bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A28: Donor lymphocyte infusion after allogeneic stem cell transplantation is a feasible therapy option with acceptable toxicity rates in patients with refractory Ewing’s sarcoma and rhabdomyosarcoma
Authors: Schober Sebastian J. , Steinhauser Maximilian , Wawer Angela , Luettichau Irene Teichert-von , Salat Christoph , Issels Rolf D. , Schwinger Wolfgang , Ussowicz Marek , Antunovic Petar , Castagna Luca , Kolb Hans-Jochem , Burdach Stefan E.G. , Thiel Uwe
Published: February 2019

Background: New therapy options are urgently needed for patients with treatment-refractory Ewing’s sarcoma (ES) or rhabdomyosarcoma (RMS). In these subgroups, the role of allogeneic stem cell transplantation (allo-SCT) to induce a graft-versus-tumor effect (GvT) remains unclear. Here, we describe our experience and the general feasibility of donor lymphocyte infusion (DLI) following allo-SCT for those patients.

Patients and Methods: We retrospectively evaluated data of eight patients with treatment-refractory ES (ES #1-4) and RMS (RMS #1-4) after DLI following allo-SCT. Data were individually evaluated for presence of graft-versus-host disease (GvHD), relapse-free survival (RFS), and overall survival (OS). Three of four ES and one of four RMS patients had received haploidentical grafts; the remaining patients received HLA-matched grafts prior to DLI. Patients received donor lymphocytes ranging from 2.5 x 104 to 1 x 108 CD3+ cells/kg body weight.

Results: ES #4 and RMS #4 developed acute GvHD after DLI. Limited chronic GvHD was observed in RMS #3. In two patients, DLI was associated with stable disease for nine (ES #2) and six months (ES #4), respectively. RMS #4 showed partial remission lasting for eight months after one dose of DLI with 1 x 106 CD3+ cells/kg combined with local hyperthermia and chemotherapy. ES #4 had residual disease before allo-SCT and was converted to CR after DLI. Altogether, seven patients died of disease and none of toxicity. RMS #2 received seven doses up to 1 x 108 CD3+ cells/kg and IL-2 including surgery and chemotherapy and remained in CR for 97 months at the date of data assessment. Median follow-up after allo-SCT was 27.5 months.

Conclusion: DLI after allo-SCT is a feasible therapy option for treatment-refractory ES and RMS patients. In this analysis, DLI-related toxicity is acceptable. These findings have to be evaluated in prospective analyses.

Citation Format: Sebastian J. Schober, Maximilian Steinhauser, Angela Wawer, Irene Teichert-von Luettichau, Christoph Salat, Rolf D. Issels, Wolfgang Schwinger, Marek Ussowicz, Petar Antunovic, Luca Castagna, Hans-Jochem Kolb, Stefan E.G. Burdach, Uwe Thiel. Donor lymphocyte infusion after allogeneic stem cell transplantation is a feasible therapy option with acceptable toxicity rates in patients with refractory Ewing’s sarcoma and rhabdomyosarcoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr A28.
Abstract A42: Prexasertib (LY2606368) is highly active as a monotherapy or in combination with chemotherapy in preclinical models of pediatric bone and soft tissue sarcoma
Authors: Lowery Caitlin D. , Blosser Wayne , Dowless Michele , Renschler Matthew , Stephens Jennifer , Beckmann Richard , Stancato Louis
Published: February 2019

The serine/threonine kinase checkpoint kinase 1 (CHK1) is a crucial mediator of replication stress and helps maintain genome integrity by governing the S-phase and G2/M cell cycle checkpoints following DNA damage, facilitating DNA repair through homologous recombination, and licensing replication forks during S phase. The CHK1 small-molecule inhibitor prexasertib (LY2606368) is currently under clinical investigation in pediatric cancer patients with recurrent or refractory solid tumors (NCT02808650). Reduced proliferation was observed at low nanomolar concentrations in a panel of pediatric bone and soft tissue sarcoma cell lines. In addition, prexasertib caused DNA damage and cell death in vitro as measured by Western blot analysis and high content imaging. Prexasertib as a monotherapy was superior to doxorubicin or actinomycin D in several cell line-derived and patient-derived xenograft (PDX) models of pediatric soft tissue sarcoma, with two models of alveolar rhabdomyosarcoma (aRMS) demonstrating a complete response to prexasertib; however, acquired resistance to prexasertib was observed. In addition, preclinical PDX mouse models of Ewing’s sarcoma and osteosarcoma typically demonstrated intrinsic resistance to single-agent prexasertib. Importantly, combination with cytotoxic chemotherapy could not only overcome intrinsic prexasertib resistance in vivo, as would be expected given the potential for prexasertib to act as a chemopotentiator, but also prevented the emergence of acquired resistance following single-agent prexasertib treatment. In vitro experiments were performed to evaluate the effect of prexasertib plus chemotherapy on cell proliferation. Future preclinical studies include genomic analysis of pediatric bone and soft tissue sarcoma models to delineate the molecular underpinnings of sensitivity and acquired resistance to prexasertib and to identify predictive biomarkers for response.

