Intracranial Transplantation Research Articles

Therapeutic Effects of Stem Cells and Substrate Reduction in Juvenile Sandhoff Mice

Sandhoff Disease (SD) involves the CNS accumulation of ganglioside GM2 and asialo-GM2 (GA2) due to inherited defects in the β-subunit gene of β-hexosaminidase A and B (Hexb gene). Substrate reduction therapy, utilizing imino sugar N-butyldeoxygalactonojirimycin (NB-DGJ), reduces ganglioside biosynthesis and levels of stored GM2 in SD mice. Intracranial transplantation of Neural Stem Cells (NSCs) can provide enzymatic cross correction, to help reduce ganglioside storage and extend life. Here we tested the effect of NSCs and NB-DGJ, alone and together, on brain β-hexosaminidase activity, GM2, and GA2 content in juvenile SD mice. The SD mice received either cerebral NSC transplantation at post-natal day 0 (p-0), intraperitoneal injection of NB-DGJ (500mg/kg/day) from p-9 to p-15, or received dual treatments. The brains were analyzed at p-15. β-galactosidase staining confirmed engraftment of lacZ-expressing NSCs in the cerebral cortex. Compared to untreated and sham-treated SD controls, NSC treatment alone provided a slight increase in Hex activity and significantly decreased GA2 content. However, NSCs had no effect on GM2 content when analyzed at p-15. NB-DGJ alone had no effect on Hex activity, but significantly reduced GM2 and GA2 content. Hex activity was slightly elevated in the NSC+drug-treated mice. GM2 and GA2 content in the dual treated mice were similar to that of the NB-DGJ treated mice. These data indicate that NB-DGJ alone was more effective in targeting storage in juvenile SD mice than were NSCs alone. No additive or synergistic effect between NSC and drug was found in these juvenile SD mice.

In Vivo Generation of Neural Tumors from Neoplastic Pluripotent Stem Cells Models Early Human Pediatric Brain Tumor Formation

Recent studies have identified gene signatures in malignant tumors that are associated with human embryonic stem cells, suggesting a molecular relationship between aggressive cancers and pluripotency. Here, we characterize neural precursors (NPs) derived from transformed human embryonic stem cells (N-t-hESCs) that exhibit neoplastic features of human brain tumors. NPs derived from t-hESCs have enhanced cell proliferation and an inability to mature toward the astrocytic lineage, compared with progeny derived from normal human embryonic stem cells (N-hESCs) independent of adherent or neurosphere outgrowth. Intracranial transplantation of NPs derived from N-t-hESCs and N-hESCs into NOD SCID mice revealed development of neuroectoderm tumors exclusively from the N-t-hESCs NPs and not from normal N-hESCs. These tumors infiltrated the ventricles and the cerebellum of recipient mice and displayed morphological, phenotypic, and molecular features associated with classic medulloblastoma including retention of a pluripotent signature. Importantly, N-t-hESCs did not exhibit cytogenetic changes associated with medulloblastoma, suggesting that aberrant cellular and molecular properties precede the acquisition of karyotypic changes thus underscoring the value of this model system of human medulloblastoma. Our study demonstrates that NPs from a starting population of neoplastic human pluripotent parent cells possess brain tumor-initiating cell capacity, thereby providing a model system to investigate initiation and progression of primitive human neural cancers that are difficult to assess using somatic sources.

428: Intracranial stem cell transplantation with and without erythropoietin facilitates motor recovery in a perinatal rat brain injury model

Peripartal brain injury in the premature infant leads to a large spectrum of clinical problems including lifelong neurodevelopmental deficits. We examined intracranial mesenchymal stem cells (MSC) transplantation as a promising therapy to facilitate motor recovery after perinatal hypoxic-ischemic brain damage with inflammatory component in a perinatal rat model.

