Duncan Stearns

Alex's Lemonade Stand Foundation Infant and Childhood Primary Brain and Central Nervous System Tumors Diagnosed in the United States in 2007-2011

The CBTRUS Statistical Report: Alexs Lemonade Stand Foundation Infant and Childhood Primary Brain and Central Nervous System Tumors Diagnosed in the United States in 2007–2011 comprehensively describes the current population-based incidence of primary malignant and non-malignant brain and CNS tumors in children ages 0–14 years, collected and reported by central cancer registries covering approximately 99.8% of the United States population (for 2011 only, data were available for 50 out of 51 registries). Overall, brain and CNS tumors are the most common solid tumor, the most common cancer, and the most common cause of cancer death in infants and children 0–14 years. This report aims to serve as a useful resource for researchers, clinicians, patients, and families.

Cdk5 disruption attenuates tumor PD-L1 expression and promotes antitumor immunity

Cyclin suppresses antitumor immunity Despite the dramatic success of cancer immunotherapy, many types of cancer do not respond. Understanding why could help us to find ways to enhance the overall responsiveness of tumors to immunotherapies. Dorand et al. report that cyclin-dependent kinase 5 (Cdk5), an enzyme that is highly expressed by neurons in many brain cancers, may dampen the ability of T cells to reject tumors. In a mouse model of medulloblastoma, if tumors were Cdk5 deficient, T cells were able to remove them. This heightened antitumor immunity correlated with reduced expression of the inhibitory molecule programmed cell death ligand 1 (PD-L1), a target of current cancer immunotherapies. Science, this issue p. 399 Cyclin-dependent kinase 5 expression by tumors inhibits antitumor immunity in human and mouse medulloblastoma. Cancers often evade immune surveillance by adopting peripheral tissue– tolerance mechanisms, such as the expression of programmed cell death ligand 1 (PD-L1), the inhibition of which results in potent antitumor immunity. Here, we show that cyclin-dependent kinase 5 (Cdk5), a serine-threonine kinase that is highly active in postmitotic neurons and in many cancers, allows medulloblastoma (MB) to evade immune elimination. Interferon-γ (IFN-γ)–induced PD-L1 up-regulation on MB requires Cdk5, and disruption of Cdk5 expression in a mouse model of MB results in potent CD4+ T cell–mediated tumor rejection. Loss of Cdk5 results in persistent expression of the PD-L1 transcriptional repressors, the interferon regulatory factors IRF2 and IRF2BP2, which likely leads to reduced PD-L1 expression on tumors. Our finding highlights a central role for Cdk5 in immune checkpoint regulation by tumor cells.

American Brain Tumor Association Adolescent and Young Adult Primary Brain and Central Nervous System Tumors Diagnosed in the United States in 2008-2012.

Case Comprehensive Cancer Center, Case Western Reserve University School of Medicine, Cleveland, OH USA Central Brain Tumor Registry of the United States, Hinsdale, IL USA Department of Pediatric Hematology-Oncology, Rainbow Babies and Children s Hospital, Cleveland, OH USA Division of Hematology, Oncology and BMT, Nationwide Children’s Hospital and The Ohio State University, Columbus, OH USA School of Public Health, University of Memphis, Memphis, TN USA Keck School of Medicine, University of Southern California, Los Angeles, CA USA Department of Pediatric Hematology and Oncology, Cleveland Clinic Children’s Hospital, Cleveland, OH USA Solon High School, Solon, OH USA

Silencing of Thrombospondin-1 Is Critical for Myc-Induced Metastatic Phenotypes in Medulloblastoma

Mechanisms by which c-Myc (Myc) amplification confers aggressive medulloblastoma phenotypes are poorly defined. Here, we show using orthotopic models that high Myc expression promotes cell migration/invasion and induces metastatic tumors, which recapitulate aggressive histologic features of Myc-amplified primary human medulloblastoma. Using ChIP-chip analysis, we identified cell migration and adhesion genes, including Tsp-1/THBS1, ING4, PVRL3, and PPAP2B, as Myc-bound loci in medulloblastoma cells. Expression of Tsp-1 was most consistently and robustly diminished in medulloblastoma cell lines and primary human tumors with high Myc expression (n = 101, P = 0.032). Strikingly, stable Tsp-1 expression significantly attenuated in vitro transformation and invasive/migratory properties of high Myc-expressing medulloblastoma cells without altering cell proliferation, whereas RNA interference-mediated Myc knockdown was consistently accompanied by increased Tsp-1 levels and reduced cell migration and invasion in medulloblastoma cells. Chromatin immunoprecipitation (ChIP) assays revealed colocalization of Myc and obligate partner Max and correlated diminished RNA polymerase II occupancy (∼3-fold decrease, P < 0.01) with increased Myc binding at a core Tsp-1 promoter. Reporter gene and/or gel shift assays confirmed direct repression of Tsp-1 transcription by Myc and also identified JPO2, a Myc interactor associated with metastatic medulloblastoma, as a cofactor in Myc-mediated Tsp-1 repression. These findings indicate the Myc-regulatory network targets Tsp-1 via multiple mechanisms in medulloblastoma transformation, and highlight a novel critical role for Tsp-1 in Myc-mediated aggressive medulloblastoma phenotypes.

