EXTH-06. ATR INHIBITORS AS MONOTHERAPY AND COMBINATORIAL THERAPY WITH TEMOZOLOMIDE IN PRECLINICAL GLIOBLASTOMA MODELS

Glioblastoma (GBM) is an aggressive brain tumor and has an extremely poor prognosis despite the use of multiple treatment modalities. DNA damage response (DDR) signaling plays an important role in inducing radiation and temozolomide (TMZ) resistance and hence has emerged as a molecular target for therapeutic development. The Ataxia Telangiectasia and Rad3-related protein (ATR) kinase is a key regulator of the DDR machinery, activated by DNA damage. Here, we show that three clinical-grade ATR inhibitors (Bay1895344, AZD6738 and Berzosertib) had similar selective sensitivity pattern across 16 glioma-like stem cell (GSC) lines tested. ATR inhibitors inhibited the growth of GSCs at low nanomolar range concentrations. Interestingly, all three ATR inhibitors showed a significant synergism with TMZ in a selective group of GSCs (Combination index and Bliss model). Importantly, we demonstrate that MGMT promoter methylation status was associated with cellular response to combination treatment with preferential inhibition of cell growth in MGMT promotor methylated (MGMT deficient) cell lines. Further, we compared the RNA-seq data from GSCs in the synergism and non-synergism group and used multiple complementary approaches to identify the response marker that confer sensitivity to combination therapy. Our preliminary data analysis identified several genes that confer sensitivity to combination treatment and studies are underway to validate the data. We also investigated the efficacy of BBB penetrant ATR inhibitor BAY 1895344 in orthotropic xenografts and administration of BAY 1895344 and TMZ combination significantly reduced tumor size and extended survival in an intracranial animal model. Combining ATR inhibitor with TMZ was well tolerated and did not confer additional toxicity as the body weights of TMZ and combination groups were comparable. The findings from this study provides a rationale to use ATR inhibitors in combination with TMZ in MGMT methylated tumors to improve therapeutic efficacy of standard of care for GBM patients.

Sensitivity analysis of the nonparametric criterion of aircraft flght control system sensors failures detection and isolation

Failures of the aircraft control system sensors can cause both deterioration of stability and controllability characteristics and the inability of safe automatic control. It is necessary to detect and isolate such failures to determine the time and place of their occurrence in order to disable failed sensors or to diagnose them subsequently for reconfiguration during the flight. The direct use of traditional parametric approaches for sensors health monitoring by using their mathematical models is impossible due to the lack of data about the true information input signals received by their sensitive elements. This leads to the necessity of solving the problem of modeling the aircraft flight dynamics with a high level of uncertainties, which makes it difficult to utilize the functional control methods and necessitate the use of excessive sensor hardware redundancy. Well-known nonparametric methods either require a priori knowledge base, preliminary training or long-term tuning on a large volume of real flight data or have low selective sensitivity for reliable detection of failed sensors. In this work, the original nonparametric criterion for detecting and isolating sensors failures is derived. Its sensitivity is analyzed by using a complete nonlinear mathematical model of aircraft flight dynamics with a regular flight control system. The theoretical value and the criterion sensitivity coefficients are determined. The formula for the automatic evaluation of the float criterion threshold value is given. A high convergence of the results with theoretical ones is shown. This makes it possible to use the obtained criterion not only for the instant detection and isolation of sensors failures, but also for preliminary diagnostics of their quantitative characteristics.

Selective sensitivity of the Black Sea microalgae to viral infection after exposure to a constant magnetic field

The change in sensitivity to viral infection of cultures of three species of Black Sea microalgae (Tetraselmis viridis, Dunaliella viridis and Phaeodactylum tricornutum) after exposure to a constant unidirectional magnetic field with a magnetic induction of 600 G is experimentally studied. The studies were carried out with using a developed laboratory installation. With the duration of the experiments 24 h and 48 h, it was recorded that being in a magnetic field resulted in an increase in the resistance to viral lysis in microalgae T. viridis, its decrease in P. tricornutum and no changes in D. viridis. We assume that the revealed changes in the sensitivity of microalgae to viral infection after exposure to a magnetic field are due to their species characteristics and selectivity.

ASXL1 mutations are associated with distinct epigenomic alterations that lead to sensitivity to venetoclax and azacytidine

The BCL2-inhibitor, Venetoclax (VEN), has shown significant anti-leukemic efficacy in combination with the DNMT-inhibitor, Azacytidine (AZA). To explore the mechanisms underlying the selective sensitivity of mutant leukemia cells to VEN and AZA, we used cell-based isogenic models containing a common leukemia-associated mutation in the epigenetic regulator ASXL1. KBM5 cells with CRISPR/Cas9-mediated correction of the ASXL1G710X mutation showed reduced leukemic growth, increased myeloid differentiation, and decreased HOXA and BCL2 gene expression in vitro compared to uncorrected KBM5 cells. Increased expression of the anti-apoptotic gene, BCL2, was also observed in bone marrow CD34+ cells from ASXL1 mutant MDS patients compared to CD34+ cells from wild-type MDS cases. ATAC-sequencing demonstrated open chromatin at the BCL2 promoter in the ASXL1 mutant KBM5 cells. BH3 profiling demonstrated increased dependence of mutant cells on BCL2. Upon treatment with VEN, mutant cells demonstrated increased growth inhibition. In addition, genome-wide methylome analysis of primary MDS samples and isogenic cell lines demonstrated increased gene-body methylation in ASXL1 mutant cells, with consequently increased sensitivity to AZA. These data mechanistically link the common leukemia-associated mutation ASXL1 to enhanced sensitivity to VEN and AZA via epigenetic upregulation of BCL2 expression and widespread alterations in DNA methylation.