Comparative morphological and immunohistochemical study of human meningioma after intracranial transplantation into nude mice

Although surgical resection of benign human meningiomas is the primary goal, in case of relapse or when they are not fully resectable, other strategies including chemotherapeutical treatment would be appropriate. The initial evaluation of chemotherapeutical agents requires an appropriate tumor model, where the natural characteristics of the original benign tumor is reflected. We here tested, whether primary cell cultures of benign human meningiomas would reliably grow after intracranial transplantation into mice, and whether they would show histomorphological and immunohistochemical characteristics of the original human tumor. Cells of 11 benign human meningiomas were transplanted into the prefrontal cortex of nude mice. After 3 months, the mice were sacrificed and their brains were histologically processed for morphological characterization and measurement of tumor volume. Additionally, the proliferation index (PI), the microvessel density, and epithelial membrane antigen (EMA) were compared between human meningiomas and tumors grown in mice by using immunohistochemical methods. Further, cyclooxygenase-2 (COX-2) expression, a possible target for pharmacological manipulation, was examined. The results showed in almost all mice (93%) a tumor formation with meningothelial histomorphology comparable to the original human tumors. The PI, vascular density and COX-2 expression were similar between human and mice meningiomas, but EMA expression was reduced in mice (P<0.01). In conclusion an implantation of benign human meningioma primary cell cultures in mice reliably results in tumor formation with morphological and immunohistological features comparable to the original human tumor. This model may therefore be suitable to test novel therapeutic agents.

The perils of proxy outcomes in evidence-based medicice: the case of recombinant Factor VIIa

There is currently a major controversy surrounding the off-label use of recombinant activated Factor VII (rFVIIa). The literature offers at best inconclusive support for its use in the management of bleeding in any situation other than in patients with Factor VII deficiency or hemophilia A or B with antibodies to factor VIII or factor IX respectively. This evidence has not, however, slowed the growth in off-label prescribing of rFVIIa in other situations including intracranial hemorrhage, cardiac surgery, trauma, transplantation and prostatectomy. We argue that the controversy surrounding such off-label use of rFVIIa stems in part from different understandings of the purposes and methods of evidence-based medicine (EBM)—in particular the use of gross, proxy outcomes such as mortality to determine the effectiveness of an intervention. We then argue that clinical impression and expert opinion should not be dismissed outright on the basis of inconclusive evidence of ineffectiveness. Those who advocate its continued use, however, have a responsibility to demonstrate the benefits that may be missed in current manifestations of EBM . We suggest that the only justified verdict in the case of rFVIIa is currently 'Not Proven'.

Intracranial transplantation of monocyte‐derived multipotential cells enhances recovery after ischemic stroke in rats

Cell transplantation has emerged as a potential therapy to reduce the neurological deficits caused by ischemic stroke. We previously reported a primitive cell population, monocyte-derived multipotential cells (MOMCs), which can differentiate into mesenchymal, neuronal, and endothelial lineages. In this study, MOMCs and macrophages were prepared from rat peripheral blood and transplanted intracranially into the ischemic core of syngeneic rats that had undergone a left middle cerebral artery occlusion procedure. Neurological deficits, as evaluated by the corner test, were less severe in the MOMC-transplanted rats than in macrophage-transplanted or mock-treated rats. Histological evaluations revealed that the number of microvessels that had formed in the ischemic boundary area by 4 weeks after transplantation was significantly greater in the MOMC-transplanted rats than in the control groups. The blood vessel formation was preceded by the appearance of round CD31(+) cells, which we confirmed were derived from the transplanted MOMCs. Small numbers of bloodvessels incorporating MOMC-derived endothelial cells expressing a mature endothelial marker RECA-1 were detected at 4 weeks after transplantation. In addition, MOMCs expressed a series of angiogenic factors, including vascular endothelial growth factor, angiopoetin-1, and placenta growth factor (PlGF). These findings provide evidence that the intracranial delivery of MOMCs enhances functional recovery by promoting neovascularization in a rat model for ischemic stroke.

Anesthesia for cerebral revascularization for adult moyamoya syndrome associated with sickle cell disease

