CSF1R+ Macrophages Sustain Pancreatic Tumor Growth through T Cell Suppression and Maintenance of Key Gene Programs that Define the Squamous Subtype.

Pancreatic ductal adenocarcinoma (PDAC) is resistant to most therapies including single-agent immunotherapy and has a dense desmoplastic stroma, and most patients present with advanced metastatic disease. We reveal that macrophages are the dominant leukocyte population both in human PDAC stroma and autochthonous models, with an important functional contribution to the squamous subtype of human PDAC. We targeted macrophages in a genetic PDAC model using AZD7507, a potent selective inhibitor of CSF1R. AZD7507 caused shrinkage of established tumors and increased mouse survival in this difficult-to-treat model. Malignant cell proliferation diminished, with increased cell death and an enhanced T cell immune response. Loss of macrophages rewired other features of the TME, with global changes in gene expression akin to switching PDAC subtypes. These changes were markedly different to those elicited when neutrophils were targeted via CXCR2. These results suggest targeting the myeloid cell axis may be particularly efficacious in PDAC, especially with CSF1R inhibitors

Intra-acinar Trypsinogen Activation Mediates Early Stages of Pancreatic Injury but Not Inflammation in Mice With Acute Pancreatitis

The role of trypsinogen activation in the pathogenesis of acute pancreatitis (AP) has not been clearly established. We generated and characterized mice lacking trypsinogen isoform 7 (T7) gene ( T −/− ). The effects of pathologic activation of trypsinogen were studied in these mice during induction of AP with cerulein. Acinar cell death, tissue damage, early intra-acinar activation of the transcription factor nuclear factor κB (NF-κB), and local and systemic inflammation were compared between T −/− and wild-type mice with AP. Deletion of T7 reduced the total trypsinogen content by 60% but did not affect physiologic function. T −/− mice lacked pathologic activation of trypsinogen, which occurs within acinar cells during early stages of AP progression. Absence of trypsinogen activation in T −/− mice led to near complete inhibition of acinar cell death in vitro and a 50% reduction in acinar necrosis during AP progression. However, T −/− mice had similar degrees of local and systemic inflammation during AP progression and comparable levels of intra-acinar NF-κB activation, which was previously shown to occur concurrently with trypsinogen activation during early stages of pancreatitis. T7 is activated during pathogenesis of AP in mice. Intra-acinar trypsinogen activation leads to acinar death during early stages of pancreatitis, which accounts for 50% of the pancreatic damage in AP. However, progression of local and systemic inflammation in AP does not require trypsinogen activation. NF-κB is activated early in acinar cells, independently of trypsinogen activation, and might be responsible for progression of AP

E2F1 positively regulates Trib2 pseudokinase expression and proliferation in acute leukaemia

Deregulation of the transcription factor E2F1 occurs in AML, and it has been shown to induce both cell cycle progression and apoptosis. In normal granulopoiesis, proliferation arrest and differentiation mediated by C/EBPalpha involves repression of E2F1 target genes. Elevated Trib2 expression has been linked with a subset of human AML and dysregulated C/EBPalpha. Using promoter assays, mutational analyses and chromatin immunoprecipitation experiments, we show that E2F1 (and E2F2, E2F3, but not E2F4 or E2F5) are bound directly to the DNA on site-specific regions on the Trib2 promoter in leukaemic cells. Trib2 expression is decreased following siRNA-mediated knockdown of E2F1, and in E2F1 knockout cells as compared to wild type cells. The reintroduction of E2F1 rescued Trib2 expression showing that E2F1 is regulating the expression of endogenous Trib2. Further analyses revealed that this activation of Trib2 by E2F1 is repressed by wild type C/EBPalpha consistent with C/EBPalpha having a negative regulatory role on E2F1, and in normal GMP cells we detect C/EBPalpha bound to the Trib2 promoter. Conversely there was synergistic activation upon coexpression of the oncogenic C/EBPalpha truncated mutant, and Trib2 expression levels were elevated in GMPs from preleukaemic mutant C/EBPalpha mice compared to wild type GMPs. Indeed a positive correlation between Trib2 and E2F1 expression in AML datasets support these findings. Finally, inhibition of the cell cycle pathway in leukaemia cells expressing high endogenous levels of Trib2 protein resulted in G1 arrest with a reduction in E2F1 levels and Trib2 protein levels. Our work indicates that the cell cycle regulator E2F1 plays a key role in the control of Trib2 expression important for the control of cell proliferation and may have important implications for normal and malignant haematopoiesis

Co-operative leukemogenesis in acute myeloid leukemia and acute promyelocytic leukemia reveals C/EBPα as a common target of TRIB1 and PML/RARA.

