bluuebunny/crossref_metadata_2025 huggingface dataset:
Performance of solar-air source heat pump heating system aided by district heat supply network research
Authors: Liu Yin , Du Chenyang , Meng Zhaofeng , Dong Suiju , Zhang Fan , Wu Feijun , Wang Mingming
Published: June 2023

To improve the usage rate of district heat supply network and solve the frost problem in solar-air heat pumps, this study used TRNSYS to construct a new solar-air source heat pump heating composite system that uses waste heat from the district heat supply network. The impact of the waste heat recovery device on the heating performance of the new composite system was analysed, and the economic and environmental performance of the composite system was compared with other heating systems. During a typical day, the average coefficient of performance of air source heat pumps (COP) of the composite system increased by 13% and the power consumption of the system decreased by 35.9%. Throughout the heating season, the power consumption of the composite system was 13.59% higher than that of traditional systems, and system coefficient of performance (SCOP) was 15.89% lower than traditional systems. Based on economic analysis, the relative operating costs of the composite system, coal-fired boiler, gas-fired boiler, oil-fired boiler and combined heat and power (CHP) are 1, 1.19, 2.33, 5.56 and 0.85, respectively. The operating cost of the composite heat source system is only 17.43% higher than that of CHP and lower than the other three heating systems. Quantitative evaluations of the environmental friendliness of these systems were conducted using the fuzzy analysis mathematical method. The evaluation results were 9.62, 5.89, 15.05, 9.10 and 6.79, respectively. It was found that the composite system is second only to the gas-fired boiler in terms of environmental friendliness and is 38.9% higher than the coal-fired boiler and 29.5% higher than the CHP.
Insignificant Difference in Early Post-injury Gene Expression Between Patients with Burns Only and Those with Inhalation Injury: A Bioinformatics Analysis
Authors: Jiang Huihao , Dou Zhe , Chen Guangyu , Zhang Guoan , Du Weili
Published: June 2023

Airway obstruction is fatal but common among burn patients in the early period after inhalation injury, during which most tracheotomies are performed within 48 h post-injury. Inflammation is common in laryngoscopy; however, the related gene expression has rarely been studied. In this study, we obtained the data of healthy control and patient samples collected within 8–48 hours post-injury from the Gene Expression Omnibus database and classified them into 10 inhalation-injury patients, 6 burn-only, and 10 healthy controls. Differential gene expression was identified between the patient groups; however, principal component analysis and cluster analysis indicated a similarity between groups. Furthermore, enrichment analysis, Kyoto Encyclopedia of Genes and Genomes, and gene set enrichment analyses showed no significant differences in immune regulation and cell adjustment between the patient groups; but differences were shown when comparing either patient group to the healthy control group, including prominent regulation in inflammatory cells, infection, and cell adjustment. Thus, the gene expression in inhalation injury and burn-only patients does not significantly differ in the early period after injury, especially in inflammation, indicating the absence of specific diagnostic markers or anti-inflammatory treatment in inhalation injury patients, with the potential to identify more subtle differences. Further research is warranted.
Morality Appraisals in Consumer Responsibilization
Authors: Barnhart Michelle , Huff Aimee Dinnín , Scott Inara
Published: June 2023

In recent decades, U.S. gun rights lobbying groups, politicians, courts, and market actors have sought to responsibilize U.S. consumers to use firearms to address the societal problem of crime. These efforts center an interpretation of the constitutional right to keep and bear arms guaranteed by the Second Amendment as an entitlement for individuals to practice armed self-defense. Using interview and online discussion data, this research investigates consumers’ responses to responsibilization for this morally fraught set of behaviors, and the role of consumers’ various understandings of the right to bear arms in these responses. Findings show that consumers consider multiple, specific armed protection scenarios and accept responsibilization in only a portion of these scenarios while rejecting it for the remainder. Acceptance is determined by their appraisals of the morality of consumer responsibilization subprocesses. Consumers’ understanding of the constitutional right serves as a heuristic in these appraisals, with some understandings leading consumers to accept responsibilization across a much larger proportion of scenarios than others. Contributions include illustrating response to consumer responsibilization as a proportionality; illuminating consumers’ active role in appraising responsibilizing efforts; and demonstrating how some consumers come to understand a responsibilized behavior as a moral entitlement.
The interplay of solvent–drug–protein interactions during albumin nanoparticles formulations for temozolomide delivery to brain cancer cells
Authors: Helal Dina O , Abdel-Mottaleb Mona M A , Kamel Amany O , Rouatbi Nadia , Han Shunping , Geneidi Ahmed-Shawky , Al-Jamal Khuloud T , Awad Gehanne A S
Published: June 2023

Objectives

Temozolomide (TMZ), the first line for glioma therapy, suffers from stability at physiological pH. TMZ was selected as a challenging model drug for loading into human serum albumin nanoparticles (HSA NPs). Our aim is to optimise the conditions for TMZ loading into HSA NPs while ensuring TMZ stability.

Methods

Blank and TMZ–HSA NPs were fabricated using the de-solvation technique and the effect of different formulation parameters was evaluated.

Key findings

For blank NPs, crosslinking time had no significant effect on NPs’ size while acetone produced significantly smaller particles than ethanol. Upon drug loading, though TMZ was stable in acetone and ethanol as single agents yet, ethanol-based NPs showed misleadingly high EE% due to drug instability in ethanol formulations as evident by the UV spectrum.

The optimum conditions for drug-loaded particles were: 10 mg/ml HSA, 4 mg TMZ using acetone, yielded NPs with 145 nm in diameter, ξ of −16.98 mV and 0.16% DL. The selected formula reduced the cell viabilities of GL261 glioblastoma cells and BL6 glioblastoma stem cells to 61.9% and 38.3%, respectively.