We report the anesthetic management of an adult patient undergoing cerebral revascularization surgery for moyamoya syndrome complicating sickle-cell disease (SCD). We present a 25 year-old male of African ethnicity with homozygous SCD who was experiencing worsening ischemic neurologic symptoms culminating in intraventricular hemorrhage from rupture of moyamoya vessels. Despite an extracranial–intracranial superficial temporal artery–middle cerebral artery bypass that was angiographically patent postoperatively, he subsequently required an intracranial omental transplant to improve cerebral blood flow to the anterior cerebral artery territory. Prior to both cerebral revascularization procedures, the patient had continued with his regularly scheduled red blood cell exchange transfusion. The importance of normothermia, normocarbia, normotension, and normovolemia is emphasized in the neuroanesthetic management. We conclude that the safe and efficacious operative treatment of moyamoya disease, using both direct and indirect revascularization procedures, is being increasingly described, and therefore anesthesiologists are likely to encounter similar cases in the future and need to be aware of the surgical procedures and perioperative implications. The overall principles of safe anesthesia (normotension, normocarbia, good oxygenation, normothermia, normovolemia) for patients with SCD also applies to patients with moyamoya. During a craniotomy, certain deviations from these are needed (hyperventilation and mannitol diuresis for brain volume reduction, induced hypothermia or manipulations of arterial blood pressures) but they can be safely used with careful monitoring of the patient.

Neural Stem Cell Transplantation as a Therapeutic Approach for Treating Lysosomal Storage Diseases

Treating the central nervous system manifestations of subjects with neuropathic lysosomal storage diseases remains a major technical challenge. This is because of the low efficiency by which lysosomal enzymes in systemic circulation are able to traverse the blood brain barrier into the central nervous system. Intracranial transplantation of neural stems cells genetically modified to overexpress the respective deficient enzymes represents a potential approach to addressing this group of diseases. The unique properties of neural stem cells and progenitor cells, such as their ability to migrate to distal sites, differentiate into various cell types and integrate within the host brain without disrupting normal function, making them particularly attractive therapeutic agents. In addition, neural stem cells are amenable to ex vivo propagation and modification by gene transfer vectors. In this regard, transplanted cells can serve not only as a source of lysosomal enzymes but also as a means to potentially repair the injured brain by replenishing the organ with healthy cells and effecting the release of neuroprotective factors. This review discusses some of the well-characterized neural stem cell types and their possible use in treating neuropathic lysosomal storage diseases such as the Niemann Pick A disease.

Mesenchymal stem cell transplantation ameliorates motor function deterioration of spinocerebellar ataxia by rescuing cerebellar Purkinje cells

BackgroundSpinocerebellar ataxia (SCA) refers to a disease entity in which polyglutamine aggregates are over-produced in Purkinje cells (PCs) of the cerebellum as well as other neurons in the central nervous system, and the formation of intracellular polyglutamine aggregates result in the loss of neurons as well as deterioration of motor functions. So far there is no effective neuroprotective treatment for this debilitating disease although numerous efforts have been made. Mesenchymal stem cells (MSCs) possess multi-lineage differentiation potentials as well as immuno-modulatory properties, and are theoretically good candidates for SCA treatment. The purpose of this study is to investigate whether transplantation of human MSCs (hMSCs) can rescue cerebellar PCs and ameliorate motor function deterioration in SCA in a pre-clinical animal model.MethodTransgenic mice bearing poly-glutamine mutation in ataxin-2 gene (C57BL/6J SCA2 transgenic mice) were serially transplanted with hMSCs intravenously or intracranially before and after the onset of motor function loss. Motor function of mice was evaluated by an accelerating protocol of rotarod test every 8 weeks. Immunohistochemical stain of whole brain sections was adopted to demonstrate the neuroprotective effect of hMSC transplantation on cerebellar PCs and engraftment of hMSCs into mice brain.ResultsIntravenous transplantation of hMSCs effectively improved rotarod performance of SCA2 transgenic mice and delayed the onset of motor function deterioration; while intracranial transplantation failed to achieve such neuroprotective effect. Immunohistochemistry revealed that intravenous transplantation was more effective in the preservation of the survival of cerebellar PCs and engraftment of hMSCs than intracranial injection, which was compatible to rotarod performance of transplanted mice.ConclusionIntravenous transplantation of hMSCs can indeed delay the onset as well as improve the motor function of SCA2 transgenic mice. The results of this preclinical study strongly support further exploration of the feasibility to transplant hMSCs for SCA patients.

