ALBERT

Description: Author(s): Manh Duc Le, Keith A. McEwen, Martin Rotter, Mathias Doerr, Alexander Barcza, Je-Geun Park, James Brooks, Eric Jobiliong, and David Fort UPd 3 is known to exhibit four antiferroquadrupolar ordered phases at low temperatures. We report measur-ements of the magnetization and magnetostriction of single crystal UPd 3 , along the principal symmetry directions, in fields up to 33 T. These results have been combined with recent inelastic neutr... [Phys. Rev. B 89, 235114] Published Tue Jun 10, 2014

Description: Crystal Growth & Design DOI: 10.1021/acs.cgd.6b01841

Description: The mechanisms linking systems-level programs of gene expression to discrete cell biological processes in vivo remain poorly understood. In this study, we have defined such a program for multi-ciliated epithelial cells (MCCs), a cell type critical for proper development and homeostasis of the airway, brain and reproductive tracts. Starting from genomic analysis of the cilia-associated transcription factor Rfx2, we used bioinformatics and in vivo cell biological approaches to gain insights into the molecular basis of cilia assembly and function. Moreover, we discovered a previously un-recognized role for an Rfx factor in cell movement, finding that Rfx2 cell-autonomously controls apical surface expansion in nascent MCCs. Thus, Rfx2 coordinates multiple, distinct gene expression programs in MCCs, regulating genes that control cell movement, ciliogenesis, and cilia function. As such, the work serves as a paradigm for understanding genomic control of cell biological processes that span from early cell morphogenetic events to terminally differentiated cellular functions.

Description: Epstein-Barr virus (EBV) is a human herpes virus that infects over 90% of the world's population and is linked with cancer development. In immune-competent individuals, EBV-infection is controlled by a highly efficient virus-specific T cell response. Following primary infection, the virus achieves lifelong persistence within the human host. Risk of EBV-driven cancers increases with immune suppression (IS). Solid organ transplant recipients receive IS medications to prevent graft rejection and are at highest risk of developing EBV-associated lymphomas known as post-transplant lymphoproliferative disease (PTLD). PTLD represents a serious complication of organ transplantation, associated with poor prognosis. Currently, no standard approach for prevention or treatment exists. Reducing the level of IS medication may control PTLD but often leads to graft-rejection. In order to promote long-term protection from EBV-driven cancers, we have developed a vaccine to bolster EBV-specific immunity by targeting the EBV immediate early protein, BZLF1. BZLF1 initiates the activation of lytic stage in EBV-infected cells and promotes B-cell transformation. Work by our group has shown BZLF1-specific T cell expansion following reduction in IS medications correlates with PTLD tumor regression and improved patient survival. Here we specifically delivered the protein (BZLF1) to dendritic cells (DCs) through its endocytic receptor DEC205. DCs were generated from HLA-B8+ donor monocytes incubated with interleukin-4 (IL4) and granulocyte-macrophage colony-stimulating factor (GM-CSF). Mature DCs were then loaded with DEC205-BZLF1 fusion protein or control protein (DEC205-Human Chorionic Gonadotropin (DEC205-HCG)). Antigen-loaded DCs were co-cultured with autologous peripheral blood mononuclear cells (PBMCs) in the presence of IL-2 for 10 days. Cells were analyzed by flow cytometry using HLA-tetramers to detect and quantify antigen-specific cytotoxic T leukocyte (CTL) response. To test the EBV vaccine in-vivo, we utilized a human-murine chimeric model of EBV-driven lymphoproliferative disease (EBV-LPD). Severe combined immune deficient (SCID) mice were engrafted with PBMCs from EBV+ donors (Hu-PBL-SCID model). The spontaneous EBV-LPD that develops in this model is comprised of human CD20+, EBV+ lymphoblasts that closely resembles PTLD. Mice were immunized with DCs loaded with DEC205-BZLF1 or DEC205-HCG at the time of PBMC transplant and received booster doses at day 14 and 28. Splenocyte from vaccinated mice were stimulated with autologous tumor (lymphoblastoid cells line, (LCL)) pulsed with BZLF1 pepmix, BZLF1 pepmix alone, and anti-CD3. Secretion of IFNg by stimulated splenocytes was detected using Human IFNg Enzyme-Linked Immunosorbent Spot (ELISpot). In vitro co-cultures treated with DEC205-BZLF1-loaded DCs showed increased expansion of EBV-specific CTLs (p-value: 0.0002) capable of abundant IFNg production and potent cytotoxicity against autologous tumor. This vaccine significantly improved survival in vaccinated mice (p-value: 0.035). Splenocytes from mice in the DEC205-BZLF1 vaccination group revealed higher responsiveness to autologous LCLs and BZLF1 pepmix compared to controls as determined by ELISpot. Human cells recovered from mouse spleen will be analyzed by mass cytometry using a multi-parametric antibody panel to evaluate central/effector memory status, CD4+ Th, CD8+ CTL, NK, and monocyte subsets. These results further support pre-clinical and clinical development of vaccine approaches utilizing the BZLF1 protein as an immunogen to harness adaptive cellular responses to prevent EBV-associated LPD in vulnerable patient populations. Disclosures No relevant conflicts of interest to declare.