Citation Format: Caitlin D. Lowery, Wayne Blosser, Michele Dowless, Matthew Renschler, Jennifer Stephens, Richard Beckmann, Louis Stancato. Prexasertib (LY2606368) is highly active as a monotherapy or in combination with chemotherapy in preclinical models of pediatric bone and soft tissue sarcoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr A42.
Abstract A46: Heterotypic nucleosomes and PRC2 role in diffuse intrinsic pontine gliomas
Authors: Piunti Andrea
Published: February 2019

Diffuse intrinsic pontine gliomas (DIPG) harbor a characteristic lysine-to-methionine mutation on histone H3 (H3K27M) that potently inhibits Polycomb Repressive Complex 2 (PRC2) enzymatic activity; however, the precise roles of H3K27M and PRC2 in tumorigenesis remain elusive. Here we perform epigenomic profiling of patient-derived H3K27M mutant DIPG cell lines and demonstrate that H3K27M resides in nucleosomes with H3K27acetylation (H3K27ac) and colocalizes with bromodomain proteins at actively transcribed genes. In contrast, PRC2 is excluded from H3K27M occupied regions but, strikingly, the PRC2 complex is required for maintaining oncogenic potential. Finally, we demonstrate that pharmacologic bromodomain inhibition suppresses tumor growth in vivo. Altogether our results indicate that H3K27M drives accumulation of H3K27ac and point to bromodomain proteins inhibition as a powerful clinical strategy against DIPG.

Citation Format: Andrea Piunti. Heterotypic nucleosomes and PRC2 role in diffuse intrinsic pontine gliomas [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr A46.
Abstract B02: Distinct CD8+ T-cell recognition profiles driven by conformational plasticity of recurrent neuroblastoma neoepitopes
Authors: Salama Sofie R. , Toor Jugmohit S. , Rao Arjun A. , McShan Andrew C. , Yarmarkovich Mark , Tripathi Sarvind , Nerli Santrupti , Madejska Ada A. , Nguyen Son , Yamaguchi Karissa , Maris John M. , Haussler David , Sgourakis Nikolaos G.
Published: February 2019

The identification of tumor neoantigens driving T-cell responses for immunotherapy poses a significant bottleneck in the development of approaches for precision cancer therapeutics. Here, we employ a bioinformatics method, Prediction Of T Cell Epitopes for Cancer Therapy (ProTECT), using sequencing data from neuroblastoma patients to identify a recurrent Anaplastic Lymphoma Kinase mutation (ALK R1275Q) that leads to two high-affinity neoepitopes when expressed in complex with common HLA alleles. Analysis of the x-ray structures of nonamer and decamer peptides bound to HLA-B*15:01 reveals that the neoepitopes adopt distinct conformations that confer measurable differences in stability in vitro, while the self-antigen is excluded from binding due to steric hindrance. MHC tetramer staining of peripheral blood mononuclear cells from HLA-matched donors shows that the two neoepitopes elicit distinct CD8+ T-cell recognition profiles. To expand the range of patients who could mount a T-cell response to our ALK neoepitopes, we additionally characterize HLA binding repertoire using Rosetta comparative modeling simulations. Subsequent determination of the x-ray structure of decamer-bound HLA-A*01:01 further validates the Rosetta predictions. This work provides a novel approach towards robust, high-throughput identification and detailed characterization of highly stable neoantigen/HLA targets with desired T-cell recognition features for cancer immunotherapy.

Citation Format: Sofie R. Salama, Jugmohit S. Toor, Arjun A. Rao, Andrew C. McShan, Mark Yarmarkovich, Sarvind Tripathi, Santrupti Nerli, Ada A. Madejska, Son Nguyen, Karissa Yamaguchi, John M. Maris, David Haussler, Nikolaos G. Sgourakis. Distinct CD8+ T-cell recognition profiles driven by conformational plasticity of recurrent neuroblastoma neoepitopes [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B02.
Abstract B09: IL6 and CXCL8 mediate redundant tumor-host signaling pathways that facilitate osteosarcoma lung metastasis
Authors: Roberts Ryan D. , Cam Hakan , Gross Amy C. , Winget Sarah , Phelps Doris , Houghton Peter J.
Published: February 2019

Osteosarcoma (OS) kills patients through aggressive metastatic spread, primarily to lung tissues. Our lab has sought to understand that tumor-host interactions that mediate this propensity for OS cells to colonize and grow in the lung. We identified two factors, IL6 and CXCL8, that become overexpressed in metastatic OS nodules relative to the same patient’s primary lesions. Expression of these factors correlates strongly with capacity to colonize lung tissue in murine models of OS metastasis. Aberrant expression of ΔNp63 drives production of IL6 and CXCL8 and can endow poorly metastatic OS tumors with increased metastatic potential. This effect can be completely reversed by blocking IL6 and CXCL8. Treating mice inoculated with circulating OS cells with inhibitors of IL6 and CXCL8 signaling reduces lung metastasis. This effect completely blocks lung colonization in some models, but only when used in combination. The effects of IL6 and CXCL8 inhibition endure for many months beyond discontinuation of therapy. Genetic manipulation studies confirm that these effects result from on-target effects of the drugs. Clinical studies performed in canine patients will help determine whether these treatments might be effective adjuvants for the treatment of patients with OS.

