The hypoxic peri-arteriolar glioma stem cell niche, an integrated concept of five types of niches in human glioblastoma.

Glioblastoma is the most lethal primary brain tumor and poor survival of glioblastoma patients is attributed to the presence of glioma stem cells (GSCs). These therapy-resistant, quiescent and pluripotent cells reside in GSC niches, which are specific microenvironments that protect GSCs against radiotherapy and chemotherapy. We previously showed the existence of hypoxic peri-arteriolar GSC niches in glioblastoma tumor samples. However, other studies have described peri-vascular niches, peri-hypoxic niches, peri-immune niches and extracellular matrix niches of GSCs. The aim of this review was to critically evaluate the literature on these five different types of GSC niches. In the present review, we describe that the five niche types are not distinct from one another, but should be considered to be parts of one integral GSC niche model, the hypoxic peri-arteriolar GSC niche. Moreover, hypoxic peri-arteriolar GSC niches are structural and functional look-alikes of hematopoietic stem cell (HSC) niches in the bone marrow. GSCs are maintained in peri-arteriolar niches by the same receptor-ligand interactions as HSCs in bone marrow. Our concept should be rigidly tested in the near future and applied to develop therapies to expel and keep GSCs out of their protective niches to render them more vulnerable to standard therapies.

Isocitrate dehydrogenase 1-mutated human gliomas depend on lactate and glutamate to alleviate metabolic stress.

Diffuse gliomas often carry point mutations in isocitrate dehydrogenase ( IDH1mut), resulting in metabolic stress. Although IDHmut gliomas are difficult to culture in vitro, they thrive in the brain via diffuse infiltration, suggesting brain-specific tumor-stroma interactions that can compensate for IDH-1 deficits. To elucidate the metabolic adjustments in clinical IDHmut gliomas that contribute to their malignancy, we applied a recently developed method of targeted quantitative RNA next-generation sequencing to 66 clinical gliomas and relevant orthotopic glioma xenografts, with and without the endogenous IDH-1R132H mutation. Datasets were analyzed in R using Manhattan plots to calculate distance between expression profiles, Ward's method to perform unsupervised agglomerative clustering, and the Mann Whitney U test and Fisher's exact tests for supervised group analyses. The significance of transcriptome data was investigated by protein analysis, in situ enzymatic activity mapping, and in vivo magnetic resonance spectroscopy of orthotopic IDH1mut- and IDHwt-glioma xenografts. Gene set enrichment analyses of clinical IDH1mut gliomas strongly suggest a role for catabolism of lactate and the neurotransmitter glutamate, whereas, in IDHwt gliomas, processing of glucose and glutamine are the predominant metabolic pathways. Further evidence of the differential metabolic activity in these cancers comes from in situ enzymatic mapping studies and preclinical in vivo magnetic resonance spectroscopy imaging. Our data support an evolutionary model in which IDHmut glioma cells exist in symbiosis with supportive neuronal cells and astrocytes as suppliers of glutamate and lactate, possibly explaining the diffuse nature of these cancers. The dependency on glutamate and lactate opens the way for novel approaches in the treatment of IDHmut gliomas.-Lenting, K., Khurshed, M., Peeters, T. H., van den Heuvel, C. N. A. M., van Lith, S. A. M., de Bitter, T., Hendriks, W., Span, P. N., Molenaar, R. J., Botman, D., Verrijp, K., Heerschap, A., ter Laan, M., Kusters, B., van Ewijk, A., Huynen, M. A., van Noorden, C. J. F., Leenders, W. P. J. Isocitrate dehydrogenase 1-mutated human gliomas depend on lactate and glutamate to alleviate metabolic stress.

Novel therapeutic strategies to target leukemic cells that hijack compartmentalized continuous hematopoietic stem cell niches.

