Chronic Myeloid Leukemia in Children and Adolescents.

Acute leukemia is the most common malignancy in childhood, while chronic myeloid leukemia is rare, accounting for only 2% to 3% of all leukemia in childhood and 9% in adolescents, with an annual incidence of 1 and 2.2 cases per million in the two groups. The goal in Pediatrics is remission and cure with tyrosine kinase inhibitors (TKIs) and monitoring closely for long-term effects of TKI use.

Anti-GD2 immunoliposomes loaded with oxamate for neuroblastoma.

Oncometabolism can be targeted for the development of less myelotoxic oncotherapeutics. Lactate dehydrogenase A (LDHA) is central to the Warburg effect, a potential oncometabolic shift in neuroblastoma (NBL). Advanced surgical, cytotoxic and cell-differentiating therapies improved survival of children with NBL. Anti-GD2 monoclonal antibodies (mAb) effectively targeting NBL are also incorporated into complex therapies. However, poor clinical outcomes of high-risk NBL require improvements. Here, we verified the pre-reported prognostic value of LDHA expression in NBL using the R2 onco-genomics platform. Kaplan-Meier curves re-demonstrated that higher tumor LDHA expression correlates with worse survival. Multivariate statistics confirmed LDHA is independent from age, stage, and MYCN amplification. In conclusion, a molecular construct is proposed with anti-GD2 mAbs utilized for the targeted delivery of liposomes containing an LDHA inhibitor, Oxamate. Development and preclinical testing of this immunoliposome may validate targeted inhibition of the Warburg effect for NBL. IMPACT: Development of therapeutics against oncometabolism. Targeted specified drug-delivery with mAb. Sparing normal tissues from profound LDHA inhibition. Immunoliposome loaded with an anti-metabolite. If preclinically successful, has translational potential.

Pediatric Patients With Sickle Cell Disease at a Public Hospital: Nutrition, Compliance and Early Experience With L-Glutamine Therapy.

BACKGROUND/AIM: Hydration and hydroxyurea (HU) can modify sickle cell disease (SCD) severity. Optimal nutrition and L-glutamine (Gln) may provide further amelioration. PATIENTS AND METHODS: Reviews of medical records and nutrition surveys were used to investigate severity of pediatric patients with SCD in relation to nutrition, growth, hematologic parameters, and diseasemodifying agents. RESULTS: Among 25 females and 25 males (9.1±7 years), beta-globin genotypes were: HbSS/Sß°, 60%; HbSC, 32%; HbSß+, 8%. The mean number of annual pain crises (APC) was 0.97±1.1. APCs increased ≥2-fold as HbF dropped to <10% with age. Proper hydration and nutrition correlated with younger ages and fewer APCs. Height and weight Z-scores were ≤-1SD in 20% of 35 surveyed patients (12±7.8 years), who had more APCs (2.5±2.5 vs. 1±1.3, p=0.03). Prealbumin levels were overall low. Twenty-two of 28 patients on HU reported ≥90% adherence - with higher mean corpuscular volume (92±9.6 vs. 74±10 f/l, p<0.01). Seventy percent of Gln prescriptions were filled. Compliance over 23 months was ≥70% in 12 patients, including 2 on chronic transfusion. Of 10 evaluable patients, 6 (8.8±2.2 years) had fewer APCs with Gln (mean 0.2 vs. 0.9, p=0.016), with increasing prealbumin levels (14.1 to 15.8 mg/dl, p=0.1). CONCLUSION: Younger, and well-nourished, well-hydrated patients have a milder clinic course. Disease severity was the worse in undernourished teenagers with suboptimal compliance. L-Glutamine with prealbumin monitoring should be considered for further evaluation in pediatric SCD.

Chemotherapy Can Synergize With Adoptive Immunotherapy to Inhibit Medulloblastoma Growth.