Citation Format: Ryan D. Roberts, Hakan Cam, Amy C. Gross, Sarah Winget, Doris Phelps, Peter J. Houghton. IL6 and CXCL8 mediate redundant tumor-host signaling pathways that facilitate osteosarcoma lung metastasis [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B09.
Abstract B17: ADGRB3 is a novel tumor suppressor epigenetically silenced in WNT medulloblastoma
Authors: Bhattacharya Debanjan , Zhu Dan , Devi Narra , Meir Erwin G. Van
Published: February 2019

Medulloblastoma (MB) is the most aggressive primary malignant brain tumor in children and arises from neural progenitor cells in the developing cerebellum. Although some subtypes of MB show a favorable prognosis with treatment, still one third of patients succumb to this disease and the children who survive suffer from long-term side effects of the aggressive treatments. ADGRB3 (formerly called BAI3) is a member of the ADGRB1-3 subfamily of adhesion GPCR transmembrane proteins, which are highly expressed in the brain specially in cerebellum and hippocampal neurons. The ADGRB3 gene was recently found to undergo somatic mutations in several cancers, but nothing is known about the functional role of ADGRB3 in medulloblastoma. Our recent findings based on analysis of a published microarray dataset and RT-PCR experiments showed that ADGRB3 mRNA expression was selectively repressed in WNT-MB tumor tissue compared to other three molecular subgroups and normal human cerebellar tissue. Using bisulfite sequencing we have detected hypermethylation of the ADGRB3 promoter exclusively in WNT-MB subgroup of human MB tissue but not in the other three molecular subgroups and normal human cerebellar tissue. These results clearly indicate that ADGRB3 is epigenetically silenced in WNT-MB via promoter hypermethylation. In vitro experiments showed that restoration of ADGRB3 expression in ADGRB3-silenced MB cell lines slows cell growth and inhibits WNT signaling targets. We further analyzed whether methyl CpG binding proteins and histone modifications modulate transcriptional repression of ADGRB3 in WNT-MB. ChIP assays revealed enrichment of repressive MBD2 protein in the ADGRB3 promoter region. siRNA mediated knockdown of ADGRB3 in ADGRB3-silent MB cell line (UW288-1) was found to reactivate ADGRB3 expression. We found that in vitro treatment with KCC-07, a recently discovered MBD2 inhibitor, reactivates ADGRB3 mRNA expression in ADGRB3-silent but not expressing MB cells. We are currently investigating whether ADGRB3 can negatively regulate the WNT/beta-catenin pathway or acts via an independent pathway that synergizes with it to facilitate transformation. Altogether, our findings define an epigenetic mechanism for ADGRB3 silencing in WNT-MB and indicate a novel role of ADGRB3 as a potent suppressor of oncogenic cerebellar transformation. Our findings also highlight the potential of epigenetic reactivation of ADGRB3 as a less toxic therapeutic intervention for the children suffering from WNT-MB.

Citation Format: Debanjan Bhattacharya, Dan Zhu, Narra Devi, Erwin G. Van Meir. ADGRB3 is a novel tumor suppressor epigenetically silenced in WNT medulloblastoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B17.
Abstract B18: Modeling renal medullary carcinomas identifies druggable vulnerabilities in SMARCB1-deficient cancers
Authors: Hong Andrew L. , Tseng Yuen-Yi , Kynnap Bryan D. , Doshi Mihir B. , Wala Jeremiah , Sandoval Gabriel , Church Alanna J. , Mullen Elizabeth , Kadoch Cigall , Roberts Charles W.M. , Beroukhim Rameen , Boehm Jesse S. , Hahn William C.
Published: February 2019

Renal medullary carcinomas (RMCs) are thought to be driven by the loss of tumor suppressor, SMARCB1. These rare kidney cancers carry a very poor prognosis and primarily affect African American adolescents and young adults with sickle cell trait. From two patients with RMC, we have identified by whole-genome sequencing mechanisms of SMARCB1 loss (e.g., inactivating fusion events involving SMARCB1). We developed in vitro models of primary and relapsed metastatic disease. We performed biochemical and functional studies to conclusively show that RMC is dependent on loss of SMARCB1, similar to rhabdoid tumors and atypical teratoid/rhabdoid tumors. Furthermore, we performed small-molecule screens, pooled CRISPR-Cas9 knockout, and RNAi suppression screens focused on druggable cancer targets. Integration of these orthogonal methods identifies a core set of targets that may provide a rational approach to therapeutic targeting for this rare kidney cancer and other SMARCB1-deficient cancers.

Citation Format: Andrew L. Hong, Yuen-Yi Tseng, Bryan D. Kynnap, Mihir B. Doshi, Jeremiah Wala, Gabriel Sandoval, Alanna J. Church, Elizabeth Mullen, Cigall Kadoch, Charles W.M. Roberts, Rameen Beroukhim, Jesse S. Boehm, William C. Hahn. Modeling renal medullary carcinomas identifies druggable vulnerabilities in SMARCB1-deficient cancers [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B18.
Abstract B21: Epigenetic modifiers MLL1 and JMJD3 regulate neuroblastoma tumorigenicity by maintaining a cancer stem cell-like population
Authors: Agarwal Saurabh , Tomolonis Julie , Vasudevan Sanjeev , Shohet Jason
Published: February 2019