Conclusions

Our results corroborated that careful manipulation of TMZ formulation processing parameters is crucial for encapsulating such chemically unstable dug while simultaneously ensuring its chemical stability.
Successful surgical removal of a cutaneous lesion compatible with gigantic vascular eccrine spiradenoma: a rare case report
Authors: Alnafee Reem A , Almasaud Aram K , Alshamrani Abdullah M , Alrasheedi Saud D , Alzahrani Abdulrahman J , Alqahtani Adel S
Published: June 2023

Giant vascular eccrine spiradenoma (GVES) is an uncommon type of eccrine spiradenoma (ES). Compared to an ES, this is characterized by a greater degree of vascularity and a bigger size overall. In clinical practice, it is frequently mistaken for a vascular or malignant tumor. To achieve an accurate diagnosis of GVES via biopsy and successfully perform the surgical removal of a cutaneous lesion in the left upper abdomen compatible with GVES. We present a 61-year-old female with the lesion accompanied by on-and-off pain, bloody discharge and skin changes surrounding the mass treated surgically. However, there was no fever, weight loss, trauma or family history of malignancy or cancer managed by surgical excision. The patient recovered well postoperatively and was discharged on the same day with a 2-week follow-up. The wound got healed, the clips were removed on Day 7 postoperatively, and there was no requirement for further follow-up.
Distal colonic atresia: a case report
Authors: Allert Tom , Schellerer Vera
Published: June 2023

Colonic atresia (CA) is a rare disease with an incidence range between one of 20 000 and one of 66 000 live births. Most CA are located within the proximal colon; distal CA are even rarer. Because of its rarity, another case shall be described herewith. A 37th week of pregnancy born child was noticed occurring multiple vomiting, a distended abdomen and additional whitish-bloody stool shortly thereafter. In the first operation, a double-barrel stoma was created. After sufficient weight gain and alignment of the stoma ends, a secondary anastomosis was created in the child after 2 months. The diagnosis can be made reliably on the basis of an X-ray and leads to a good outcome with prompt surgical intervention. However, accompanying malformations should always be considered.
The nuclear-localized RNA helicase 13 is essential for chloroplast development in Arabidopsis thaliana
Authors: Luo Pan , Shi Ce , Zhou Yi , Zhou Jiao , Zhang Xuecheng , Wang Yukun , Yang Yong , Peng Xiongbo , Xie Tingting , Tang Xingchun
Published: June 2023

The chloroplast is a semi-autonomous organelle with a double membrane structure, and its structural stability is a prerequisite for its correct function. Chloroplast development is regulated by known nuclear-encoded chloroplast proteins or proteins encoded within the chloroplast itself. However, the mechanism of chloroplast development regulated by other organelles remains largely unknown. Here, we report that the nuclear-localized DEAD-box RNA helicase 13 (RH13) is essential for chloroplast development in Arabidopsis thaliana. RH13 is widely expressed in tissues and localized to the nucleolus. A homozygous rh13 mutant shows abnormal chloroplast structure and leaf morphogenesis. Proteomic analysis showed that the expression levels of photosynthesis-related proteins in chloroplasts were reduced due to loss of RH13. Furthermore, RNA-sequencing and proteomics data revealed decreases in the expression levels of these chloroplast-related genes, which undergo alternative splicing events in the rh13 mutant. Taken together, we propose that nucleolus-localized RH13 is critical for chloroplast development in Arabidopsis.
GRB 221009A afterglow from a shallow angular structured jet
Authors: Gill Ramandeep , Granot Jonathan
Published: June 2023

Exceptionally bright gamma-ray burst (GRB) afterglows can reveal the angular structure of their jets. GRB jets appear to have a narrow core (of half-opening angle θc), beyond which their kinetic energy drops as a power-law with angle θ from the jet’s symmetry axis, $E_{\mathrm{ k},\rm iso}(\theta)\propto [1+(\theta /\theta _\mathrm{ c})^2]^{-a/2}$. The power-law index a reflects the amount of mixing between the shocked jet and confining medium, which depends on the jet’s initial magnetization. Weakly magnetized jets undergo significant mixing, leading to shallow (a ≲ 2) angular profiles. We use the exquisite multiwaveband afterglow observations of GRB 221009A to constrain the jet angular structure using a dynamical model that accounts for both the forward and reverse shocks, for a power-law external density profile, next ∝ R−k. Both the forward shock emission, that dominates the optical and X-ray flux, and the reverse shock emission, that produces the radio afterglow, require a jet with a narrow core (θc ≈ 0.021) and a shallow angular structure (a ≈ 0.8) expanding into a stellar wind (k ≈ 2). Moreover, these data appear to favour a small fraction (ξe ≈ 10−2) of shock heated electrons forming a power-law energy distribution in both shocks.
ATRT-03. THE ORAL IRON-MIMETIC GALLIUM MALTOLATE INHIBITS ATRT IN VIVO – IMAGING AND HISTOLOGICAL CHARACTERIZATION
Authors: Al-Gizawiy Mona , Wujek Robert T , Zoss Casey J , Tanaka Ryuma , Mirza Shama P , Chitambar Christopher R , Schmainda Kathleen M
Published: June 2023

BACKGROUND

Atypical teratoid rhabdoid tumor (ATRT) is a highly aggressive CNS tumor that is associated with poor outcome. This dire prognosis is due in part to its poor response to the limited treatment options available. Iron and iron proteins play a key role in the growth of many solid cancers, including brain tumors. Our published studies show that the ferrobiology of brain tumors can be exploited for therapeutic purposes. Here, we further demonstrate the inhibitory effects of oral gallium maltolate (GaM) in pediatric ATRT in vivo.