BACKGROUND/AIM: In the age of ever-increasing developments in targeted cancer treatments, new immune-based approaches for brain tumor therapy represent an attractive avenue. Despite encouraging pre-clinical data, results in patients have been sub-optimal, likely due to tumor-induced immune suppression and intrinsic resistance to immune attack. Chemotherapy and biologic agents may be able to disrupt these mechanisms and restore tumor sensitivity to immune attack. In this study, we explore whether a combination of gemcitabine and rapamycin can sensitize medulloblastoma cells to immunotherapy in vitro and in vivo. MATERIALS AND METHODS: With the commercial medulloblastoma cell line, Daoy, we explored the concentrations of combinations of Gemcitabine with rapamycin needed to induce cytotoxicity. Next, we used flow cytometry to assess the cytotoxicity of chemotherapy-treated Daoy cells with the addition of anti-tumor T-cells, generated from naive T-cells stimulated in the presence of Daoy lysate-pulsed dendritic cells. Then, we examined the efficacy of chemotherapy alone versus chemotherapy plus immunotherapy in tumor growth inhibition of subcutaneous medulloblastoma xenografts. RESULTS: Rapamycin alone at <1,000 nM had moderate activity against Daoy cells in vitro and IC50 was >1,000 nM. Gemcitabine had a 3-day IC50 alone of 10 nM but in combination with 100 nM rapamycin, it decreased to 1 nM, suggesting increased cytotoxicity with combined therapy. Stimulated T-cells mediated in-vitro cytotoxicity, although background cytotoxicity of unstimulated "naïve" T-cells was also significant. Finally, established subcutaneous Daoy cell xenografts in SCID mice were treated with chemotherapy alone or chemotherapy plus adoptive immunotherapy (stimulated and non-stimulated). Gemcitabine and rapamycin alone significantly slowed tumor growth, but the addition of immunotherapy further augmented inhibition. CONCLUSION: Combining immunotherapy and chemo-biologic therapy inhibit medulloblastoma cell and xenograft growth, and may offer an effective treatment for patients with medulloblastoma.

Novel ELANE Gene Mutation in a Newborn with Severe Congenital Neutropenia: Case Report and Literature Review.

Severe neutropenia is defined as an absolute neutrophil count (ANC) of less than 0.5 × 10 9 /L. Severe congenital neutropenia (SCN) is an inborn disorder with maturation arrest of granulocytes due to various genetic abnormalities, which may lead to immunodeficiency. Among several associated genetic mutations, the variants or heterozygous mutations of the ELANE gene coding neutrophil elastase comprise approximately 50% of the genetic causes of SCN. We present a newborn (male) with severe neutropenia due to a novel ELANE gene mutation. The newborn was born at 38 6/7 weeks gestation to a 25-year-old mother with hypertension and morbid obesity. Pregnancy and delivery were uncomplicated but the baby obtained a complete blood count (CBC) on day of life 2 for a work up of hyperbilirubinemia. He was noted to initially have an ANC of 0.2 × 10 9 /L and 0 on subsequent blood counts. A bone marrow biopsy showed a left shift and consistent with myeloid maturation arrest. In direct DNA sequencing analysis, we found an ELANE gene mutation (Val119Glu, V119E), which may be a new gene mutation to cause SCN. The diagnosis of SCN in newborns is usually based on neutropenia identified on a routine CBC. Sufficient awareness and high suspicion of this rare disease can prevent missed or delayed diagnosis of SCN. Our analysis also suggests a new pathological mutation in the ELANE gene and supports the important role of molecular testing in SCN.

Primary Renal Ewing Sarcoma in Children and Young Adults.

The Ewing sarcoma family of tumors (ESFT) are high-grade small round blue cell malignancies traditionally presenting in children and adolescents. The most common site of primary disease is bone, though extraskeletal primary sites are well-recognized. We present 6 cases of primary ESFT of the kidney and 1 case of the adrenal gland. Patients were 11 to 18 years of age at diagnosis. Metastases at diagnosis were present in most cases (n=6). All patients underwent surgery, and most received radiation (n=5). Five patients relapsed after initial remission. Comprehensive review of the primary renal ESFT literature was used to analyze various factors, including age, sex, disease metrics, metastases at diagnoses, and overall survival in a total of 362 cases. Notably, while the general ESFT population has reported rates of metastasis at diagnosis of 20% to 25%, this rate in the renal ESFT population was 53% with a rate of 59% in adolescent and young-adult patients (11 to 24 y). Nodal disease at diagnosis was present in 24% of renal ESFT cases compared with 3.2% in patients with primary skeletal ESFT. While this malignant process may share histologic and molecular features with its bone and soft tissue counterparts, primary renal ESFT presentations seem to be more aggressive and have worse outcomes.

A Case of Langerhans Cell Histiocytosis With Multifocal, Single-System GI Tract Involvement and Literature Review.

