Genomic analysis and preclinical xenograft model development identify potential therapeutic targets for MYOD1-mutant soft-tissue sarcoma of childhood.

The myogenic differentiation 1 gene (MYOD1) p.L122R somatic mutation was first discovered in a subset of clinically aggressive embryonal rhabdomyosarcomas and has since been described in both pediatric and adult spindle cell/sclerosing rhabdomyosarcomas. Relatively little is known about the clinical, molecular, and histopathological features of these tumors in children. In order to further characterize the genomic and clinical features of pediatric MYOD1-mutant sarcomas, we evaluated a cohort of soft-tissue sarcoma patients treated at Texas Children's Hospital. Tumor DNA was subjected to next-generation panel sequencing and/or Sanger sequencing of the MYOD1 hotspot mutation. The MYOD1 p.L122R mutation was identified in six tumors, with a variant allele fraction greater than 0.8 in three cases, suggestive of loss of heterozygosity. One sclerosing rhabdomyosarcoma lacking the MYOD1 hotspot mutation was observed to have a MYOD1 copy number gain, also with evidence of loss of heterozygosity. Cancer gene panel sequencing revealed potentially targetable alterations in six of seven (86%) patients with MYOD1 alterations, including four patients with an alteration in the PI3K-AKT pathway: two hotspot PIK3CA mutations and deletions in PTEN and TSC2. On histopathologic review, MYOD1-altered tumors exhibited spindle and/or round cells and varying degrees of hyaline sclerosis. At last follow-up, six patients had died of disease and the seventh progressed early and was subsequently lost to follow-up. Both pre- and post-therapy patient-derived xenograft models were generated from one patient's tumor. These models were confirmed to harbor the MYOD1 and PIK3CA mutations seen in the primary tumor and were shown to be sensitive to PI3K/mTOR inhibition in vitro and in vivo. In conclusion, this study adds to recent reports describing the clinicopathologic and genomic features of MYOD1-altered soft-tissue sarcomas in children, including dismal prognosis and potential molecular targets for therapy. The novel preclinical models developed will facilitate further biological and preclinical study of this rare and aggressive tumor. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Integrated DNA Copy Number and Expression Profiling Identifies IGF1R as a Prognostic Biomarker in Pediatric Osteosarcoma.

Osteosarcoma is a primary malignant bone tumor arising from bone-forming mesenchymal cells in children and adolescents. Despite efforts to understand the biology of the disease and identify novel therapeutics, the survival of osteosarcoma patients remains dismal. We have concurrently profiled the copy number and gene expression of 226 osteosarcoma samples as part of the Strategic Partnering to Evaluate Cancer Signatures (SPECS) initiative. Our results demonstrate the heterogeneous landscape of osteosarcoma in younger populations by showing the presence of genome-wide copy number abnormalities occurring both recurrently among samples and in a high frequency. Insulin growth factor receptor 1 (IGF1R) is a receptor tyrosine kinase which binds IGF1 and IGF2 to activate downstream pathways involved in cell apoptosis and proliferation. We identify prevalent amplification of IGF1R corresponding with increased gene expression in patients with poor survival outcomes. Our results substantiate previously tenuously associated copy number abnormalities identified in smaller datasets (13q34+, 20p13+, 4q35-, 20q13.33-), and indicate the significance of high fibroblast growth factor receptor 2 (FGFR2) expression in distinguishing patients with poor prognosis. FGFR2 is involved in cellular proliferation processes such as division, growth and angiogenesis. In summary, our findings demonstrate the prognostic significance of several genes associated with osteosarcoma pathogenesis.

Integrated DNA and RNA sequencing reveals targetable alterations in metastatic pediatric papillary thyroid carcinoma.

