Multi-Inlet Spheroid Generator for High-Throughput Combinatorial Drug Screening Based on the Tumor Microenvironment.

Tumor spheroids are powerful tools for drug screening and understanding tumor physiology. Among spheroid formation methods, the hanging drop method is considered most suitable for high-throughput screening (HTS) of anticancer drugs because it does not require surface treatment. However, it still needs to increase the liquid-holding capacity because hanging drops often fall due to the increased pressure caused by the addition of drugs, cells, etc. Here, we report a multi-inlet spheroid generator (MSG) enabling the stable addition of liquid-containing drugs or cells into a spheroid through its side inlet. The MSG was able to load additional solutions through the side inlet without increasing the force applied to the hanging drop. The volume of the additional liquid was easily controlled by varying the diameter of the side inlet. Furthermore, the sequences of the solution injections were manipulated using multiple side inlets. The feasibility of the MSG in clinical application was demonstrated by testing the efficacy of drugs in patient-derived cancer (PDC) cells and controlling the stromal cell ratio in the tumor microenvironment (TME) containing spheroids. Our results suggest that the MSG is a versatile platform for HTS of anticancer drugs and recapitulating the TME.

Methylation and molecular profiles of ependymoma: Influence of patient age and tumor anatomic location.

Ependymomas are tumors of the central nervous system that can occur in patients of all ages. Guidelines from the World Health Organization (WHO) for the grading of ependymomas consider patient age, tumor resection range, tumor location and histopathological grade. However, recent studies have suggested that a greater focus on both tumor location and patient age in terms of transcriptomic, genetic, and epigenetic analyses may provide a more accurate assessment of clinical prognosis than the grading system proposed by WHO guidelines. The current study identified the differences and similarities in ependymoma characteristics using three different molecular analyses and methylation arrays. Primary intracranial ependymoma tissues were obtained from 13 Korean patients (9 adults and 4 children), after which whole-exome sequencing (WES), ion-proton comprehensive cancer panel (CCP) analysis, RNA sequencing, and Infinium HumanMethylation450 BeadChip array analysis was performed. Somatic mutations, copy number variations, and fusion genes were identified. It was observed that the methylation status and differentially expressed genes were significantly different according to tumor location and patient age. Several novel gene fusions and somatic mutations were identified, including a yes-associated protein 1 fusion mutation in a child with a good prognosis. Moreover, the methylation microarray revealed that genes associated with neurogenesis and neuron differentiation were hypermethylated in the adult group, whereas genes in the homeobox gene family were hypermethylated in the supratentorial (ST) group. The results confirmed the existence of significantly differentially expressed tumor-specific genes based on tumor location and patient age. These results provided valuable insight into the epigenetic and genetic profiles of intracranial ependymomas and uncovered potential strategies for the identification of location- and age-based ependymoma-related prognostic factors.

Brain tumor diagnostic model and dietary effect based on extracellular vesicle microbiome data in serum.

The human microbiome has been recently associated with human health and disease. Brain tumors (BTs) are a particularly difficult condition to directly link to the microbiome, as microorganisms cannot generally cross the blood-brain barrier (BBB). However, some nanosized extracellular vesicles (EVs) released from microorganisms can cross the BBB and enter the brain. Therefore, we conducted metagenomic analysis of microbial EVs in both serum (152 BT patients and 198 healthy controls (HC)) and brain tissue (5 BT patients and 5 HC) samples based on the V3-V4 regions of 16S rDNA. We then developed diagnostic models through logistic regression and machine learning algorithms using serum EV metagenomic data to assess the ability of various dietary supplements to reduce BT risk in vivo. Models incorporating the stepwise method and the linear discriminant analysis effect size (LEfSe) method yielded 12 and 29 significant genera as potential biomarkers, respectively. Models using the selected biomarkers yielded areas under the curves (AUCs) >0.93, and the model using machine learning resulted in an AUC of 0.99. In addition, Dialister and [Eubacterium] rectale were significantly lower in both blood and tissue samples of BT patients than in those of HCs. In vivo tests showed that BT risk was decreased through the addition of sorghum, brown rice oil, and garlic but conversely increased by the addition of bellflower and pear. In conclusion, serum EV metagenomics shows promise as a rich data source for highly accurate detection of BT risk, and several foods have potential for mitigating BT risk.

Tumor treating fields plus temozolomide for newly diagnosed glioblastoma: a sub-group analysis of Korean patients in the EF-14 phase 3 trial.