METHODS

Pediatric CHLA-266 ATRT cells were stereotactically implanted into the right striatum of male athymic rats. Advanced MR imaging at 9.4T was carried out weekly starting two weeks after implantation. Daily oral GaM (50mg/kg) or vehicle were provided on tumor confirmation. Longitudinal advanced MRI parameters were processed for enhancing tumor ROIs in OsiriX 8.5.1 (lite) with Imaging Biometrics Software (Imaging Biometrics LLC). Statistical analyses included Kaplan-Meier survival plots, linear mixed model comparisons, and t-statistic for slopes comparison (as indicator of tumor growth rate).

RESULTS

In concordance with our previous in vitro work, in vivo ATRT xenografts were found to be highly sensitive to oral GaM. Median overall survival was 89 days in the control group and 170 days in the treatment group (p=0.011). GaM-treated xenograft tumors grew at a significantly slower rate than control tumors (p<0.0001). On MRI, untreated ATRT xenograft tumors, as their clinical counterparts, tended to localize near the ventricle and occasionally invade it. Histologically, xenograft tumors in our rat model recapitulated the histological features of human ATRT. A significant reduction in MIB-1% and mitotic index (p<0.001 and p<0.05, respectively) was seen in the treatment group. Transferrin receptor and H-ferritin expression in GaM-treated tumors illustrated cellular iron deprivation.

CONCLUSION

Monotherapy with GaM profoundly inhibited ATRT growth and prolonged survival in vivo.
ATRT-06. ETMR TUMORIGENESIS MIRRORS RADIAL GLIAL DEVELOPMENT
Authors: Yao Fupan , Mumal Iqra , Raghuram Nikhil , Xu Liming , Liu Laurie , Ho Ben , Huang Annie
Published: June 2023

Embryonal tumors with multilayered rosettes (ETMRs) are rare pediatric brain tumors defined primarily by overexpression of the C19MC microRNA cluster. These tumors affect mostly children under the age of 3. However, little is known about the origin and developmental contexts of ETMRs. We hypothesized that the early onset of these tumors suggests an embryonic tumorigenic event. To elucidate the embryonic contexts of which ETMRs arise, we curated three mouse embryogenesis datasets representing the full spectrum of murine embryonic development. We generated timepoint structure signatures of this atlas and projected them onto a single nuclei RNA sequencing dataset of 8 primary ETMRs. Consistent with existing literature, we discovered that ETMRs resemble a spectrum of radial glial, neuronal, and oligodendrocytic cell types. We also recapitulated potential ETMR driver mechanisms such as overexpression of WNT and SHH signaling by gene set enrichment analysis. We validated the enrichment of lineage and driver markers through GSEA and marker gene enrichment in a separate cohort of ETMRs characterized by bulk RNA sequencing. Taken together, the embryonic context of ETMR development may play crucial roles in the early identification and treatments of this lethal disease.
ATRT-10. SINGLE-CELL TRANSCRIPTOME ANALYSIS REVEALS CELLULAR HIERARCHIES AND THERAPEUTIC VULNERABILITIES OF ETMR
Authors: Beck Alexander , Gabler Lisa , Cruzeiro Gustavo A V , Lambo Sander , Englinger Bernhard , Shaw McKenzie L , Hack Olivia A , Liu Ilon , Junior Carlos A O de Biagi , Haase Rebecca D , Klenner Marbod , Freidel Pia , Madlener Sibylle , Mayr Lisa , Senfter Daniel , Peyrl Andreas , Slavc Irene , Lötsch Daniela , Dorfer Christian , Haberler Christine , Pfister Stefan M , Baird Lissa , Chi Susan N , Alexandrescu Sanda , Gojo Johannes , Kool Marcel , Hovestadt Volker , Filbin Mariella G
Published: June 2023

Embryonal tumors with multilayered rosettes (ETMR) are malignant brain tumors that occur predominantly in infants and young children. Most patients die within two years of diagnosis, and more effective, targeted therapies are urgently needed. To better characterize the oncogenic mechanisms of key driver alterations and to identify novel therapeutic targets, we set out to study the cellular heterogeneity of ETMR using single-cell RNA sequencing.

Analyses conducted on >4,000 high-quality cells collected from eleven primary and relapse specimens revealed a common cellular hierarchy across all tumors: A highly proliferative neural stem cell-like population (SOX2+) that gives rise to intermediate progenitors (NEUROD1/NEUROG1+) and more differentiated neuron-like cells (STMN2/4+). These malignant cell populations closely overlap with histological patterns of ETMRs, as confirmed by multiplexed immunofluorescence microscopy on patients’ tumors.

Comparison to single-cell datasets from human fetuses indicated high resemblance to normal cortical neurogenesis but also revealed key tumor-specific differences. These include expression of the chromosome 19 miRNA cluster (C19MC, the presumed driver in ~90% of ETMRs), which was restricted to the malignant stem cell-like population. Investigating oncogenic mechanisms of C19MC (comprising 46 miRNA genes) through transcriptome-wide RNA immunoprecipitation analysis, we identified extensive target gene regulation for most C19MC members, including distinct regulators of cell cycle, pluripotency, and neuronal differentiation. Silencing of C19MC families with antisense oligonucleotides resulted in pronounced reduction of ETMR cell line growth, indicating potential avenues for therapeutic targeting in the future.