Acute myeloid leukemia and acute lymphoblastic leukemia cells hijack hematopoietic stem cell (HSC) niches in the bone marrow and become leukemic stem cells (LSCs) at the expense of normal HSCs. LSCs are quiescent and resistant to chemotherapy and can cause relapse of the disease. HSCs in niches are needed to generate blood cell precursors that are committed to unilineage differentiation and eventually production of mature blood cells, including red blood cells, megakaryocytes, myeloid cells and lymphocytes. Thus far, three types of HSC niches are recognized: endosteal, reticular and perivascular niches. However, we argue here that there is only one type of HSC niche, which consists of a periarteriolar compartment and a perisinusoidal compartment. In the periarteriolar compartment, hypoxia and low levels of reactive oxygen species preserve the HSC pool. In the perisinusoidal compartment, hypoxia in combination with higher levels of reactive oxygen species enables proliferation of progenitor cells and their mobilization into the circulation. Because HSC niches offer protection to LSCs against chemotherapy, we review novel therapeutic strategies to inhibit homing of LSCs in niches for the prevention of dedifferentiation of leukemic cells into LSCs and to stimulate migration of leukemic cells out of niches. These strategies enhance differentiation and proliferation and thus sensitize leukemic cells to chemotherapy. Finally, we list clinical trials of therapies that tackle LSCs in HSC niches to circumvent their protection against chemotherapy.

Study protocol of a phase II clinical trial of oral metformin for the intravesical treatment of non-muscle invasive bladder cancer.

BACKGROUND: Non-muscle-invasive bladder cancer (NMIBC) is the most common neoplasm of the urinary tract and requires life-long invasive surveillance to detect disease recurrence. Currently, there are no effective oral therapies that delay disease recurrence or progression. We recently demonstrated that in mice, metformin accumulates unchanged in the urine. Urothelial cells are exposed to metformin concentrations ~ 240-fold higher than in serum. This was effective in the treatment of mouse bladder cancer models. METHODS: We describe the protocol of a multi-centre, open-label, phase II clinical trial of metformin in up to 49 evaluable patients with intermediate-risk NMIBC with the aim to determine the overall response to administration of oral metformin for 3 months on a marker tumour deliberately left following transurethral resection of multiple, papillary NMIBC tumours. All patients will receive metformin orally at doses up to 3000 mg per day. Metformin treatment will start within 2 weeks following transurethral resection of all tumours except one marker lesion. After 3 months of metformin treatment, the effect of metformin on the marker lesion is evaluated by cystoscopy and biopsy under anaesthesia. Residual tumour, if present at this evaluation, will be resected. In case of complete disappearance of the marker lesion, the former tumour area will be biopsied. The primary outcome is the complete response rate of the marker lesion, as determined by decentralised scoring of pre- and post-treatment cystoscopy images by expert independent urologists. Secondary outcomes are the partial response rate, overall safety of metformin and the duration of the time to recurrence. DISCUSSION: Preclinical studies show the potential role of oral metformin treatment in the management of NMIBC. It could offer an alternative to current adjuvant intravesical treatment. If positive, the reported results of this study could warrant further phase III trials to compare the efficacy of metformin against current treatments of intravesical installations with chemotherapy or Bacillus Calmette-Guérin (BCG). TRIAL REGISTRATION: This trial is registered in ClinicalTrials.gov under NCT03379909.

IDH1-mutant cancer cells are sensitive to cisplatin and an IDH1-mutant inhibitor counteracts this sensitivity.

Isocitrate dehydrogenase ( IDH1)-1 is mutated in various types of human cancer, and the presence of this mutation is associated with improved responses to irradiation and chemotherapy in solid tumor cells. Mutated IDH1 (IDH1MUT) enzymes consume NADPH to produce d-2-hydroxyglutarate (d-2HG) resulting in the decreased reducing power needed for detoxification of reactive oxygen species (ROS), for example. The objective of the current study was to investigate the mechanism behind the chemosensitivity of the widely used anticancer agent cisplatin in IDH1MUT cancer cells. Oxidative stress, DNA damage, and mitochondrial dysfunction caused by cisplatin treatment were monitored in IDH1MUT HCT116 colorectal cancer cells and U251 glioma cells. We found that exposure to cisplatin induced higher levels of ROS, DNA double-strand breaks (DSBs), and cell death in IDH1MUT cancer cells, as compared with IDH1 wild-type ( IDH1WT) cells. Mechanistic investigations revealed that cisplatin treatment dose dependently reduced oxidative respiration in IDH1MUT cells, which was accompanied by disturbed mitochondrial proteostasis, indicative of impaired mitochondrial activity. These effects were abolished by the IDH1MUT inhibitor AGI-5198 and were restored by treatment with d-2HG. Thus, our study shows that altered oxidative stress responses and a vulnerable oxidative metabolism underlie the sensitivity of IDH1MUT cancer cells to cisplatin.-Khurshed, M., Aarnoudse, N., Hulsbos, R., Hira, V. V. V., van Laarhoven, H. W. M., Wilmink, J. W., Molenaar, R. J., van Noorden, C. J. F. IDH1-mutant cancer cells are sensitive to cisplatin and an IDH1-mutant inhibitor counteracts this sensitivity.