High-risk neuroblastoma (NB) represents a major clinical challenge in pediatric oncology. Despite significant dose escalation of intense therapies, long-term survival for NB patients remains poor (<45%), and the disease accounts for almost 15% of all pediatric cancer deaths. Relapse of metastatic, drug-resistant disease and treatment-related toxicities mandates the development of new therapeutic strategies. Recently, we discovered a highly tumorigenic, chemoresistant, and self-renewing subpopulation in NB with features similar to cancer stem cells (CSCs). This G-CSF receptor (CD114)-expressing subpopulation can escape initial therapy and cause aggressively invasive relapsed disease. Therefore, developing direct targeting strategies and understanding for the molecular mechanisms maintaining this NB CSC-like subpopulation is essential to delineate effective therapeutics for NB patients. By using siRNA screening approach and low-density pathway arrays, we found that the epigenetic regulators mixed-lineage leukemia-1 (MLL1; KMT2A; a H3K4me3 methyltransferase) and Jumonji D3 (JMJD3; KDM6B; a H3K27me3 demethylase) are important epigenetic regulators for NB proliferation and growth. Interestingly, we found that epigenetic regulators MLL1 and JMJD3 are aberrantly expressed in CD114+ subpopulation and regulate the expression of the G-CSF receptor gene (CSF3R) itself, by maintaining active histone modifications at the promoter locus. Inhibition of MLL1 and JMJD3 with specific small-molecule inhibitors reverses the histone patterns at CSF3R promoter and blocks gene expression, induces apoptosis selectively in CD114+ cells, and inhibits overall NB proliferation in vitro. Furthermore, inhibiting MLL1 and JMJD3 leads to dramatic tumor regression (p<0.001) and reduction in incidences of metastasis (p<0.001) in vivo. Most interestingly, both of these inhibitors increase the overall survival of the treated mice in vivo. As expected, reduction in tumor size was significantly correlated with the reduction in tumor CD114+ cells. Taken together, these data highlight that: a) neuroblastoma is maintained by the complex interplay of epigenetic modifiers, and b) direct targeting of these epigenetic modifiers and combining this approach with current therapy is a novel therapeutic approach to high-risk NB.

Citation Format: Saurabh Agarwal, Julie Tomolonis, Sanjeev Vasudevan, Jason Shohet. Epigenetic modifiers MLL1 and JMJD3 regulate neuroblastoma tumorigenicity by maintaining a cancer stem cell-like population [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B21.
Abstract B22: Antitumor effects of selective PRMT1 inhibitors on neuroblastoma in vitro and in vivo
Authors: Hansen Jeanne N. , Lotta Louis T. , Eberhardt Allison , Zheng Yujun George , Ji Sen , Yang Shengyong , Schor Nina F. , Li Xingguo
Published: February 2019

The MYC family of transcription factors is one of the most frequently overexpressed oncogenes in human malignancies, including tumors that are derived from the nervous system. MYCN plays a causative role in neuroblastoma, a common and fatal childhood cancer of the developing sympathetic nervous system. Thus, the MYC family has been considered as a promising cancer target; however, so far, no small molecules can directly target MYC or MYCN in vivo. We have recently reported that PRMT1 functions as a key regulator of the stability and oncogenicity of MYCN. In this study, we have performed the structure-activity relationship analysis of diamidine-related PRMT1-seletive inhibitors. In correlation with their inhibition against PRMT1 in biochemical assays, these compounds showed remarkable anticancer properties both in neuroblastoma cell lines in vitro and in a preclinical mouse model of neuroblastoma in vivo. Mechanistically, these potent PRMT1 inhibitors destabilized MYCN, caused cell cycle arrest, and restored normal developmental signals of the developing sympathetic neuroblast precursors. Together, our results demonstrate that PRMT1-seletive inhibitors are promising anti-neuroblastoma agents and may represent a general strategy to target certain MYC/MYCN-driven cancers.

Citation Format: Jeanne N. Hansen, Louis T. Lotta, Allison Eberhardt, Yujun George Zheng, Sen Ji, Shengyong Yang, Nina F. Schor, Xingguo Li. Antitumor effects of selective PRMT1 inhibitors on neuroblastoma in vitro and in vivo [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B22.
Abstract B24: Characterization of somatically acquired regulatory mutations in high-risk neuroblastoma
Authors: Harenza Jo Lynne , Chen Gregory , Maris John M.
Published: February 2019

Neuroblastoma is a childhood cancer of the developing peripheral nervous system and is uniquely characterized by a broad spectrum of phenotypes ranging from spontaneous tumor regression to relentless progression. The 5-year survival rate for children in the low- and intermediate-risk groups is over 95%; however, children with high-risk disease have a 40-50% likelihood of survival despite aggressive multimodal therapy. High-risk neuroblastomas harbor relatively few protein coding mutations at diagnosis, with the oncogenic mechanisms underlying MYCN amplification and ALK activation remaining obscure. We therefore hypothesized that characterizing high-risk neuroblastoma noncoding genetic variation will define gene regulatory mechanisms driving tumor progression, and that somatically acquired noncoding mutations alter critical neurodevelopmental gene regulatory regions. Here, we have integrated neuroblastoma cell line histone ChIP-Seq (N = 10), ATAC-Seq (N = 12), and RNA-Seq (N = 39) with high-risk patient (N = 105) whole-genome mutation data to define the landscape of noncoding somatic events dysregulating enhancers and promoters in high-risk neuroblastoma. We found a significantly higher number of mutations per patient in high-risk and stage 4 patients, compared to other risk groups (Wilcox Whigh-low = 129.5, p = 1.47e-07; Whigh-int = 70, p = 3.66e-08) and stages (WStage4-Stage3 = 86, p = 2.68e-3; WStage4-Stage4s= 65.5, p = 1.21e-12). We found a median of 1,809 somatic noncoding single nucleotide mutations per patient, and a subset of these recurrently mutate sequences underlying super-enhancer peaks of 102 genes. An additional subset lie within CCCTC-binding factor (CTCF) motifs (a median of 5 mutations per patient), suggesting potential dysregulation of chromosome looping in neuroblastoma. Understanding noncoding drivers of oncogenic gene expression and translating these basic science discoveries to the clinic is a critical step towards changing the devastating course of this disease.