Impaired anaplerosis is a major contributor to glycolysis inhibitor toxicity in glioma

Background Reprogramming of metabolic pathways is crucial to satisfy the bioenergetic and biosynthetic demands and maintain the redox status of rapidly proliferating cancer cells. In tumors, the tricarboxylic acid (TCA) cycle generates biosynthetic intermediates and must be replenished (anaplerosis), mainly from pyruvate and glutamine. We recently described a novel enolase inhibitor, HEX, and its pro-drug POMHEX. Since glycolysis inhibition would deprive the cell of a key source of pyruvate, we hypothesized that enolase inhibitors might inhibit anaplerosis and synergize with other inhibitors of anaplerosis, such as the glutaminase inhibitor, CB-839. Methods We analyzed polar metabolites in sensitive (ENO1-deleted) and resistant (ENO1-WT) glioma cells treated with enolase and glutaminase inhibitors. We investigated whether sensitivity to enolase inhibitors could be attenuated by exogenous anaplerotic metabolites. We also determined the synergy between enolase inhibitors and the glutaminase inhibitor CB-839 in glioma cells in vitro and in vivo in both intracranial and subcutaneous tumor models. Results Metabolomic profiling of ENO1-deleted glioma cells treated with the enolase inhibitor revealed a profound decrease in the TCA cycle metabolites with the toxicity reversible upon exogenous supplementation of supraphysiological levels of anaplerotic substrates, including pyruvate. ENO1-deleted cells also exhibited selective sensitivity to the glutaminase inhibitor CB-839, in a manner rescuable by supplementation of anaplerotic substrates or plasma-like media PlasmaxTM. In vitro, the interaction of these two drugs yielded a strong synergistic interaction but the antineoplastic effects of CB-839 as a single agent in ENO1-deleted xenograft tumors in vivo were modest in both intracranial orthotopic tumors, where the limited efficacy could be attributed to the blood-brain barrier (BBB), and subcutaneous xenografts, where BBB penetration is not an issue. This contrasts with the enolase inhibitor HEX, which, despite its negative charge, achieved antineoplastic effects in both intracranial and subcutaneous tumors. Conclusion Together, these data suggest that at least for ENO1-deleted gliomas, tumors in vivo—unlike cells in culture—show limited dependence on glutaminolysis and instead primarily depend on glycolysis for anaplerosis. Our findings reinforce the previously reported metabolic idiosyncrasies of in vitro culture and suggest that cell culture media nutrient composition more faithful to the in vivo environment will more accurately predict in vivo efficacy of metabolism targeting drugs.

Person agreement with inherent case DPs in Chirag Dargwa

The article argues that the syntactic behavior of non-absolutive subjects of finite clauses in the Nakh-Daghestanian language Chirag Dargwa is a result of their interaction with two different functional heads in a clause: v and T. Discussing empirical data from Chirag, I present the puzzling behavior of person agreement, which shows selective sensitivity to arguments in the ergative, dative, and genitive cases. The primary evidence comes from the periphrastic causative, which displays some typologically unusual properties in case marking and agreement. I show that the ability to trigger person agreement is not an intrinsic property of ergative, dative, and genitive DPs in Chirag, but rather is endowed to the highest DP in T’s c-command domain over the course of the derivation. I propose that all non-absolutive subjects start out as DPs assigned inherent case and a theta-role by v, and that T further assigns structural nominative case to the DP in Spec,vP, thus making it accessible to φ-probes.

MYCN mediates TFRC-dependent ferroptosis and reveals vulnerabilities in neuroblastoma

MYCN amplification is tightly associated with the poor prognosis of pediatric neuroblastoma (NB). The regulation of NB cell death by MYCN represents an important aspect, as it directly contributes to tumor progression and therapeutic resistance. However, the relationship between MYCN and cell death remains elusive. Ferroptosis is a newly identified cell death mode featured by lipid peroxide accumulation that can be attenuated by GPX4, yet whether and how MYCN regulates ferroptosis are not fully understood. Here, we report that MYCN-amplified NB cells are sensitive to GPX4-targeting ferroptosis inducers. Mechanically, MYCN expression reprograms the cellular iron metabolism by upregulating the expression of TFRC, which encodes transferrin receptor 1 as a key iron transporter on the cell membrane. Further, the increased iron uptake promotes the accumulation of labile iron pool, leading to enhanced lipid peroxide production. Consistently, TFRC overexpression in NB cells also induces selective sensitivity to GPX4 inhibition and ferroptosis. Moreover, we found that MYCN fails to alter the general lipid metabolism and the amount of cystine imported by System Xc(−) for glutathione synthesis, both of which contribute to ferroptosis in alternative contexts. In conclusion, NB cells harboring MYCN amplification are prone to undergo ferroptosis conferred by TFRC upregulation, suggesting that GPX4-targeting ferroptosis inducers or TFRC agonists can be potential strategies in treating MYCN-amplified NB.