BACKGROUND: Pediatric papillary thyroid carcinoma (PTC) is clinically and biologically distinct from adult PTC. We sequenced a cohort of clinically annotated pediatric PTC cases enriched for high-risk tumors to identify genetic alterations of relevance for diagnosis and therapy. METHODS: Tumor DNA and RNA were extracted from FFPE tissue and subjected to next-generation sequencing (NGS) library preparation using a custom 124-gene hybridization capture panel and the 75-gene Archer Oncology Research Panel, respectively. NGS libraries were sequenced on an Illumina MiSeq. RESULTS: Thirty-six pediatric PTC cases were analyzed. Metastases were frequently observed to cervical lymph nodes (29/36, 81%), with pulmonary metastases less commonly found (10/36, 28%). Relapsed or refractory disease occurred in 18 patients (18/36, 50%). DNA sequencing revealed targetable mutations in 8 of 31 tumors tested (26%), most commonly BRAF p.V600E (n = 6). RNA sequencing identified targetable fusions in 13 of 25 tumors tested (52%): RET (n = 8), NTRK3 (n = 4), and BRAF. Mutually exclusive targetable alterations were discovered in 15 of the 20 tumors (75%) with both DNA and RNA analyzed. Fusion-positive PTC was associated with multifocal disease, higher tumor staging, and higher American Thyroid Association risk levels. Both BRAF V600E mutations and gene fusions were correlated with the presence of cervical metastases. CONCLUSIONS: Targetable alterations were identified in 75% of pediatric PTC cases with both DNA and RNA evaluated. Inclusion of RNA sequencing for detection of fusion genes is critical for evaluation of these tumors. Patients with fusion-positive tumors were more likely to have features of high-risk disease.

No neuron is an island: Homeostatic plasticity and over-constraint in a neural circuit.

To support computation the activity of neurons must vary within a useful range, which highlights one potential value of homeostatic plasticity. The interconnectedness of the brain, however, introduces the possibility that combinations of homeostatic mechanisms can produce over-constraint in which not all set points can be satisfied. We use a simulation of the cerebellum to investigate the potential for such conflict and its consequences. In this instance the conflict produces perpetual drift and eventual saturation of synaptic weights. We show that these problems can be resolved for this network by a particular combination of sites and rules for plasticity. We also show that simulations that implement these rules for homeostatic plasticity are more resistant to forgetting. These results illustrate the general principle that homeostatic plasticity within a system must not set up conflicts in which mutually exclusive set points exist and that one consequence can be perpetual induction of plasticity.

Intratumoral morphologic and molecular heterogeneity of rhabdoid renal cell carcinoma: challenges for personalized therapy.

Rhabdoid histology in clear-cell renal cell carcinoma is associated with a poor prognosis. The prognosis of patients with clear-cell renal cell carcinoma may also be influenced by molecular alterations. The aim of this study was to evaluate the association between histologic features and salient molecular changes in rhabdoid clear-cell renal cell carcinoma. We macrodissected the rhabdoid and clear-cell epithelioid components from 12 cases of rhabdoid clear-cell renal cell carcinoma. We assessed cancer-related mutations from eight cases using a clinical next-generation exome-sequencing platform. The transcriptome of rhabdoid clear-cell renal cell carcinoma (n=8) and non-rhabdoid clear-cell renal cell carcinoma (n=37) was assessed by RNA-seq and gene expression microarray. VHL (63%) showed identical mutations in all regions from the same tumor. BAP1 (38%) and PBRM1 (13%) mutations were identified in the rhabdoid but not in the epithelioid component and were mutually exclusive in 3/3 cases and 1 case, respectively. SETD2 (63%) mutations were discordant between different histologic regions in 2/5 cases, with mutations called only in the epithelioid and rhabdoid components, respectively. The transcriptome of rhabdoid clear-cell renal cell carcinoma was distinct from advanced-stage and high-grade clear-cell renal cell carcinoma. The diverse histologic components of rhabdoid clear-cell renal cell carcinoma, however, showed a similar transcriptomic program, including a similar prognostic gene expression signature. Rhabdoid clear-cell renal cell carcinoma is transcriptomically distinct and shows a high rate of SETD2 and BAP1 mutations and a low rate of PBRM1 mutations. Driver mutations in clear-cell renal cell carcinoma are often discordant across different morphologic regions, whereas the gene expression program is relatively stable. Molecular profiling of clear-cell renal cell carcinoma may improve by assessing for gene expression and sampling tumor foci from different histologic regions.