Description: Mantle cell lymphoma (MCL) is an incurable B-cell malignancy characterized by genetic dysregulation of cyclin D1 and activation of signaling pathways driving uncontrolled MCL cell proliferation and survival. Ibrutinib is an FDA-approved irreversible inhibitor of Bruton's tyrosine kinase (BTK), a downstream target of the B-cell receptor (BCR) pathway. While ibrutinib exhibits significant single-agent therapeutic activity in patients with relapsed/refractory MCL, the vast majority of MCL patients on ibrutinib progress with aggressive disease and short survival (3-8 mo). Although ~80% of chronic lymphocytic leukemia patients with acquired ibrutinib resistance have mutations in BTK and PLCγ2, this is uncommon in MCL suggesting alternative mechanisms driving this resistant phenotype. Understanding drug-resistance mechanisms and developing effective therapies for ibrutinib resistant (IR) MCL are urgently needed. The major type II protein arginine methyltransferase enzyme, PRMT5, catalyzes symmetric dimethylation of arginine residues on histone tails (H3R8 and H4R3) and other proteins. PRMT5 regulates a vast array of biologic functions including RNA processing, DNA damage response, signal transduction, and gene expression. Amplified PRMT5 activity drives the expression and activity of key oncogenes (MYC, CYCLIND1, NOTCH1) while silencing expression and activity of tumor suppressors (ST7, RBL2, and p53). Our group has shown PRMT5 is overexpressed and dysregulated in MCL and strategies aimed at selectively targeting PRMT5 show anti-tumor activity in preclinical lymphoma models. Here we describe the development of a novel patient derived xenograft (PDX) of IR-MCL and explore PRMT5 inhibition as an alternative therapeutic option to circumvent IR. Peripheral blood mononuclear cells from a 75 yo male patient diagnosed with acquired classic IR-MCL were engrafted intravenously into NSG mice. After 5 passages, all mice engrafted with 107 MCL cells developed histologically confirmed MCL infiltrating kidney, lymph nodes, bone marrow, spleen and peripheral blood. Circulating human CD5+/CD19+ cells were detectable and quantifiable by flow cytometry by day 21 post-engraftment. Karyotype analysis confirmed the hallmark t(11;14)(q13;q32) of MCL while retaining nearly all cytogenetic abnormalities present in the patient's primary tumor including a deletion of chromosome 9, associated with deletion of MTAP, a therapeutic vulnerability for PRMT5-targeted therapy. Whole exome sequencing confirmed genomic stability with successive passages. Ex vivo cytotoxicity assays and protein pathway analysis further confirmed resistance to ibrutinib (IC50 〉1 µM) with maintained hyper-phosphorylation of AKT (Ser473) and ERK (Thr202/Tyr204). Western blot analysis showed elevated levels of c-MYC, CYCLIND1, BCL2, and pERK. After validation of circulating disease at day 25 post engraftment, mice were treated with either a novel small molecule inhibitor of PRMT5 (PRT382, 10 mg/kg orally 4 days on 3 days off) or ibrutinib (75 mg/kg administered in drinking water, n=5 mice per treatment group). Treatment of this PDX model with PRT382 resulted in significantly decreased disease burden and improved median survival compared to control animals from 48 to 83 days, respectively (p=0.0045). We found no significant difference in survival (p= 0.6540) or circulating disease burden with ibrutinib therapy compared to control group. The full BTK occupancy of ibrutinib treated mice was validated using fluorescence resonance energy transfer-based assay. Ex vivo PDX MCL cells from PRT382-treated mice showed loss of symmetric dimethyl arginine with preservation of asymmetric dimethyl arginine levels, reduced H4(Sme2)R3 epigenetic marks, and elevated levels of BCL2, MYC, and pAKT/pERK. We developed a cell line (SEFA) allowing for in vitro mechanistic studies. We are currently investigating potential mechanisms responsible for circumventing IR-MCL by integrating genome-wide changes to chromatin accessibility and whole transcriptome analysis. This IR-MCL PDX mouse model serves as a useful tool to investigate mechanisms of drug resistance, provides a platform to explore novel pre-clinical therapeutic strategies to circumvent IR and demonstrates the therapeutic activity of PRMT5 targeted therapy in this aggressive disease. Disclosures Byrd: Pharmacyclics LLC, an AbbVie Company: Other: Travel Expenses, Research Funding, Speakers Bureau; Janssen: Consultancy, Other: Travel Expenses, Research Funding, Speakers Bureau; Ohio State University: Patents & Royalties: OSU-2S; Genentech: Research Funding; BeiGene: Research Funding; Janssen: Consultancy, Other: Travel Expenses, Research Funding, Speakers Bureau; TG Therapeutics: Other: Travel Expenses, Research Funding, Speakers Bureau; Gilead: Other: Travel Expenses, Research Funding, Speakers Bureau; Novartis: Other: Travel Expenses, Speakers Bureau; Genentech: Research Funding; Acerta: Research Funding; Acerta: Research Funding; Ohio State University: Patents & Royalties: OSU-2S; BeiGene: Research Funding; Genentech: Research Funding; BeiGene: Research Funding; Janssen: Consultancy, Other: Travel Expenses, Research Funding, Speakers Bureau; Novartis: Other: Travel Expenses, Speakers Bureau; Pharmacyclics LLC, an AbbVie Company: Other: Travel Expenses, Research Funding, Speakers Bureau; Gilead: Other: Travel Expenses, Research Funding, Speakers Bureau; Gilead: Other: Travel Expenses, Research Funding, Speakers Bureau; Novartis: Other: Travel Expenses, Speakers Bureau; Pharmacyclics LLC, an AbbVie Company: Other: Travel Expenses, Research Funding, Speakers Bureau; TG Therapeutics: Other: Travel Expenses, Research Funding, Speakers Bureau; Acerta: Research Funding; Ohio State University: Patents & Royalties: OSU-2S; TG Therapeutics: Other: Travel Expenses, Research Funding, Speakers Bureau. Vaddi:Prelude Therapeutics: Employment. Scherle:Prelude Therapeutics: Employment. Baiocchi:Prelude: Consultancy.

