Novel analogs of sulfasalazine as system xc− antiporter inhibitors: Insights from the molecular modeling studies

System x_c⁻ (Sx_c⁻), a cystine-glutamate antiporter, is established as an interesting target for the treatment of several pathologies including epileptic seizures, glioma, neurodegenerative diseases, and multiple sclerosis. Erastin, sorafenib, and sulfasalazine (SSZ) are a few of the established inhibitors of Sx_c⁻. However, its pharmacological inhibition with novel and potent agents is still very much required due to potential issues, for example, potency, bioavailability, and blood–brain barrier (BBB) permeability, with the current lead molecules such as SSZ. Therefore, in this study, we report the synthesis and structure–activity relationships (SAR) of SSZ derivatives along with molecular docking and dynamics simulations using the developed homology model of xCT chain of Sx_c⁻ antiporter. The generated homology model attempted to address the limitations of previously reported comparative protein models, thereby increasing the confidence in the computational modeling studies. The main objective of the present study was to derive a suitable lead structure from SSZ eliminating its potential issues for the treatment of glioblastoma multiforme (GBM), a deadly and malignant grade IV astrocytoma. The designed compounds with favorable Sx_c⁻ inhibitory activity following in vitro Sx_c⁻ inhibition studies, showed moderately potent cytotoxicity in patient-derived human glioblastoma cells, thereby generating potential interest in these compounds. The xCT-ligand model can be further optimized in search of potent lead molecules for novel drug discovery and development studies.     

Type 1 collagen as a potential niche component for CD133‐positive glioblastoma cells

Cancer stem cells are thought to be closely related to tumor progression and recurrence, making them attractive therapeutic targets. Stem cells of various tissues exist within niches maintaining their stemness. Glioblastoma stem cells (GSCs) are located at tumor capillaries and the perivascular niche, which are considered to have an important role in maintaining GSCs. There were some extracellular matrices (ECM) on the perivascular connective tissue, including type 1 collagen. We here evaluated whether type 1 collagen has a potential niche for GSCs. Imunohistochemical staining of type 1 collagen and CD133, one of the GSCs markers, on glioblastoma (GBM) tissues showed CD133‐positive cells were located in immediate proximity to type 1 collagen around tumor vessels. We cultured human GBM cell lines, U87MG and GBM cells obtained from fresh surgical tissues, T472 and T555, with serum‐containing medium (SCM) or serum‐free medium with some growth factors (SFM) and in non‐coated (Non‐coat) or type 1 collagen‐coated plates (Col). The RNA expression levels of CD133 and Nestin as stem cell markers in each condition were examined. The Col condition not only with SFM but SCM made GBM cells more enhanced in RNA expression of CD133, compared to Non‐coat/SCM. Semi‐quantitative measurement of CD133‐positive cells by immunocytochemistry showed a statistically significant increase of CD133‐positive cells in Col/SFM. In addition, T472 cell line cultured in the Col/SFM had capabilities of sphere formation and tumorigenesis. Type 1 collagen was found in the perivascular area and showed a possibility to maintain GSCs. These findings suggest that type 1 collagen could be one important niche component for CD133‐positive GSCs and maintain GSCs in adherent culture.     

Regional and Voxel‐Wise Comparisons of Blood Flow Measurements Between Dynamic Susceptibility Contrast Magnetic Resonance Imaging (DSC‐MRI) and Arterial Spin Labeling (ASL) in Brain Tumors

The objective of the current study was to evaluate the regional and voxel‐wise correlation between dynamic susceptibility contrast (DSC) and arterial spin labeling (ASL) perfusion magnetic resonance imaging (MRI) measurement of cerebral blood flow (CBF) in patients with brain tumors. Thirty patients with histologically verified brain tumors were evaluated in the current study. DSC‐MRI was performed by first using a preload dose of gadolinium contrast, then collecting a dynamic image acquisition during a bolus of contrast, followed by posthoc contrast agent leakage correction. Pseudocontinuous ASL was collected using 30 pairs of tag and control acquisition using a 3‐dimensional gradient‐echo spin‐echo (GRASE) acquisition. All images were registered to a high‐resolution anatomical atlas. Average CBF measurements within regions of contrast‐enhancement and T2 hyperintensity were evaluated between the two modalities. Additionally, voxel‐wise correlation between CBF measurements obtained with DSC and ASL were assessed. Results demonstrated a positive linear correlation between DSC and ASL measurements of CBF when regional average values were compared; however, a statistically significant voxel‐wise correlation was only observed in around 30‐40% of patients. These results suggest DSC and ASL may provide regionally similar, but spatially different measurements of CBF.     