Nonredundant Functions for Akt Isoforms in Astrocyte Growth and Gliomagenesis in an Orthotopic Transplantation Model

The AKT family, comprising three highly homologous kinases, is an essential mediator of the PTEN/PI3K pathway, which is deregulated in many human cancers. A thorough understanding of the specific activities of each isoform in normal and disease tissues is lacking. We evaluated the role of each Akt isoform in gliomagenesis by using a model system driven by common glioma abnormalities, loss of function of p53 and Pten, and expression of EGFRvIII. Both Pten deletion and EGFRvIII expression accelerated the proliferation of p53-null primary murine astrocytes. All three Akt isoforms were expressed and phosphorylated in astrocytes, with significantly higher activation in Pten-null cells. Despite substantial compensation in many contexts when individual Akt isoforms were inhibited, isoform-specific effects were also identified. Specifically, loss of Akt1 or Akt2 decreased proliferation of Pten wild-type astrocytes, whereas combined loss of multiple isoforms was needed to inhibit proliferation of Pten-null astrocytes. In addition, Akt3 was required for anchorage-independent growth of transformed astrocytes and human glioma cells, and Akt3 loss inhibited invasion of transformed astrocytes. EGFRvIII expression transformed p53-null astrocytes with or without Pten deletion, causing rapid development of high-grade astrocytoma on intracranial transplantation. Furthermore, tumorigenesis of Pten;p53-null astrocytes expressing EGFRvIII was delayed by Akt1 loss and accelerated by Akt2 loss. Taken together, these results indicate context-dependent roles for individual Akt isoforms and suggest that there may be heterogeneous tumor response to isoform-specific inhibitors.

Abstract 3402: Up-regulation and invasive role of MUC18/MCAM in glioma sphere culture

Abstract Glioblastomas are characterized by aggressive invasion into normal brain tissues, thus limiting complete removal by surgical resection resulting in recurrence. Our identified molecule involved in glioma invasion is important for developing targeted effective therapies in the malignant glioma. Cancer stem cells are capable for self-renewing and are thought to be responsible for tumor metastasis and dissemination. However, the invasive activities of cancer stem cells have not been fully understood. To investigate a role of glioma stem cells in the tumor invasion, tumor sphere culture was established from mouse glioma RSV-M cells. The high tumorigeneity and aggressive migration of RSV-M tumor sphere were observed in intracranial transplantation. Quantitative RT-PCR analysis showed the enhanced expression of metastasis-related genes in RSV-M tumor sphere. Of these, MUC18/MCAM also highly expressed in tumor sphere derived from human glioma cell lines. Importantly, CD133-positive fraction of glioma sphere culture isolated from glioblastoma patients contained a cell population expressing MUC18 but CD133-negative fraction not. To examine a function of MUC18 in glioma cells, the expression or knockdown construct for the gene were introduced into human glioma cells. Over-expression of MUC18 in the glioma cells increased their invasive capacity and clonogenicity in comparison to the parent or knockdown cells. In addition, expression of some metastasis-related genes was up-regulated in MUC18-over-expressing cells. These results suggest that enhanced MUC18 expression of glioma stem cells may contribute to the aggressive invasion of this tumor. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3402. doi:10.1158/1538-7445.AM2011-3402

Frizzled 4 Regulates Stemness and Invasiveness of Migrating Glioma Cells Established by Serial Intracranial Transplantation

One of the most detrimental hallmarks of glioblastoma multiforme (GBM) is cellular invasiveness, which is considered a potential cause of tumor recurrence. Infiltrated GBM cells are difficult to completely eradicate surgically and with local therapeutic modalities. Although much effort has focused on understanding the various mechanisms controlling GBM invasiveness, its nature remains poorly understood. In this study, we established highly serial intracranial transplantation. U87R4 cells were highly invasive and displayed stem cell-like properties, as compared to noninvasive but proliferative U87L4 cells. Microarray analysis during serial transplantation revealed that apoptosis-inducing genes (caspase3 and PDCD4) were downregulated whereas several cancer stem cell-relevant genes [Frizzled 4 (FZD4) and CD44] were upregulated in more invasive cells. U87R4 cells were resistant to anticancer drug-induced cell death, partly due to downregulation of caspase3 and PDCD4, and they retained activated Wnt/β-catenin signaling due to upregulation of Frizzled 4, which was sufficient to control neurosphere formation. We also found that FZD4 promoted expression of the epithelial to mesenchymal transition regulator SNAI1, along with acquisition of a mesenchymal phenotype. Taken together, our results argue that Frizzled 4 is a member of the Wnt signaling family that governs both stemness and invasiveness of glioma stem cells, and that it may be a major cause of GBM recurrence and poor prognosis.