Citation Format: Jo Lynne Harenza, Gregory Chen, John M. Maris. Characterization of somatically acquired regulatory mutations in high-risk neuroblastoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B24.
Abstract B36: Targeting MDM4 in neuroblastoma to reactivate p53 signaling
Authors: Woodfield Sarah E. , Patel Roma H. , Ibarra Aryana M. , Chen Zhenghu , Vasudevan Sanjeev A.
Published: February 2019

Introduction: Neuroblastoma is the most common pediatric extracranial solid tumor. Despite aggressive therapeutic regimens, long-term survival of patients with high-risk disease is less than 50%. The TP53 tumor suppressor gene is wild-type and functional in 99% of neuroblastoma cases at diagnosis. Thus, we hypothesized that targeting the p53 regulator MDM4 to reactivate p53 signaling would be effective in neuroblastoma.

Methods: Levels of gene expression of MDM4 and its association with event-free and overall survival in neuroblastoma patients were analyzed using the publically available patient datasets in the R2 Genomics Analysis and Visualization Platform. MDM4 gene and protein expression were also examined in five neuroblastoma cell lines (IMR-32, SH-SY5Y, NB-19, SK-N-AS, and LAN-1) with qPCR and immunoblotting assays. The effects of MDM4 inhibitor NSC207895 on TP53 wild-type IMR-32 and SH-SY5Y, TP53 mutant SK-N-AS, and TP53 null LAN-1 cells were analyzed with MTT assays for cytotoxicity and qPCR experiments to look at p53 targets.

Results: MDM4 expression is significantly correlated with both event-free and overall survival of neuroblastoma patients in the Kocak (n=476) and Oberthuer (n=251) datasets in the R2 Genomics Analysis and Visualization Platform, with lower expression correlated with increased survival (p<0.01). All five neuroblastoma cell lines showed gene and protein expression of MDM4. NSC207895 had strong cytotoxic effects at very low concentrations (0.08-0.09 μM) after only 24 hours of treatment with TP53 wild-type IMR-32 and SH-SY5Y cells. NSC207895 exposure also led to upregulation of p53 targets Bax (3.7- to 4.3-fold), Puma (5.7- to 21.6-fold), and CDKN1A (14.5- to 53.1-fold) in these cells but not in TP53 mutant SK-N-AS or TP53 null LAN-1 cells.

Conclusions: Inhibition of MDM4 shows efficacy in preclinical models of neuroblastoma by upregulating p53 tumor suppressor signaling in TP53 wild-type cells and is a promising therapeutic strategy for high-risk patients with wild-type TP53.

Citation Format: Sarah E. Woodfield, Roma H. Patel, Aryana M. Ibarra, Zhenghu Chen, Sanjeev A. Vasudevan. Targeting MDM4 in neuroblastoma to reactivate p53 signaling [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B36.
Abstract B41: Modeling bone development and cancer predisposition in Diamond Blackfan anemia (DBA)
Authors: Hom Jimmy , Dulmovits Brian , Peters Luanne , Vlachos Adrianna , Lipton Jeffrey M. , Blanc Lionel
Published: February 2019

DBA is an inherited bone marrow failure syndrome characterized by pure red cell aplasia, congenital physical anomalies, and cancer predisposition, in particular an increased incidence of osteogenic sarcoma (O:E 42.44). In order to investigate the mechanism of defective bone development and ultimately oncogenesis, we used an in vitro model consisting of mouse embryonic stem cells gene trapped for the Ribosomal Protein 19 gene (Rps19) and developed an in vivo transgenic conditional mouse model with floxed Rps19 alleles and a bone-specific expressing Col1a1-CreER. Using a 23-day stepwise osteoblast differentiation model, haploinsufficient Rps19+/- mouse ES cells did not show differences in the ability to develop into mesodermal progenitors based on qPCR analysis of mesodermal transcription factors, but displayed a decreased ability to produce mature osteoblasts based upon decreased alizarin red staining. In addition, higher levels of the Trp53 protein were found in Rps19+/- mouse ES cells at both the pluripotent and mesodermal stages of differentiation. Further, Rps19+/- mouse ES cells presented with aberrant WNT signaling, as represented by decreased levels of β-catenin early in osteoblast differentiation and rising to higher levels towards late osteoblast differentiation. We assessed the hypothesis of WNT signaling involvement in lineage specification and found that Rps19+/- mouse ES cell-derived osteoblasts show decreased levels of Runx2 and increased levels of Sox9. In the transgenic mouse model, tamoxifen-induced recombination of Rps19 at E15-17 did not show gross morphologic skeletal defects at E20.5 for Rps19fl/+ embryos, although Rps19fl/fl embryos displayed severe skeletal defects through alizarin red and alcian blue staining. In conclusion, haploinsufficient levels of Rps19 found in mice contribute to improper skeletal development and probably reflect the clinical scenario. We plan to utilize these in vitro and in vivo models to provide clues into the mechanism of ribosomal protein haploinsufficiency in tumor development, which may suggest surveillance and treatment strategies.