BACKGROUND: Tumor treating fields (TTFields) are anti-mitotic, non-invasive loco-regional cancer therapy comprising low intensity, intermediate frequency alternating electric fields. TTFields plus Temozolomide (TTFields/TMZ) extended survival versus TMZ alone in newly diagnosed glioblastoma (GBM) patients in the EF-14 trial. We report on Korean newly diagnosed GBM patients who participated in the EF-14 trial. METHODS: Thirty-nine participants of the EF-14 trial were enrolled at 8 sites in South Korea. Patients (24 TTFields/TMZ; 14 TMZ alone) received: TTFields (200 kHz) for > 18 h/day; TMZ at 120-150 mg for 5 days per a 28 day cycle. Safety and efficacy were assessed. RESULTS: Patient baseline characteristics were balanced in the 2 arms and the mean age was 52.1 years, 66.7% were male with a mean KPS of 90. Safety incidence was comparable between the 2 arms. In the TTFields/TMZ arm, 30% suffered from skin irritation versus 52% in the entire study population. No TTFields-related serious adverse events were reported. The median progression-free survival (PFS) in the TTFields/TMZ arm was 6.2 months (95% CI 4.2-12.2) versus 4.2 (95% CI 1.9-11.2) with TMZ alone (p = 0.67). Median overall survival was 27.2 months (95% CI 21-NA) with TTFields/TMZ versus 15.2 months (95% CI 7.5-24.1; HR 0.27, p = 0.01) with TMZ alone. CONCLUSION: Median OS and 1- and 2-year survival rates were higher with TTFields/TMZ and similar to the entire EF-14 population. About 30% of patients reported skin irritation, a lower rate than seen in the entire EF-14 population. These results demonstrate the efficacy and safety of TTFields in Korean newly diagnosed glioblastoma patients. CLINICAL TRIALS: Clinicaltrials.gov Identifier: NCT00916409.

Repositioning of the antipsychotic trifluoperazine: Synthesis, biological evaluation and in silico study of trifluoperazine analogs as anti-glioblastoma agents.

Repositioning of the antipsychotic drug trifluoperazine for treatment of glioblastoma, an aggressive brain tumor, has been previously suggested. However, trifluoperazine did not increase the survival time in mice models of glioblastoma. In attempt to identify an effective trifluoperazine analog, fourteen compounds have been synthesized and biologically in vitro and in vivo assessed. Using MTT assay, compounds 3dc and 3dd elicited 4-5 times more potent inhibitory activity than trifluoperazine with IC50 = 2.3 and 2.2 µM against U87MG glioblastoma cells, as well as, IC50 = 2.2 and 2.1 µM against GBL28 human glioblastoma patient derived primary cells, respectively. Furthermore, they have shown a reasonable selectivity for glioblastoma cells over NSC normal neural cell. In vivo evaluation of analog 3dc confirmed its advantageous effect on reduction of tumor size and increasing the survival time in brain xenograft mouse model of glioblastoma. Molecular modeling simulation provided a reasonable explanation for the observed variation in the capability of the synthesized analogs to increase the intracellular Ca2+ levels. In summary, this study presents compound 3dc as a proposed new tool for the adjuvant chemotherapy of glioblastoma.

Giant Magnetic Heat Induction of Magnesium-Doped γ-Fe2 O3 Superparamagnetic Nanoparticles for Completely Killing Tumors.

Magnetic fluid hyperthermia has been recently considered as a Renaissance of cancer treatment modality due to its remarkably low side effects and high treatment efficacy compared to conventional chemotheraphy or radiotheraphy. However, insufficient AC induction heating power at a biological safe range of AC magnetic field (Happl ·fappl < 3.0-5.0 × 109 A m-1 s-1 ), and highly required biocompatibility of superparamagnetic nanoparticle (SPNP) hyperthermia agents are still remained as critical challenges for successful clinical hyperthermia applications. Here, newly developed highly biocompatible magnesium shallow doped γ-Fe2 O3 (Mg0.13 -γFe2 O3 ) SPNPs with exceptionally high intrinsic loss power (ILP) in a range of 14 nH m2 kg-1 , which is an ≈100 times higher than that of commercial Fe3 O4 (Feridex, ILP = 0.15 nH m2 kg-1 ) at Happl ·fappl = 1.23 × 109 A m-1 s-1 are reported. The significantly enhanced heat induction characteristics of Mg0.13 -γFe2 O3 are primarily due to the dramatically enhanced out-of-phase magnetic susceptibility and magnetically tailored AC/DC magnetic softness resulted from the systematically controlled Mg2+ cations distribution and concentrations in octahedral site Fe vacancies of γ-Fe2 O3 instead of well-known Fe3 O4 SPNPs. In vitro and in vivo magnetic hyperthermia studies using Mg0.13 -γFe2 O3 nanofluids are conducted to estimate bioavailability and biofeasibility. Mg0.13 -γFe2 O3 nanofluids show promising hyperthermia effects to completely kill the tumors.