Targeting glutaminolysis in chondrosarcoma in context of the IDH1/2 mutation.

INTRODUCTION: Chondrosarcoma is a malignant cartilage-forming bone tumour in which mutations in IDH1 and IDH2 frequently occur. Previous studies suggest an increased dependency on glutaminolysis in IDH1/2 mutant cells, which resulted in clinical trials with the drugs CB-839, metformin and chloroquine. In this study, the preclinical rationale for using these drugs as a treatment for chondrosarcoma was evaluated. METHODS: Expression of glutaminase was determined in 120 cartilage tumours by immunohistochemistry. Ten chondrosarcoma cell lines were treated with the metabolic compounds CB-849, metformin, phenformin (lipophilic analogue of metformin) and chloroquine. RESULTS: A difference in glutaminase expression levels between the different tumour grades (p = 0.001, one-way ANOVA) was identified, with the highest expression observed in high-grade chondrosarcomas. Treatment with CB-839, metformin, phenformin or chloroquine revealed that chondrosarcoma cell lines are sensitive to glutaminolysis inhibition. Metformin and phenformin decreased mTOR activity in chondrosarcoma cells, and metformin decreased LC3B-II levels, which is counteracted by chloroquine. CONCLUSION: Targeting glutaminolysis with CB-839, metformin, phenformin or chloroquine is a potential therapeutic strategy for a subset of high-grade chondrosarcomas, irrespective of the presence or absence of an IDH1/2 mutation.

A phase Ib study of everolimus combined with metformin for patients with advanced cancer.

Background The efficacy to monotherapy with the mTOR inhibitor everolimus in advanced cancer is often limited due to therapy resistance. Combining everolimus with metformin may decrease the chance of therapy resistance. Methods Patients received everolimus and metformin in a 3 + 3 dose-escalation scheme. Objectives were to determine the dose-limiting toxicities (DLTs), maximum tolerated dose, toxic effects, pharmacokinetics and anti-tumour efficacy. Results 9 patients received study treatment for a median duration of 48 days (range: 4-78). 6 patients discontinued due to toxicity and 3 patients because of progressive disease. At the starting dose level of 10 mg everolimus qd and 500 mg metformin bid, 3 out of 5 patients experienced a DLT. After de-escalation to 5 mg everolimus qd and 500 mg metformin bid, considerable toxicity was still observed and patient enrollment was terminated. In pharmacokinetic analyses, metformin was eliminated slower when co-administered with everolimus than as single-agent. After 9 weeks of treatment, 3 patients were still on study and all had stable disease. Conclusion The combination of everolimus and metformin is poorly tolerated in patients with advanced cancer. The pharmacokinetic interaction between everolimus and metformin may have implications for diabetic cancer patients that are treated with these drugs. Our results advocate for future clinical trials with combinations of other mTOR inhibitors and biguanides.

Effects of the Green Tea Polyphenol Epigallocatechin-3-Gallate on Glioma: A Critical Evaluation of the Literature.

The review discusses the effects of Epigallocatechin-3-gallate Gallate (EGCG) on glioma as a basis for future research on clinical application of EGCG. Epidemiological studies on the effects of green tea or EGCG on the risk of glioma is inconclusive due to the limited number of studies, the inclusion of all tea types in these studies, and the focus on caffeine rather than EGCG. In vivo experiments using EGCG monotherapy are inconclusive. Nevertheless, EGCG induces cell death, prevents cellular proliferation, and limits invasion in multiple glioma cell lines. Furthermore, EGCG enhances the efficacy of anti-glioma therapies, including irradiation, temozolomide, carmustine, cisplatin, tamoxifen, and TNF-related apoptosis-inducing ligand, but reduces the effect of bortezomib. Pro-drugs, co-treatment, and encapsulation are being investigated to enhance clinical applicability of EGCG. Mechanisms of actions of EGCG have been partly elucidated. EGCG has both anti-oxidant and oxidant properties. EGCG inhibits pro-survival proteins, such as telomerase, survivin, GRP78, PEA15, and P-gp. EGCG inhibits signaling of PDGFR, IGF-1R, and 67LR. EGCG reduces invasiveness of cancer cells by inhibiting the activities of various metalloproteinases, cytokines, and chemokines. Last, EGCG inhibits some NADPH-producing enzymes, thus disturbing redox status and metabolism of glioma cells. In conclusion, EGCG may be a suitable adjuvant to potentiate anti-glioma therapies.

