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Tumor recurrence developing from therapy resistance, immune escape and metastasis is the leading cause of death in medulloblastoma, the most frequent malignant pediatric brain tumor. Amplification of MYC genes is the most common genetic alteration in Group 3 and Group 4 subgroups that constitute two thirds of medulloblastoma. SOX9 is a transcription factor present in stem cells in the normal brain but is limited to rare, quiescent cells in medulloblastoma patients with MYC gene amplifications. By studying paired primary-recurrent patient samples and patient-derived xenografts we here identified significant accumulation of SOX9-positive cells in Group 3 and Group 4 relapses. To follow relapse at the single cell level we developed an inducible dual Tet model of MYC-driven MB, where MYC was re-directed from the treatment-sensitive bulk cells to resistant, dormant SOX9-positive cells by doxycycline. In this model, distant recurrent tumors and spinal metastases developed. SOX9 promoted immune escape, DNA repair suppression and was essential for recurrence. Tumor cell dormancy was non-hierarchical, migratory and depended on MYC suppression by SOX9 to promote relapse. By using computational modeling and treatment we also showed how doxorubicin and MGMT inhibitors were specifically targeting recurrent cells that could be of potential use in future treatments for patients affected by these fatal relapses. more...

// Prasoon Agarwal 1, * , Mohammad Alzrigat 1, * , Alba Atienza Parraga 1 , Stefan Enroth 1 , Umashankar Singh 2 , Johanna Ungerstedt 3 , Anders Osterborg 4 , Peter J. Brown 5 , Anqi Ma 6 , Jian Jin 6 , Kenneth Nilsson 1 , Fredrik Oberg 1 , Antonia Kalushkova 1, ‡ , Helena Jernberg-Wiklund 1, ‡ 1 Science for Life Laboratory, Department of Immunology, Genetics and Pathology, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden 2 Department of Biological Engineering, Indian Institute of Technology, Gandhinagar, Gujarat, India 3 Department of Medicine, Center for Hematology and Regenerative Medicine (HERM), Karolinska Institute Huddinge, Stockholm, Sweden 4 Department of Oncology-Pathology, Karolinska University Hospital Solna, Stockholm, Sweden 5 Structural Genomics Consortium, University of Toronto, Toronto, Ontario, Canada 6 Departments of Structural and Chemical Biology, Oncological Sciences, and Pharmacology and Systems Therapeutics, Icahn School of Medicine at Mount Sinai, New York, NY, USA * These authors have contributed equally to this work ‡ Corresponding last co-authors Correspondence to: Helena Jernberg-Wiklund, e-mail: helena.jernberg_wiklund@igp.uu.se Antonia Kalushkova, e-mail: antonia.kalushkova@igp.uu.se Keywords: multiple myeloma, Polycomb, EZH2, H3K27me3, UNC1999 Received: June 03, 2015 Accepted: December 29, 2015 Published: January 7, 2016 ABSTRACT Multiple myeloma (MM) is a malignancy of the antibody-producing plasma cells. MM is a highly heterogeneous disease, which has hampered the identification of a common underlying mechanism for disease establishment as well as the development of targeted therapy. Here we present the first genome-wide profiling of histone H3 lysine 27 and lysine 4 trimethylation in MM patient samples, defining a common set of active H3K4me3-enriched genes and silent genes marked by H3K27me3 (H3K27me3 alone or bivalent) unique to primary MM cells, when compared to normal bone marrow plasma cells. Using this epigenome profile, we found increased silencing of H3K27me3 targets in MM patients at advanced stages of the disease, and the expression pattern of H3K27me3-marked genes correlated with poor patient survival. We also demonstrated that pharmacological inhibition of EZH2 had anti-myeloma effects in both MM cell lines and CD138+ MM patient cells. In addition, EZH2 inhibition decreased the global H3K27 methylation and induced apoptosis. Taken together, these data suggest an important role for the Polycomb repressive complex 2 (PRC2) in MM, and highlights the PRC2 component EZH2 as a potential therapeutic target in MM. more...

AKN-028 is a novel tyrosine kinase inhibitor with preclinical activity in acute myeloid leukemia (AML), presently undergoing investigation in a phase I/II study. It is a potent inhibitor of the FMS-like kinase 3 (FLT3) but shows in vitro activity in a wide range of AML samples. In the present study, we have characterized the effects of AKN-028 on AML cells in more detail. AKN-028 induced a dose-dependent G0/1 arrest in AML cell line MV4-11. Treatment with AKN-028 caused significantly altered gene expression in all AML cell types tested (430 downregulated, 280 upregulated transcripts). Subsequent gene set enrichment analysis revealed enrichment of genes associated with the proto-oncogene and cell cycle regulator c-Myc among the downregulated genes in both AKN-028 and midostaurin treated cells. Kinase activity profiling in AML cell lines and primary AML samples showed that tyrosine kinase activity, but not serine/threonine kinase activity, was inhibited by AKN-028 in a dose dependent manner in all samples tested, reaching approximately the same level of kinase activity. Cells sensitive to AKN-028 showed a higher overall tyrosine kinase activity than more resistant ones, whereas serine/threonine kinase activity was similar for all primary AML samples.In summary, AKN-028 induces cell cycle arrest in AML cells, downregulates Myc-associated genes and affect several signaling pathways. AML cells with high global tyrosine kinase activity seem to be more sensitive to the cytotoxic effect of AKN-028 in vitro. more...