Culture of mouse amniotic fluid-derived cells on irradiated STO feeders results in the generation of primitive endoderm cell lines capable of self-renewal in vitro.

The cells present in amniotic fluid (AF) are currently used for prenatal diagnosis of fetal anomalies but are also a potential source of cells for cell therapy. To better characterize putative progenitor cell populations present in AF, we used culture conditions that support self-renewal to determine if these promoted the generation of stable cell lines from AF-derived cells (AFC). Cells isolated from E11.5 mouse were cultured on irradiated STO fibroblast feeder layers in human embryonic germ cell derivation conditions. The cultures grew multicellular epithelial colonies that could be repropagated from single cells. Reverse transcription semiquantitative polymerase chain reaction of established cell lines revealed that they belonged to the extraembryonic endoderm (ExEn) expressing high levels of Gata6, Gata4, Sox17, Foxa2 and Sox7 mRNA. Hierarchical clustering based on the whole transcriptome expression profile of the AFC lines (AFCL) shows significant correlation between transcription profiles of AFCL and blastocyst-derived XEN, an ExEn cell line. In vitro differentiation of AFCL results in the generation of cells expressing albumin and α-fetoprotein (AFP), while intramuscular injection of AFCL into immunodeficient mice produced AFP-positive tumors with primitive endodermal appearance. Hence, E11.5 mouse AF contains cells that efficiently produce XEN lines. These AF-derived XEN lines do not spontaneously differentiate into embryonic-type cells but are phenotypically stable and have the capacity for extensive expansion. The lack of requirement for reprogramming factors to turn AF-derived progenitor cells into stable cell lines capable of massive expansion together with the known ability of ExEn to contribute to embryonic tissue suggests that this cell type may be a candidate for banking for cell therapies.

A subtraction mechanism of temporal coding in cerebellar cortex.

The temporally specific learning displayed by the cerebellum facilitates mechanistic analysis of neural timing and temporal coding. We report evidence for a subtraction-like mechanism of temporal coding in cerebellar cortex in which activity in a subset of granule cells specifically codes the interval between the offset of two mossy fiber inputs. In a large-scale cerebellar simulation, cessation of one of two ongoing mossy fiber inputs produces a robust temporal code in the population of granule cells. This activity supports simulation learning in response to temporal patterns of stimuli, even when those same stimuli do not support learning when presented individually. Using stimulation of mossy fiber inputs to the cerebellum as training stimuli in rabbits, we confirmed these unusual predictions in a cerebellum-dependent form of learning. Analysis of the simulations reveals a specific working hypothesis for this temporal subtraction process that involves interactions between granule cells and the inhibitory Golgi cells. The results suggest how feedforward inhibition, such as that present in the cerebellar cortex, can contribute to temporal coding.

A Validation Framework for Somatic Copy Number Detection in Targeted Sequencing Panels.

Somatic copy number alterations (SCNAs) in tumors are clinically significant diagnostic, prognostic, and predictive biomarkers. SCNA detection from targeted next-generation sequencing panels is increasingly common in clinical practice; however, detailed descriptions of optimization and validation of SCNA pipelines for small targeted panels are limited. This study describes the validation and implementation of a tumor-only SCNA pipeline using CNVkit, augmented with custom modules and optimized for clinical implementation by testing reference materials and clinical tumor samples with different classes of copy number variation (CNV; amplification, single copy loss, and biallelic loss). Using wet-bench and in silico methods, various parameters impacting CNV calling, including assay-intrinsic variables (establishment of normal reference and sequencing coverage), sample-intrinsic variables (tumor purity and sample quality), and CNV algorithm-intrinsic variables (bin size), were optimized. The pipeline was trained and tested on an optimization cohort and validated using an independent cohort with a sensitivity and specificity of 100% and 93%, respectively. Using custom modules, intragenic CNVs with breakpoints within tumor suppressor genes were uncovered. Using the validated pipeline, re-analysis of 28 pediatric solid tumors that had been previously profiled for mutations identified SCNAs in 86% (24/28) samples, with 46% (13/28) samples harboring findings of potential clinical relevance. Our report highlights the importance of rigorous establishment of performance characteristics of SCNA pipelines and presents a detailed validation framework for optimal SCNA detection in targeted sequencing panels.

