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  • TERT and DNMT1 expression predict sensitivity to decitabine in gliomas. Neuro-oncology Park, J., Sahm, F., Steffl, B., Arrillaga-Romany, I., Cahill, D., Monje, M., Herold-Mende, C., Wick, W., Turcan, S. 2020

    Abstract

    BACKGROUND: Decitabine (DAC) is an FDA approved DNA methyltransferase (DNMT) inhibitor that is used in the treatment of patients with myelodysplastic syndromes. Previously, we showed that DAC has marked antitumor activity against gliomas with isocitrate dehydrogenase 1 (IDH1) mutations. Based on promising preclinical results, a clinical trial has been launched to determine the effect of DAC in IDH-mutant gliomas. The next step is to comprehensively assess the efficacy and potential determinants of response to DAC in malignant gliomas.METHODS: The expression and activity of TERT and DNMT1 were manipulated in patient-derived IDH1-mutant and -wildtype glioma lines, followed by assessment of cell proliferation with DAC treatment alone or in combination with telomerase inhibitors. RNA sequencing, KEGG enrichment and correlation analysis were performed.RESULTS: IDH1-mutant glioma tumorspheres with hemizygous co-deletion of chromosome arms 1p/19q were particularly sensitive to DAC and showed significant inhibition of DNA replication genes. Our transcriptome analysis revealed that DAC induced CDKN1A/p21 expression, along with down-regulation of TERT. These molecular changes were also observed following doxorubicin treatment, supporting the importance of DAC-induced DNA damage in contributing to this effect. We demonstrated that knockdown of p21 led to TERT up-regulation. Strikingly, TERT overexpression increased DNMT1 levels and DAC sensitivity via a telomerase-independent mechanism. Furthermore, RNAi targeting of DNMT1 abrogated DAC response in TERT-proficient glioma cells.CONCLUSIONS: DAC down-regulates TERT through p21 induction. Our data points to TERT and DNMT1 levels as potential determinants of response to DAC treatment.

    View details for DOI 10.1093/neuonc/noaa207

    View details for PubMedID 32882013

  • Epigenetic Reprogramming for Targeting IDH-Mutant Malignant Gliomas. Cancers Park, J. W., Turcan, Ş. n. 2019; 11 (10)

    Abstract

    Targeting the epigenome has been considered a compelling treatment modality for several cancers, including gliomas. Nearly 80% of the lower-grade gliomas and secondary glioblastomas harbor recurrent mutations in isocitrate dehydrogenase (IDH). Mutant IDH generates high levels of 2-hydroxyglutarate (2-HG) that inhibit various components of the epigenetic machinery, including histone and DNA demethylases. The encouraging results from current epigenetic therapies in hematological malignancies have reinvigorated the interest in solid tumors and gliomas, both preclinically and clinically. Here, we summarize the recent advancements in epigenetic therapy for lower-grade gliomas and discuss the challenges associated with current treatment options. A particular focus is placed on therapeutic mechanisms underlying favorable outcome with epigenetic-based drugs in basic and translational research of gliomas. This review also highlights emerging bridges to combination treatment with respect to epigenetic drugs. Given that epigenetic therapies, particularly DNA methylation inhibitors, increase tumor immunogenicity and antitumor immune responses, appropriate drug combinations with immune checkpoint inhibitors may lead to improvement of treatment effectiveness of immunotherapy, ultimately leading to tumor cell eradication.

    View details for DOI 10.3390/cancers11101616

    View details for PubMedID 31652645

    View details for PubMedCentralID PMC6826741

  • Single-nucleus chromatin accessibility reveals intratumoral epigenetic heterogeneity in IDH1 mutant gliomas. Acta neuropathologica communications Al-Ali, R. n., Bauer, K. n., Park, J. W., Al Abdulla, R. n., Fermi, V. n., von Deimling, A. n., Herold-Mende, C. n., Mallm, J. P., Herrmann, C. n., Wick, W. n., Turcan, Ş. n. 2019; 7 (1): 201