Effect of Tumor-Treating Fields Plus Maintenance Temozolomide vs Maintenance Temozolomide Alone on Survival in Patients With Glioblastoma: A Randomized Clinical Trial.

Importance: Tumor-treating fields (TTFields) is an antimitotic treatment modality that interferes with glioblastoma cell division and organelle assembly by delivering low-intensity alternating electric fields to the tumor. Objective: To investigate whether TTFields improves progression-free and overall survival of patients with glioblastoma, a fatal disease that commonly recurs at the initial tumor site or in the central nervous system. Design, Setting, and Participants: In this randomized, open-label trial, 695 patients with glioblastoma whose tumor was resected or biopsied and had completed concomitant radiochemotherapy (median time from diagnosis to randomization, 3.8 months) were enrolled at 83 centers (July 2009-2014) and followed up through December 2016. A preliminary report from this trial was published in 2015; this report describes the final analysis. Interventions: Patients were randomized 2:1 to TTFields plus maintenance temozolomide chemotherapy (n = 466) or temozolomide alone (n = 229). The TTFields, consisting of low-intensity, 200 kHz frequency, alternating electric fields, was delivered (≥ 18 hours/d) via 4 transducer arrays on the shaved scalp and connected to a portable device. Temozolomide was administered to both groups (150-200 mg/m2) for 5 days per 28-day cycle (6-12 cycles). Main Outcomes and Measures: Progression-free survival (tested at α = .046). The secondary end point was overall survival (tested hierarchically at α = .048). Analyses were performed for the intent-to-treat population. Adverse events were compared by group. Results: Of the 695 randomized patients (median age, 56 years; IQR, 48-63; 473 men [68%]), 637 (92%) completed the trial. Median progression-free survival from randomization was 6.7 months in the TTFields-temozolomide group and 4.0 months in the temozolomide-alone group (HR, 0.63; 95% CI, 0.52-0.76; P < .001). Median overall survival was 20.9 months in the TTFields-temozolomide group vs 16.0 months in the temozolomide-alone group (HR, 0.63; 95% CI, 0.53-0.76; P < .001). Systemic adverse event frequency was 48% in the TTFields-temozolomide group and 44% in the temozolomide-alone group. Mild to moderate skin toxicity underneath the transducer arrays occurred in 52% of patients who received TTFields-temozolomide vs no patients who received temozolomide alone. Conclusions and Relevance: In the final analysis of this randomized clinical trial of patients with glioblastoma who had received standard radiochemotherapy, the addition of TTFields to maintenance temozolomide chemotherapy vs maintenance temozolomide alone, resulted in statistically significant improvement in progression-free survival and overall survival. These results are consistent with the previous interim analysis. Trial Registration: clinicaltrials.gov Identifier: NCT00916409.

The cost-effectiveness of deep brain stimulation for patients with treatment-resistant obsessive-compulsive disorder.

BACKGROUND: Obsessive-compulsive disorder (OCD) is a chronic neuropsychiatric disorder with a 2% to 3% lifetime prevalence; in addition, 10% of OCD patients are resistant to conventional therapy. Deep brain stimulation (DBS) has been an effective treatment for treatment resistant OCD patients (TROCD). We aimed to determine the cost-effectiveness of DBS for TROCD. METHODS: We used a Markov model to estimate the cost-effectiveness of DBS compared to conventional treatment for TROCD with a 10-year time horizon. Published data were used to estimate the rates of treatment response and complications. Costs were calculated from the perspective of the third-party payer. Data on quality of life were obtained from a literature review and a survey of OCD patients. We applied the model separately to Korea and the United Kingdom (UK) to enhance the validity. RESULTS: Base-case analysis showed an incremental cost-effectiveness ratio of US$37,865 per quality-adjusted life-year in Korea and US$34,462 per quality-adjusted life-year in the UK. According to the World Health Organization's criteria, DBS for TROCD was "cost-effective" in Korea (<3x GDP per capita) and "highly cost-effective" in the UK (

Magnetically softened iron oxide (MSIO) nanofluid and its application to thermally-induced heat shock proteins for ocular neuroprotection.