A decrementing form of plasticity apparent in cerebellar learning.

Long-term synaptic plasticity is believed to underlie the capacity for learning and memory. In the cerebellum, for example, long-term plasticity contributes to eyelid conditioning and to learning in eye movement systems. We report evidence for a decrementing form of cerebellar plasticity as revealed by the behavioral properties of eyelid conditioning in the rabbit. We find that conditioned eyelid responses exhibit within-session changes that recover by the next day. These changes, which increase with the interstimulus interval, involve decreases in conditioned response magnitude and likelihood as well as increases in latency to onset. Within-subject comparisons show that these changes differ in magnitude depending on the type of training, arguing against motor fatigue or changes in motor pathways downstream of the cerebellum. These phenomena are also observed when stimulation of mossy fibers substitutes for the conditioned stimulus, suggesting that changes take place within the cerebellum or in downstream efferent pathways. Together, these observations suggest a plasticity mechanism in the cerebellum that is induced during training sessions and fades within 23 h. To formalize this hypothesis more specifically, we show that incorporating a short-lasting potentiation at the granule cell to Purkinje cell synapses in a computer simulation of the cerebellum reproduces these behavioral effects. We propose the working hypothesis that the presynaptic form of long-term potentiation observed at these synapses is reversed by time rather than by a corresponding long-term depression. These results demonstrate the utility of eyelid conditioning as a means to identify and characterize the rules that govern input to output transformations in the cerebellum.

Theta modulation in the medial and the lateral entorhinal cortices.

Hippocampal neurons show a strong modulation by theta frequency oscillations. This modulation is thought to be important not only for temporal encoding and decoding of information in the hippocampal system, but also for temporal ordering of neuronal activities on timescales at which physiological mechanisms of synaptic plasticity operate. The medial entorhinal cortex (MEC), one of the two major cortical inputs to the hippocampus, is known to show theta modulation. Here, we show that the local field potentials (LFPs) in the other major cortical input to the hippocampus, the lateral entorhinal cortex (LEC), show weaker theta oscillations than those shown in the MEC. Neurons in LEC also show weaker theta modulation than that of neurons in MEC. These findings suggest that LEC inputs are integrated into hippocampal representations in a qualitatively different manner than the MEC inputs. Furthermore, MEC grid cells increase the scale of their periodic spatial firing patterns along the dorsoventral axis, corresponding to the increasing size of place fields along the septotemporal axis of the hippocampus. We show here a corresponding gradient in the tendency of MEC neural firing to skip alternate theta cycles. We propose a simple model based on interference of delta oscillations with theta oscillations to explain this behavior.

Germline POLE mutation in a child with hypermutated medulloblastoma and features of constitutional mismatch repair deficiency.

Ultra-hypermutation (>100 mutations/Mb) is rare in childhood cancer genomes and has been primarily reported in patients with constitutional mismatch repair deficiency (CMMRD) caused by biallelic germline mismatch repair (MMR) gene mutations. We report a 5-yr-old child with classic clinical features of CMMRD and an ultra-hypermutated medulloblastoma with retained MMR protein expression and absence of germline MMR mutations. Mutational signature analysis of tumor panel sequencing data revealed a canonical DNA polymerase-deficiency-associated signature, prompting further genetic testing that uncovered a germline POLE p.A456P missense variant, which has previously been reported as a recurrent somatic driver mutation in cancers. This represents the earliest known onset of malignancy in a patient with a germline mutation in the POLE proofreading polymerase. The clinical features in this child, virtually indistinguishable from those of CMMRD, suggest that polymerase-proofreading deficiency should be considered in the differential diagnosis of CMMRD patients with retained MMR function.

Effect of Brain Tumor Presence During Radiation on Tissue Toxicity: Transcriptomic and Metabolic Changes.