Citation Format: Jimmy Hom, Brian Dulmovits, Luanne Peters, Adrianna Vlachos, Jeffrey M. Lipton, Lionel Blanc. Modeling bone development and cancer predisposition in Diamond Blackfan anemia (DBA) [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B41.
Abstract IA04: Identification of new vulnerabilities in fusion-driven pediatric cancers with functional and chemical genomic approaches
Authors: Stegmaier Kimberly
Published: February 2019

Fusion-driven pediatric cancers remain a therapeutic challenge. On one hand, the genomic simplicity of these malignancies and the tumor selectivity of their expressed fusion oncoproteins represent a therapeutic opportunity. These fusion oncoproteins, however, are typically transcription factors, and make for particularly challenging drug targets. Novel approaches to targeting these malignancies are needed. The EWS/ETS-driven pediatric solid tumor Ewing sarcoma is one exemplary childhood cancer. Our laboratory is integrating next-generation sequencing, functional genomic screening (shRNA and CRISPR/Cas9), and chemical genomic screening to identify, validate, and mechanistically dissect new targets in Ewing sarcoma. Emerging data from these efforts will be presented.

Citation Format: Kimberly Stegmaier. Identification of new vulnerabilities in fusion-driven pediatric cancers with functional and chemical genomic approaches [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr IA04.
Abstract PR11: Synthetic lethality between ATRX mutations and MYCN amplification in neuroblastoma
Authors: Zeineldin Maged , Federico Sara M. , Griffith Lyra , Jeon Jongyre , Chen Xiang , Easton John , Wu Jianrong , Mao Shenghua , Liu Yanling , Naranjo Arlene , Pappo Alberto S. , Hogarty Michael D. , Dyer Michael A.
Published: February 2019

Neuroblastoma is a clinically heterogeneous tumor responsible for 15% of cancer-related deaths in children. Older age at diagnosis and MYCN amplification are associated with poor prognosis in neuroblastoma. Previously, sequencing stage 4 neuroblastoma tumors revealed recurrent mutations in the chromatin remodeler ATRX that are significantly associated with older age and mutually exclusive with MYCN amplification. Here, we expanded our genetic analysis of neuroblastoma across all ages and stages. We sequenced 475 neuroblastoma tumors and found that the incidence of ATRX mutations is correlated with older age at diagnosis and advanced stages of the disease. We also directly studied the mutual exclusivity between ATRX mutations and MYCN amplification using both cultured cells and orthotopic mouse models. We found that targeting ATRX using shRNA or CRISPR-Cas-9 reduces cell viability in MYCN-amplified cells. Additionally, induction of MYCN overexpression results in cell death in ATRX-mutant neuroblastoma cells. To characterize the molecular basis of the ATRX-mutations/MYCN-amplification incompatibility, we studied epigenetic, metabolic, and replicative changes associated with MYCN amplification in these cells. We discovered that induction of MYCN overexpression provokes mitochondrial abnormalities, metabolic reprograming, and replicative stress in ATRX mutant cells. Collectively, these data show synthetic lethality between ATRX mutations and MYCN amplification and suggest that the chromatin remodeler ATRX is important in reducing replicative stress associated with MYCN amplification in neuroblastoma. These data will also help in identifying therapeutic targets, which will reduce the morbidity and mortality associated with high-risk neuroblastoma.

Citation Format: Maged Zeineldin, Sara M. Federico, Lyra Griffith, Jongyre Jeon, Xiang Chen, John Easton, Jianrong Wu, Shenghua Mao, Yanling Liu, Arlene Naranjo, Alberto S. Pappo, Michael D. Hogarty, Michael A. Dyer. Synthetic lethality between ATRX mutations and MYCN amplification in neuroblastoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr PR11.
Abstract PR12: Hedgehog pathway drives fusion-negative rhabdomyosarcoma initiated from nonmyogenic endothelial progenitors
Authors: Drummond Catherine J. , Hanna Jason A. , Garcia Matthew R. , Devine Daniel J. , Heyrana Alana J. , Finkelstein David , Rehg Jerold E. , Hatley Mark E.
Published: February 2019

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. Despite aggressive chemotherapy, radiotherapy, and surgery, clinical outcomes for RMS have not improved for three decades, emphasizing the need to uncover the molecular underpinnings of the disease. RMS has been presumed to originate from derailed muscle progenitors based on the histologic appearance and gene expression pattern of the tumors. However, an origin restricted to skeletal muscle does not explain RMS occurring in tissues devoid of skeletal muscle, such as the prostate, bladder, salivary gland, biliary tree, and the omentum. Previously, we described that activation of Sonic Hedgehog signaling through expression of a conditional, constitutively active Smoothened allele, SmoM2, under control of an adipocyte-restricted adipose protein 2 (aP2)-Cre recombinase transgene in mice gives rise to aggressive skeletal muscle tumors that display the histologic and molecular characteristics of human embryonal RMS (ERMS). With the short latency and anatomic restricted tumor location in the neck, we sought to leverage this model to explore the cell of origin of ERMS.