Long-Term Safety and Efficacy of Deferasirox (Exjade®) In Transfused Patients with Sickle Cell Disease Treated for up to 5 Years

AbstractAbstract 845 Background:Transfusion therapy can effectively treat many complications associated with sickle cell disease (SCD), but iron overload will develop without iron chelation therapy. Despite long-term transfusion requirements, long-term data for iron chelation in SCD are limited. The oral iron chelator, deferasirox, effectively reduced iron burden in SCD patients aged ≥2 years during 1 year of treatment (Vichinsky et al. Br J Haematol 2007). This 5-year follow-up is the first report of long-term deferasirox treatment in SCD patients. Methods:Eligible patients that completed the 1-year core study (randomized to deferasirox [deferasirox cohort] or deferoxamine [crossover cohort]) entered the 4- year extension. All received deferasirox while continuing their regular transfusion regimen. Deferasirox dose in the extension was initially based on serum ferritin (SF) trends in the core study (deferasirox cohort) or transfusion requirements (crossover cohort). Dose adjustments were based on monthly SF and safety assessments (investigator-reported adverse events [AEs] and centrally processed lab parameters). Growth and sexual development (Tanner staging) were assessed annually. Results:Of the patients in the deferasirox (n=132) and crossover (n=53) cohorts that received '1 deferasirox dose during the core or extension, 43 (33%) and 19 (36%) patients, respectively, completed the extension. Reasons for discontinuation included withdrawal of consent (n=44, 23.8%), loss to follow-up (n=17, 9.2%) and AEs (n=14, 7.6%). Three deaths occurred, all in the extension (intraventricular hemorrhage, n=2; intracranial bleed post liver transplantation, n=1); none considered by investigators to be deferasirox-related. Mean dose during the study was 18.7 ± 6.5 and 21.2 ± 5.3 mg/kg/day in the deferasirox and crossover cohorts, respectively. Investigator-assessed drug-related AEs (≥5% overall) included nausea (14.6%), diarrhea (10.8%), increased blood creatinine (5.9%) and vomiting (5.4%). Generally, these AEs were manageable and transient, and decreased in frequency over time. Serious AEs were reported in 70.8% of patients overall and were mostly related to the underlying disease. Serious investigator-assessed drug-related AEs were reported in 8 patients (6.1%) in the deferasirox cohort and 1 patient (1.9%) in the crossover cohort. In the deferasirox and crossover cohorts respectively, 9 (6.8%) patients and 1 (1.9%) patient had 2 consecutive serum creatinine level increases >33% above baseline and >upper limit of normal (ULN). Median creatinine clearance remained stable within normal range throughout the study. One patient from each cohort had alanine aminotransferase (ALT) >10 × ULN on 2 consecutive visits; both had ALT values ≤ULN at the start of deferasirox treatment. In 37 patients with data available before and after dose increases to ≥30 mg/kg/day, no clinically relevant differences were observed in AE profile or laboratory parameters before and after dose increase. Deferasirox had no adverse effect on pediatric growth and adolescent sexual development. Overall, median SF levels in patients who received deferasirox treatment for ≥4 years decreased significantly from 3410 to 3108 ng/mL at end of study (median absolute change, –591 ng/mL, P=0.027; n=67). Decreases in SF were more pronounced when mean deferasirox dose increased to >20 mg/kg/day (Figure 1). In the deferasirox cohort, the median absolute change in SF levels from start of deferasirox to end-of-study was greater in patients aged ≥16 than 2–<16 years (–476 vs –156 ng/mL) reflecting low 1st-year deferasirox doses and conservative dose escalation in pediatric patients. Conclusions:This is the first study to report that deferasirox can significantly decrease SF levels over the long-term in SCD patients without evidence of renal toxicity. Treatment for up to 5 years was associated with a clinically manageable safety profile, without progressive decreases in median creatinine clearance, which remained within normal range. The modest, but statistically significant, decrease in SF is most likely due to under-dosing in the first 2 years and variability in SF related to disease factors. Patients titrated to an appropriate dose had a clinically relevant decrease in SF, highlighting the need to adjust dose, usually to >20 mg/kg/day, to achieve negative iron balance. [Display omitted] Disclosures:Vichinsky:Novartis: Consultancy, Research Funding, Speakers Bureau; Apotex: Consultancy, Research Funding; Hemaquest: Consultancy, Membership on an entity's Board of Directors or advisory committees. Bernaudin:Novartis: Investigator for SCD deferasirox (Exjade). Forni:Novartis: Research Funding. Gardner:Novartis: Research Funding. Hassell:Novartis: Research Funding. Heeney:Novartis: Research Funding. Kutlar:Novartis: Research Funding. Lane:Novartis: Research Funding. Mathias:Novartis: Research Funding. Porter:Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Tebbi:Novartis: Speakers Bureau. Wilson:Novartis: Honoraria, Research Funding, Speakers Bureau. Griffel:Novartis: Employment. Deng:Novartis: Employment. Giannone:Novartis: Employment. Coates:Novartis: Research Funding, Speakers Bureau.