    Abstract

    The presence of genome-wide DNA hypermethylation is a hallmark of lower grade gliomas (LGG) with isocitrate dehydrogenase (IDH) mutations. Further molecular classification of IDH mutant gliomas is defined by the presence (IDHmut-codel) or absence (IDHmut-noncodel) of hemizygous codeletion of chromosome arms 1p and 19q. Despite the DNA hypermethylation seen in bulk tumors, intra-tumoral heterogeneity at the epigenetic level has not been thoroughly analyzed. To address this question, we performed the first epigenetic profiling of single cells in a cohort of 5 gliomas with IDH1 mutation using single nucleus Assay for Transposase-Accessible Chromatin with high-throughput sequencing (snATAC-seq). Using the Fluidigm HT IFC microfluidics platform, we generated chromatin accessibility maps from 336 individual nuclei, and identified variable promoter accessibility of non-coding RNAs in LGGs. Interestingly, local chromatin structures of several non-coding RNAs are significant factors that contribute to heterogeneity, and show increased promoter accessibility in IDHmut-noncodel samples. As an example for clinical significance of this result, we identify CYTOR as a poor prognosis factor in gliomas with IDH mutation. Open chromatin assay points to differential accessibility of non-coding RNAs as an important source of epigenetic heterogeneity within individual tumors and between molecular subgroups. Rare populations of nuclei that resemble either IDH mutant molecular group co-exist within IDHmut-noncodel and IDHmut-codel groups, and along with non-coding RNAs may be an important issue to consider for future studies, as they may help guide predict treatment response and relapse.A web-based explorer for the data is available at shiny.turcanlab.org.

    View details for DOI 10.1186/s40478-019-0851-y

    View details for PubMedID 31806013

    View details for PubMedCentralID PMC6896263

  • Subcellular Localization of Sprouty2 in Human Glioma Cells. Frontiers in molecular neuroscience Hausott, B. n., Park, J. W., Valovka, T. n., Offterdinger, M. n., Hess, M. W., Geley, S. n., Klimaschewski, L. n. 2019; 12: 73

    Abstract

    Sprouty proteins act ubiquitously as signaling integrators and inhibitors of receptor tyrosine kinase (RTK) activated pathways. Among the four Sprouty isoforms, Sprouty2 is a key regulator of growth factor signaling in several neurological disorders. High protein levels correlate with reduced survival of glioma patients. We recently demonstrated that abrogating its function inhibits tumor growth by overstimulation of ERK and induction of DNA replication stress. The important role of Sprouty2 in the proliferation of malignant glioma cells prompted us to investigate its subcellular localization applying super-resolution fluorescence and immunoelectron microscopy. We found that cytoplasmic Sprouty2 is not homogenously distributed but localized to small spots (<100 nm) partly attached to vimentin filaments and co-localized with activated ERK. The protein is associated with early, late and recycling endosomes in response to but also independently of growth factor stimulation. The subcellular localization of Sprouty2 in all areas exhibiting strong RTK activities may reflect a protective response of glioma cells to limit excessive ERK activation and to prevent cellular senescence and apoptosis.

    View details for DOI 10.3389/fnmol.2019.00073

    View details for PubMedID 30983969

    View details for PubMedCentralID PMC6449699

  • Sprouty2 enhances the tumorigenic potential of glioblastoma cells. Neuro-oncology Park, J. W., Wollmann, G. n., Urbiola, C. n., Fogli, B. n., Florio, T. n., Geley, S. n., Klimaschewski, L. n. 2018; 20 (8): 1044–54

    Abstract

    Sprouty2 (SPRY2), a feedback regulator of receptor tyrosine kinase (RTK) signaling, has been shown to be associated with drug resistance and cell proliferation in glioblastoma (GBM), but the underlying mechanisms are still poorly defined.SPRY2 expression and survival patterns of patients with gliomas were analyzed using publicly available databases. Effects of RNA interference targeting SPRY2 on cellular proliferation in established GBM or patient-derived GBM stemlike cells were examined. Loss- or gain-of-function of SPRY2 to regulate the tumorigenic capacity was assessed in both intracranial and subcutaneous xenografts.SPRY2 was found to be upregulated in GBM, which correlated with reduced survival in GBM patients. SPRY2 knockdown significantly impaired proliferation of GBM cells but not of normal astrocytes. Silencing of SPRY2 increased epidermal growth factor-induced extracellular signal-regulated kinase (ERK) and Akt activation causing premature onset of DNA replication, increased DNA damage, and impaired proliferation, suggesting that SPRY2 suppresses DNA replication stress. Abrogating SPRY2 function strongly inhibited intracranial tumor growth and led to significantly prolonged survival of U87 xenograft-bearing mice. In contrast, SPRY2 overexpression promoted tumor propagation of low-tumorigenic U251 cells.The present study highlights an antitumoral effect of SPRY2 inhibition that is based on excessive activation of ERK signaling and DNA damage response, resulting in reduced cell proliferation and increased cytotoxicity, proposing SPRY2 as a promising pharmacological target in GBM patients.

