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Glioblastoma (GBM) still presents as one of the most aggressive tumours in the brain, which despite enormous research efforts, remains incurable today. As many theories evolve around the persistent recurrence of this malignancy, the assumption of a small population of cells with a stem-like phenotype remains a key driver of its infiltrative nature. In this article, we research Chordin-like 1 (CHRDL1), a secreted protein, as a potential key regulator of the glioma stem-like cell (GSC) phenotype. It has been shown that CHRDL1 antagonizes the function of bone morphogenic protein 4 (BMP4), which induces GSC differentiation and, hence, reduces tumorigenicity. We, therefore, employed two previously described GSCs spheroid cultures and depleted them of CHRDL1 using the stable transduction of a CHRDL1-targeting shRNA. We show with in vitro cell-based assays (MTT, limiting dilution, and sphere formation assays), Western blots, irradiation procedures, and quantitative real-time PCR that the depletion of the secreted BMP4 antagonist CHRDL1 prominently decreases functional and molecular stemness traits resulting in enhanced radiation sensitivity. As a result, we postulate CHRDL1 as an enforcer of stemness in GSCs and find additional evidence that high CHRDL1 expression might also serve as a marker protein to determine BMP4 susceptibility.
Glioblastoma (GBM) is the most common and most aggressive primary brain tumor, with a very high rate of recurrence and a median survival of 15 months after diagnosis. Abundant evidence suggests that a certain sub-population of cancer cells harbors a stem-like phenotype and is likely responsible for disease recurrence, treatment resistance and potentially even for the infiltrative growth of GBM. GBM incidence has been negatively correlated with the serum levels of 25-hydroxy-vitamin D3, while the low pH within tumors has been shown to promote the expression of the vitamin D3-degrading enzyme 24-hydroxylase, encoded by the CYP24A1 gene. Therefore, we hypothesized that calcitriol can specifically target stem-like glioblastoma cells and induce their differentiation. Here, we show, using in vitro limiting dilution assays, quantitative real-time PCR, quantitative proteomics and ex vivo adult organotypic brain slice transplantation cultures, that therapeutic doses of calcitriol, the hormonally active form of vitamin D3, reduce stemness to varying extents in a panel of investigated GSC lines, and that it effectively hinders tumor growth of responding GSCs ex vivo. We further show that calcitriol synergizes with Temozolomide ex vivo to completely eliminate some GSC tumors. These findings indicate that calcitriol carries potential as an adjuvant therapy for a subgroup of GBM patients and should be analyzed in more detail in follow-up studies.
Background: Dexamethasone (Dex) is the most common corticosteroid to treat edema in glioblastoma (GBM) patients. Recent studies identified the addition of Dex to radiation therapy (RT) to be associated with poor survival. Independently, Tumor Treating Fields (TTFields) provides a novel anti-cancer modality for patients with primary and recurrent GBM. Whether Dex influences the efficacy of TTFields, however, remains elusive.
Methods: Human GBM cell lines MZ54 and U251 were treated with RT or TTFields in combination with Dex and the effects on cell counts and cell death were determined via flow cytometry. We further performed a retrospective analysis of GBM patients with TTFields treatment +/- concomitant Dex and analysed its impact on progression-free (PFS) and overall survival (OS).
Results: The addition of Dex significantly reduced the efficacy of RT in U251 and MZ54 cells. TTFields (200 kHz/250 kHz) induced massive cell death in both cell lines. Concomitant treatment of TTFields and Dex did not reduce the overall efficacy of TTFields. Further, in our retrospective clinical analysis, we found that the addition of Dex to TTFields therapy did not influence PFS nor OS.
Conclusion: Our translational investigation indicates that the efficacy of TTFields therapy in patients with GBM and primary GBM cell lines is not affected by the addition of Dex.
Glioblastoma is a very aggressive tumor and represents the most common primary brain malignancy. Key characteristics include its high resistance against conventional treatments, such as radio- and chemotherapy and its diffuse tissue infiltration, preventing complete surgical resection. The analysis of migration and invasion processes in a physiological microenvironment allows for enhanced understanding of these processes and can lead to improved therapeutic approaches. Here, we combine two state-of-the-art techniques, adult organotypic brain tissue slice culture (OTC) and light sheet fluorescence microscopy (LSFM) of cleared tissues in a combined method termed OTCxLSFM. Using this methodology, we can show that glioblastoma tissue infiltration can be effectively blocked through treatment with arsenic trioxide, as well as genetic depletion of the tetraspanin, transmembrane receptor CD9. With our analysis-pipeline we gain single-cell level, three-dimensional information, as well as insights into the morphological appearance of the tumor cells.
Glioblastoma (GBM) is a devastating disease and the most common primary brain malignancy of adults with a median survival barely exceeding one year. Recent findings suggest that the antipsychotic drug pimozide triggers an autophagy-dependent, lysosomal type of cell death in GBM cells with possible implications for GBM therapy. One oncoprotein that is often overactivated in these tumors and associated with a particularly dismal prognosis is Signal Transducer and Activator of Transcription 3 (STAT3). Here, we used isogenic human and murine GBM knockout cell lines, advanced fluorescence microscopy, transcriptomic analysis and FACS-based assessment of cell viability to show that STAT3 has an underappreciated, context-dependent role in drug-induced cell death. Specifically, we demonstrate that depletion of STAT3 significantly enhances cell survival after treatment with Pimozide, suggesting that STAT3 confers a particular vulnerability to GBM. Furthermore, we show that active STAT3 has no major influence on the early steps of the autophagy pathway, but exacerbates drug-induced lysosomal membrane permeabilization (LMP) and release of cathepsins into the cytosol. Collectively, our findings support the concept of exploiting the pro-death functions of autophagy and LMP for GBM therapy and to further determine whether STAT3 can be employed as a treatment predictor for highly apoptosis-resistant, but autophagy-proficient cancers.