. When the independent cell line was again grown in this same conditioned medium, it showed that it increased in colony formation. A proteomic analysis was performed for this conditioned medium, and we found that IL-11 was the key significant cytokine, which was the key mediator, and also it was associated with poor prognosis. When we sorted the tumor into different cells, we found, and we studied the IL-11 expression levels, we found that microglia was the principal source of this IL-11. Also, to find whether IL-11 was necessary, whether it was sufficient for glioblastoma tumorigenicity and enhanced resistance, we used IL-11 neutralizing antibodies, and we found when we used these antibodies, it abrogated the enhancing effect of the conditioned medium. When we used IL-11 treatment in a concentration similar to the IL-11 concentration found in glioblastoma specimens, it increased the sphere formation in vitro. Also, if you can see from these two, we can see that the noctopic expression IL-11, it increased the tumorigenicity and also enhanced the temozolomide resistance. What pathway mediates this effect of IL-11? We studied this, and we found that IL-11 causes phosphorylation of STAT3 in a time-dependent fashion. Also, it increased the level of MYC, which is a downstream effector of STAT3, by threefold. When we used agents, we inhibited the activation of STAT3 found that there was a decreased effect for the levels of IL-11 and also for MYC. When we tried to produce a heat map, we found that this IL-11 also increased the expression for MYC in vivo, as evidenced by this heat map. From a recently published study, this study concluded that the role of PI3K gamma, it is a lipid kinase secreted from the macrophages, the leukocytes, causes the accumulation of microglia in the tumor microenvironment. It acts as a critical switch, a molecular switch between immune stimulation and suppression, and activation of this actually causes suppression. We wondered, what if we inhibit this PI3K gamma? Will it reduce the accumulation of microglia in the tumor microenvironment? Will it convert a short-term survival to a long-term survival? We used both the genetic and the pharmacologic approaches. We used TG100-115, which is a PI3K gamma inhibitor, which is a pharmacologic agent. When we used the knockout mice, the PI3K gamma mice, we found that there was a significant decreased expression of these inflammatory, the clinically pertinent inflammatory signatures. The short-term survival was now a long-term survival in these mice, which were PI3K gamma negative mice available to us by genetic approach. And when we tried to inhibit PI3K gamma using the drug TG100-115, we found it decreased the microglia density by twofold. Also, it decreased the levels of IL-11. And there was significant increase in survival in the mouse cohort in which this drug was used. The PI3K gamma inhibition also decreased the expression of MYC in microglia when it was used. Moreover, it sensitized the glioblastoma to the effects of temozolomide. It augmented the antineoplastic effects for temozolomide. In a nutshell, in glioblastoma model, if you talk, we say that there is secretion of IL-11, which acts on the receptors, the microglia secrete IL-11, acts on the receptors, activates TAT3-MYC signaling axis. It induces glioblastoma cells to adopt a stem cell-like state, which have an enhanced resistance against temozolomide and increased tumor density, right? But this signaling axis, we found it is absent in exceptional responders. Also, this signaling axis could be inhibited by PI3K gamma inhibitor, like we used TG100-115. In a nutshell, I would say that we described the non-cell autonomous contribution of microglia secreted IL-11 to the chemotherapeutic response and the clinical survival. In aggregate, our study demonstrates that microglia-glioblastoma interaction, it determines the clinical survival, and also, I think it highlights the translation potential for personalized oncology care if we target this interaction. Thank you very much. Thank you.

microglia-glioblastoma interactions

clinical survival

isocitrate dehydrogenase mutations

inflammatory signatures

stem-cell-like state