The driver and passenger effects of isocitrate dehydrogenase 1 and 2 mutations in oncogenesis and survival prolongation.

Mutations in isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) are key events in the development of glioma, acute myeloid leukemia (AML), chondrosarcoma, intrahepatic cholangiocarcinoma (ICC), and angioimmunoblastic T-cell lymphoma. They also cause D-2-hydroxyglutaric aciduria and Ollier and Maffucci syndromes. IDH1/2 mutations are associated with prolonged survival in glioma and in ICC, but not in AML. The reason for this is unknown. In their wild-type forms, IDH1 and IDH2 convert isocitrate and NADP(+) to α-ketoglutarate (αKG) and NADPH. Missense mutations in the active sites of these enzymes induce a neo-enzymatic reaction wherein NADPH reduces αKG to D-2-hydroxyglutarate (D-2HG). The resulting D-2HG accumulation leads to hypoxia-inducible factor 1α degradation, and changes in epigenetics and extracellular matrix homeostasis. Such mutations also imply less NADPH production capacity. Each of these effects could play a role in cancer formation. Here, we provide an overview of the literature and discuss which downstream molecular effects are likely to be the drivers of the oncogenic and survival-prolonging properties of IDH1/2 mutations. We discuss interactions between mutant IDH1/2 inhibitors and conventional therapies. Understanding of the biochemical consequences of IDH1/2 mutations in oncogenesis and survival prolongation will yield valuable information for rational therapy design: it will tell us which oncogenic processes should be blocked and which "survivalogenic" effects should be retained.

Mutational profiling of kinases in glioblastoma.

BACKGROUND: Glioblastoma is a highly malignant brain tumor for which no cure is available. To identify new therapeutic targets, we performed a mutation analysis of kinase genes in glioblastoma. METHODS: Database mining and a literature search identified 76 kinases that have been found to be mutated at least twice in multiple cancer types before. Among those we selected 34 kinase genes for mutation analysis. We also included IDH1, IDH2, PTEN, TP53 and NRAS, genes that are known to be mutated at considerable frequencies in glioblastoma. In total, 174 exons of 39 genes in 113 glioblastoma samples from 109 patients and 16 high-grade glioma (HGG) cell lines were sequenced. RESULTS: Our mutation analysis led to the identification of 148 non-synonymous somatic mutations, of which 25 have not been reported before in glioblastoma. Somatic mutations were found in TP53, PTEN, IDH1, PIK3CA, EGFR, BRAF, EPHA3, NRAS, TGFBR2, FLT3 and RPS6KC1. Mapping the mutated genes into known signaling pathways revealed that the large majority of them plays a central role in the PI3K-AKT pathway. CONCLUSIONS: The knowledge that at least 50% of glioblastoma tumors display mutational activation of the PI3K-AKT pathway should offer new opportunities for the rational development of therapeutic approaches for glioblastomas. However, due to the development of resistance mechanisms, kinase inhibition studies targeting the PI3K-AKT pathway for relapsing glioblastoma have mostly failed thus far. Other therapies should be investigated, targeting early events in gliomagenesis that involve both kinases and non-kinases.

Recent advances in the molecular understanding of glioblastoma.

Glioblastoma is the most common and most aggressive primary brain tumor. Despite maximum treatment, patients only have a median survival time of 15 months, because of the tumor's resistance to current therapeutic approaches. Thus far, methylation of the O (6)-methylguanine-DNA methyltransferase (MGMT) promoter has been the only confirmed molecular predictive factor in glioblastoma. Novel "genome-wide" techniques have identified additional important molecular alterations as mutations in isocitrate dehydrogenase 1 (IDH1) and its prognostic importance. This review summarizes findings and techniques of genetic, epigenetic, transcriptional, and proteomic studies of glioblastoma. It provides the clinician with an up-to-date overview of current identified molecular alterations that should ultimately lead to new therapeutic targets and more individualized treatment approaches in glioblastoma.

