Effective re-induction regimen for children with recurrent medulloblastoma.

Background: There is no standard treatment for the recurrence of medulloblastoma, the most common malignant childhood brain tumor, and prognosis remains dismal. In this study, we introduce a regimen that is well-tolerated and effective at inducing remission. Methods: The primary objectives of this study were to assess tolerability of the regimen and overall response rate (ORR). A retrospective chart review of patients with recurrent medulloblastoma, treated at two institutions with a re-induction regimen of intravenous irinotecan and cyclophosphamide with oral temozolomide and etoposide, was performed. Demographic, clinicopathologic, toxicity, and response data were collected and analyzed. Results: Nine patients were identified. Median age was 5.75 years. Therapy was well-tolerated with no therapy-limiting toxicities and no toxic deaths. Successful stem cell collection was achieved in all 5 patients in whom it was attempted. ORR after 2 cycles was 78%. Three patients had a complete response, 4 patients had a partial response, 1 patient had stable disease, and 1 patient had progressive disease. Four patients are alive with no evidence of disease (NED), 2 patients are alive with disease, 2 patients have died of disease, and 1 patient died of toxicity related to additional therapy (NED at time of death). Conclusions: This regimen is well-tolerated and effective. Tumor response was noted in the majority of cases, allowing patients to proceed to additional treatment with no or minimal disease. Further study of this regimen in a clinical trial setting is an important next step.

Pediatric Chordoma: A Tale of Two Genomes.

Little is known regarding the genomic alterations in chordoma, with the exception of loss of SMARCB1, a core member of the SWI/SNF complex, in poorly differentiated chordomas. A TBXT duplication and rs2305089 polymorphism, located at 6q27, are known genetic susceptibility loci. A comprehensive genomic analysis of the nuclear and mitochondrial genomes in pediatric chordoma has not yet been reported. In this study, we performed whole exome and mitochondrial DNA (mtDNA) genome sequencing on 29 chordomas from 23 pediatric patients. Findings were compared with that from whole genome sequencing datasets of 80 adult skull base chordoma patients. In the pediatric chordoma cohort, 81% percent of the somatic mtDNA mutations were observed in NADH complex genes, which is significantly enriched compared to the rest of the mtDNA genes (p=0.001). In adult chordomas, mtDNA mutations were also enriched in the NADH complex genes (p<0.0001). Furthermore, a progressive increase in heteroplasmy of non-synonymous mtDNA mutations was noted in patients with multiple tumors (p=0.0007). In the nuclear genome, rare likely germline in-frame indels in ARID1B, a member of the SWI/SNF complex located at 6q25.3, were observed in five pediatric patients (22%) and four patients in the adult cohort (5%). The frequency of rare ARID1B indels in the pediatric cohort is significantly higher than that of the adult cohort (p=0.0236, Fisher's exact test), but they were both significantly higher than that in the ethnicity-matched populations (p<5.9e-07 and p<0.0001174, respectively). Implications: germline ARID1B indels and mtDNA aberrations appear important for chordoma genesis, especially in pediatric chordoma.

Disease Evolution Monitored by Serial Cerebrospinal Fluid Liquid Biopsies in Two Cases of Recurrent Medulloblastoma.

Medulloblastoma is the most common malignant brain tumor in childhood. Initial treatment generally includes surgery, irradiation, and chemotherapy. Approximately 20-30% of patients will experience a recurrence, which portends a very poor prognosis. The current standard of care for evaluation for relapse includes radiographic surveillance with magnetic resonance imaging at regular intervals. The presence of circulating tumor DNA in the cerebrospinal fluid has been demonstrated to be a predictor of a higher risk of progression in a research setting for patients with medulloblastoma treated on a prospective single institution clinical trial. We have previously published and clinically validated a liquid-biopsy-based genetic assay utilizing low-pass whole genome sequencing to detect copy number alterations in circulating tumor DNA. Here, we present two teenage patients with posterior fossa medulloblastoma with recurrent disease who have been monitored with serial liquid biopsies showing tumor evolution over time, demonstrating the clinical utility of these approaches.

Final report of the phase II NEXT/CNS-GCT-4 trial: GemPOx followed by marrow-ablative chemotherapy for recurrent intracranial germ cell tumors.

Background: Patients with relapsed intracranial germinoma can achieve durable remission with standard chemotherapy regimens and/or reirradiation; however, innovative therapies are required for patients with relapsed and/or refractory intracranial nongerminomatous germ cell tumors (NGGCTs) due to their poor prognosis. Improved outcomes have been reported using reinduction chemotherapy to achieve minimal residual disease, followed by marrow-ablative chemotherapy (HDCx) with autologous hematopoietic progenitor cell rescue (AuHPCR). We conducted a phase II trial evaluating the response and toxicity of a 3-drug combination developed for recurrent intracranial germ cell tumors consisting of gemcitabine, paclitaxel, and oxaliplatin (GemPOx). Methods: A total of 9 patients with confirmed relapsed or refractory intracranial GCT were enrolled after signing informed consent, and received at least 2 cycles of GemPOx, of which all but 1 had relapsed or refractory NGGCTs. One patient with progressive disease was found to have pathologically confirmed malignant transformation to pure embryonal rhabdomyosarcoma (without GCT elements), hence was ineligible and not included in the analysis. Patients who experienced sufficient responses proceeded to receive HDCx with AuHPCR. Treatment response was determined based on radiographic tumor assessments and tumor markers. Results: A total of 7 patients achieved sufficient response and proceeded with HDCx and AuHPCR, and 5 subsequently received additional radiotherapy. A total of 2 patients developed progressive disease while receiving GemPOx. Myelosuppression and transaminitis were the most common treatment-related adverse events. With a mean follow-up of 44 months, 4 patients (3 NGGCTs, 1 germinoma) are alive without evidence of disease. Conclusions: GemPOx demonstrates efficacy in facilitating stem cell mobilization, thus facilitating the feasibility of both HDCx and radiotherapy.

