IDH1 mutation activates mTOR signaling pathway, promotes cell proliferation and invasion in glioma cells.

BACKGROUND: Glioma is the most common type of brain tumors and isocitrate dehydrogenase (IDH1) gene is the most prominent molecular marker about the disease prognosis, response to therapy and patient survival. There are conflicting data about the effect of IDH1 mutation on glial cell proliferation, invasion and migration characteristics. The effect of IDH1 mutation on mTOR signaling pathway, which has key roles in tumorigenesis process, is limited and previous data is controversial. We aimed to explore the effect of wild type and mutant IDH1 overexpression on glioma cells and investigated the correlation with mTOR signaling pathway associated genes. METHODS AND RESULTS: U87-MG and A172 cells were transfected with different IDH1 mutant gene overexpressing (R132H, R132L, R132S, R132C) viral vectors. Cell proliferation, cell invasion and migration analysis as well as quantitative PCR analysis with the mutant glioma cell lines were performed. Forty-two patient derived glioma cells were obtained from patients with different glioma subtypes and cancer cells were enriched by culturing cells. Overexpression of both mutant and wild type IDH1 gene promoted the cell proliferation, but only IDH1 mutation increased cell invasion and migration. The expression of IDH1 mutation activated mTOR signaling via upregulation of WNTA, PRKAA2, GSK3B and MTOR genes as well as phosphorylated mTOR protein level. CONCLUSIONS: Our results highlighted IDH1 mutation upregulate mTOR signaling pathway and promote cell proliferation, invasion and migration.

A prelude to the proximity interaction mapping of CXXC5.

CXXC5 is a member of the zinc-finger CXXC family proteins that interact with unmodified CpG dinucleotides through a conserved ZF-CXXC domain. CXXC5 is involved in the modulation of gene expressions that lead to alterations in diverse cellular events. However, the underlying mechanism of CXXC5-modulated gene expressions remains unclear. Proteins perform their functions in a network of proteins whose identities and amounts change spatiotemporally in response to various stimuli in a lineage-specific manner. Since CXXC5 lacks an intrinsic transcription regulatory function or enzymatic activity but is a DNA binder, CXXC5 by interacting with proteins could act as a scaffold to establish a chromatin state restrictive or permissive for transcription. To initially address this, we utilized the proximity-dependent biotinylation approach. Proximity interaction partners of CXXC5 include DNA and chromatin modifiers, transcription factors/co-regulators, and RNA processors. Of these, CXXC5 through its CXXC domain interacted with EMD, MAZ, and MeCP2. Furthermore, an interplay between CXXC5 and MeCP2 was critical for a subset of CXXC5 target gene expressions. It appears that CXXC5 may act as a nucleation factor in modulating gene expressions. Providing a prelude for CXXC5 actions, our results could also contribute to a better understanding of CXXC5-mediated cellular processes in physiology and pathophysiology.

Development of a Rapid and Sensitive IDH1/2 Mutation Detection Method for Glial Tumors and a Comparative Mutation Analysis of 236 Glial Tumor Samples.