PURPOSE: Radiation therapy (RT) causes functional and transcriptomic changes in the brain; however, most studies have been carried out in normal rodent brains. Here, the long-term effect of irradiation and tumor presence during radiation was investigated. METHODS AND MATERIALS: Male Wistar rats ∼7 weeks old were divided into 3 groups: sham implant, RT+sham implant, and RT+tumor implant (C6 glioma). Hypofractionated irradiation (8 or 6 Gy/day for 5 days) was localized to a 1-cm strip of cranium starting 5 days after implantation, resulting in complete tumor regression and prolonged survival. Biopsy of tissue was performed in the implant area 65 days after implantation. RNA was hybridized to GeneChip Rat Exon 1.0 ST array. Data were analyzed using significant analysis of microarrays and ingenuity pathway analysis. 1H magnetic resonance spectroscopy (1H-MRS) imaging was performed in the implantation site 65 to 70 days after implantation using a 9.4 T Biospec magnetic resonance imaging scanner with a quadrature rat brain array. Immunohistochemical staining for astrogliosis, HMG-CoA synthase 2, γ-aminobutyric acid (GABA) and taurine was performed at ∼65 days after implantation. RESULTS: Eighty-four genes had a false discovery rate <3.5%. We compared RT+tumor implant with RT+sham implant animals. The tumor presence affected networks associated with cancer/cell morphology/tissue morphology. 1H-MRS showed significant reduction in taurine levels (P<.04) at the implantation site in both groups. However, the RT+tumor group also showed significant increase in levels of neurotransmitter GABA (P=.02). Hippocampal taurine levels were only significantly reduced in the RT+tumor group (P=.03). HMG-CoA synthase 2, GABA and taurine levels were confirmed using staining. Glial fibrillary acidic protein staining demonstrated a significant increase in inflammation that was heightened in the RT+tumor group. CONCLUSIONS: Our data indicate that tumor presence during radiation significantly affects long-term functional transcriptomics landscape and neurotransmitter levels at the tumor implantation site/normal tissue, accompanied by increased inflammation (astrogliosis).

High-resolution Single Particle Analysis from Electron Cryo-microscopy Images Using SPHIRE.

SPHIRE (SPARX for High-Resolution Electron Microscopy) is a novel open-source, user-friendly software suite for the semi-automated processing of single particle electron cryo-microscopy (cryo-EM) data. The protocol presented here describes in detail how to obtain a near-atomic resolution structure starting from cryo-EM micrograph movies by guiding users through all steps of the single particle structure determination pipeline. These steps are controlled from the new SPHIRE graphical user interface and require minimum user intervention. Using this protocol, a 3.5 Å structure of TcdA1, a Tc toxin complex from Photorhabdus luminescens, was derived from only 9500 single particles. This streamlined approach will help novice users without extensive processing experience and a priori structural information, to obtain noise-free and unbiased atomic models of their purified macromolecular complexes in their native state.

Biphasic components of sarcomatoid clear cell renal cell carcinomas are molecularly similar to each other, but distinct from, non-sarcomatoid renal carcinomas.

Sarcomatoid transformation, wherein an epithelioid carcinomatous tumour component coexists with a sarcomatoid histology, is a predictor of poor prognosis in clear cell renal cell carcinoma. Our understanding of sarcomatoid change has been hindered by the lack of molecular examination. Thus, we sought to characterize molecularly the biphasic epithelioid and sarcomatoid components of sarcomatoid clear cell renal cell carcinoma and compare them to non-sarcomatoid clear cell renal cell carcinoma. We examined the transcriptome of the epithelioid and sarcomatoid components of advanced stage sarcomatoid clear cell renal cell carcinoma (n=43) and non-sarcomatoid clear cell renal cell carcinoma (n=37) from independent discovery and validation cohorts using the cDNA microarray and RNA-seq platforms. We analyzed DNA copy number profiles, generated using SNP arrays, from patients with sarcomatoid clear cell renal cell carcinoma (n=10) and advanced non-sarcomatoid clear cell renal cell carcinoma (n=155). The epithelioid and sarcomatoid components of sarcomatoid clear cell renal cell carcinoma had similar gene expression and DNA copy number signatures that were, however, distinct from those of high-grade, high-stage non-sarcomatoid clear cell renal cell carcinoma. Prognostic clear cell renal cell carcinoma gene expression profiles were shared by the biphasic components of sarcomatoid clear cell renal cell carcinoma and the sarcomatoid component showed a partial epithelial-to-mesenchymal transition signature. Our genome-scale microarray-based transcript data were validated in an independent set of sarcomatoid and non-sarcomatoid clear cell renal cell carcinomas using RNA-seq. Sarcomatoid clear cell renal cell carcinoma is molecularly distinct from non-sarcomatoid clear cell renal cell carcinoma, with its genetic programming largely shared by its biphasic morphological components. These data explain why a low percentage of sarcomatoid histology augurs a poor prognosis; suggest the need to modify the pathological grading system and introduce the potential for candidate biomarkers to detect sarcomatoid change preoperatively without specifically sampling the histological sarcomatoid component.