Langerhans cell histiocytosis (LCH) is the most common histiocytic disorder, characterized by the pathologic clonal proliferation and accumulation of immature Langerhans cells within organs. Multiple organ systems can be affected, resulting in a spectrum of clinical manifestations. Isolated gastrointestinal involvement in LCH is rare and usually presents in childhood as a multisystem disease and usually has poor outcomes. We describe a 20-year-old Hispanic female with multifocal, single-system gastrointestinal LCH. Initially diagnosed from a CD1a, S100, and CD207 (Langerin) positive appendix tissue after an appendectomy and confirmed multifocal with an endoscopy. She had a full clinical and endoscopic resolution of disease with cytarabine therapy.

The humanin analogue (HNG) prevents temozolomide-induced male germ cell apoptosis and other adverse effects in severe combined immuno-deficiency (SCID) mice bearing human medulloblastoma.

Subfertility is a major concern of long-term cancer survivors at the reproductive age. We have previously demonstrated that a potent humanin analogue, HNG, protected chemotherapy-induced apoptosis in germ cells but not cancer cells in a metastatic melanoma allograft model. In this study, we utilized severe combined immuno-deficiency (SCID) mice bearing human medulloblastoma to study the effect of HNG in Temozolomide (TMZ) induced male germ cell apoptosis and white blood cell (WBC) suppression. Human medulloblastoma DAOY cells were injected subcutaneously into the right flank of male SCID mice. Three weeks later, groups of tumor-bearing mice received one of the following treatments: vehicle, HNG, TMZ, or TMZ + HNG. 24 h after last injection, the tumors weights, complete blood counts, liver and spleen weights, male germ cell apoptosis was assessed. HNG did not affect TMZ's significant anti-tumor action. HNG significantly prevented TMZ-induced germ cell apoptosis and attenuated the suppressed total WBC and granulocyte counts in SCID mice with or without TMZ treatment. HNG also attenuated TMZ-induced body weight loss and decrease of spleen and liver weights. In conclusion, HNG ameliorated TMZ-induced germ cell apoptosis; WBC and granulocytes loss; and decreased body/organ weights without compromising the TMZ's anti-cancer action on medulloblastoma xenografts in SCID mice.

A Phase 3 Trial of l-Glutamine in Sickle Cell Disease.

BACKGROUND: Oxidative stress contributes to the complex pathophysiology of sickle cell disease. Oral therapy with pharmaceutical-grade l-glutamine (USAN, glutamine) has been shown to increase the proportion of the reduced form of nicotinamide adenine dinucleotides in sickle cell erythrocytes, which probably reduces oxidative stress and could result in fewer episodes of sickle cell-related pain. METHODS: In a multicenter, randomized, placebo-controlled, double-blind, phase 3 trial, we tested the efficacy of pharmaceutical-grade l-glutamine (0.3 g per kilogram of body weight per dose) administered twice daily by mouth, as compared with placebo, in reducing the incidence of pain crises among patients with sickle cell anemia or sickle ß0-thalassemia and a history of two or more pain crises during the previous year. Patients who were receiving hydroxyurea at a dose that had been stable for at least 3 months before screening continued that therapy through the 48-week treatment period. RESULTS: A total of 230 patients (age range, 5 to 58 years; 53.9% female) were randomly assigned, in a 2:1 ratio, to receive l-glutamine (152 patients) or placebo (78 patients). The patients in the l-glutamine group had significantly fewer pain crises than those in the placebo group (P=0.005), with a median of 3.0 in the l-glutamine group and 4.0 in the placebo group. Fewer hospitalizations occurred in the l-glutamine group than in the placebo group (P=0.005), with a median of 2.0 in the l-glutamine group and 3.0 in the placebo group. Two thirds of the patients in both trial groups received concomitant hydroxyurea. Low-grade nausea, noncardiac chest pain, fatigue, and musculoskeletal pain occurred more frequently in the l-glutamine group than in the placebo group. CONCLUSIONS: Among children and adults with sickle cell anemia, the median number of pain crises over 48 weeks was lower among those who received oral therapy with l-glutamine, administered alone or with hydroxyurea, than among those who received placebo, with or without hydroxyurea. (Funded by Emmaus Medical; ClinicalTrials.gov number, NCT01179217 .).