Low-pass whole-genome and targeted sequencing of cell-free DNA from cerebrospinal fluid in pediatric patients with central nervous system tumors.

Background: Central nervous system tumors are the most common pediatric solid tumors and the most frequent cause of cancer-related morbidity in childhood. Significant advances in understanding the molecular features of these tumors have facilitated the development of liquid biopsy assays that may aid in diagnosis and monitoring response to therapy. In this report, we describe our comprehensive liquid biopsy platform for detection of genome-wide copy number aberrations, sequence variants, and gene fusions using cerebrospinal fluid (CSF) from pediatric patients with brain, spinal cord, and peripheral nervous system tumors. Methods: Cell-free DNA was isolated from the CSF from 55 patients, including 47 patients with tumors and 8 controls. Results: Abnormalities in cell-free DNA were detected in 24 (51%) patients including 11 with copy number alterations, 9 with sequence variants, and 7 with KIAA1549::BRAF fusions. Positive findings were obtained in patients spanning histologic subtypes, tumor grades, and anatomic locations. Conclusions: This study demonstrates the feasibility of employing this platform in routine clinical care in upfront diagnostic and monitoring settings. Future studies are required to determine the utility of this approach for assessing response to therapy and long-term surveillance.

Combined low-pass whole genome and targeted sequencing in liquid biopsies for pediatric solid tumors.

We designed a liquid biopsy (LB) platform employing low-pass whole genome sequencing (LP-WGS) and targeted sequencing of cell-free (cf) DNA from plasma to detect genome-wide copy number alterations (CNAs) and gene fusions in pediatric solid tumors. A total of 143 plasma samples were analyzed from 19 controls and 73 patients, including 44 bone or soft-tissue sarcomas and 12 renal, 10 germ cell, five hepatic, and two thyroid tumors. cfDNA was isolated from plasma collected at diagnosis, during and after therapy, and/or at relapse. Twenty-six of 37 (70%) patients enrolled at diagnosis without prior therapy (radiation, surgery, or chemotherapy) had circulating tumor DNA (ctDNA), based on the detection of CNAs from LP-WGS, including 18 of 27 (67%) patients with localized disease and eight of 10 (80%) patients with metastatic disease. None of the controls had detectable somatic CNAs. There was a high concordance of CNAs identified by LP-WGS to CNAs detected by chromosomal microarray analysis in the matching tumors. Mutations identified in tumor samples with our next-generation sequencing (NGS) panel, OncoKids®, were also detected by LP-WGS of ctDNA in 14 of 26 plasma samples. Finally, we developed a hybridization-based capture panel to target EWSR1 and FOXO1 fusions from patients with Ewing sarcoma or alveolar rhabdomyosarcoma (ARMS), respectively. Fusions were detected in the plasma from 10 of 12 patients with Ewing sarcoma and in two of two patients with ARMS. Combined, these data demonstrate the clinical applicability of our LB platform to evaluate pediatric patients with a variety of solid tumors.

The spectrum of mitochondrial DNA (mtDNA) mutations in pediatric CNS tumors.

BACKGROUND: We previously established the landscape of mitochondrial DNA (mtDNA) mutations in 23 subtypes of pediatric malignancies, characterized mtDNA mutation profiles among these subtypes, and provided statistically significant evidence for a contributory role of mtDNA mutations to pediatric malignancies. METHODS: To further delineate the spectrum of mtDNA mutations in pediatric central nervous system (CNS) tumors, we analyzed 545 tumor-normal paired whole-genome sequencing datasets from the Children's Brain Tumor Tissue Consortium. RESULTS: Germline mtDNA variants were used to determine the haplogroup, and maternal ancestry, which was not significantly different among tumor types. Among 166 (30.5%) tumors we detected 220 somatic mtDNA mutations, primarily missense mutations (36.8%), as well as 22 loss-of-function mutations. Different pediatric CNS tumor subtypes had distinct mtDNA mutation profiles. The number of mtDNA mutations per tumor ranged from 0.20 (dysembryoplastic neuroepithelial tumor [DNET]) to 0.75 (meningiomas). The average heteroplasmy was 10.7%, ranging from 4.6% in atypical teratoid/rhabdoid tumor (AT/RT) to 26% in diffuse intrinsic pontine glioma. High-grade gliomas had a significant higher number of mtDNA mutations per sample than low-grade gliomas (0.6 vs 0.27) (P = .004), with almost twice as many missense mtDNA mutations per sample (0.24 vs 0.11), and higher average heteroplasmy levels (16% vs 10%). Recurrent mtDNA mutations may represent hotspots which may serve as biologic markers of disease. CONCLUSIONS: Our findings demonstrate varying contributions of mtDNA mutations in different subtypes of CNS tumors. Sequencing the mtDNA genome may ultimately be used to characterize CNS tumors at diagnosis and monitor disease progression.

