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Introduction: Recently we found that RELA ependymomas (EPN) demonstrate inappropriate activation of the Hedgehog (Hh) pathway. However, how this activation is modulated remains to be investigated. Primary cilia are essential to positive and negative modulation of Hh signaling and changes in ciliation seem to be linked to Hh drug resistance in pediatric cancers. In contrast, targeting cilia integrity by molecules that prevents ciliary disassembly, such as Aurora-A (AURKA), has been shown to overcome this resistance. However, the role of the Hh pathway and the prevalence and function of cilia in pediatric RELA ependymoma (EPN) subgroup have not been examined. Thus, we explored the effect of Hh inhibitor, sonidegib, alone or in combination with Alisertib, AURKA inhibitor, in EPN RELA cell line. Methods: Hh pathway activation of RELA cell line (BXD-1425) was evaluated by immunostaining using ARL13B, a marker specific for ciliary membranes, proto-oncogene Smoothened (SMO) and Hh effector, GLI, antibodies in untreated cells or after stimulation with SMO agonist (SAG). Western blot was performed for protein expression of GLI1, GLI2, and full-length (FL) and repressor form (R) of Hh transcriptional program GLI3. Cell proliferation was assessed by CCK8 assay. Cell apoptosis was detected with Annexin V-FITC by flow cytometry. Results: We showed that EPN RELA cell line is frequently ciliated. Fluorescence intensity showed SMO and GLI recruitment to cilia in both BXD untreated cells and after stimulation with SAG, implicating in activation of Hh signals. Surprisingly, sonidegib treatment reduced ciliary formation in BXD cell line. Cilia loss was followed by increase of GLI recruitment to cilia and GLI2 expression, potentially by reduction of GLI3R. Our results indicate that impaired GLI processing probably by cilia disruption could explain persistent activation of downstream Hh pathway. To test this hypothesis, we targeted the cilia integrity through the combination of sonidegib with alisertib, to see if the cilia maintenance might affect Hh function and overcome sonidegib resistance. The combination prevented the primary cilia disruption followed by upregulation of GLI3R and downregulation of GLI recruitment to cilia, GLI2, and GLI3FL expression. Our data point towards to a synergistic combination that enhances sonidegib efficiency by primary cilia preservation, reducing cell proliferation and increasing apoptosis BXD cells compared to drugs alone treatment. Conclusion: Our findings suggest a synergistic combination of sonidegib with alisertib that enhances the inhibition of Hh major effectors such as GLIs (GLI2 and GLI3FL) and induces GLI3R expression. Our data suggest that combination of these agents may represent a novel approach for treatment of ST-EPN-RELA tumors to overcome chemoresistance and these mechanisms are potentially mediated by primary cilia, a key regulator for Hh activity. Citation Format: Taciani de Almeida Magalhães, Gustavo Alencastro Veiga Cruzeiro, Kleiton Silva Borges, Cherry Liu Yulu, Graziella Ribeiro de Souza, Keteryne Rodrigues da Silva, Carlos Alberto Scridelli, Adrian Salic, Luiz Gonzaga Tone. Alisertib acts synergistically with sonidegib by modulating primary cilia assembly in a pediatric RELA ependymoma cell line [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr A30.

Introduction Gold nanoparticles (GNPs) have drawn immense attention in cancer diagnosis and therapy during the past few years. Nanoparticles preferably leak and accumulate inside the tumour due to many reasons including high vascular density, increased vessel permeability, and defective lymphatic draining of tumours via a process called the enhanced permeability and retention (EPR) effect. Very small particles (<30 nm), although can enter the tumour tissue by the EPR effect, they also may leave it again by passive diffusion. This size-dependent cellular uptake thus limits the use of very small nanoparticles in cancer therapy. This study was conducted to improve the retention of small nanoparticles and overcome its size limitation by linking it to a noncoding DNA fragment. Material and methods A noncoding DNA fragment was amplified by PCR using a 5' thiol-labelled forward primer and an unmodified 3' reverse primer. After amplification, the PCR product was purified then conjugate to citrate stabilized10 nm GNPs. MCF-7 cells were treated with GNP-DNA conjugate using metafectene as a transfection agent for cytotoxicity studies. The GNP-DNA conjugate chemosensitization of MCF-7 cells to doxorubicin (Adriamycin ) was also tested. Results and discussions Treatment of MCF-7 cells with GNP-DNA conjugate resulted in a significant reduction in cell viability when compared to both control and unmodified GNP groups. The cell toxicity tests revealed that the conjugation of GNP with DNA lead to a 69.4-fold decrease in the IC 50 of GNP in MCF-7 cells. Furthermore, GNP-DNA conjugate showed a synergistic effect with doxorubicin leading to a significant reduction in cell viability at concentrations as low as 70 mM GNP-DNA and 1.6 mM doxorubicin. Conclusion The results of this study shed light on the marked effect of the modification of small GNP with DNA fragment on the effect of GNP in cancer cells and hence enables studying the effects of GNP size on cellular chemosensitivity without being restricted by GNP size-related differences in uptake efficiency. Introduction For Triple Negative Breast Cancers (TNBC) there are no effective specific targeted therapy readily available. CD44 is found overexpressed in many tumours, in particular TNBC, making this an attractive receptor for therapeutic targeting. Besides, Hsp90 inhibitors (Hsp90i) have been shown as promising molecules to treat cancer. Here we show our both drug delivery approaches to target TNBC. First, we describe a unique non-cationic liposome-based siRNA delivery system with a core composed of siRNA:protamine complexes and a shell designed for the active targeting of CD44-expressing cells using for the first time an anti-CD44 aptamer as targeting ligand. This was evaluated for the silencing of the luciferase reporter gene (luc2) in a TNBC breast cancer model in vitro and in vivo (orthotopically implanted) (Alshaer, 2015(Alshaer, , 2018. Secondly, we succeeded to use the inhibition of the chaperone Hsp90 against breast cancer. A promising Hsp90i derived from Novobiocin (...