Efficacy of Asparaginase Erwinia chrysanthemi With and Without Temozolomide Against Glioma Cells and Intracranial Mouse Medulloblastoma.

BACKGROUND: Anti-metabolites are less-myelosuppressive than DNA-damaging anticancer drugs and may be useful against brain tumors. MATERIALS AND METHODS: We evaluated the asparagine/glutamine-deaminating agent Erwinaze with/without temozolomide against brain tumor cells and mouse medulloblastomas. RESULTS: Erwinaze treatment of cell lines and neurospheres led to dose-dependent reductions of cells (reversible by L-glutamine), with half maximal inhibitory concentrations (IC50s) of 0.12->10 IU/ml. Erwinaze at <1 IU/ml reduced temozolomide IC50s by 3.6- to 13-fold (300-1,200 µM to 40-330 µM). Seven-week-old SMO/SMO mice treated with Erwinaze (regardless of temozolomide treatment) had better survival 11 weeks post-therapy, compared to those not treated with Erwinaze (81.25% vs. 46.15, p=0.08). Temozolomide-treated mice developed 10% weight loss, impairing survival. All 16 mice treated with temozolomide (regardless of Erwinaze treatment) succumbed by 40-weeks of age, whereas 5/8 animals treated with Erwinaze alone and 2/6 controls survived (p=0.035). CONCLUSION: Erwinaze enhances cytotoxicity of temozolomide in vitro, and improves survival in SMO/SMO mice, likely by reducing cerebrospinal fluid glutamine. Temozolomide-associated toxicity prevented demonstration of any potential combinatorial advantage with Erwinaze in vivo.

In search of druggable targets for GBM amino acid metabolism.

BACKGROUND: Amino acid (AA) pathways may contain druggable targets for glioblastoma (GBM). Literature reviews and GBM database ( http://r2.amc.nl ) analyses were carried out to screen for such targets among 95 AA related enzymes. METHODS: First, we identified the genes that were differentially expressed in GBMs (3 datasets) compared to non-GBM brain tissues (5 datasets), or were associated with survival differences. Further, protein expression for these enzymes was also analyzed in high grade gliomas (HGGs) (proteinatlas.org). Finally, AA enzyme and gene expression were compared among the 4 TCGA (The Cancer Genome Atlas) subtypes of GBMs. RESULTS: We detected differences in enzymes involved in glutamate and urea cycle metabolism in GBM. For example, expression levels of BCAT1 (branched chain amino acid transferase 1) and ASL (argininosuccinate lyase) were high, but ASS1 (argininosuccinate synthase 1) was low in GBM. Proneural and neural TCGA subtypes had low expression of all three. High expression of all three correlated with worse outcome. ASL and ASS1 protein levels were mostly undetected in high grade gliomas, whereas BCAT1 was high. GSS (glutathione synthetase) was not differentially expressed, but higher levels were linked to poor progression free survival. ASPA (aspartoacylase) and GOT1 (glutamic-oxaloacetic transaminase 1) had lower expression in GBM (associated with poor outcomes). All three GABA related genes -- glutamate decarboxylase 1 (GAD1) and 2 (GAD2) and 4-aminobutyrate aminotransferase (ABAT) -- were lower in mesenchymal tumors, which in contrast showed higher IDO1 (indoleamine 2, 3-dioxygenase 1) and TDO2 (tryptophan 2, 3-diaxygenase). Expression of PRODH (proline dehydrogenase), a putative tumor suppressor, was lower in GBM. Higher levels predicted poor survival. CONCLUSIONS: Several AA-metabolizing enzymes that are higher in GBM, are also linked to poor outcome (such as BCAT1), which makes them potential targets for therapeutic inhibition. Moreover, existing drugs that deplete asparagine and arginine may be effective against brain tumors, and should be studied in conjunction with chemotherapy. Last, AA metabolism is heterogeneous in TCGA subtypes of GBM (as well as medulloblastomas and other pediatric tumors), which may translate to variable responses to AA targeted therapies.

Clinical aggressiveness of malignant gliomas is linked to augmented metabolism of amino acids.

