Glioblastoma (GB) is a highly invasive type of brain cancer exhibiting poor prognosis. As such, its microenvironment plays a crucial role in its progression. Among the brain stromal cells, the microglia were shown to facilitate GB invasion and immunosuppression. However, the reciprocal mechanisms by which GB cells alter microglia/macrophages behavior are not fully understood. We propose that these mechanisms involve adhesion molecules such as the Selectins family. These proteins are involved in immune modulation and cancer immunity. We show that P-selectin mediates microglia-enhanced GB proliferation and invasion by altering microglia/macrophages activation state. We demonstrate these findings by pharmacological and molecular inhibition of P-selectin which leads to reduced tumor growth and increased survival in GB mouse models. Our work sheds light on tumor-associated microglia/macrophage function and the mechanisms by which GB cells suppress the immune system and invade the brain, paving the way to exploit P-selectin as a target for GB therapy.

Glioblastoma (GB) is the most lethal type of primary tumor in the central nervous system. Current treatments include surgical resection followed by chemotherapy and radiotherapy. With this therapeutic regimen, the median survival is less than two years. However, these treatments do not much improve the overall survival of GB patients. GBs are highly angiogenic and invasive tumors and often acquire resistance to therapy. The invasive nature of the disease limits the ability to achieve complete resection of the tumor and the majority of GB patients will experience disease relapse. Moreover, GB is highly heterogeneous, harboring different mutations and presenting different phenotypes. As the brain is considered to be an immune‐privileged tissue, GB is defined as a cold tumor for which current immunotherapies have not yet been demonstrated to improve survival. On top of these challenges, the blood brain barrier (BBB) restricts the uptake of drugs by the brain, thus limiting the therapeutic options. Therefore, enormous efforts are being dedicated to the development of novel nanomedicines, which will be able to cross the BBB and specifically target the cancer cells. Here, the current achievements in drug delivery and novel therapeutic approaches for GB therapy are discussed.

During sleep our brain switches between two starkly different brain states - slow wave sleep (SWS) and rapid eye movement (REM) sleep. While this two-state sleep pattern is abundant across birds and mammals, its existence in other vertebrates is not universally accepted, its evolutionary emergence is unclear and it is undetermined whether it is a fundamental property of vertebrate brains or an adaptation specific to homeotherms. To address these questions, we conducted electrophysiological recordings in the Agamid lizard, Laudakia vulgaris during sleep. We found clear signatures of two-state sleep that resemble the mammalian and avian sleep patterns. These states switched periodically throughout the night with a cycle of ~90 seconds and were remarkably similar to the states previously reported in Pogona vitticeps. Interestingly, in contrast to the high temperature sensitivity of mammalian states, state switches were robust to large variations in temperature. We also found that breathing rate, micro-movements and eye movements were locked to the REM state as they are in mammals. Collectively, these findings suggest that two-state sleep is abundant across the agamid family, shares physiological similarity to mammalian sleep, and can be maintain in poikilothems, increasing the probability that it existed in the cold-blooded ancestor of amniotes.

Development of chemo-resistance is a major challenge in glioblastoma (GB) treatment. This phenomenon is often driven by increased activation of genes associated with DNA repair, such as the alkyl-removing enzyme O6-methylguanine-DNA methyltransferase (MGMT) in combination with overexpression of canonical genes related to cell proliferation and tumor progression, such as Polo-like kinase 1 (Plk1). Hereby, we attempt to sensitize resistant GB cells using our established amphiphilic poly(α)glutamate (APA): small interfering RNA (siRNA) polyplexes, targeting Plk1. Furthermore, we improved brain-targeting by decorating our nanocarrier with sulfonate groups. Our sulfonated nanocarrier showed superior selectivity towards P-selectin (SELP), a transmembrane glycoprotein overexpressed in GB and angiogenic brain endothelial cells. Self-assembled polyplexes of sulfonated APA and siPlk1 internalized into GB cells and into our unique 3-dimensional (3D) GB spheroids inducing specific gene silencing. Moreover, our RNAi nanotherapy efficiently reduced the cell viability of both chemo-sensitive and chemo-resistant GB cells. Our developed sulfonated amphiphilic poly(α)glutamate nanocarrier has the potential to target siRNA to GB brain tumors. Our findings may strengthen the therapeutic applications of siRNA for chemo-resistant GB tumors, or as a combination therapy for chemo-sensitive GB tumors.

Glioblastoma (GB) is an aggressive type of brain cancer with high mortality rate. It is a highly angiogenic tumor exhibiting an extremely invasive nature. As such, its brain microenvironment plays a crucial role in its progression. Microglia are the brain resident immune cells which have been shown to facilitate GB cell invasion and immune suppression. The mechanism by which GB cells alter microglia behavior is yet to be fully understood. One proposed mechanism involves adhesion molecules such as the Selectins family of proteins which are expressed on the surface of endothelial and immune cells and are involved in immune modulation and cancer immunity. We have previously shown that P-Selectin (SELP) is expressed by GB cells. Here, we investigated the factional role of SELP in GB-microglia interactions. First, we found that microglia cells facilitate the expression and secretion of SELP by GB cells, and that GB cells facilitate the expression of P-Selectin ligand by microglia. We then showed that SELP mediates microglia-enhanced GB invasion and proliferation in 2D and 3D in vitro models and has a role in microglia activation state. These findings were validated in vivo, showing that inhibition or downregulation of SELP leads to reduced tumor growth, increased overall survival and improved immune response. Single-Cells RNA-seq analysis of the tumors revealed an increase in pro-inflammatory microglia signature, reduction in cancer cell tumorigenesis potential and improved T cell activation. Our results indicated that SELP has an important role in GB progression and microenvironment activation. This work can improve our understanding of tumor-associated microglia function and the mechanisms by which GB cells suppress the immune system and invade the brain tissue. Citation Format: Eilam Yeini, Paula Ofek, Sabina Pozzi, Nitzan Albeck, Dikla Ben-Shushan, Galia Tiram, Sapir Golan, Ron Kleiner, Ron Sheinin, Shlomit Reich-Zeliger, Rachel Grossman, Zvi Ram, Henry Brem, Thomas Hyde, Prerna Magod, Dinorah Friedmann-Morvinski, Asaf Madi, Ronit Satchi-Fainaro. P-selectin axis plays a key role in microglia immunophenotype and glioblastoma progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2716.