To identify more immediately actionable targets, we investigated inter-cellular signaling between malignant cell populations of ETMRs. Interestingly, we identified marked FGFR and NOTCH receptor-ligand interactions common to all tumors. An in vitro screen of experimental and approved small molecule inhibitors designed to target these interactions nominated several promising candidates for clinical evaluation. Our unpublished results provide much needed insight into targeting ETMR cellular states using multiple modes of action.
ATRT-11. DEVELOPMENT OF NOVEL PRECLINICAL MODELS AND THERAPEUTIC STRATEGIES FOR ETMR
Authors: Cocito Carolina , Arias-Stella Esteban Uceda , Zhang Xiaohu , McKnight Crystal , Itkin Zina , Klumpp-Thomas Carleen , Cruzeiro Gustavo Alencastro Veiga , Chi Susan N , Pisapia David J , Filbin Mariella G , Dahmane Nadia , Thomas Craig J , Greenfield Jeffrey P
Published: June 2023

Embryonal Tumor with Multilayered Rosettes (ETMR) is a very rare, aggressive pediatric brain tumor with a dismal 5-year overall survival rate. Few genetic aberrations in miRNA clusters and processing machinery have been detected in ETMRs with 90% of ETMR patients having the C19MC amplification. ETMRs show heterogenous histopathological features, including regions of abundant neuropil, hypercellular regions with multilayered rosettes, and papillary structures resembling primitive neuroectoderm. This unique histology, together with the detection of C19MC or DICER alterations, are necessary for diagnosis. The cellular and molecular mechanisms underlying ETMR development is poorly understood, and targeted therapeutics have not been developed due to scarcity of preclinical models.

To expand knowledge of ETMR biology and accelerate the design of novel, it is thus essential to develop preclinical models. We have generated a novel patient-derived ETMR cell line from a resected patient tumor sample (CTBP-03A) and created two patient-derived xenograft (PDX) models (CTBP-03A and CBTP-38). We have used these unique patient-derived cell lines to identify mechanisms driving ETMR development and progression. To identify novel therapeutic targets, we conducted high-throughput drug screening (HTDS) utilizing 2480 approved and investigational drugs, against the CBTP-03A cell line. Among the candidate drugs showing an inhibitory effect on CBTP-03, 63 compounds were selected based on their activity, CNS penetration ability, mechanism of action, and bioavailability, if known. The 63 potency-selected hits were screened in an all-versus-all format, totaling 1953 combinations. The rapidity of action of the top choice compounds used as single agent or in double and triple combination was assessed by caspase glow assay. This initial combination screen identified DNA replication stress and FGF signaling as potential new drug combinations for ETMR.

We will thus describe the first combinatorial HTDS on two novel ETMR patient-derived cell lines and potential drug combination strategies for the treatment of ETMR.
ATRT-13. ASSESSMENT OF MITOCHONDRIAL BIOENERGETICS AND FISSION-FUSION DYNAMICS IN ETMR
Authors: Liapis Evangelos , Pamreddy Annapurna , Maas Allison , Hanson Derek , Carter Claire
Published: June 2023

BACKGROUND

Embryonal tumor with multilayered rosettes (ETMR) is a deadly pediatric brain tumor with limited available therapeutic options. Dysregulated mitochondrial bioenergetics and dynamics have been linked to the initiation and progression of diverse human cancers, however, their role in ETMR pathobiology remains unknown. We used the primary ETMR cell line, BT183, and an in-house developed human neural stem cell (NSC) line to evaluate mitochondrial morphology, bioenergetics and fission-fusion dynamics involved in ETMR growth and aggression.

METHODS

Mitochondrial morphology was determined by widefield immunofluorescence and dSTORM super-resolution microscopy in BT183 and NSC monolayers and by TEM in BT183 and NSCs grown as neurospheres. Relative expression of mitochondrial proteins were quantified in BT183 and NSC lysates by western blotting.

RESULTS

Widefield immunofluorescence and super-resolution imaging revealed profound differences in mitochondrial morphology in BT183 compared to NSCs. BT183 mitochondria appear as fragmented, rounded structures, with little interconnectivity, while NSC mitochondria present a typical elongated, filamentous morphology. These findings were corroborated by TEM image analysis of BT183 and NSC neurospheres, which revealed additional morphological alterations in BT183 mitochondria at the ultrastructural level, including aberrant cristae structure. Western blot analysis of BT183 and NSC lysates showed up to 5-fold differential expression of mitochondrial OXPHOS, glycolytic and fission-fusion proteins in the two cell types. Fission promoters phospho-Drp1 and MFF displayed significantly higher expression in BT183 relative to NSCs, whereas fusion-inducing proteins Opa1 and Miro2 showed significantly lower differential expression in BT183.

CONCLUSIONS

BT183 cells exhibited striking mitochondrial fragmentation and differential expression of proteins involved in fission-fusion, OXPHOS and glycolysis when compared to normal human NSCs. Ongoing studies include the analysis of mitochondrial structure and morphology in patient samples to validate our in-vitro findings, in addition to in-vitro metabolic flux analysis and targeted protein depletion studies to identify druggable pathways based on mitochondrial function and dynamics.
ATRT-14. ASSESSMENT OF MITOCHONDRIAL BIOENERGETICS AND FISSION-FUSION DYNAMICS IN ETMR
Authors: Liapis Evangelos , Pamreddy Annapurna , Maas Allison , Hanson Derek , Carter Claire
Published: June 2023

BACKGROUND

Embryonal tumor with multilayered rosettes (ETMR) is a deadly pediatric brain tumor with limited available therapeutic options. Dysregulated mitochondrial bioenergetics and dynamics have been linked to the initiation and progression of diverse human cancers, however, their role in ETMR pathobiology remains unknown. We used the primary ETMR cell line, BT183, and an in-house developed human neural stem cell (NSC) line to evaluate mitochondrial morphology, bioenergetics and fission-fusion dynamics involved in ETMR growth and aggression.