Glutamine, glutamate, asparagine, and aspartate are involved in an enzyme-network that controls nitrogen metabolism. Branched-chain-amino-acid aminotransferase-1 (BCAT1) promotes proliferation of gliomas with wild-type IDH1 and is closely connected to the network. We hypothesized that metabolism of asparagine, glutamine, and branched-chain-amino-acids is associated with progression of malignant gliomas. Gene expression for asparagine synthetase (ASNS), glutaminase (GLS), and BCAT1 were analyzed in 164 gliomas from 156 patients [33-anaplastic gliomas (AG) and 131-glioblastomas (GBM), 64 of which were recurrent GBMs]. ASNS and GLS were twofold higher in GBMs versus AGs. BCAT1 was also higher in GBMs. ASNS expression was twofold higher in recurrent versus new GBMs. Five patients had serial samples: 4-showed higher ASNS and 3-higher GLS at recurrence. We analyzed grade and treatment in 4 groups: (1) low ASNS, GLS, and BCAT1 (n = 96); (2) low ASNS and GLS, but high BCAT1 (n = 26); (3) high ASNS or GLS, but low BCAT1 (n = 25); and (4) high ASNS or GLS and high BCAT1 (n = 17). Ninety-one  % of patients (29/32) with grade-III lesions were in group 1. In contrast, 95 % of patients (62/65) in groups 2-4 had GBMs. Treatment was similar in 4 groups (radiotherapy-80 %; temozolomide-30 %; other chemotherapy-50 %). High expression of ASNS, GLS, and BCAT1 were each associated with poor survival in the entire group. The combination of lower ASNS, GLS, and BCAT1 levels correlated with better survival for newly diagnosed GBMs (66 patients; P = 0.0039). Only tumors with lower enzymes showed improved outcome with temozolomide. IDH1(WT) gliomas had higher expression of these genes. Manipulation of amino acid metabolism in malignant gliomas may be further studied for therapeutics development.

Allergic reactions and antiasparaginase antibodies in children with high-risk acute lymphoblastic leukemia: A children's oncology group report.

BACKGROUND: The objectives of this study were to assess the incidence of clinical allergy and end-induction antiasparaginase (anti-ASNase) antibodies in children with high-risk acute lymphoblastic leukemia treated with pegylated (PEG) Escherichia coli ASNase and to determine whether they carry any prognostic significance. METHODS: Of 2057 eligible patients, 1155 were allocated to augmented arms in which PEG ASNase replaced native ASNase postinduction. Erwinia chrysanthemi (Erwinia) ASNase could be used to replace native ASNase after allergy, if available. Allergy and survival data were complete for 990 patients. End-induction antibody titers were available for 600 patients. RESULTS: During the consolidation phase, 289 of 990 patients (29.2%) had an allergic reaction. There were fewer allergic reactions to Erwinia ASNase than to native ASNase (odds ratio, 4.33; P < .0001) or PEG ASNase (odds ratio, 3.08; P < .0001) only during phase 1 of interim maintenance. There was no significant difference in 5-year event-free survival (EFS) between patients who received PEG ASNase throughout the entire study postinduction versus those who developed an allergic reaction to PEG ASNase during consolidation phase and subsequently received Erwinia ASNase (80.8% ± 2.8% and 81.6% ± 3.8%, respectively; P = .66). Patients who had positive antibody titers postinduction were more likely to have an allergic reaction to PEG ASNase (odds ratio, 2.4; P < .001). The 5-year EFS rate between patients who had negative versus positive antibody titers (80% ± 2.6% and 77.7% ± 4.3%, respectively; P = .68) and between patients who did not receive any ASNase postconsolidation and those who received PEG ASNase throughout the study (P = .22) were significantly different. CONCLUSIONS: The current results demonstrate differences in the incidence rates of toxicity between ASNase preparations but not in EFS. The presence of anti-ASNase antibodies did not affect EFS.

Clinicocytopathologic correlation in an atypical presentation of lymphadenopathy with review of literature.