// Mohammad Alzrigat 1 , Alba Atienza Parraga 1, * , Prasoon Agarwal 2, * , Hadil Zureigat 3 , Anders Osterborg 4 , Hareth Nahi 5 , Anqi Ma 6 , Jian Jin 6 , Kenneth Nilsson 1 , Fredrik Oberg 1 , Antonia Kalushkova 1 , Helena Jernberg-Wiklund 1 1 Science for Life Laboratory, Department of Immunology, Genetics and Pathology, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden 2 Department of Laboratory Medicine, Division of Clinical Immunology, Karolinska University Hospital, Huddinge, Stockholm, Sweden 3 Department of Medicine, Faculty of Medicine, University of Jordan, Amman, Jordan 4 Department of Oncology-Pathology, Karolinska University Hospital, Solna, Stockholm, Sweden 5 Department of Medicine, Unit of Hematology, Karolinska University Hospital, Huddinge, Stockholm, Sweden 6 Department of Pharmacological Sciences and Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA * These authors have contributed equally to this work Correspondence to: Helena Jernberg-Wiklund, email: helena.jernberg_wiklund@igp.uu.se Mohammad Alzrigat, email: mohammad.alzrigat@igp.uu.se Keywords: multiple myeloma, EZH2, H3K27me3, microRNA, UNC1999 Received: October 09, 2016 Accepted: December 15, 2016 Published: December 30, 2016 ABSTRACT Multiple Myeloma (MM) is a plasma cell tumor localized to the bone marrow (BM). Despite the fact that current treatment strategies have improved patients' median survival time, MM remains incurable. Epigenetic aberrations are emerging as important players in tumorigenesis making them attractive targets for therapy in cancer including MM. Recently, we suggested the polycomb repressive complex 2 (PRC2) as a common denominator of gene silencing in MM and presented the PRC2 enzymatic subunit enhancer of zeste homolog 2 (EZH2) as a potential therapeutic target in MM. Here we further dissect the anti-myeloma mechanisms mediated by EZH2 inhibition and show that pharmacological inhibition of EZH2 reduces the expression of MM-associated oncogenes; IRF-4, XBP-1, PRDM1/BLIMP-1 and c-MYC. We show that EZH2 inhibition reactivates the expression of microRNAs with tumor suppressor functions predicted to target MM-associated oncogenes; primarily miR-125a-3p and miR-320c. ChIP analysis reveals that miR-125a-3p and miR-320c are targets of EZH2 and H3K27me3 in MM cell lines and primary cells. Our results further highlight that polycomb-mediated silencing in MM includes microRNAs with tumor suppressor activity. This novel role strengthens the oncogenic features of EZH2 and its potential as a therapeutic target in MM. more...

Background and aim: Known colorectal cancer syndromes, such as familial adenomatous polyposis and hereditary non-polyposis colorectal cancer, have been identified in only a small proportion of cases with a family history of disease. In an attempt to identify loci harbouring novel predisposing genes, we have performed a genome wide linkage analysis in 18 colorectal cancer families recruited from the Department of Clinical Genetics at Karolinska Hospital, Sweden. Methods: Multipoint parametric and non-parametric linkage analyses were performed using two affected status criteria, stringent and less stringent. Parametric analysis was performed under the assumption of locus homogeneity and locus heterogeneity. Results: The initial scan performed using the less stringent affected status criteria revealed regions of interest on chromosome 11 (marker D11S1314: heterogeneity logarithm of odds (HLOD) score 1.96, non-parametric LOD (NPL) score 1.28; and marker D11S908: HLOD score 2.10, NPL score 2.16) and chromosome 14 (marker D14S258: HLOD score 2.61, NPL score 2.88). Using the stringent affected status criteria, a locus on chromosome 22 was suggested in the parametric analysis (marker D22S315: HLOD score 1.26). After finemapping of the regions on chromosomes 11 and 14, HLOD and NPL scores were reduced but still within the range of suggestive linkage. Haplotype analysis revealed overlapping regions between D11S987 and D11S4207 (proximal region), D11S4120 and D11S4090 (distal region), on chromosome 11, and between D14S1038 and D14S1069 on chromosome 14. Conclusion: Our study provides evidence of genetic heterogeneity among Swedish colorectal cancer families. Three novel regions were suggested to be of interest in a proportion of families analysed. Further studies are needed to confirm this result. more...

Dysregulated NF-κB signaling appears to be particularly important in B-cell malignancies, with recurrent mutations identified within both the canonical and non-canonical NF-κB pathways, as well as in components of the B-cell receptor (BcR) and Toll-like receptor (TLR) signaling pathways. In chronic lymphocytic leukemia (CLL), although recurrent mutations have been identified in MYD88 (TLR signaling) and BIRC3 (non-canonical NF-κB pathway), their frequency is low (10% for the mutant allele, we identified mutations (n=35) within 30/124 (24%) patients in 14/18 NF-κB genes analyzed. IκB genes, which encode for cytoplasmic inhibitor proteins, accounted for 20/35 (57%) mutations, with IκBε (encoded by NFKBIE) mutated in 8 patients; notably, 3/8 cases carried an identical 4bp deletion within exon 1 of NFKBIE. Prompted by these findings, we proceeded to validate our findings in an independent CLL cohort (n=168) using the same methodology as above and primarily focusing on cases with poor-prognostic features. We identified 30 mutations within 28 CLL patients in 11/18 NF-κB genes analyzed. Strikingly, 13/30 mutations were found within IκBε, with 10/13 patients carrying the same 4bp NFKBIE deletion. Notably, investigations into whether additional cases (within both the discovery and validation cohort) may harbor mutations of low clonal abundance ( Disclosures No relevant conflicts of interest to declare. more...