Lineage tracing experiments identified aP2-Cre labeled cells are distinctly nonmyogenic and were identified as endothelial cells found in the interstitium between muscle fibers. We illustrate that aP2-Cre is not expressed in the quiescent or activated muscle stem cells or satellite cells. Expression of oncogenic SmoM2 with aP2-Cre results in proliferation and expansion of the aP2-Cre labeled muscle interstitial cells and myogenic transdifferentiation resulting in ERMS. Activation of the Hedgehog pathway in aP2-Cre labeled endothelial progenitors results in Tbx1 expression, which is a skeletal muscle specification factor in the head and neck. We illustrate that endothelium and skeletal muscle within the head and neck arise from Kdr (Vegfr2) expressing progenitors. Hedgehog pathway activation in committed KDR+ endothelial progenitors results in Tbx1 expression and subsequent Myod1 expression driving a partially myogenic program characteristic of ERMS. Our work identifies reprogramming cell fate as a mechanism of transformation in pediatric sarcoma.

Citation Format: Catherine J. Drummond, Jason A. Hanna, Matthew R. Garcia, Daniel J. Devine, Alana J. Heyrana, David Finkelstein, Jerold E. Rehg, Mark E. Hatley. Hedgehog pathway drives fusion-negative rhabdomyosarcoma initiated from nonmyogenic endothelial progenitors [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr PR12.
Abstract A007: Methyl jasmonate and finasteride synergistically reduce testosterone-induced benign prostatic hyperplasia through regulation of inflammatory and apoptotic processes in rats
Authors: Adaramoye Oluwatosin A. , Owumi Solomon E. , Akanni Olubukola , Olapade-Olaopa Oluwabunmi E. , Abiola Olusoji J. , Akinloye Oluyemi
Published: February 2019

Background: Phytotherapy is becoming a treatment option in management of diseases, including benign prostatic hyperplasia (BPH). We have shown previously that methyl jasmonate (MeJA) ameliorated BPH; however, the underlying mechanism of action remains unknown. This study was designed to investigate in mechanistic terms the protective role of MeJA in BPH.

Methods: BPH was induced by daily injections of testosterone propionate (TP) (3 mg/kg) for 28 days.

Results: The weight and organo-somatic weight of prostate in BPH rats were 6.8 and 5.1 times higher than castrated-control group, respectively. Inflammatory markers, prostatic myeloperoxidase,and total nitric oxide were significantly increased in BPH group. The activity of aniline hydroxylase (Phase I drug metabolizing enzyme) was significantly increased in BPH rats by 22%. In BPH group, immunohistochemistry revealed strong expression of prostatic inducible nitric oxide synthase, cyclooxygenase-2 and Bcl2, while mild expression of p53 and Bax WAS seen. Serum triglyceride and total cholesterol were significantly increased, while HDL-c was decreased in BPH. Interestingly, MeJA and finasteride (singly or combination) attenuated inflammatory and apoptotic indices in BPH rats.

Conclusion: MeJA and finasteride synergistically protect against TP-induced BPH via mechanisms that involve antiinflammation, induction of apoptosis, and inhibition of phase I drug metabolizing enzyme.

Citation Format: Oluwatosin A. Adaramoye, Solomon E. Owumi, Olubukola Akanni, Oluwabunmi E. Olapade-Olaopa, Olusoji J. Abiola, Oluyemi Akinloye. Methyl jasmonate and finasteride synergistically reduce testosterone-induced benign prostatic hyperplasia through regulation of inflammatory and apoptotic processes in rats [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A007.
Abstract A042: Modulation of translation regulation by N6-methyladenosine in prostate cancer
Authors: Cotter Kellie A. , Meyer Kate D. , Benelli Matteo , Cohen Sandra , Jaffrey Samie , Demichelis Francesca , Rubin Mark
Published: February 2019

Recent evidence has highlighted the role of N6-methyladenosine (m6A) in post-transcriptional control of mRNA, in particular in altering translation efficiency. The demethylase (ALKBH5) and a methyltransferase (METTL3), which control m6A placement, are down- and upregulated, respectively, in prostate adenocarcinoma (PCa). This leads to the hypothesis that PCa exhibits alterations in m6A, and by extension changes in the translation efficiency of those transcripts. In this study we seek to produce an epitranscriptome map of m6A in PCa using single-nucleotide m6A mapping (miCLIP) of benign (RWPE) and PCa (LNCaP) cell lines. Distribution of m6A marks within transcripts mimics those seen in previous datasets: most peaks map to regions in cds or 3′ UTR, and show enrichment surrounding the TSS and stop codon. Overall miCLIP identifies over 10,000 transcripts with at least one m6A in the two cell lines, with many androgen-signaling genes and oncogenes exhibiting a high number of m6A sites. In order to assess the role of m6A in alterations in translation efficiency, we combined ribosome footprint profiling with RNA-seq in cells with and without METTL3 knockdown. This analysis identified several oncogenes as targets for translational regulation by m6A along with multiple genes related to cell migration, metastasis, and integrin signaling. These results validate m6A as a potentially important additional mechanism of gene regulation at the level of translation in PCa. Further, by allowing for the identification of changes in expression at the protein level previously undetectable by gene expression analysis alone, these findings may lead to the discovery of novel biomarkers or pathways as potential targets for treatment.