OBJECTIVES: To present a clinicocytopathologic correlation of an atypical case of cat scratch disease (CSD) involving retroperitoneal lymph nodes, with emphasis on communication between service teams for managing lymphadenopathy of unknown origin. We consider clinical and cytologic differential diagnoses and review the literature on atypical cases of CSD, with emphasis on abdominal presentation and cytologic findings. METHODS: Clinical services met with the cytology service to review clinical and pathologic features. Literature was reviewed via PubMed search (Harbor-UCLA subscriptions). Immunohistochemistry and Steiner silver stains were performed by Harbor-UCLA Department of Pathology. Enzyme-linked immunosorbent assay IgG and IgM Bartonella henselae titers were carried out by Quest Nichols Institute. RESULTS: Fine-needle aspirate Diff-Quik and Papanicolaou smears and H&E-stained cell block showed abundant histiocytes, monocytoid B cells, and numerous neutrophils associated with necrosis corresponding to a late stage of CSD infection. Silver stain was positive for clumps of pleomorphic organisms. IgM and IgG antibody titers were elevated. CONCLUSIONS: The cytologic findings of CSD in an atypical abdominal presentation are similar to those of a classic presentation. Laboratory workup for atypical CSD should include at least two other modalities aside from cytomorphologic features. Close clinical and cytologic correlation avoided potentially unnecessary and harmful surgery and enabled timely treatment.

Asparagine depletion potentiates the cytotoxic effect of chemotherapy against brain tumors.

UNLABELLED: Targeting amino acid metabolism has therapeutic implications for aggressive brain tumors. Asparagine is an amino acid that is synthesized by normal cells. However, some cancer cells lack asparagine synthetase (ASNS), the key enzyme for asparagine synthesis. Asparaginase (ASNase) contributes to eradication of acute leukemia by decreasing asparagine levels in serum and cerebrospinal fluid. However, leukemic cells may become ASNase-resistant by upregulating ASNS. High expression of ASNS has also been associated with biologic aggressiveness of other cancers, including gliomas. Here, the impact of enzymatic depletion of asparagine on proliferation of brain tumor cells was determined. ASNase was used as monotherapy or in combination with conventional chemotherapeutic agents. Viability assays for ASNase-treated cells demonstrated significant growth reduction in multiple cell lines. This effect was reversed by glutamine in a dose-dependent manner--as expected, because glutamine is the main amino group donor for asparagine synthesis. ASNase treatment also reduced sphere formation by medulloblastoma and primary glioblastoma cells. ASNase-resistant glioblastoma cells exhibited elevated levels of ASNS mRNA. ASNase cotreatment significantly enhanced gemcitabine or etoposide cytotoxicity against glioblastoma cells. Xenograft tumors in vivo showed no significant response to ASNase monotherapy and little response to temozolomide alone. However, combinatorial therapy with ASNase and temozolomide resulted in significant growth suppression for an extended duration of time. Taken together, these findings indicate that amino acid depletion warrants further investigation as adjunctive therapy for brain tumors. IMPLICATIONS: Findings have potential impact for providing adjuvant means to enhance brain tumor chemotherapy.

Autologous tumor lysate-pulsed dendritic cell immunotherapy for pediatric patients with newly diagnosed or recurrent high-grade gliomas.

Immunotherapy has the potential to improve clinical outcomes with little toxicity for pediatric patients with brain tumors. We conducted a pilot feasibility study of tumor lysate-pulsed dendritic cell (DC) vaccination in pediatric patients (1 to 18 years old) with newly diagnosed or recurrent high-grade glioma (HGG). A total of nine DC vaccine doses, each containing 1 × 10(6) cells per dose were administered to three out of the seven originally enrolled patients. Toxicities were limited to mild side-effects, except in one case of elevated alkaline phosphatase, which resolved without clinical consequences. Two patients with primary lesions amongst the three vaccinated were alive at the time of writing, both without evidence of disease. Pre- and post-vaccination tumor samples from a patient with an anaplastic oligoastrocytoma that recurred failed to demonstrate immune cell infiltration by immunohistochemistry. Peripheral cytokine levels were evaluated in one patient following DC vaccination and demonstrated some changes in relation to vaccination. DC vaccine is tolerable and feasible with some limitations for pediatric patients with HGG. Dendritic cell based immunotherapy may provide some clinical benefit in pediatric patients with glioma, especially for patients with minimal residual disease, but further investigation of this modality is required.

A molecular screening approach to identify and characterize inhibitors of glioblastoma stem cells.