Hierarchical cognitive maps.

We describe a computational model of spatial navigation based on experimental studies conducted with human participants. The model builds and uses a hierarchical cognitive map of a large environment. Computer simulations show that the model correctly describes experimental results including hierarchical organization of space and distance estimation. Furthermore, the model predicts that reaction time for distance estimation varies nonlinearly with distance.

Silencing BMI1 eliminates tumor formation of pediatric glioma CD133+ cells not by affecting known targets but by down-regulating a novel set of core genes.

Clinical outcome of children with malignant glioma remains dismal. Here, we examined the role of over-expressed BMI1, a regulator of stem cell self-renewal, in sustaining tumor formation in pediatric glioma stem cells. Our investigation revealed BMI1 over-expression in 29 of 54 (53.7%) pediatric gliomas, 8 of 8 (100%) patient derived orthotopic xenograft (PDOX) mouse models, and in both CD133+ and CD133- glioma cells. We demonstrated that lentiviral-shRNA mediated silencing of suppressed cell proliferation in vitro in cells derived from 3 independent PDOX models and eliminated tumor-forming capacity of CD133+ and CD133- cells derived from 2 PDOX models in mouse brains. Gene expression profiling showed that most of the molecular targets of BMI1 ablation in CD133+ cells were different from that in CD133- cells. Importantly, we found that silencing BMI1 in CD133+ cells derived from 3 PDOX models did not affect most of the known genes previously associated with the activated BMI1, but modulated a novel set of core genes, including RPS6KA2, ALDH3A2, FMFB, DTL, API5, EIF4G2, KIF5c, LOC650152, C20ORF121, LOC203547, LOC653308, and LOC642489, to mediate the elimination of tumor formation. In summary, we identified the over-expressed BMI1 as a promising therapeutic target for glioma stem cells, and suggest that the signaling pathways associated with activated BMI1 in promoting tumor growth may be different from those induced by silencing BMI1 in blocking tumor formation. These findings highlighted the importance of careful re-analysis of the affected genes following the inhibition of abnormally activated oncogenic pathways to identify determinants that can potentially predict therapeutic efficacy.

Mouse Lymphoblastic Leukemias Induced by Aberrant Prdm14 Expression Demonstrate Widespread Copy Number Alterations Also Found in Human ALL.

Aberrant expression and activation of oncogenes in somatic cells has been associated with cancer initiation. Required for reacquisition of pluripotency in the developing germ cell, PRDM14 initiates lymphoblastic leukemia when misexpressed in murine bone marrow. Activation of pluripotency in somatic cells can lead to aneuploidy and copy number alterations during iPS cell generation, and we hypothesized that PRDM14-induced lymphoblastic leukemias would demonstrate significant chromosomal damage. High-resolution oligo array comparative genomic hybridization demonstrated infrequent aneuploidy but frequent amplification and deletion, with amplifications occurring in a 5:1 ratio with deletions. Many deletions (i.e., Cdkn2a, Ebf1, Pax5, Ikzf1) involved B-cell development genes and tumor suppressor genes, recapitulating deletions occurring in human leukemia. Pathways opposing senescence were frequently deactivated via Cdkn2a deletion or Tbx2 amplification, with corollary gene expression. Additionally, gene expression studies of abnormal pre-leukemic B-precursors showed downregulation of genes involved in chromosomal stability (i.e., Xrcc6) and failure to upregulate DNA repair pathways. We propose a model of leukemogenesis, triggered by pluripotency genes like Prdm14, which involves ongoing DNA damage and failure to activate non-homologous end-joining secondary to aberrant gene expression.