Reusable Sensor for Strontium Sulfate Scale Monitoring in Seawater

The onset of scaling in oil pipelines can halt or drastically reduce oil production, causing huge financial losses and delays. Current methods used to monitor scaling can take weeks, while the scaling process only takes few hours. The proposed sensor is designed for online monitoring of strontium ions concentration in seawater as an early scaling indicator. The sensor operates in the GHz range by probing the shift in the resonance frequency due to changes in the ionic concentrations of the medium. The results show selective sensitivity to changes in the strontium ions concentration even in the presence of many other ions found in seawater. The measured sensitivity is found to be stable and linear with a detection level of better than 0.08% (0.042 mol/L) of strontium ions in seawater. This work demonstrates a robust GHz sensor for strontium sulfate scale monitoring and early detection, which could be used in the oil industry to prevent huge production losses. These results could also be extended further to target the monitoring of other ions in different industrial sectors.

Features of Recognizing Facial Expressions in Short Time Intervals: the Level of Anxiety and Professional Affiliation

The aim of this study was to investigate how differences in anxiety levels relate to selective sensitivity to basic emotions (emotional bias) with minimal exposure time. Masked pictures of happiness, angry, fear, disgust, surprise, sad and neutral facial expressions were presented to 298 men at exposure times in intervals 16ms, 34ms, 49ms, 66ms. After presenting each image, the participants chose on the screen by pressing a key the name of an emotion suitable, in their opinion, Taylor Manifest Anxiety Scale (TMAS) was used to measure of trait anxiety. There were subjects of various professional groups (firefighters, military, athletes, psychologists, mathematicians). We found that Selective sensitivity to basic emotions at exposure times up to 49ms is determined by internal interpolation of the perceiver’s personality. Highly anxious men are unconsciously more likely to choose fear, anger and disgust. The increase in anxiety are accompanied by decreased preference of anger and happiness. Low-anxious men unconsciously ignore fear, anger, disgust, and preferred neutral face. Men of different professions are differed in the level of anxiety and emotional bias in basic emotions. Firefighters have the lowest level of anxiety, mathematics have the highest.

Impaired Anaplerosis Is a Major Contributor to Glycolysis Inhibitor Toxicity in Glioma

Reprogramming of metabolic pathways is crucial to satisfy the bioenergetic and biosynthetic demands and maintain the redox status of rapidly proliferating cancer cells. In tumors, the tricarboxylic acid (TCA) cycle generates biosynthetic intermediates by oxidation of anaplerotic substrates, such as glucose-derived pyruvate and glutamine20 derived glutamate. We have previously documented that a subset of tumors with 1p36 homozygous deletion exhibit co-deletion of ENO1, in turn becoming extremely dependent on its redundant isoform ENO2 and sensitive to an overall enzymatic deficiency of enolase. Metabolomic profiling of ENO1-deleted glioma cells treated with an enolase inhibitor revealed a profound decrease in TCA cycle metabolites, which correlated with cell-line specific sensitivity to enolase inhibition, highlighting the importance of glycolysis derived pyruvate for anaplerosis. Correspondingly, the toxicity of the enolase inhibitor was significantly attenuated by exogenous supplementation of supraphysiological levels of anaplerotic substrates including pyruvate. These findings led us to hypothesize that cancer cells with ENO1 homozygous deletions treated with an enolase inhibitor might show exceptional sensitivity to inhibition of glutaminolysis because of reduced anaplerotic flow from glycolysis. We found that ENO1-deleted cells indeed exhibited selective sensitivity to the glutaminase inhibitor CB-839, and this sensitivity was also attenuated by exogenous supplementation of anaplerotic substrates including pyruvate. Despite these promising in vitro results, the antineoplastic effects of CB-839 as a single agent in ENO1-deleted xenograft tumors in vivo were modest in both intracranial orthotopic tumors, where the limited efficacy could be attributed to the blood brain barrier (BBB), and subcutaneous xenografts, where BBB penetration is not an issue. This contrasts with the enolase inhibitor HEX, which, despite its negative charge, achieved antineoplastic effects in both intracranial and subcutaneous tumors. Together, these data suggest that at least for 1p36-deleted gliomas, tumors in vivo—unlike cells in culture—show limited dependence on glutaminolysis and instead primarily depend on glycolysis for anaplerosis. Our findings reinforce the previously reported metabolic idiosyncrasies of the in vitro and in vivo environments as the potential reasons for the differential efficacy of metabolism targeted therapies in in vitro and in vivo systems.