DDRE-46. REDUCED CANCER CELL SENSITIVITY TO TUMOR TREATING FIELDS (TTFields) THROUGH ACTIVATION OF THE PI3K/AKT/mTOR SIGNALING PATHWAY CAN BE MITIGATED USING PI3K INHIBITORS OR PI3K/mTOR DUAL INHIBITORS

INTRODUCTION Tumor Treating Fields (TTFields) therapy is an approved anti-cancer treatment modality applied non-invasively and loco-regionally to the tumor region. TTFields have been demonstrated to extend life, however, most patients do eventually progress. The current study aimed to identify potential molecular mechanisms involved in reduced cellular sensitivity to TTFields. METHODS Cancer cells that exhibit reduced sensitivity to TTFields were generated by continuous long duration application of TTFields (7 or 13 days, depending on the cell line). Changes in cellular signaling pathways were evaluated in ovarian A2780 and glioblastoma U-87 MG cancer cells exposed to long-term relative to short-term (3 or 7 days, depending on the cell line) treatment with TTFields using Luminex multiplex assay followed by Western blot validation. The relevance of the affected pathways was confirmed through evaluation of the response to long-term application of TTFields in combination with pharmacological pathway inhibitors by measuring cell counts, apoptosis, and clonogenicity. Relevant pathway markers in tumor sections from tumor-bearing rats treated with TTFields were examined using immunohistochemistry. RESULTS Continuous long-term application of TTFields reduced cellular sensitivity to TTFields and was accompanied by increased levels of phosphorylated AKT, mTOR and additional proteins from the PI3K/AKT/mTOR signaling pathway. Increased phosphorylation of AKT was also evident in tumor sections from rats treated with TTFields. Concomitant use of TTFields with PI3K inhibitors or PI3K/mTOR dual inhibitors sensitized A2780 cells to long-term TTFields application. CONCLUSION Our study demonstrates that decreased cancer cell sensitivity to long-term TTFields application is mediated by activation of the PI3K/AKT/mTOR signaling pathway and provides a rationale for further examining the potential benefit of combining TTFields with PI3K or PI3K/mTOR dual inhibitors.

Study on the mechanism of Jiawei Linggui Zhugan Decoction in the treatment of myelodysplastic syndrome based on network pharmacological analysis

the research method of network pharmacology is used to explore the material basis and mechanism of modified Linggui Zhugan Decoction in the treatment of myelodysplastic syndrome. Methods: the main active components of 8 traditional Chinese medicines of Jiawei Linggui Zhugan Decoction were searched through tcmsp database, and the target was predicted. The relevant targets of myelodysplastic syndrome were searched through geo database, and the common action targets were obtained by intersection of traditional Chinese medicine targets and disease targets. The core targets were selected by topological analysis with Cytoscape software. Finally, go-bp biological function enrichment and KEGG pathway analysis were carried out based on R software. Results: according to the database analysis, there were 248 active compounds and 3695 targets in the modified Linggui Zhugan decoction, of which 34 were common targets with metabolic syndrome; Through the topological analysis of common targets, 9 core targets were selected. Go-bp biological function enrichment and KEGG pathway analysis found that it can play its therapeutic role through p53, AGE-RAGE, cellular sensitivity, NF KB and other signal pathways. Conclusion: modified Linggui Zhugan decoction may play a therapeutic role through p53 signaling pathway, AGE-RAGE signaling pathway, cellular sensitivity, NF kappa B signaling pathway and cell cycle, so as to provide a new scientific basis for its clinical and basic research.