Citation Format: Kellie A. Cotter, Kate D. Meyer, Matteo Benelli, Sandra Cohen, Samie Jaffrey, Francesca Demichelis, Mark Rubin. Modulation of translation regulation by N6-methyladenosine in prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A042.
Abstract A043: Bypass kinase pathways lead to acquired CDK4/6 inhibitor resistance in prostate cancer
Authors: Leeuw Renee de , McNair Christopher M. , Schiewer Matthew J. , Neupane Neermala P. , Augello Michael A. , Li Zhen , Cheng Larry , Yoshida Akihiro , Diehl J. Alan , Hazard E Starr , McCourtney Sean M. , Hardiman Gary , Hussain Maha H. , Drake Justin M. , Kelly Wm. Kevin , Knudsen Karen E.
Published: February 2019

Cyclin dependent kinase-4/6 (CDK4/6) kinase inhibitors have shown clinical benefit in treatment of solid tumor types, including breast cancer. However, resistance is common, and the underpinning mechanisms of action are not well understood. Given the dependence of CDK4/6 inhibitors on retinoblastoma tumor suppressor (RB) function for activity, this class of agents may be particularly effective in tumor types for which RB loss is infrequent or occurs late in tumor progression. Here, models of acquired palbociclib resistance were generated in early-stage, RB-positive cancers, wherein it was shown that acquired palbociclib resistance resulted in cross-resistance to other CDK4/6 inhibitors under clinical testing. Furthermore, cells showing acquired resistance exhibited aggressive in vitro and in vivo phenotypes without genetic loss of RB or RB pathway members, including enhanced proliferative capacity, migratory potential, and characteristics of epithelial-to-mesenchymal transition. Further analyses through integration of RNA sequencing and phospho-proteomics identified activation of the MAPK signaling pathway as a mediator of CDK4/6 inhibitor resistance, capable of bypassing CDK4/6 activity. However, this altered kinase dependence resulted in sensitization to MEK inhibitors, suggestive of new clinical opportunities in CDK4/6-resistant tumors. In sum, the studies herein not only identify activation of the MAPK pathway as capable of bypassing the CDK4/6 requirement and promoting aggressive tumor characteristics, but also nominate MEK inhibitors as potential mechanisms to treat or prevent CDK4/6 inhibitor resistance.

Citation Format: Renee de Leeuw, Christopher M. McNair, Matthew J. Schiewer, Neermala P. Neupane, Michael A. Augello, Zhen Li, Larry Cheng, Akihiro Yoshida, J. Alan Diehl, E Starr Hazard, Sean M. McCourtney, Gary Hardiman, Maha H. Hussain, Justin M. Drake, Wm. Kevin Kelly, Karen E. Knudsen. Bypass kinase pathways lead to acquired CDK4/6 inhibitor resistance in prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A043.
Abstract A056: Is non-mutated GPR89A protein a potential drug target in prostate cancer?
Authors: Ma Yuanjun , Liu Yanling , Nilsson Sten , Li Chunde
Published: February 2019

Mutated genes as drug targets to treat cancer are a widely known concept; however, the effect is moderate. Targeting nonmutated genes is an alternative and probably has potential to benefit patients. GPR89A, as a nonmutated protein in prostate cancer, was evaluated as a drug target in this project. Data showed that GPR89A knockdown could induce high cancer cell death rate. Combination use of GPR89A siRNA and docetaxel triggered synergism effect, which implicates a possibility that GPR89A inhibitors could be used together with docetaxel. The high-throughput screening discovered 39 drug candidates that could reduce cancer cell viability and GPR89A expression simultaneously. These findings highlight the importance of nonmutated genes in cancer progression. Targeting nonmutated genes, e.g., GPR89A, should be evaluated further to find significant association between these genes/proteins and cancers.

Citation Format: Yuanjun Ma, Yanling Liu, Sten Nilsson, Chunde Li. Is non-mutated GPR89A protein a potential drug target in prostate cancer? [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A056.
Abstract A070: Targeting N-myristoyltransferase 1 inhibits prostate cancer progression
Authors: Alsaidan Omar , Kim Sungjin , Goodwin Octavia , Li Qianjin , Bai Aiping , Bielawska Alicja , Zheng George , Lebedyeva Iryna , Albers Thomas , Norris James S. , Pegan Scott D. , Cai Houjian
Published: February 2019

N-myristoyltransferase (NMT) facilitates co/post-translational myristoylation of several oncogenic proteins, regulating their functions in tumor progression. N-myristoyltransferase (NMT), which has been shown to be upregulated in a variety of cancers, is overexpressed in androgen-independent prostate cancer cells. We demonstrate that genetically, ablation of NMT1 inhibited proliferation of prostate cancer cells, tumor growth, and suppressed myristoylation profile of prostate cancer cells. Screening a panel of myristoyl-CoA analogs against purified human NMT1 protein leads to identifying the B13 as an inhibitor for NMT1. B13 significantly suppressed prostate cancer cell proliferation, migration, and invasion by cell cycle arrest, and inhibited the growth of prostate xenograft tumors with minimal pathologic effect on major organs in vivo. Structure activity relationship based optimization of B13 led to LCL204, which showed better inhibitory properties towards NMT1. We demonstrate that targeting protein myristoylation is a potential therapeutic approach to inhibit prostate tumor progression.

Citation Format: Omar Alsaidan, Sungjin Kim, Octavia Goodwin, Qianjin Li, Aiping Bai, Alicja Bielawska, George Zheng, Iryna Lebedyeva, Thomas Albers, James S. Norris, Scott D. Pegan, Houjian Cai. Targeting N-myristoyltransferase 1 inhibits prostate cancer progression [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A070.