O34. TGF-β signaling maintains tumorigenicity of glioma-initiating cells through the Sox4–Sox2 axis

Recent studies have identified essential signaling pathways for retention of cancer stem cells in various types of cancer. We show here that autocrine TGF-β signaling plays essential roles in retention of stemness of glioma-initiating cells. TGF-β induced expression of Sox2, and this induction was mediated by Sox4, a novel, direct target gene of TGF-β. Inhibitors of TGF-β signaling resulted in deprivation of tumorigenicity of glioma-initiating cells through induction of their differentiation, and these effects were attenuated in glioma-initiating cells transduced with exogenous Sox2 or Sox4. Furthermore, glioma-initiating cells pre-treated with a TGF-β signaling inhibitor exhibited less tumorigenic activity and lethal potency in an intracranial transplantation assay. To target cancer stem cells in clinical situations, elucidation of differences between cancer stem cells and normal stem cells, and identification of novel targets which can eradicate cancer stem cells without affecting normal stem cells will be important. We show that the regulatory mechanism of the Sox axis in glioma-initiating cells is distinct from that in neural progenitor cells. Our findings suggest important differences between glioma-initiating cells and neural progenitor cells, and may open the way to depriving glioma-initiating cells of the tumorigenic activity with minimally affecting normal brain tissues (Ikushima et al., 2009). Ikushima, H., et al., 2009. Autocrine TGF-βsignaling maintains tumorigenicity of glioma-initiating cells through Sry-related HMG-box factors. Cell Stem Cell 5 (5), 504–514.

SNAI2/Slug promotes growth and invasion in human gliomas

BackgroundNumerous factors that contribute to malignant glioma invasion have been identified, but the upstream genes coordinating this process are poorly known.MethodsTo identify genes controlling glioma invasion, we used genome-wide mRNA expression profiles of primary human glioblastomas to develop an expression-based rank ordering of 30 transcription factors that have previously been implicated in the regulation of invasion and metastasis in cancer.ResultsUsing this approach, we identified the oncogenic transcriptional repressor, SNAI2/Slug, among the upper tenth percentile of invasion-related transcription factors overexpressed in glioblastomas. SNAI2 mRNA expression correlated with histologic grade and invasive phenotype in primary human glioma specimens, and was induced by EGF receptor activation in human glioblastoma cells. Overexpression of SNAI2/Slug increased glioblastoma cell proliferation and invasion in vitro and promoted angiogenesis and glioblastoma growth in vivo. Importantly, knockdown of endogenous SNAI2/Slug in glioblastoma cells decreased invasion and increased survival in a mouse intracranial human glioblastoma transplantation model.ConclusionThis genome-scale approach has thus identified SNAI2/Slug as a regulator of growth and invasion in human gliomas.

Abstract 1557: Corticorelin acetate exhibits preclinical antitumor activity, alone and with bevacizumab, against human brain tumor models