METHODS

Mitochondrial morphology was determined by widefield immunofluorescence and dSTORM super-resolution microscopy in BT183 and NSC monolayers and by TEM in BT183 and NSCs grown as neurospheres. Relative expression of mitochondrial proteins were quantified in BT183 and NSC lysates by western blotting.

RESULTS

Widefield immunofluorescence and super-resolution imaging revealed profound differences in mitochondrial morphology in BT183 compared to NSCs. BT183 mitochondria appear as fragmented, rounded structures, with little interconnectivity, while NSC mitochondria present a typical elongated, filamentous morphology. These findings were corroborated by TEM image analysis of BT183 and NSC neurospheres, which revealed additional morphological alterations in BT183 mitochondria at the ultrastructural level, including aberrant cristae structure. Western blot analysis of BT183 and NSC lysates showed up to 5-fold differential expression of mitochondrial OXPHOS, glycolytic and fission-fusion proteins in the two cell types. Fission promoters phospho-Drp1 and MFF displayed significantly higher expression in BT183 relative to NSCs, whereas fusion-inducing proteins Opa1 and Miro2 showed significantly lower differential expression in BT183.

CONCLUSIONS

BT183 cells exhibited striking mitochondrial fragmentation and differential expression of proteins involved in fission-fusion, OXPHOS and glycolysis when compared to normal human NSCs. Ongoing studies include the analysis of mitochondrial structure and morphology in patient samples to validate our in-vitro findings, in addition to in-vitro metabolic flux analysis and targeted protein depletion studies to identify druggable pathways based on mitochondrial function and dynamics.
ATRT-18. ELUCIDATING THE ETMR MICROENVIRONMENT USING SPATIAL MULTI-OMICS TECHNOLOGIES
Authors: Liapis Evangelos , Pamreddy Annapurna , O’Neill Kelly , Maas Allison , Hanson Derek , Carter Claire
Published: June 2023

Embryonal tumor with multilayered rosettes (ETMR) is a highly aggressive CNS neoplasm that occurs almost exclusively in infants under 4 years of age and is associated with an extremely poor prognosis. A bottleneck in the development of new and curative treatments for ETMR stems from our limited understanding of the spatiotemporal biological heterogeneity within patient tumor samples and a lack of preclinical models that adequately reflect the entire biological spectrum. Novel lipid-based therapeutics that encompass ganglioside-directed immunotherapy, targeting lipid metabolism and membrane lipid therapy, have shown efficacy in the treatment of brain tumors but have yet to be evaluated in the treatment of ETMR. We carried out mass spectrometry imaging on ETMR patient samples to identify the histology-specific accumulation of lipids within the diverse cell populations of the tumor microenvironment. Our preliminary studies identified the accumulation of glycosphingolipids, cardiolipins, phosphatidylinositols, and ceramide-1-phosphates within tumor cells, including the characteristic multilayered rosettes, and within different microvascular proliferations. All data were correlated to histopathology and Ki67 proliferation index. We next evaluated the lipid profiles in the patient-derived ETMR cell line, BT183, compared to control neural stem cells (NSCs), grown as 3D tumorspheres and neurospheres, respectively. From this data we identified lipids that accumulated in our patient samples and the ETMR cell line but were significantly reduced or not detected in the control NSCs. Analysis aimed at a systems biology approach to study the ETMR microenvironment using imaging mass cytometry for multiplex IHC and spatial transcriptomics to correlate lipidomic phenotype to distinct cell populations within the microenvironment are ongoing. Preclinical model development based on these findings and drug screening of therapeutics targeting these lipidomic pathways are also ongoing.
BIOL-07. DISTINCTIVE FEATURES OF HIGH-GRADE GLIOMA MOUSE MODELS REVEALED BY SINGLE-NUCLEUS RNA-SEQUENCING GUIDE PRE-CLINICAL MODEL SELECTION
Authors: Ernst Kati , Okonechnikov Konstantin , von Soosten Laura , Hofmann Nina , Mack Norman , Schwalm Benjamin , Wechsler-Reya Robert J , Pfister Stefan M , Thongjuea Supat , Jones David T W , Zuckermann Marc
Published: June 2023

Glioma is the most common pediatric central nervous system tumor, with high-grade gliomas (HGG) having one of the worst prognoses of all human cancers. In order to develop better diagnostics and therapies, it is essential to use faithful disease models. Currently, highly passaged patient-derived xenograft (PDX) models are most widely used in preclinical studies, although alternative immunocompetent models are becoming increasingly available. Here, we compare several in vivo glioma models on a single-cell level and investigate their similarity to primary human tumors. Single-nucleus sequencing was used to analyze >125,000 nuclei of primary patient samples, xenografts, autochthonous mouse tumors and related allografts including early and late in vivo passages – with a focus on MET-fusion-driven as well as H3 K27M-mutant HGG. Transcriptomic profiles of single tumor cells and associated stromal/immune components reveal insights into model-specific intratumoral heterogeneity, tumor evolution, and similarities between mouse models and patient samples. In addition, matched tumors engrafted into immunocompromised and immunocompetent animals are used to examine tumor-immune crosstalk and the modifying role of the tumor microenvironment. This improved understanding of how the analyzed model systems evolve over time and how they reflect patient’s tumor compositions will allow to prioritize and refine modern preclinical trials.
BIOL-15. PATIENT-DERIVED XENOGRAFTS OF MATCHED PRIMARY-RECURRENT PEDIATRIC BRAIN TUMORS AS A RESOURCE FOR IDENTIFYING THERAPY RESISTANCE PROFILES
Authors: Zhao Miao , Olsen Thale , Bergström Tobias , Rosén Gabriela , Dalmo Erika , Vikström Elin , Larsson Ida , Lin Tina , Sundström Anders , Holmberg Karl , Weishaupt Holger , Nelander Sven , Giraud Geraldine , Swartling Fredrik
Published: June 2023

Tumor recurrence developing from therapy resistance, immune escape and metastasis is the leading cause of death of malignant pediatric brain tumors. By studying paired primary-recurrent patient samples we previously identified significant accumulation of SOX9-positive cells in Group 3 medulloblastoma that was similarly induced following long-term irradiation in transgenic mouse models of MYC-induced medulloblastoma.