Description: Neural tube closure defects are a major cause of infant mortality, with exencephaly accounting for nearly one-third of cases. However, the mechanisms of cranial neural tube closure are not well understood. Here we show that this process involves a tissue-wide pattern of apical constriction controlled by Sonic hedgehog (Shh) signaling. Midline cells in the mouse midbrain neuroepithelium are short with large apical surfaces, whereas lateral cells are taller and undergo synchronous apical constriction, driving neural fold elevation. Embryos lacking the Shh effector Gli2 fail to produce appropriate midline cell architecture, whereas embryos with expanded Shh signaling, including the IFT-A complex mutants Ift122 and Ttc21b and embryos expressing activated Smoothened, display apical constriction defects in lateral cells. Disruption of lateral, but not midline, cell remodeling results in exencephaly. These results reveal a morphogenetic program of patterned apical constriction governed by Shh signaling that generates structural changes in the developing mammalian brain.

Description: Premature fusion of the cranial sutures (craniosynostosis), affecting 1 in 2000 newborns, is treated surgically in infancy to prevent adverse neurologic outcomes. To identify mutations contributing to common non-syndromic midline (sagittal and metopic) craniosynostosis, we performed exome sequencing of 132 parent-offspring trios and 59 additional probands. Thirteen probands (7%) had damaging de novo or rare transmitted mutations in SMAD6, an inhibitor of BMP – induced osteoblast differentiation (p