Glioblastoma (GBM) is among the most lethal of all cancers. GBM consist of a heterogeneous population of tumor cells among which a tumor-initiating and treatment-resistant subpopulation, here termed GBM stem cells, have been identified as primary therapeutic targets. Here, we describe a high-throughput small molecule screening approach that enables the identification and characterization of chemical compounds that are effective against GBM stem cells. The paradigm uses a tissue culture model to enrich for GBM stem cells derived from human GBM resections and combines a phenotype-based screen with gene target-specific screens for compound identification. We used 31,624 small molecules from 7 chemical libraries that we characterized and ranked based on their effect on a panel of GBM stem cell-enriched cultures and their effect on the expression of a module of genes whose expression negatively correlates with clinical outcome: MELK, ASPM, TOP2A, and FOXM1b. Of the 11 compounds meeting criteria for exerting differential effects across cell types used, 4 compounds showed selectivity by inhibiting multiple GBM stem cells-enriched cultures compared with nonenriched cultures: emetine, n-arachidonoyl dopamine, n-oleoyldopamine (OLDA), and n-palmitoyl dopamine. ChemBridge compounds #5560509 and #5256360 inhibited the expression of the 4 mitotic module genes. OLDA, emetine, and compounds #5560509 and #5256360 were chosen for more detailed study and inhibited GBM stem cells in self-renewal assays in vitro and in a xenograft model in vivo. These studies show that our screening strategy provides potential candidates and a blueprint for lead compound identification in larger scale screens or screens involving other cancer types.

High-dose chemotherapy with autologous hematopoietic stem-cell rescue for pediatric brain tumor patients: a single institution experience from UCLA.

Background. Dose-dependent response makes certain pediatric brain tumors appropriate targets for high-dose chemotherapy with autologous hematopoietic stem-cell rescue (HDCT-AHSCR). Methods. The clinical outcomes and toxicities were analyzed retrospectively for 18 consecutive patients ≤19 y/o treated with HDCT-AHSCR at UCLA (1999-2009). Results. Patients' median age was 2.3 years. Fourteen had primary and 4 recurrent tumors: 12 neural/embryonal (7 medulloblastomas, 4 primitive neuroectodermal tumors, and a pineoblastoma), 3 glial/mixed, and 3 germ cell tumors. Eight patients had initial gross-total and seven subtotal resections. HDCT mostly consisted of carboplatin and/or thiotepa ± etoposide (n = 16). Nine patients underwent a single AHSCR and nine ≥3 tandems. Three-year progression-free and overall survival probabilities were 60.5% ± 16 and 69.3% ± 11.5. Ten patients with pre-AHSCR complete remissions were alive/disease-free, whereas 5 of 8 with measurable disease were deceased (median followup: 2.3 yrs). Nine of 13 survivors avoided radiation. Single AHSCR regimens had greater toxicity than ≥3 AHSCR (P < .01). Conclusion. HDCT-AHSCR has a definitive, though limited role for selected pediatric brain tumors with poor prognosis and pretransplant complete/partial remissions.

Clinical outcome in pediatric glial and embryonal brain tumors correlates with in vitro multi-passageable neurosphere formation.

BACKGROUND: Cultured brain tumors can form neurospheres harboring tumorigenic cells with self renewal and differentiation capacities. Renewable neurosphere formation has clinical predictive value in adult malignant gliomas, yet its prognostic role for pediatric brain tumors is unknown. METHODS: Established neurosphere conditions were used for culturing samples from glial, embryonal and mixed glioneuronal tumors from 56 pediatric patients. Potential associations between neurosphere formation and clinical outcome were analyzed retrospectively. RESULTS: Thirty-seven percent of all samples formed renewable neurospheres. Analysis of available clinical outcome data from 51 patients demonstrated significantly increased hazard ratios (HR) for both disease progression (HR = 9.9, P < 0.001) and death (HR = 16.6, P < 0.01) in the neurosphere forming group. Furthermore, neurosphere formation correlated with adverse progression free survival (PFS) in glial and embryonal tumors, but not in mixed glioneuronal tumors. Overall survival (OS) was significantly worse for neurosphere-forming patients with embryonal tumors, as a group and amongst the subgroup with medulloblastoma, but not in the glial group. Multivariate analysis showed that neurosphere formation was associated with diminished PFS and OS independent of age, gender, or treatment. Neurosphere formation was an independent predictor of diminished PFS of glial tumors after adjusting for grade. Multivariate analysis, adjusting for both Ki67 staining and neurosphere formation, demonstrated that neurosphere formation remained predictive of progression whereas Ki67 did not. CONCLUSIONS: Neurosphere formation is more predictive of pediatric brain tumor progression than semi-quantitative Ki67 staining. Pediatric brain tumor derived neurospheres may provide a predictive model for preclinical explorations.