The PML/RARA fusion protein occurs as a result of the t(15;17) translocation in the acute promyelocytic leukemia subtype of human acute myeloid leukemia. Gain of chromosome 8 is the most common chromosomal gain in human acute myeloid leukemia, including acute promyelocytic leukemia. We previously demonstrated that gain of chromosome 8-containing MYC is of central importance in trisomy 8, but the role of the nearby TRIB1 gene has not been experimentally addressed in this context. We have now tested the hypothesis that both MYC and TRIB1 have functional roles underlying leukemogenesis of trisomy 8 by using retroviral vectors to express MYC and TRIB1 in wild-type bone marrow and in marrow that expressed a PML/RARA transgene. Interestingly, although MYC and TRIB1 readily co-operated in leukemogenesis for wild-type bone marrow, TRIB1 provided no selective advantage to cells expressing PML/RARA. We hypothesized that this lack of co-operation between PML/RARA and TRIB1 reflected a common pathway for their effect: both proteins targeting the myeloid transcription factor C/EBPα. In support of this idea, TRIB1 expression abrogated the all-trans retinoic acid response of acute promyelocytic leukemia cells in vitro and in vivo Our data delineate the common and redundant inhibitory effects of TRIB1 and PML/RARA on C/EBPα providing a potential explanation for the lack of selection of TRIB1 in human acute promyelocytic leukemia, and highlighting the key role of C/EBPs in acute promyelocytic leukemia pathogenesis and therapeutic response. In addition, the co-operativity we observed between MYC and TRIB1 in the absence of PML/RARA show that, outside of acute promyelocytic leukemia, gain of both genes may drive selection for trisomy 8

Radio sound as material culture in the home

SIGLEAvailable from British Library Document Supply Centre-DSC:DXN024189 / BLDSC - British Library Document Supply CentreGBUnited Kingdo

Ein rechnergestuetztes Verfahren fuer den Generalplanentwurf von Schiffen

Available from TIB Hannover: RA 489(517) / FIZ - Fachinformationszzentrum Karlsruhe / TIB - Technische InformationsbibliothekSIGLEDEGerman

A Synthetic Lethal Approach to Eradicate AML via Synergistic Activation of Pro-Apoptotic p53 By MDM2 and BET Inhibitors

Acute Myeloid Leukemia (AML) is a typically-lethal molecularly heterogeneous disease, with few broad-spectrum therapeutic targets. Unusually compared to many other cancers, over 90% of AML patients retain wild type TP53, encoding pro-apoptotic tumor suppressor p53. However, wild-type p53 functions are frequently suppressed by MDM2, an E3 ubiquitin ligase that targets p53 for proteasomal degradation. MDM2 inhibitors (MDM2i), which activate wild-type p53, show encouraging pre-clinical activity, but limited clinical activity. In an effort to find targets that synergize with p53 activation via MDM2i and minimize toxicity, we performed a cell-based synthetic lethal drug screen and a CRISPR viability screen. These screens identified BRD4 inhibition as a candidate synthetic lethal partner of MDM2i. BRD4 is a member of the Bromodomains and Extraterminal (BET) family of proteins, a transcriptional co-activator and already a candidate AML therapeutic target. Surprisingly, we found inhibition of BRD4 alone induces expression of some of p53 target genes. We unexpectedly reveal that BRD4 binds to p53 target genes and acts as a transcriptional repressor of these genes. Synergistic cell killing by the drug combination (MDM2i + BET inhibitor (BETi)) depends on synergistic activation of p53 target genes, such as PUMA and NOXA, due to simultaneous stabilization of p53 by MDM2i and relief of BRD4-mediated repression by BETi. Our combined therapy of MDM2i and BETi is synergistically lethal to human AML cell lines harboring wild type TP53in vitro, against two mouse models of AML in vivo, and against primary human patient blasts in vitro. Furthermore, we used BET PROTACs to selectively and completely induce degradation of BRD4 in cells. Consistent with results from BETi, BET degraders and MDM2i synergize to suppress cell viability with superior potency. Taken together, our data show BRD4 represses p53-mediated transcription activation and apoptosis in AML. Therefore, co-targeting wild-type TP53 and a transcriptional repressor function of BRD4 represent a novel synthetic lethal vulnerability in AML

3102 – A Synthetic Lethality Approach to Eradicate AML via Synergistic Activation of Pro-Apoptotic p53 By MDM2 and BET Inhibitors

Acute Myeloid Leukemia (AML) is a typically-lethal molecularly heterogeneous disease, with few broad-spectrum therapeutic targets. Unusually, over 90% of AML patients retain wild type TP53, encoding pro-apoptotic tumor suppressor p53. However, wild-type p53 functions are frequently suppressed by MDM2, an E3 ubiquitin ligase that targets p53 for proteasomal degradation. MDM2 inhibitors (MDM2i), which activate wild-type p53, show encouraging pre-clinical activity, but limited clinical activity. In an effort to find targets that synergize with p53 activation via MDM2i and minimize toxicity, we performed a cell-based synthetic lethal drug screen and a CRISPR viability screen. These screens identified BRD4 inhibition as a candidate synthetic lethal partner of MDM2i. BRD4 is a member of the Bromodomains and Extraterminal (BET) family of proteins, a transcriptional co-activator and already an AML therapeutic target. Surprisingly, we found inhibition of BRD4 alone induces expression of some of p53 target genes. We unexpectedly reveal that BRD4 binds to p53 target genes and acts as a transcriptional repressor of these genes. Synergistic cell killing by the drug combination depends on synergistic activation of p53 target genes, such as PUMA and NOXA, due to simultaneous stabilization of p53 by MDM2i and relief of BRD4-mediated repression by BETi (BET inhibitors). Our combined therapy of MDM2i and BETi is synergistically lethal to human AML cell lines harboring wild type TP53 in vitro, against two mouse models of AML in vivo, and against primary human patient blasts in vitro. Taken together, our data show BRD4 represses p53-mediated transcription activation and apoptosis in AML. Therefore, co-targeting wild-type TP53 and a transcriptional repressor function of BRD4 represent a novel synthetic lethal vulnerability in AML