Transcriptomic Alterations Induced by Vemurafenib After Treatment of Melanoma: A Comprehensive Bioinformatics Analysis

Backgrounds Melanoma is a highly aggressive kind of cancer with very poor prognosis. B-raf inhibitor vemurafenib has indeed harvested substantial clinical benefits. Nevertheless, its drug resistance has also hampered scientists effort towards successful melanoma treatment. In this study, we used data derived from the GEO database to analyze the effect on vemurafenib sensitive cell lines after vemurafenib treatment. GEO datasets GSE42872 (cohort1), GSE127988 (cohort2), GSE110054 (cohort3) were included in the analysis. Results We found 25 common Differentially Expressed Genes(DEGs) in 3 datasets, including 10 upregulated genes and 15 downregulated genes after vemurefenib application. Analysis using web tool TIMER showed significant correlation of the upregulated genes with immune infiltration level in skin cell melanoma. GO enrichment analysis showed that after vemurafenib treatment, all datasets showed downregulation in DNA replication and cell cycle arrest. Meanwhile, genes related to neuro-generation, extracellular matrix and cell-cell adhesion were significantly enriched in all three datasets. KEGG analysis showed that pathways like P53, PI3K-Akt, and Rap signaling pathways were enriched in DEGs after vemurafenib administration.Conlusions The findings of the candidate DEGs and pathways may not only reveal the cellular sensitivity to vemurafenib treatment, but also give rise to a better understanding of the mechanism of cancer cell cycle arrest and cellular resistance towards vemurafenib targeted therapy.

Transcriptomic Alterations Induced by Vemurafenib After Treatment of Melanoma: A Comprehensive Bioinformatics Analysis

Backgrounds: Melanoma is a highly aggressive kind of cancer with very poor prognosis. B-raf inhibitor vemurafenib has indeed harvested substantial clinical benefits. Nevertheless, its drug resistance has also hampered scientists effort towards successful melanoma treatment. In this study, we used data derived from the GEO database to analyze the effect on vemurafenib sensitive cell lines after vemurafenib treatment. GEO datasets GSE42872 (cohort1), GSE127988 (cohort2), GSE110054 (cohort3) were included in the analysis. Results: We found 25 common Differentially Expressed Genes(DEGs) in 3 datasets, including 10 upregulated genes and 15 downregulated genes after vemurefenib application. Analysis using web tool TIMER showed significant correlation of the upregulated genes with immune infiltration level in skin cell melanoma. GO enrichment analysis showed that after vemurafenib treatment, all datasets showed downregulation in DNA replication and cell cycle arrest. Meanwhile, genes related to neuro-generation, extracellular matrix and cell-cell adhesion were significantly enriched in all three datasets. KEGG analysis showed that pathways like P53, PI3K-Akt, and Rap signaling pathways were enriched in DEGs after vemurafenib administration.Conlusions: The findings of the candidate DEGs and pathways may not only reveal the cellular sensitivity to vemurafenib treatment, but also give rise to a better understanding of the mechanism of cancer cell cycle arrest and cellular resistance towards vemurafenib targeted therapy.

N6-Methyladenosine, DNA Repair, and Genome Stability

N6-methyladenosine (m6A) modification in mRNAs and non-coding RNAs is a newly identified epitranscriptomic mark. It provides a fine-tuning of gene expression to serve as a cellular response to endogenous and exogenous stimuli. m6A is involved in regulating genes in multiple cellular pathways and functions, including circadian rhythm, cell renewal, differentiation, neurogenesis, immunity, among others. Disruption of m6A regulation is associated with cancer, obesity, and immune diseases. Recent studies have shown that m6A can be induced by oxidative stress and DNA damage to regulate DNA repair. Also, deficiency of the m6A eraser, fat mass obesity-associated protein (FTO) can increase cellular sensitivity to genotoxicants. These findings shed light on the novel roles of m6A in modulating DNA repair and genome integrity and stability through responding to DNA damage. In this mini-review, we discuss recent progress in the understanding of a unique role of m6As in mRNAs, lncRNAs, and microRNAs in DNA damage response and regulation of DNA repair and genome integrity and instability.

Signatures of immune dysfunction in HIV and HCV infection share features with chronic inflammation in aging and persist after viral reduction or elimination

Chronic inflammation is thought to be a major cause of morbidity and mortality in aging, but whether similar mechanisms underlie dysfunction in infection-associated chronic inflammation is unclear. Here, we profiled the immune proteome, and cellular composition and signaling states in a cohort of aging individuals versus a set of HIV patients on long-term antiretroviral therapy therapy or hepatitis C virus (HCV) patients before and after sofosbuvir treatment. We found shared alterations in aging-associated and infection-associated chronic inflammation including T cell memory inflation, up-regulation of intracellular signaling pathways of inflammation, and diminished sensitivity to cytokines in lymphocytes and myeloid cells. In the HIV cohort, these dysregulations were evident despite viral suppression for over 10 y. Viral clearance in the HCV cohort partially restored cellular sensitivity to interferon-α, but many immune system alterations persisted for at least 1 y posttreatment. Our findings indicate that in the HIV and HCV cohorts, a broad remodeling and degradation of the immune system can persist for a year or more, even after the removal or drastic reduction of the pathogen load and that this shares some features of chronic inflammation in aging.