Global gene expression profiling confirms the molecular fidelity of primary tumor-based orthotopic xenograft mouse models of medulloblastoma.

We previously showed that primary tumor-based orthotopic xenograft mouse models of medulloblastoma replicated the histopathological phenotypes of patients' original tumors. Here, we performed global gene expression profiling of 11 patient-specific xenograft models to further determine whether the xenograft tumors were molecularly accurate during serial subtransplantations in mouse brains and whether they represented all the molecular subtypes of medulloblastoma that were recently described. Analysis of the transcriptomes of 9 pairs of matched passage I xenografts and patients' tumors revealed high correlation coefficients (r(2) > 0.95 in 5 models, > 0.9 in 3 models, and > 0.85 in 1 model) and only identified 69 genes in which expressions were altered (FDR = 0.0023). Subsequent pair-wise comparisons between passage I, III, and V xenografts from the 11 models further showed that no dramatic alterations were introduced (r(2) > 0.9 in 8 models and > 0.8 in 3 models). The genetic abnormalities of each model were then identified through comparison with control RNAs from 5 normal cerebella and 2 fetal brains. Hierarchical clustering using 3 previously published molecular signatures showed that our models span the whole spectrum of molecular subtypes, including SHH (n = 2), WNT (n = 2), and the most recently identified group C (n = 4) and group D (n = 3). In conclusion, we demonstrated that the 11 orthotopic medulloblastoma xenograft models were molecularly faithful to the primary tumors, and our comprehensive collection of molecularly distinct animal models should serve as a valuable resource for the development of new targeted therapies for medulloblastoma.

A clinically relevant orthotopic xenograft model of ependymoma that maintains the genomic signature of the primary tumor and preserves cancer stem cells in vivo.

Limited availability of in vitro and in vivo model systems has hampered efforts to understand tumor biology and test novel therapies for ependymoma, the third most common malignant brain tumor that occurs in children. To develop clinically relevant animal models of ependymoma, we directly injected a fresh surgical specimen from a 9-year-old patient into the right cerebrum of RAG2/severe complex immune deficiency (SCID) mice. All five mice receiving the initial transplantation of the patient tumor developed intracerebral xenografts, which have since been serially subtransplanted in vivo in mouse brains for 4 generations and can be cryopreserved for long-term maintenance of tumorigenicity. The xenograft tumors shared nearly identical histopathological features with the original tumors, harbored 8 structural chromosomal abnormalities as detected with spectral karyotyping, maintained gene expression profiles resembling that of the original patient tumor with the preservation of multiple key genetic abnormalities commonly found in human ependymomas, and contained a small population (<2.2%) of CD133(+) stem cells that can form neurospheres and display multipotent capabilities in vitro. The permanent cell line (BXD-1425EPN), which was derived from a passage II xenograft tumor and has been passaged in vitro more than 70 times, expressed similar differentiation markers of the xenograft tumors, maintained identical chromosomal abnormalities, and formed tumors in the brains of SCID mice. In conclusion, direct injection of primary ependymoma tumor cells played an important role in the generation of a clinically relevant mouse model IC-1425EPN and a novel cell line, BXD-1425EPN. This cell line and model will facilitate the biological studies and preclinical drug screenings for pediatric ependymomas.

Latent learning, shortcuts and detours: a computational model.

Voicu and Schmajuk (Rob. Auto. Syst. 35 (2001a) 23) described a model of spatial navigation and exploration that includes an action system capable of guiding, with the help of a cognitive system, the search for specific goals as determined by a motivation system. Whereas in the original model the cognitive map stores information about the connectivity between places in the environment, in the present version the cognitive map also stores information about the paths traversed by the agent. Computer simulations show that the network correctly describes experimental results including latent learning in a maze, detours in a maze, and shortcuts in an open field. In addition, the model generates novel predictions about detours and shortcuts in an open field.