Hepatoblastoma in an 11-year-old: Case report and a review of the literature

Rationale: Hepatoblastoma is a rare malignancy. Approximately 100 cases are diagnosed yearly in the United States. The highest incidence occurs in infants and in children younger than 5 years. Cases involving patients older than 5 years are very rare. We describe the case of a patient who was diagnosed with hepatoblastoma at an atypical age of presentation for this type of malignancy. We also performed Ovid MEDLINE search for hepatoblastoma and epidemiology reports occurring in children between the ages of 5 and 18 years. In this article we review the epidemiology and summarize case reports published between 1997 and 2012 of patients with hepatoblastoma, who were older than 5 years. Patient concerns and diagnosis: Our patient is an 11 year old boy with stage IV hepatoblastoma with lung and omental metastases at diagnosis. Interventions: The patient received 7 cycles of chemotherapy following the treatment plan of COG protocol AHEP 0731, off study. He also had tumor resection and omentectomy and achieved complete remission. Outcomes: He later had disease recurrence and after undergoing treatment with different modalities, ultimately died of his disease. Review of SEER program data shows that the incidence of hepatoblastoma in children above the age of 5 years is too infrequent to be calculated. Literature review revealed 13 cases of patients diagnosed at age older than 5 years. Most cases were published due to unusual associations and/or complications. There are no obvious unifying characteristics for these cases, although there may be a slight male preponderance and many patients in this selected series presented with elevated Alpha-fetoprotein. Lessons: The reported case is rare, given the very low incidence of hepatoblastoma outside of infancy. A systematic review of characteristics and outcomes for patients older than 5 years who are enrolled in cooperative group hepatoblastoma trials may reveal important information about the epidemiology and tumor biology in this rare patient population.

DONOR SKIN ALLOGRAFT SURVIVAL AFTER BONE MARROW TRANSPLANTATION: CASE REPORT AND SYSTEMATIC REVIEW OF THE LITERATURE

BACKGROUND We present a case of skin allograft survival in a patient who previously received a bone marrow transplant from the same HLA-matched donor. DNA fingerprinting of skin biopsies showed mixed cellularity originating from the donor and recipient (68% and 32% donor DNA in the allograft skin and the native recipients skin, respectively). Histologic sections demonstrated both grade 3/4 rejection and graft-versus-host-disease. We have conducted a systematic review in search for other cases of donor skin allograft survival after a bone marrow or hematopoietic stem cell transplantation. METHODS All reported cases in English, Spanish, French, and German were captured using the electronic databases. Bibliographies of relevant articles were manually searched. RESULTS Nineteen patients (12 females) who received skin allografts from their bone marrow or hematopoietic stem cell donors were identified. Average age was 27.2 years (range: 5 months to 64 years). Skin allografts were used to treat graft-versus-host-disease, Herlitz junctional epidermolysis bullosa, and to test tolerance before a kidney transplantation from the same donor. Eight cases were not receiving immunosuppressive therapy. Allografts survived in all patients. In three patients, skin punch biopsies were taken, and these biopsies demonstrated mixed donor and recipient cellularity. The pathology result is specified in two more cases, with no signs of rejection. CONCLUSIONS The same donor skin allografts may be a safe option to treat severe cutaneous conditions in recipients of a bone marrow/hematopoietic stem cell transplantation. However, future studies are needed to confirm these results.

Abstract A050: Cdk5 disruption attenuates tumor PD-L1 expression via regulation of IFN-γ signaling components and promotes antitumor immunity