Magnetically softened iron oxide (MSIO) nanofluid, PEGylated (Mn0.5Zn0.5)Fe2O4, was successfully developed for local induction of heat shock proteins (HSPs) 72 in retinal ganglion cells (RGCs) for ocular neuroprotection. The MSIO nanofluid showed significantly enhanced alternating current (AC) magnetic heat induction characteristics including exceptionally high SLP (Specific Loss Power, > 2000 W/g). This phenomenon was resulted from the dramatically improved AC magnetic softness of MSIO caused by the magnetically tailored Mn(2+) and Zn(2+) distributions in Fe3O4. In addition, the MSIO nanofluid with ultra-thin surface coating layer thickness and high monodispersity allowed for a higher cellular uptake up to a 52.5% with RGCs and enhancing "relaxation power" for higher AC heating capability. The RGCs cultured with MSIO nanofluid successfully induced HSPs 72 by magnetic nanofluid hyperthermia (MNFH). Moreover, it was interestingly observed that systematic control of "AC magnetically-induced heating up rate" reaching to a constant heating temperature of HSPs 72 induction allowed to achieve maximized induction efficiency at the slowest AC heating up rate during MNFH. In addition to in-vitro experimental verification, the studies of MSIO infusion behavior using animal models and a newly designed magnetic coil system demonstrated that the MSIO has promising biotechnical feasibility for thermally-induced HSPs agents in future glaucoma clinics.

Effect of peripheral nerve tetanic stimulation on the inter-trial variability and accuracy of transcranial motor-evoked potential in brain surgery.

OBJECTIVES: The aim of this study was to evaluate and compare the advantages of post-tetanic motor-evoked potential (p-MEP) and conventional motor-evoked potential (c-MEP) in terms of MEP inter-trial variability and accuracy. METHODS: c-MEP and p-MEP were quantified in subjects who underwent brain surgery. c-MEP was generated by transcranial electrical stimulation (TES). p-MEP was generated using a preconditioning process involving tetanic stimulation at the left tibial nerve followed by TES. The presence of significant MEP deterioration was monitored during major surgical process. An additional 5-8 MEP obtained after major surgical process were used to analyze amplitude parameters such as mean, standard deviation, range, coefficient of variation (CV), and range to mean ratio. RESULTS: When only irreversible MEP deteriorations were considered as positive results, the false-positive rate was identical for p-MEP and c-MEP. When total MEP deteriorations were considered as positive results, the false-positive rate of p-MEP was lower and p-MEP had higher specificity than c-MEP. The mean amplitude of p-MEP was significantly higher than that of c-MEP. The CV and range to mean ratio of p-MEP were less than those of c-MEP. CONCLUSION: The p-MEP technique is useful for augmenting MEP amplitude and reducing inter-trial variability. SIGNIFICANCE: p-MEP has clinical significance as a useful technique for intraoperative monitoring.

Physical contribution of Néel and Brown relaxation to interpreting intracellular hyperthermia characteristics using superparamagnetic nanofluids.

In this work, the AC magnetically-induced heating characteristics of various viscous nanofluids with either soft ferrite (Fe3O4) or hard ferrite (CoFe2O4) superparamagnetic nanoparticles (SPNPs) were investigated to empirically and physically interpret the contribution of "Néel relaxation loss power, P(Néel relaxation loss)," or "Brown relaxation loss power, P(Brown relaxation loss)," to the total AC heat generation of intracellular hyperthermia or in-vivo hyperthermia. It was found that the contribution of P(Brown relaxation loss) to the total AC heating power, P(totaI), and the specific loss power (SLP) was severely affected by the surrounding environment (or in-vivo environment) while the contribution of P(Néel relaxation loss) to the P(total) was independent of the variation of surrounding environment. Furthermore, all the theoretical and experimental results strongly suggested that highly efficacious intracellular hyperthermia (or in-vivo hyperthermia) modality can be achieved by enhancing the P(Néel relaxation loss) rather than the P(Brown relaxation loss) of SPNP agents in nanofluids.