To follow relapse biology in malignant pediatric brain tumors at the single cell level we have established patient-derived xenografts (PDXs) from a number of matched primary-relapse samples, including pediatric high-grade glioma, medulloblastoma and ependymoma profiled with DNA methylation analysis. We next used scRNA-Seq to identify clones with particular expression of genes and distinct signaling pathways enriched in recurrent tumor cells as compared to their matched primary tumor cells. We further used a lentivirus-based barcoding strategy coupled to scRNA-Seq to study the molecular evolution of cell clones arising from genetically modified mouse models and PDXs that survive long-term fractionated irradiation, cisplatin or temozolomide treatment in vitro and in vivo.

By using computational modeling we identified means of specific targeting of recurrent cells that could be of potential use in future treatments for patients affected by fatal relapses. Our results suggest that pediatric brain tumors of distinct tumor entities use different ways to acquire therapy resistance in patients which is often coupled to an expansion of clones that are developing therapy-induced resistance.
BIOL-16. OBESOGENIC DIET EXPOSURE INCREASES NF1-OPTIC PATHWAY GLIOMA FORMATION IN PART THROUGH EFFECTS ON THE CELL OF ORIGIN
Authors: Chan Ambrose , Zhang Kailong , Martin Gemma , Gutmann David , Brossier Nicole
Published: June 2023

Pediatric low-grade glioma incidence has been rising in the U.S. over the last 20 years, concurrent with a rising rate of obesity in both the adult and pediatric populations. Recently, children of obese mothers have been demonstrated to have increased rates of several tumors, including brain tumors. Importantly, obesity in the U.S. is driven in large part by diet, given the abundance and accessibility of high-fat, high-sugar food choices. High-fat diet exposure has been previously demonstrated to promote proliferation and glial differentiation of embryonic neuroglial progenitor cells (NPCs) around the third ventricular zone (TVZ), suggesting that in utero exposure to an obesogenic diet might affect the formation of pediatric tumors derived from these cells, such as Neurofibromatosis Type 1 (NF1)-related optic pathway glioma (OPG). Based on these data, we hypothesized that maternal obesogenic exposure would increase Nf1-OPG formation through intrinsic effects on the tumor cell of origin. We demonstrated that progeny from obese dams exposed to HFHS diet during and preceding gestation demonstrated increased proliferation and glial differentiation of WT and Nf1-heterozygous TVZ NPCs in vivo. Progeny of non-obese dams exposed to this diet during gestation demonstrated a similar phenotype, suggesting these effects were related to maternal diet rather than weight. We then assessed how obesogenic diet exposure affected tumor formation. We determined that this exposure increases glioma penetrance in two low-penetrance models of NF1-OPG. Finally, we demonstrated that obesogenic diet exposure resulted in earlier tumor onset in a high-penetrance Nf1-OPG model. Taken together, these findings suggest that obesogenic diet exposure increases pediatric glioma formation, in part through effects on the tumor cell of origin while still in utero. It may also have implications for clinical prognosis, as earlier age of tumor onset has been negatively associated with visual outcomes in NF1-OPG.
BIOL-17. DISSECTING THE EXPRESSION AND FUNCTION OF ANGIOPOIETIN-1 IN PEDIATRIC BRAIN TUMORS
Authors: Langhnoja Jaldeep , Wei Xin , Arounleut Phonepasong , Kundu Ipsita , Phoenix Timothy
Published: June 2023

BACKGROUND

Angiopoietin-1 (Angpt1) is a secreted protein that promotes angiogenesis and vascular stability in development and certain disease states through activation of the Tie2 receptor. However, little is known about its expression or function within pediatric brain tumors.

METHODS

We utilized available datasets from patient samples and created murine models of pediatric high-grade glioma and ependymoma by in utero electroporation. Using a combination of transcriptomics and Angpt1-GFP reporter tumor models we evaluated Angpt1 expression across different types of pediatric brain tumors. We also applied inducible Angpt1 floxed mice to generate tumor-specific knock-out models.

RESULTS

Angpt1 is upregulated in glial brain tumors, including high-grade gliomas and ependymomas. We find Angpt1 is expressed by specific cell populations within gliomas and ependymomas, along with non-tumor reactive astrocytes and perivascular mural cells. Leveraging our in vivo brain tumor platform we have created tumor specific Angpt1 knock-out models by including a plasmid expressing Cre-recombinase. Ongoing studies aim to delineate the function of Angpt1 in tumor pathogenesis, including tumor angiogenesis and blood-brain barrier function.