Pediatric brain tumors: the era of molecular diagnostics, targeted and immune-based therapeutics, and a focus on long term neurologic sequelae.

Pediatric brain tumors are the second most common malignancy of childhood after acute leukemia and the number one cause of cancer-related mortality1. Over the past decade, advanced molecular diagnostics have led to the discovery of new molecularly-defined tumor types with prognostic and therapeutic implications. Methylation profiles, whole genome sequencing, and transcriptomics have defined subgroups and revealed heterogeneity within commonly defined tumor entities2,3. These findings have also revealed important differences between adult and pediatric brain tumors of similar histology. The majority of pediatric low grade gliomas (pLGG) are defined by alterations in the mitogen-activated protein kinase (MAPK) pathway including BRAFV600E point mutation, K1AA1549-BRAF fusion, and FGFR1 alterations as opposed to IDH1/2 mutations and 1p/19q co-deletion seen more frequently in adult low grade gliomas4. These findings have led to targeted therapies, namely BRAF and MEK inhibitors, which are currently being evaluated in phase III clinical trials and may soon supplant chemotherapy as standard of care for pLGG's. While targeted therapy trials for pediatric brain tumors have had significant success, immunotherapy remains a challenge in a group of tumors with generally lower mutational burden compared to adult tumors and relatively "cold" immune microenvironment. Despite this, a wide array of immunotherapy trials including vaccine therapies, immune checkpoint blockade, chimeric antigen receptor (CAR) T cells, and viral therapies are on-going. Unique to pediatrics, multiple clinical trials have sought to answer the question of whether the most malignant pediatric brain tumors in the youngest patients can be successfully treated with high dose chemotherapy in lieu of radiation to avoid devastating long-term neurocognitive deficits. Due to the collaborative work of multiple pediatric neuro-oncology consortiums, the recent history of pediatric brain tumor research is one of efficient translation from bench to bedside in a rare group of tumors resulting in significant progress in the field. Here, advances in the areas of molecular characterization, targeted and immune-based therapies, and reduction in long term co-morbidities will be reviewed.

Transient juvenile myelomonocytic leukemia in the setting of PTPN11 mutation and Noonan syndrome with secondary development of monosomy 7.

Juvenile myelomonocytic leukemia (JMML) is a rare childhood neoplasm with poor prognosis except in the setting of Noonan syndrome, where prognosis is generally favorable. We present the case of a child with JMML in the setting of germline PTPN11 mutation and Noonan syndrome with suspected secondary development of monosomy 7 in the bone marrow. Diagnosed shortly after birth, she has been managed with active surveillance alone. Myeloblast percentages initially fluctuated; however, bone marrow biopsy at 4 years of age showed spontaneous remission despite persistence of the monosomy 7 clone, supporting a cautious approach in similar cases.

Personalizing the treatment of pediatric medulloblastoma: Polo-like kinase 1 as a molecular target in high-risk children.

Medulloblastoma is the most common malignant brain tumor in children. This disease is heterogeneous and is composed of four subtypes of medulloblastoma [WNT, Sonic Hedgehog (SHH), Group 3, and Group 4]. An immediate goal is to identify novel molecular targets for the most aggressive forms of medulloblastoma. Polo-like kinase 1 (PLK1) is an oncogenic kinase that controls cell cycle and proliferation, making it a strong candidate for medulloblastoma treatment. In this study, pediatric medulloblastomas were subtyped in two patient cohorts (discovery cohort, n = 63 patients; validation cohort, n = 57 patients) using NanoString nCounter analysis and PLK1 mRNA was assessed. We determined that the SHH and Group 3 subtypes were independently associated with poor outcomes in children as was PLK1 using Cox regression analyses. Furthermore, we screened a library of 129 compounds in clinical trials using a model of pediatric medulloblastoma and determined that PLK1 inhibitors were the most promising class of agents against the growth of medulloblastoma. In patient-derived primary medulloblastoma isolates, the PLK1 small-molecule inhibitor BI2536 suppressed the self-renewal of cells with high PLK1 but not low PLK1 expression. PLK1 inhibition prevented medulloblastoma cell proliferation, self-renewal, cell-cycle progression, and induced apoptosis. In contrast, the growth of normal neural stem cells was unaffected by BI2536. Finally, BI2536 extended survival in medulloblastoma-bearing mice with efficacy comparable with Headstart, a standard-of-care chemotherapy regimen. We conclude that patients with medulloblastoma expressing high levels of PLK1 are at elevated risk. These preclinical studies pave the way for improving the treatment of medulloblastoma through PLK1 inhibition.