In order to escape normal immune surveillance, tumors mimic peripheral tissue tolerance mechanisms such as the expression of programmed cell death-ligand 1 (PD-L1), the inhibition of which can lead to potent anti-tumor responses. Here we show the contribution of cyclin dependent kinase 5 (Cdk5) to immune evasion. Cdk5 is a proline-directed serine/threonine kinase, which is highly expressed in post-mitotic neurons and directs a variety of homeostatic functions from cytoskeletal rearrangement to neurotransmitter signaling. Studies in xenograft models have linked Cdk5 activity to the generation and metabolic activity of primary tumors within the central nervous system (CNS). Here we show that Cdk5, expressed in both murine and human medulloblastoma (MB) cell lines, plays a major role in the ability of MB to avoid immune detection. First, we observed that decreased Cdk5 expression was associated with an increase in the number of infiltrating CD3+ cells in the tumor mass of clinical MB samples. Additionally, using publicly available datasets, we found that decreased Cdk5 expression is correlated with better overall survival or fewer distant metastasis in melanoma, breast cancer, glioma, and lung adenocarcinoma. Next, using a CRISPR/Cas-9 approach, we silenced Cdk5 in a murine model of Sonic Hedgehog (SHH) pathway MB. Rather than causing intrinsic growth defects in MB, interference of Cdk5 activity sensitizes tumors to killing by the normal host immune system in a CD4+ T cell-dependent manner. This rejection is associated with increased IFN-γ expression in the tumor microenvironment, as well as increased PD-L1 expression by myeloid populations in both subcutaneous and intracranial tumors. Mechanistically, we observed an attenuated response to IFN-γ stimulated expression of programmed death ligand 1 (PD-L1) on MB cells. This blunted response was recapitulated when MB cells were treated with Roscovitine, a non-selective Cdk5 inhibitor. Furthermore, using a quantitative global phosphoproteomics approach, we found that Cdk5 knockdown also increased phosphorylation of S-440 and S-443 on interferon regulatory factor binding protein 2 (IRF2BP2), an upstream co-repressor of interferon regulatory factor 2 (IRF-2). Increased phosphorylation of IRF2BP2 corresponded with stable expression of IRF-2 with a concomitant decrease in surface expression of PD-L1. These observations highlight a critical role for Cdk5 in the immune escape mechanisms of primary CNS tumors and provides new therapeutic targets for PD-1/PD-L1 directed immunotherapy. Citation Format: Rodney D. Dorand, Jr., Joseph Nthale, Jay T. Myers, Deborah S. Barkauskas, Stefanie Avril, Steven M. Chirieleison, Tej K. Pareek, Derek W. Abbott, Duncan S. Stearns, John J. Letterio, Alex Y. Huang, Agne Petrosiute. Cdk5 disruption attenuates tumor PD-L1 expression via regulation of IFN-γ signaling components and promotes antitumor immunity [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A050.

Abstract B158: Cyclin dependent kinase-5 regulates IFNγ induced PD-L1 expression via IRF-1 in medulloblastoma

Immune checkpoint blockade offers an exciting, new adjuvant therapy to both solid and liquid tumors. However, the underlying mechanisms by which tumors alter their expression of immune checkpoint blockade ligands, such as programmed death ligand 1 (PD-L1), to achieve adaptive immune resistance needs to be further studied. We sought to investigate the mechanisms by which PD-L1 expression is regulated in response to IFNγ stimulation in medulloblastoma, a primary CNS neoplasm. MB is the most common pediatric CNS malignancy that is capable of metastasis to distant sites. MB is classified into four different molecular subgroups. Patched (PTCH), a signaling molecule of the Sonic Hedgehog (SHH) pathway, represents one subgroup and accounts for 25% of all sporadic human MB. Because MB develops from primitive cerebellar granule neuron precursor cells, it is thought that errors in development or migration of these neurons contribute to tumorigenesis. Cyclin Dependent Kinase 5 (CDK5) is a non-canonical member of the CDK family that does not require cyclin for activity but relies on binding to its obligate activators, p35 and p39 which are mostly expressed in post mitotic neurons. CDK5 is required for normal Central Nervous System (CNS) development. Utilizing both short hairpin RNA (shRNA) approach and the bacterial CRISPR-Cas9 (Cr) system we silenced Cdk5 expression in a C57BL/6 (B6) syngeneic MB line derived from p53-/- PTCH+/- (MM1). In vitro proliferation experiments revealed that MM1-WT, MM1 cells transduced with shRNA targeting CDK5 (MM1-shCDK5), and MM1 cells transduced with non-silencing, scrambled RNA sequences (MM1-shNS) had similar growth kinetics. Since there were no intrinsic differences in in vitro growth kinetics, we subcutaneously injected immunocompetent C57BL/B6 mice with 5x104 tumor cells and measured tumor growth every three days. In our first experiment, 19 days after injection 7.7% (N=13) of mice injected with MM1-WT and 0% (N=7) injected with MM1-shNS were tumor free compared with 76.5% (N=13) of animals injected with MM1-shCDK5 Comparison of tumor free survival revealed statistically significant differences between the MM1-WT with MM1-shCDK5 groups (p In summary, we have shown that Cdk5 is expressed in Type 2 MB and that silencing of its expression with shRNA results in an altered growth phenotype in vivo but not in vitro. Furthermore, eliminating the ability of MM1 tumors to express PD-L1 might lead to effective immune rejection of MM1-shCDK5. Mechanistically, we showed that this enhanced tumor rejection may be due in part to the failure of Cdk5 deficient MB cells to alter IRF-1 expression which serves as a transcription factor for PD-L1. These novel findings point to CDK5 as a potential immunotherapeutic target in MB. Citation Format: Rodney Dixon Dorand, Jr., Agne Petrosiute, Joseph Nthale, Tej Pareek, Francesca Scrimieri, Jay Myers, Deborah Barkauskas, John J. Letterio, Duncan S. Stearns, Alex Y. Huang. Cyclin dependent kinase-5 regulates IFNγ induced PD-L1 expression via IRF-1 in medulloblastoma. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B158.