CONCLUSION

Current work finds that Angpt1 is upregulated in glial brain tumors and nominates it as a potential modulator of brain tumor vascular biology, including tumor angiogenesis and permeability.
BIOL-21. PATIENT-DERIVED, THREE-DIMENSIONAL ORGANOID PLATFORM FOR PEDIATRIC BRAIN TUMOR MODELING AND THERAPEUTIC TESTING
Authors: Beale David , Baubet Valerie , Goel Peeyush , Patterson Luke , Jacob Fadi , Santi-vicini Maria-Rita , Viaene Angela , Ming Guo-li , Song Hongjun , Madsen Peter , Foster Jessica , Griffin Crystal , Stern Ally , Storm Phillip , Koptyra Mateusz , Resnick Adam
Published: June 2023

Brain tumors are the leading cause of cancer-related death in children. An important hurdle to scientific and clinical progress in the field has been the limited availability of preclinical tumor models. In recent years, the Children’s Brain Tumor Network (CBTN) has accelerated the development of patient-derived cell lines and xenografts, offering these resources to the community through open-source access. While these models are extremely valuable, their development process can be lengthy and result in clonally selected lines which presents a challenge for studying complex tumor biology. To address the need for three-dimensional tissue culture, our group in conjunction with CBTN, generates organoids from fresh tissue specimens obtained directly from surgical resection. This effort resulted in numerous organoid models generated for high-grade glioma (HGG), medulloblastoma, ependymoma, atypical teratoid-rhabdoid tumor, and low-grade glioma tumors. The tissue was processed within an hour post-extraction and cultured in growth factor-free media. Organoid formation and growth were observed as early as in 2 weeks post initiation and continued for up to three months. Banked organoids established growth upon return to culture and resembled similar cell composition to those in the original culture. This provides an excellent preclinical testing model for precision medicine-driven trials. To represent this, we utilized CAR-T testing approach and treated GD2-positive HGG organoids with GD2 CAR-T cells and control CD19 CAR-T cells. 72h exposure to GD2 CAR-T induced morphological changes and significant increase in dead cells within organoids, which was not observed in CD19 CAR-T treated counterparts (p<0.05). Here we provide a simple and efficient workflow for the generation and testing of three-dimensional tumor organoid models generated from fresh surgical tissue. The platform can accelerate not only tumor biology explorations but also empower a diverse array of precision medicine-driven testing for pediatric brain tumors.
CNSC-02. INVESTIGATING THE ROLE OF THE WNT SIGNALING TRANSDUCTION PATHWAY IN GLIOMA BLOOD-BRAIN-BARRIER INTEGRITY
Authors: Tang Kayen , Conrad Christina , Lim Sanghee , Kwak Minhye , Cesaire Melissa , Jackson Sadhana
Published: June 2023

The WNT signaling transduction pathway has been linked to cancer stem cell self-renewal, differentiation, and blood brain barrier (BBB) development. Previous studies have shown that WNT pathway inhibition increased BBB permeability, specifically in WNT medulloblastoma. Glioblastoma, an aggressive malignant tumor with poor prognosis, has also been shown to have high WNT expression and variable BBB permeability. We hypothesize that activating WNT pathway inhibition in glioblastoma stem cells (GSCs) will inhibit glioma progression, increase BBB permeability within the tumor microenvironment, and enhance overall treatment responsiveness. We transduced primary pediatric derived glioma stem cells with overexpressing vectors for WNT inhibitors (DKK1 or WIF1). To characterize effects of DKK1 or WIF1 overexpression, we evaluated cell migration patterns via RTCA xCelligence, cell cycle progression via Annexin 5 assays, and cell viability via cell-titer glo. Brain endothelial cell integrity was evaluated with the treatment of CHIR99021, WNT pathway activator, or GSC condition media with immunoblotting for junctional protein expression (Claudin-5, Claudin-3, Occludin, ZO-1, VE-Cadherin). Verification of WNT inhibitory overexpressing GSCs for DKK1 and WIF1 was performed using RNAseq, proteomics, and immunoblotting. DKK1-GSCs demonstrated a statistically significant impairment in the migratory slope compared to empty vector GSCs. Cell cycle analysis also revealed that DKK1-GSCs were arrested in G0/1 phase while WIF1-GSCs were arrested in S/G2M phase. CHIR99021 endothelial treatment resulted in increased secretion of WNT inhibitors DKK1 and SFRP1 and resultantly, there was a decrease in junctional protein expression. Overall, WNT inhibitor overexpressing GSCs exhibited unique cellular patterns and indirectly disrupted endothelial cell integrity. Future studies will evaluate orthotopic WNT inhibitor overexpressing transplant rodent models to determine BBB integrity, drug permeability, model survival, and chemotherapeutic treatment response. By enhancing our understanding of the WNT pathway in GSCs, we anticipate understanding the use of future therapeutic WNT pathway targeting to increase chemotherapeutic responsiveness and improve glioblastoma prognosis.
CNSC-03. EXPLORING STRUCTURAL DISRUPTION OF TRICELLULAR JUNCTIONS TO ENHANCE BLOOD-BRAIN-BARRIER PERMEABILITY
Authors: Cesaire Melissa , Dalmage Mahalia , Lim Sanghee , Minhye Kwak , Tang Kayen , Conrad Christina , Karim Baktiar , Butcher Donna , Jackson Sadhana
Published: June 2023