    View details for DOI 10.1093/neuonc/noy028

    View details for PubMedID 29635363

    View details for PubMedCentralID PMC6280149

  • Brain cancer stem-like cell genesis from p53-deficient mouse astrocytes by oncogenic Ras BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Lee, J., Gil, J., Kim, J., Kim, T., Jin, X., Oh, S., Sohn, Y., Jeon, H., Park, H., Park, J., Shin, Y., Chung, Y. G., Lee, J., You, S., Kim, H. 2008; 365 (3): 496-502

    Abstract

    Here, we show that H-ras(V12) causes the p53-knockout mouse astrocytes (p53-/- astrocytes) to be transformed into brain cancer stem-like cells. H-ras(V12) triggers the p53-/- astrocytes to express a Nestin and a Cd133, which are expressed in normal and cancer neural stem cells. H-ras(V12) also induces the formation of a single cell-derived neurosphere under neural stem cell culture conditions. Furthermore, H-ras(V12)-overexpressing p53-/- astrocytes (p53-/-ast-H-ras(V12)) possess an in vitro self-renewal capacity, and are aberrantly differentiated into Tuj1-positve neurons both in vitro and in vivo. Amongst a variety of Ras-mediated canonical signaling pathways, we demonstrated that the MEK/ERK signaling pathway is responsible for neurosphere formation in p53-deficient astrocytes, whereas the PI3K/AKT signaling pathway is involved in oncogenic transformation in these cells. These findings suggest that the activation of Ras signaling pathways promotes the generation of brain cancer stem-like cells from p53-deficient mouse astrocytes by changing cell fate and transforming cell properties.

    View details for DOI 10.1016/j.bbrc.2007.11.005

    View details for Web of Science ID 000251663100016

    View details for PubMedID 18021740

  • Selective cell death of oncogenic Akt-transduced brain cancer cells by etoposide through reactive oxygen species-mediated damage MOLECULAR CANCER THERAPEUTICS Oh, S., Sohn, Y., Park, J., Park, H., Jeon, H., Kim, T., Lee, J., Jung, J., Jin, X., Chung, Y. G., Choi, Y., You, S., Lee, J., Kim, H. 2007; 6 (8): 2178-2187

    Abstract

    We have established several glioma-relevant oncogene-engineered cancer cells to reevaluate the oncogene-selective cytotoxicity of previously well-characterized anticancer drugs, such as etoposide, doxorubicin, staurosporine, and carmustine. Among several glioma-relevant oncogenes (activated epidermal growth factor receptor, Ras, and Akt, as well as Bcl-2 and p53DD used in the present study), the activated epidermal growth factor receptor, Ras, and Akt exerted oncogenic transformation of Ink4a/Arf(-/-) murine astrocyte cells. We identified that etoposide, a topoisomerase II inhibitor, caused selective killing of myristylated Akt (Akt-myr)-transduced Ink4a/Arf(-/-) astrocytes and U87MG cells in a dose- and time-dependent manner. Etoposide-selective cytotoxicity in the Akt-myr-transduced cells was shown to be caused by nonapoptotic cell death and occurred in a p53-independent manner. Etoposide caused severe reactive oxygen species (ROS) accumulation preferentially in the Akt-myr-transduced cells, and elevated ROS rendered these cells highly sensitive to cell death. The etoposide-selective cell death of Akt-myr-transduced cells was attenuated by pepstatin A, a lysosomal protease inhibitor. In the present study, we show that etoposide might possess a novel therapeutic activity for oncogenic Akt-transduced cancer cells to kill preferentially through ROS-mediated damage.

    View details for Web of Science ID 000248663000005

    View details for PubMedID 17699715