Inhibition of eukaryotic initiation factor 4E by tomivosertib suppresses angiogenesis,growth, and survival of glioblastoma and enhances chemotherapy's efficacy

Glioblastoma is characterized by extensive vascularization and is highly resistant to current therapy. Identification of drugs that target tumor directly and angiogenesis processes present an effective therapeutic strategy for glioblastoma. Mnk kinase is required for the activation of eukaryotic initiation factor 4E (eIF4E), which mediates translation of oncogenic proteins. We investigated the effects of tomivosertib, a novel MAPK-interacting kinase (MNK) inhibitor, on glioblastoma angiogenesis, growth, and survival. We found that tomivosertib inhibited growth and induced caspase-dependent apoptosis in various glioblastoma cell lines. Tomivosertib disrupted glioblastoma endothelial cell capillary network formation, growth, and survival. Mechanistically, tomivosertib acted on glioblastoma via suppressing MNK-dependent eIF4E phosphorylation and activation in tumor and endothelial cells. We further found that temozolomide activated eIF4E and this was reversed by tomivosertib. Using glioblastoma xenograft mouse model, we demonstrated that temozolomide and tomivosertib combination had higher efficacy than tomivosertib alone. Of note, tomivosertib inhibited glioblastoma angiogenesis and decreased p-eIF4E level in mice. We finally showed that p-eIF4E activation was a common molecular feature in glioblastoma patients. Our pre-clinical findings suggest that tomivosertib is a useful addition to the treatment armamentarium for glioblastoma and that targeting MNK-eIF4E pathway represents a therapeutic strategy to overcome glioblastoma chemoresistance.     

High linear-energy-transfer radiation can overcome radioresistance of glioma stem-like cells to low linear-energy-transfer radiation

Ionizing radiation is applied as the standard treatment for glioblastoma multiforme (GBM). However, radiotherapy remains merely palliative, not curative, because of the existence of glioma stem cells (GSCs), which are regarded as highly radioresistant to low linear-energy-transfer (LET) photons. Here we analyzed whether or not high-LET particles can overcome the radioresistance of GSCs. Glioma stem-like cells (GSLCs) were induced from the GBM cell line A172 in stem cell culture medium. The phenotypes of GSLCs and wild-type cells were confirmed using stem cell markers. These cells were irradiated with ⁶⁰Co gamma rays or reactor neutron beams. Under neutron-beam irradiation, high-LET proton particles can be produced through elastic scattering or nitrogen capture reaction. Radiosensitivity was assessed by a colony-forming assay, and the DNA double-strand breaks (DSBs) were assessed by a histone gamma-H2AX focus detection assay. In stem cell culture medium, GSLCs could form neurosphere-like cells and express neural stem cell markers (Sox2 and Musashi) abundantly in comparison with their parental cells. GSLCs were significantly more radioresistant to gamma rays than their parental cells, but neutron beams overcame this resistance. There were significantly fewer gamma-H2AX foci in the A172 GSLCs 24 h after irradiation with gamma rays than in their parental cultured cells, while there was no apparent difference following neutron-beam irradiation. High-LET radiation can overcome the radioresistance of GSLCs by producing unrepairable DNA DSBs. High-LET radiation therapy might have the potential to overcome GBM''s resistance to X-rays in a clinical setting.     

Phospholipase D1 inhibition sensitizes glioblastoma to temozolomide and suppresses its tumorigenicity

Resistance of glioblastoma to the chemotherapeutic compound temozolomide is associated with the presence of glioblastoma stem cells in glioblastoma and is a key obstacle for the poor prognosis of glioblastoma. Here, we show that phospholipase D1 is elevated in CD44^High glioblastoma stem cells and in glioblastoma, especially recurring glioblastoma. Phospholipase D1 elevation positively correlated with the level of CD44 and poor prognosis in glioblastoma patients. Temozolomide significantly upregulated the expression of phospholipase D1 in the low and moderate CD44 populations of glioblastoma stem cells, but not in the CD44^High population in which phospholipase D1 is highly expressed. Phospholipase D1 conferred resistance to temozolomide in CD44^High glioblastoma stem cells and increased their self-renewal capacity and maintenance. Phospholipase D1 expression significantly correlated with levels of temozolomide resistance factors, which were suppressed by microRNA-320a and -4496 induced by phospholipase D1 inhibition. Genetic and pharmacological targeting of phospholipase D1 attenuated glioblastoma stem cell-derived intracranial tumors of glioblastoma using the microRNAs, and improved survival. Treatment solely with temozolomide produced no benefits on the glioblastoma, whereas in combination, phospholipase D1 inhibition sensitized glioblastoma stem cells to temozolomide and reduced glioblastoma tumorigenesis. Together, these findings indicate that phospholipase D1 inhibition might overcome resistance to temozolomide and represents a potential treatment strategy for glioblastoma. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.     

Angiosarcomatous component in gliosarcoma: case report and consideration of diagnostic challenge and hemorrhagic propensity

An angiosarcomatous component in gliosarcoma may be associated with an increased intraoperative hemorrhagic risk and preoperative diagnostic challenge. We report a unique case of gliosarcoma with an angiosarcomatous component in a 61-year-old man. His brain MRI demonstrated a well-demarcated right occipital tumor with multiple flow voids and rim-like enhancement as well as intratumoral strip and nodular enhancements. He underwent a craniotomy for tumor resection. Intraoperatively, significant tumor hemorrhage required greater efforts to control intraoperative bleeding and to maintain hemostasis. Pathological examination of the tumor revealed alternating gliomatous and sarcomatous/angiosarcomatous components with intratumoral hemorrhage. He was postoperatively treated with chemoradiation. The tumor recurred at 9 months, for which the second resection was performed with similarly greater efforts to achieve hemostasis. The recurrent tumor was pathologically similar despite treatment-associated changes. Awareness of this angiosarcomatous component in gliosarcoma with the hemorrhagic risk is important for both the preoperative diagnosis and surgical management.