The blood-brain barrier (BBB) hinders CNS chemotherapy entry for malignant glioma treatment. It is predominantly composed of brain endothelium linked by bicellular (BJ) and tricellular (TJ) tight junctions. Previous studies demonstrated,angubindin-1 inhibited TJ angulin-1 to increase brain and CSF targeted therapy in rodent lung cancer models. We hypothesize angubindin-1 can transiently decrease BBB junctional integrity to increase chemotherapy permeability and prolong rodent glioma model survival. Rat brain endothelial cells were treated with TJ and BJ inhibitors against angulin-1 (angubindin-1 600µg/mL), claudin-3 (C-CPE 200µg/mL) or claudin-5 (C-CPE-MT 200µg/mL). Endothelial integrity was assessed by immunoblotting and cell-cell electrical impedance. Effects of angubindin-1 on efflux transporter P-glycoprotein (PGP) and migration were studied using rat and human derived glioma cells. Rat glioma models were treated with doxil (3 mg/kg), angubindin-1 (10 mg/kg or 30 mg/kg), or combination therapy for assessments on tumor volume, BBB permeability and survival. We observed decreased angulin-1 expression 5 hours after angubindin-1 treatment, with return to baseline by 24 hours (p<0.05). Angubindin-1, CCPE, and CCPE-mt globally reduced endothelial cell-cell integrity, maximally at 4 hours with a return to baseline by 12 hours; with angubindin-1 demonstrating the largest decreased cell-adhesion (angubindin-1 vs control, p< 0.0001). Angubindin-1 also decreased PGP efflux of rhodamine in both endothelial and glioma cells, along with demonstrating a pro-migratory dose-dependent effect on rat glioma cells. Combined angubindin-1 and doxil increased survival in rat glioma models compared with doxil alone (24 days vs. 18 days, p < 0.0001). Day 14 tumor volume was significantly decreased with angubindin-1 and combination treatment respectively (77.5 % vs 81.6 %, p<0.05). Ongoing studies are exploring, angubindin-1’s effect on large CNS drug permeability, TJ localization post BBB disruption and additional combinational treatments in glioma models. Collectively, these findings pose a unique opportunity to disrupt BBB integrity and improve malignant glioma treatment options.
CNSC-04. EVALUATING LONGITUDINAL WORKING MEMORY PERFORMANCE IN PEDIATRIC PATIENTS WITH CRANIOPHARYNGIOMA VIA FMRI
Authors: Chang Chih-Chiang , Zou Ping , Scoggins Matthew A , Conklin Heather , Hua Chia-Ho , Merchant Thomas E , Sitaram Ranganatha
Published: June 2023

Craniopharyngioma is a type of benign brain tumor that develops in the suprasellar region of the brain and can lead to long-term neurological, endocrine, and visual impairments as well as neurocognitive deficits resulting from the tumor and its treatment. The contemporary treatment options for this tumor are radical surgery or limited surgery in conjunction with radiotherapy. The current understanding of the impact of brain injury related to the tumor and its treatment such as proton radiotherapy (PRT) on neurocognitive function is limited.

The study (NCT01419067) was approved by the IRB and informed consent was obtained from patients or their parents. Functional magnetic resonance imaging (fMRI) was performed in a 3T MRI imaging system (Siemens MAGNETOM Prisma, Erlangen, Germany) and used to evaluate the neurocognitive performance of 58 pediatric patients (31 females and 27 males) before and after proton radiotherapy (PRT). The patients underwent serial imaging at four-time points (baseline, 18, 36, and 60 months after PRT) and performed a working memory (0-back, 1-back, and 2-back) task-based fMRI. A full factorial model was used for longitudinal group analysis at different time points through statistical parametric mapping. Omission errors, commission errors, and reaction time were measured for each task.

The results showed robust detection of the working memory network with increased activation (FDR<0.05) in the precuneus, mid-temporal gyrus, and angular gyrus at 36 months compared to 18 months. The behavioral test results showed a gradual decrease in omission errors in the 2-back task group and an increase in reaction time for all three task groups over time.

These findings suggest that PRT has a transient effect on brain activation and behavioral performance, potentially indicating improved executive functioning, recovery from the mass effect of the tumor and/or surgery, and the minimal negative impact from PRT.
DIPG-03. IDENTIFICATION OF NEW THERAPEUTIC TARGETS BY ARRAYED CRISPRA-BASED APPROACH TO MAP RADIORESISTANCE IN DMG
Authors: Fernandez Solène , Laternser Sandra , Sellitto Stefano , Neupane Sandesh , Petrovic Antonela , Ciani Ludovica , Kritzer Bettina , Hasler Kerstin , Bamert David , Nazarian Javad , Aguzzi Adriano
Published: June 2023

Pediatric Diffuse Midline Glioma (DMG) is amongst the most aggressive childhood brain tumors. The dismal prognosis is less than a year for children diagnosed. Radiotherapy (RT) remains the only standard treatment that provides only transient relief of clinical symptoms of DMG patients. Poor survival is associated with radio-resistance due in part to P53 mutational status and to invasive tumor properties related to epigenetic dysregulation. Over 80% of DMG patients harbor oncohistone 3 mutations (H3K27M) leading to global genome hypomethylation and transcriptional disruption. Transcriptional factors are known as master regulators of the genome but their implication in response to radiation treatment remains unclear.

We developed an arrayed-CRISPR screen strategy to understand the role of transcription factors in radioresistance mechanisms in DMG-patient-derived cells.

We performed arrayed-CRISPRa screening in multiwell plates with DMG cells expressing the dCas9-VP64, infected with gRNA lentiviral library to target each transcription factor (one transcription factor overexpressed per well). Each plate was irradiated fractionally (2Gy/day) and after irradiation stopped a comparison between cells overexpressing one targeted transcription to internal controls (non-targeted-gRNA infected cells) has been conducted. Transcription factor “hits” identification is based on Log2FC (logarithm 2 fold-change) with a value higher than 0.6, considering these targets as potentially involved in DMG cell radio-resistance and/or proliferation mechanisms. We identified 82 “hits” with more than 30% described as involved in radio-resistance mechanisms and/or tumorigenesis process, sustaining our screen results. We focus on “hits” with a Log2FC higher than 1 to identify the most robust targets playing a role in resistance to radiation. Currently, DMG cells are being treated with RT combination with pharmacological inhibitors to validate our candidate target involvement in DMG radio-resistance. Our goal is to achieve a better understanding of resistance mechanisms to identify therapeutic treatments to combine with radiotherapy to overcome radio-resistance developed by DMG cells.