IDH1/2 Mutations in Cancer Stem Cells and Their Implications for Differentiation Therapy.

Isocitrate dehydrogenase 1 and 2 (IDH1/2) are enzymes recurrently mutated in various types of cancer, including glioma, cholangiocarcinoma, chondrosarcoma, and acute myeloid leukemia. Mutant IDH1/2 induce a block in differentiation and thereby contribute to the stemness and oncogenesis of their cells of origin. Recently, small-molecule inhibitors of mutant IDH1/2 have been Food and Drug Administration-approved for the treatment of IDH1/2-mutated acute myeloid leukemia. These inhibitors decrease the stemness of the targeted IDH1/2-mutated cancer cells and induce their differentiation to more mature cells. In this review, we elucidate the mechanisms by which mutant IDH1/2 induce a block in differentiation and the biological and clinical effects of the release into differentiation by mutant-IDH1/2 inhibitors. (J Histochem Cytochem 70:83-97, 2022).

Hyperthermia as a Potential Cornerstone of Effective Multimodality Treatment with Radiotherapy, Cisplatin and PARP Inhibitor in IDH1-Mutated Cancer Cells.

Mutations in the isocitrate dehydrogenase 1 (IDH1MUT) gene occur in various types of malignancies, including ~60% of chondrosarcomas, ~30% of intrahepatic cholangiocarcinomas and >80% of low-grade gliomas. IDH1MUT are causal in the development and progression of these types of cancer due to neomorphic production of the oncometabolite D-2-hydroxyglutarate (D-2HG). Intracellular accumulation of D-2HG has been implicated in suppressing homologous recombination and renders IDH1MUT cancer cells sensitive to DNA-repair-inhibiting agents, such as poly-(adenosine 5'-diphosphate−ribose) polymerase inhibitors (PARPi). Hyperthermia increases the efficacy of DNA-damaging therapies such as radiotherapy and platinum-based chemotherapy, mainly by inhibition of DNA repair. In the current study, we investigated the additional effects of hyperthermia (42 °C for 1 h) in the treatment of IDH1MUT HCT116 colon cancer cells and hyperthermia1080 chondrosarcoma cancer cells in combination with radiation, cisplatin and/or a PARPi on clonogenic cell survival, cell cycle distribution and the induction and repair of DNA double-strand breaks. We found that hyperthermia in combination with radiation or cisplatin induces an increase in double-strand breaks and cell death, up to 10-fold in IDH1MUT cancer cells compared to IDH1 wild-type cells. This vulnerability was abolished by the IDH1MUT inhibitor AGI-5198 and was further increased by the PARPi. In conclusion, our study shows that IDH1MUT cancer cells are sensitized to hyperthermia in combination with irradiation or cisplatin and a PARPi. Therefore, hyperthermia may be an efficacious sensitizer to cytotoxic therapies in tumors where the clinical application of hyperthermia is feasible, such as IDH1MUT chondrosarcoma of the extremities.

Cell Biology Meets Cell Metabolism: Energy Production Is Similar in Stem Cells and in Cancer Stem Cells in Brain and Bone Marrow.

Energy production by means of ATP synthesis in cancer cells has been investigated frequently as a potential therapeutic target in this century. Both (an)aerobic glycolysis and oxidative phosphorylation (OXPHOS) have been studied. Here, we review recent literature on energy production in glioblastoma stem cells (GSCs) and leukemic stem cells (LSCs) versus their normal counterparts, neural stem cells (NSCs) and hematopoietic stem cells (HSCs), respectively. These two cancer stem cell types were compared because their niches in glioblastoma tumors and in bone marrow are similar. In this study, it became apparent that (1) ATP is produced in NSCs and HSCs by anaerobic glycolysis, whereas fatty acid oxidation (FAO) is essential for their stem cell fate and (2) ATP is produced in GSCs and LSCs by OXPHOS despite the hypoxic conditions in their niches with FAO and amino acids providing its substrate. These metabolic processes appeared to be under tight control of cellular regulation mechanisms which are discussed in depth. However, our conclusion is that systemic therapeutic targeting of ATP production via glycolysis or OXPHOS is not an attractive option because of its unwanted side effects in cancer patients.