Abstract Corticorelin acetate is a synthetic form of corticotropin-releasing factor undergoing clinical trials in the treatment of peritumoral brain edema. We investigated preclinically its potential as an antitumor agent against orthotopically implanted human brain tumors, and its ability to enhance the therapeutic activity of bevacizumab in one of these models. Intracranial (ic) tumor transplantation into the right cerebrum was performed using two patient-derived human tumor xenografts: D-341MED pediatric medulloblastoma and D-456MG pediatric multiforme glioblastoma. Mice were randomized 3 days after ic tumor implantation at which time all treatments were begun. Corticorelin acetate was administered at 0.1 and/or 0.2 mg mg/kg/dose, sc, twice daily (bid) depending upon the experiment. Bevacizumab was administered at a dose of 5 mg/kg/day, intraperitoneally (ip), twice weekly. Treatments were continued until a median day of death was reached for each group. In both studies antitumor effects were evaluated based on a survival analysis (Kaplan-Meier). In the D-341MED, comparing treatments vs control, the low dose corticorelin acetate produced a 56% increase in life span (ILS) (p&amp;lt;0.001); the higher dose of corticorelin acetate yielded a 20% ILS which was also statistically different than the control group (p&amp;lt;0.05). In the D-456MG ic-implanted tumor experiment, corticorelin acetate (0.2 mg/kg/dose, bid, sc) achieved a 42% ILS (p&amp;lt;0.001) while bevacizumab produced a 64% ILS (p&amp;lt;0.001). When the two drugs on these same regimens were applied in combination, the resulting ILS of 92% (p&amp;lt;0.001 versus control) represented a therapeutic enhancement that was statistically different (p&amp;lt;0.05) from either solo drug treatment. Conclusion: Corticorelin acetate significantly increased the lifespan of mice implanted orthotopically with two different pediatric brain tumor models. In the D-456MG model, in which corticorelin acetate and bevacizumab were evaluated, the combination produced a therapeutic outcome superior to that found using either of the two agents alone. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1557.

Intracranial Transplant of Olfactory Ensheathing Cells in Children and Adolescents with Cerebral Palsy: A Randomized Controlled Clinical Trial

Successful repair of damage in cerebral palsy (CP) needs effective clinical interventions other than simply symptomatic treatments. To elucidate the feasibility of using olfactory ensheathing cells (OECs) to treat CP in children and adolescents, we conducted a randomized controlled clinical trial (RCT) on 33 patients. The patients were randomly assigned into two groups (treatment group, n = 18; control group, n = 15), and OECs derived from aborted fetal tissue were injected into the bilateral corona radiata in the frontal lobes (a key point for neural network restoration, KPNNR). The Gross Motor Function Measure (GMFM-66) and the Caregiver Questionnaire Scale were used to evaluate the patients' neurological function and overall health status. Among the 14 patients who completed the 6-month study, six received the cell transplantation and the other eight served as controls. In OEC treatment group, GMFM-66 scores were 26.67 +/- 25.33 compared with 19.00 +/- 20.00 for the control group. Concurrently, the Caregiver Questionnaire Scale score decreased to 77.83 +/- 15.99 in the treatment group in comparison to 138.66 +/- 64.06 of the control group. This trial, albeit small in sample size, indicates OEC KPNNR transplantation is effective for functional improvement in children and adolescents with CP, yet without obvious side effects. This small-scale study suggests that the procedure may be a plausible alternative method to treat this not yet curable disorder, and we urge further evaluation with a large-scale RCT.

Autocrine TGF-β Signaling Maintains Tumorigenicity of Glioma-Initiating Cells through Sry-Related HMG-Box Factors

Despite aggressive surgery, radiotherapy, and chemotherapy, treatment of malignant glioma remains formidable. Although the concept of cancer stem cells reveals a new framework of cancer therapeutic strategies against malignant glioma, it remains unclear how glioma stem cells could be eradicated. Here, we demonstrate that autocrine TGF-beta signaling plays an essential role in retention of stemness of glioma-initiating cells (GICs) and describe the underlying mechanism for it. TGF-beta induced [corrected] expression of Sox2, a stemness gene, and this induction was mediated by Sox4, a direct TGF-beta target gene. Inhibitors of TGF-beta signaling drastically deprived tumorigenicity of GICs by promoting their differentiation, and these effects were attenuated in GICs transduced with Sox2 or Sox4. Furthermore, GICs pretreated with TGF-beta signaling inhibitor exhibited less lethal potency in intracranial transplantation assay. These results identify an essential pathway for GICs, the TGF-beta-Sox4-Sox2 pathway, whose disruption would be a therapeutic strategy against gliomas.