BACKGROUND: The presence of mutations in the isocitrate dehydrogenase 1 and 2 genes (IDH1/2) in glioma tumors is correlated with good prognosis upon standard-of-care treatment. Therefore, information on whether the glioma tumor has IDH1/2 mutations could be used in the correct diagnosis and management of glial tumors. The two most common techniques used to detect IDH1/2 mutations, immunohistochemistry (IHC) and Sanger sequencing, are prone to missing these mutations, especially if the tumor cells that carry the mutations constitute a small minority of the tumor itself. OBJECTIVES: We developed and validated a rapid method (3-mismatch-amplification refractory mutation system [3m-ARMS]) that can be used for pre-, intra- and postoperative detection of the most common IDH1/2 mutations in glial tumors with high specificity and sensitivity. We also conducted a comprehensive IDH1/2 mutation analysis in 236 glial tumor samples comparing 3m-ARMS, IHC and Sanger sequencing. METHODS: 3m-ARMS was optimized and validated for the specific and sensitive detection of the most common IDH1 and IDH2 mutations. We then analyzed 236 glial tumor samples for the presence of IDH1/2 mutations using 3m-ARMS, Sanger sequencing and IHC techniques. We then analyzed and compared the results, evaluating the diagnostic and screening potential of 3m-ARMS. RESULTS: Comparison of the three techniques used in the mutation analysis showed that 3m-ARMS-based IDH1/2 mutation detection was superior to IHC and Sanger sequencing-based IDH1/2 mutation detection in terms of accuracy, specificity and sensitivity, especially for tumor samples in which only a small minority of the cell population carried the mutation. 3m-ARMS could detect the presence of femtogram levels of IDH1/2 mutant DNA in DNA samples in which the mutant DNA-to-wild-type DNA ratio was as low as 1:100,000. CONCLUSION: Sanger sequencing and IHC-based methods have shortcomings when detecting mutations in glial tumors so can miss IDH1/2 mutations in glial tumors when used alone without proper modifications. 3m-ARMS-based mutation detection is fast and simple with potential for use as a diagnostic test for the majority of hot spot mutations in IDH1/2 genes. It can detect IDH1/2 mutations within an hour so can be adapted for intraoperative diagnosis.

Dynamic transcriptional events mediated by estrogen receptor alpha.

17beta-estradiol (E2), the main circulating estrogen hormone, is involved in a wide variety of physiological functions ranging from the development to the maintenance of many tissues and organs. The effects of E2 on cells are primarily conveyed by the transcription factors, estrogen receptor (ER) alpha and beta. The regulation of responsive genes by the well-defined ER alpha in response to E2 relies on complex and highly organized processes that dynamically integrate functions of many transcription regulators to induce spatiotemporal alterations in chromatin state and structure. Changes in gene expressions result in cell-specific responses that include proliferation, differentiation and death. Deregulation of E2-ER alpha signaling contributes to the initiation and progression of target tissue malignancies. We aim here to provide a review of recent findings on dynamic transcriptional events mediated by E2-ER alpha with the anticipation that a better understanding of complex regulatory mechanisms underlying ER actions would be a critical basis for the development of effective prognostic tools for and therapeutic interventions against estrogen target tissue malignancies.

Integration of multi-scale molecular modeling approaches with experiments for the in silico guided design and discovery of novel hERG-Neutral antihypertensive oxazalone and imidazolone derivatives and analysis of their potential restrictive effects on cell proliferation.

AT1 antagonists is the most recent drug class of molecules against hypertension and they mediate their actions through blocking detrimental effects of angiotensin II (A-II) when acts on type I (AT1) A-II receptor. The effects of AT1 antagonists are not limited to cardiovascular diseases. AT1 receptor blockers may be used as potential anti-cancer agents - due to the inhibition of cell proliferation stimulated by A-II. Therefore, AT1 receptors and the A-II biosynthesis mechanisms are targets for the development of new synthetic drugs and therapeutic treatment of various cardiovascular and other diseases. In this work, multi-scale molecular modeling approaches were performed and it is found that oxazolone and imidazolone derivatives reveal similar/better interaction energy profiles compared to the FDA approved sartan molecules at the binding site of the AT1 receptor. In silico-guided designed hit molecules were then synthesized and tested for their binding affinities to human AT1 receptor in radioligand binding studies, using [125I-Sar1-Ile8] AngII. Among the compounds tested, 19d and 9j molecules bound to receptor in a dose response manner and with relatively high affinities. Next, cytotoxicity and wound healing assays were performed for these hit molecules. Since hit molecule 19d led to deceleration of cell motility in all three cell lines (NIH3T3, A549, and H358) tested in this study, this molecule is investigated in further tests. In two cell lines (HUVEC and MCF-7) tested, 19d induced G2/M cell cycle arrest in a concentration dependent manner. Adherent cells detached from the plates and underwent cell death possibly due to apoptosis at 19d concentrations that induced cell cycle arrest.