Facilitatory and inhibitory effects of SCN5A mutations on atrial fibrillation in Brugada syndrome.

AIMS: Brugada syndrome (BrS) is associated with increased risk for atrial fibrillation (AFib). However, the role of SCN5A mutations in the occurrence of AFib remains unclear. Cardiac sodium current reduction caused by SCN5A mutations may facilitate AFib by slowing intra-atrial conduction and inducing structural changes, but also prevent it by suppressing atrial ectopic activity. Here, we examined the relation between SCN5A mutations, atrial conduction velocity, atrial structural changes, and atrial ectopic activity in BrS. METHODS AND RESULTS: Data from 214 BrS patients [78 with an SCN5A mutation (patients with an SCN5A mutation, BrSSCN5A+) and 136 without an SCN5A mutation (patients without an SCN5A mutation, BrSSCN5A-)] were collected. Intra-atrial conduction velocity was assessed by measuring P-wave durations at baseline and during sodium channel provocation testing. Atrial structural changes were assessed by measuring atrial dimensions using cardiac magnetic resonance imaging. Atrial ectopic activity was assessed by determining the incidence of atrial ectopic beats using 24 h Holter recordings. Clinical characteristics (including AFib occurrence) did not differ between BrSSCN5A+ and BrSSCN5A-. Baseline P-wave durations were longer in BrSSCN5A+ than in BrSSCN5A-, but lengthened markedly in BrSSCN5A- during provocation testing. Atrial dimensions did not differ. Atrial ectopic beats occurred more often in BrSSCN5A-, and the proportion of patients experiencing one or more atrial ectopic beats was larger in BrSSCN5A- than in BrSSCN5A+. CONCLUSION: In BrS, the presence of an SCN5A mutation is associated with intra-atrial conduction slowing and suppressed atrial ectopic activity. Intra-atrial conduction slowing may provide a plausible substrate for AFib maintenance, while reduced atrial ectopic activity may constitute inhibition of the trigger for AFib initiation.

Energy Metabolism in IDH1 Wild-Type and IDH1-Mutated Glioblastoma Stem Cells: A Novel Target for Therapy?

Cancer is a redox disease. Low levels of reactive oxygen species (ROS) are beneficial for cells and have anti-cancer effects. ROS are produced in the mitochondria during ATP production by oxidative phosphorylation (OXPHOS). In the present review, we describe ATP production in primary brain tumors, glioblastoma, in relation to ROS production. Differentiated glioblastoma cells mainly use glycolysis for ATP production (aerobic glycolysis) without ROS production, whereas glioblastoma stem cells (GSCs) in hypoxic periarteriolar niches use OXPHOS for ATP and ROS production, which is modest because of the hypoxia and quiescence of GSCs. In a significant proportion of glioblastoma, isocitrate dehydrogenase 1 (IDH1) is mutated, causing metabolic rewiring, and all cancer cells use OXPHOS for ATP and ROS production. Systemic therapeutic inhibition of glycolysis is not an option as clinical trials have shown ineffectiveness or unwanted side effects. We argue that systemic therapeutic inhibition of OXPHOS is not an option either because the anti-cancer effects of ROS production in healthy cells is inhibited as well. Therefore, we advocate to remove GSCs out of their hypoxic niches by the inhibition of their binding to niches to enable their differentiation and thus increase their sensitivity to radiotherapy and/or chemotherapy.

Immunohistochemical Detection of Neural Stem Cells and Glioblastoma Stem Cells in the Subventricular Zone of Glioblastoma Patients.

Glioblastoma usually recurs after therapy consisting of surgery, radiotherapy, and chemotherapy. Recurrence is at least partly caused by glioblastoma stem cells (GSCs) that are maintained in intratumoral hypoxic peri-arteriolar microenvironments, or niches, in a slowly dividing state that renders GSCs resistant to radiotherapy and chemotherapy. Because the subventricular zone (SVZ) is a major niche for neural stem cells (NSCs) in the brain, we investigated whether GSCs are present in the SVZ at distance from the glioblastoma tumor. We characterized the SVZ of brains of seven glioblastoma patients using fluorescence immunohistochemistry and image analysis. NSCs were identified by CD133 and SOX2 but not CD9 expression, whereas GSCs were positive for all three biomarkers. NSCs were present in all seven samples and GSCs in six out of seven samples. The SVZ in all samples were hypoxic and expressed the same relevant chemokines and their receptors as GSC niches in glioblastoma tumors: stromal-derived factor-1α (SDF-1α), C-X-C receptor type 4 (CXCR4), osteopontin, and CD44. In conclusion, in glioblastoma patients, GSCs are present at distance from the glioblastoma tumor in the SVZ. These findings suggest that GSCs in the SVZ niche are protected against radiotherapy and chemotherapy and protected against surgical resection due to their distant localization and thus may contribute to tumor recurrence after therapy.