Imaging of Peripheral Benzodiazepine Receptor Expression as Biomarkers of Detrimental versus Beneficial Glial Responses in Mouse Models of Alzheimer's and Other CNS Pathologies

We demonstrate the significance of peripheral benzodiazepine receptor (PBR) imaging in living mouse models of Alzheimer's disease (AD) as biomarkers and functional signatures of glial activation. By radiochemically and immunohistochemically analyzing murine models of the two pathological hallmarks of AD, we found that AD-like Abeta deposition is concurrent with astrocyte-dominant PBR expression, in striking contrast with nonastroglial PBR upregulation in accumulations of AD-like phosphorylated tau. Because tau-induced massive neuronal loss was distinct from the marginal neurodegeneration associated with Abeta plaques in these models, cellular localization of PBR reflected deleterious and beneficial glial reactions to tau versus Abeta pathologies, respectively. This notion was subsequently examined in models of various non-AD neuropathologies, revealing the following reactive glial dynamics underlying differential PBR upregulation: (1) PBR(-) astrogliosis uncoupled with microgliosis or coupled with PBR(+) microgliosis associated with irreversible neuronal insults; and (2) PBR(+) astrogliosis coupled with PBR(- or +/-) microgliosis associated with minimal or reversible neuronal toxicity. Intracranial transplantation of microglia also indicated that nontoxic microglia drives astroglial PBR expression. Moreover, levels of glial cell line-derived neurotrophic factor (GDNF) in astrocytes were correlated with astroglial PBR, except for increased GDNF in PBR(-) astrocytes in the model of AD-like tau pathology, thereby suggesting that PBR upregulation in astrocytes is an indicator of neurotrophic support. Together, PBR expressions in astrocytes and microglia reflect beneficial and deleterious glial reactions, respectively, in diverse neurodegenerative disorders including AD, pointing to new applications of PBR imaging for monitoring the impact of gliosis on the pathogenesis and treatment of AD.

Glioma stem cells are more aggressive in recurrent tumors with malignant progression than in the primary tumor, and both can be maintained long-term in vitro

BackgroundDespite the advances made during decades of research, the mechanisms by which glioma is initiated and established remain elusive. The discovery of glioma stem cells (GSCs) may help to elucidate the processes of gliomagenesis with respect to their phenotype, differentiation and tumorigenic capacity during initiation and progression. Research on GSCs is still in its infancy, so no definitive conclusions about their role can yet be drawn. To understand the biology of GSCs fully, it is highly desirable to establish permanent and biologically stable GSC lines.MethodsIn the current study, GSCs were isolated from surgical specimens of primary and recurrent glioma in a patient whose malignancy had progressed during the previous six months. The GSCs were cryopreserved and resuscitated periodically during long-term maintenance to establish glioma stem/progenitor cell (GSPC) lines, which were characterized by immunofluorescence, flow cytometry and transmission electronic microscopy. The primary and recurrent GSPC lines were also compared in terms of in vivo tumorigenicity and invasiveness. Molecular genetic differences between the two lines were identified by array-based comparative genomic hybridization and further validated by real-time PCR.ResultsTwo GSPC lines, SU-1 (primary) and SU-2 (recurrent), were maintained in vitro for more than 44 months and 38 months respectively. Generally, the potentials for proliferation, self-renewal and multi-differentiation remained relatively stable even after a prolonged series of alternating episodes of cryopreservation and resuscitation. Intracranial transplantation of SU-1 cells produced relatively less invasive tumor mass in athymic nude mice, while SU-2 cells led to much more diffuse and aggressive lesions strikingly recapitulated their original tumors. Neither SU-1 nor SU-2 cells reached the terminal differentiation stage under conditions that would induce terminal differentiation in neural stem cells. The differentiation of most of the tumor cells seemed to be blocked at the progenitor cell phase: most of them expressed nestin but only a few co-expressed differentiation markers. Transmission electron microscopy showed that GSCs were at a primitive stage of differentiation with low autophagic activity. Array-based comparative genomic hybridization revealed genetic alterations common to both SU-1 and SU-2, including amplification of the oncogene EGFR and deletion of the tumor suppressor PTEN, while some genetic alterations such as amplification of MTA1 (metastasis associated gene 1) only occurred in SU-2.ConclusionThe GSPC lines SU-1 and SU-2 faithfully retained the characteristics of their original tumors and provide a reliable resource for investigating the mechanisms of formation and recurrence of human gliomas with progressive malignancy. Such investigations may eventually have major impacts on the understanding and treatment of gliomas.