DNA Damage Response during Replication Correlates with CIN70 Score and Determines Survival in HNSCC Patients

Aneuploidy is a consequence of chromosomal instability (CIN) that affects prognosis. Gene expression levels associated with aneuploidy provide insight into the molecular mechanisms underlying CIN. Based on the gene signature whose expression was consistent with functional aneuploidy, the CIN70 score was established. We observed an association of CIN70 score and survival in 519 HNSCC patients in the TCGA dataset; the 15% patients with the lowest CIN70 score showed better survival (p = 0.11), but association was statistically non-significant. This correlated with the expression of 39 proteins of the major repair complexes. A positive association with survival was observed for MSH2, XRCC1, MRE11A, BRCA1, BRCA2, LIG1, DNA2, POLD1, MCM2, RAD54B, claspin, a negative for ERCC1, all related with replication. We hypothesized that expression of these factors leads to protection of replication through efficient repair and determines survival and resistance to therapy. Protein expression differences in HNSCC cell lines did not correlate with cellular sensitivity after treatment. Rather, it was observed that the stability of the DNA replication fork determined resistance, which was dependent on the ATR/CHK1-mediated S-phase signaling cascade. This suggests that it is not the expression of individual DNA repair proteins that causes therapy resistance, but rather a balanced expression and coordinated activation of corresponding signaling cascades.

Extensive crosstalk of G protein-coupled receptors with the Hedgehog signalling pathway

Hedgehog (Hh) ligands orchestrate tissue patterning and growth by acting as morphogens, dictating different cellular responses depending on ligand concentration. Cellular sensitivity to Hh ligands is influenced by heterotrimeric G protein activity, which controls production of the second messenger 3',5'-cyclic adenosine monophosphate (cAMP). cAMP in turn activates Protein kinase A (PKA), which functions as an inhibitor and (uniquely in Drosophila) an activator of Hh signalling. A few mammalian Gαi- and Gαs-coupled G protein-coupled receptors (GPCRs) have been shown to influence Sonic Hh (Shh) responses in this way. To determine if this is a more general phenomenon, we carried out an RNAi screen targeting GPCRs in Drosophila. RNAi-mediated depletion of more than 40% of GPCRs tested either decreased or increased Hh responsiveness in the developing Drosophila wing, closely matching the effects of Gαs and Gαi depletion, respectively. Genetic analysis indicated that the orphan GPCR Mthl5 lowers cAMP levels to attenuate Hh responsiveness. Our results identify Mthl5 as a new Hh signalling pathway modulator in Drosophila and suggest that many GPCRs may crosstalk with the Hh pathway in mammals.

Control of topoisomerase II activity and chemotherapeutic inhibition by TCA cycle metabolites

SUMMARYTopoisomerase II (topo II) is essential for disentangling newly replicated chromosomes. DNA unlinking involves the physical passage of one DNA duplex through another and depends on the transient formation of double-strand DNA breaks, a step exploited by frontline chemotherapeutics to kill cancer cells. Although anti-topo II drugs are efficacious, they also elicit cytotoxic side effects in normal cells; insights into how topo II is regulated in different cellular contexts is essential to improve their targeted use. Using chemical fractionation and mass spectrometry, we have discovered that topo II is subject to metabolic control through the TCA cycle. We show that TCA metabolites stimulate topo II activity in vitro and that levels of TCA flux modulate cellular sensitivity to anti-topo II drugs in vivo. Our works reveals an unanticipated connection between the control of DNA topology and cellular metabolism, a finding with important ramifications for the clinical use of anti-topo II therapies.