A Phase Ib Clinical Trial of Metformin and Chloroquine in Patients with IDH1-Mutated Solid Tumors.

BACKGROUND: Mutations in isocitrate dehydrogenase 1 (IDH1) occur in 60% of chondrosarcoma, 80% of WHO grade II-IV glioma and 20% of intrahepatic cholangiocarcinoma. These solid IDH1-mutated tumors produce the oncometabolite D-2-hydroxyglutarate (D-2HG) and are more vulnerable to disruption of their metabolism. METHODS: Patients with IDH1-mutated chondrosarcoma, glioma and intrahepatic cholangiocarcinoma received oral combinational treatment with the antidiabetic drug metformin and the antimalarial drug chloroquine. The primary objective was to determine the occurrence of dose-limiting toxicities (DLTs) and the maximum tolerated dose (MTD). Radiological and biochemical tumor responses to metformin and chloroquine were investigated using CT/MRI scans and magnetic resonance spectroscopy (MRS) measurements of D-2HG levels in serum. RESULTS: Seventeen patients received study treatment for a median duration of 43 days (range: 7-74 days). Of twelve evaluable patients, 10 patients discontinued study medication because of progressive disease and two patients due to toxicity. None of the patients experienced a DLT. The MTD was determined to be 1500 mg of metformin two times a day and 200 mg of chloroquine once a day. A serum D/L-2HG ratio of ≥4.5 predicted the presence of an IDH1 mutation with a sensitivity of 90% and a specificity of 100%. By utilization of digital droplet PCR on plasma samples, we were able to detect tumor-specific IDH1 hotspot mutations in circulating tumor DNA (ctDNA) in investigated patients. CONCLUSION: Treatment of advanced IDH1-mutated solid tumors with metformin and chloroquine was well tolerated but did not induce a clinical response in this phase Ib clinical trial.

A Phase II Study Demonstrates No Feasibility of Adjuvant Treatment with Six Cycles of S-1 and Oxaliplatin in Resectable Esophageal Adenocarcinoma, with ERCC1 as Biomarker for Response to SOX.

We assessed the feasibility of adjuvant S-1 and oxaliplatin following neoadjuvant chemoradiotherapy (nCRT) and esophagectomy. Patients treated with nCRT (paclitaxel, carboplatin) and esophagectomy received six 21-day cycles with oxaliplatin (130 mg/m2) on day 1 and S-1 (25 mg/m2 twice daily) on days 1-14. The primary endpoint was feasibility, defined as ≥50% completing treatment. We performed exploratory propensity-score matching to compare survival, ERCC1 and Thymidylate Synthase (TS) immunohistochemistry analyses, proteomics biomarker discovery and 5-FU pharmacokinetic analyses. Forty patients were enrolled and 48% completed all adjuvant cycles. Median dose intensity was 98% for S-1 and 62% for oxaliplatin. The main reason for early discontinuation was toxicity (67%). The median recurrence-free and overall survival were 28.3 months and 40.8 months, respectively (median follow-up 29.1 months). Survival was not significantly prolonged compared to a matched cohort (p = 0.09). Patients with ERCC1 negative tumor expression had significantly better survival compared to ERCC1 positivity (p = 0.01). Our protein signature model was predictive of survival [p = 0.04; Area under the curve (AUC) 0.80]. Moreover, 5-FU pharmacokinetics significantly correlated with treatment-related toxicity. To conclude, six cycles adjuvant S-1 and oxaliplatin were not feasible in pretreated esophageal adenocarcinoma. Although the question remains whether additional treatment with chemotherapy should be provided in the adjuvant setting, subgroups such as patients with ERCC1 negativity could potentially benefit from adjuvant SOX based on our exploratory biomarker research.