Pancreatic ductal adenocarcinoma (PDAC) offers an optimal model for discovering “druggable” molecular pathways that participate in inflammation-associated cancer development. Chronic pancreatitis, a common prolonged inflammatory disease, behaves as a well-known premalignant condition that contributes to PDAC development. Although the mechanisms underlying the pancreatitis-to-cancer transition remain to be fully elucidated, emerging evidence supports the hypothesis that the actions of proinflammatory mediators on cells harboring Kras mutations promote neoplastic transformation. Recent elegant studies demonstrated that the IL-17 pathway mediates this phenomenon and can be targeted with antibodies, but the downstream mechanisms by which IL-17 functions during this transition are currently unclear. In this study, we demonstrate that IL-17 induces the expression of REG3β, a well-known mediator of pancreatitis, during acinar-to-ductal metaplasia and in early PanIN lesions. Furthermore, we found that REG3β promotes cell growth and decreases sensitivity to cell death through activation of the gp130-JAK2-STAT3-dependent pathway. Genetic inactivation of REG3β in the context of oncogenic Kras-driven PDAC resulted in reduced PanIN formation, an effect that could be rescued by administration of exogenous REG3β. Taken together, our findings provide mechanistic insight into the pathways underlying inflammation-associated pancreatic cancer, revealing a dual and contextual pathophysiological role for REG3β during pancreatitis and PDAC initiation.

SummaryBone marrow mesenchymal stem/stromal cells (BM-MSCs) are key components of the hematopoietic niche thought to have a direct role in leukemia pathogenesis. BM-MSCs from patients with acute myeloid leukemia (AML) have been poorly characterized due to disease heterogeneity. We report a functional, genetic, and immunological characterization of BM-MSC cultures from 46 AML patients, stratified by molecular/cytogenetics into low-risk (LR), intermediate-risk (IR), and high-risk (HR) subgroups. Stable MSC cultures were successfully established and characterized from 40 of 46 AML patients irrespective of the risk subgroup. AML-derived BM-MSCs never harbored tumor-specific cytogenetic/molecular alterations present in blasts, but displayed higher clonogenic potential than healthy donor (HD)-derived BM-MSCs. Although HD- and AML-derived BM-MSCs equally provided chemoprotection to AML cells in vitro, AML-derived BM-MSCs were more immunosuppressive/anti-inflammatory, enhanced suppression of lymphocyte proliferation, and diminished secretion of pro-inflammatory cytokines. Multivariate analysis revealed that the level of interleukin-10 produced by AML-derived BM-MSCs as an independent prognostic factor negatively affected overall survival. Collectively our data show that AML-derived BM-MSCs are not tumor related, but display functional differences contributing to therapy resistance and disease evolution.

Mixed-lineage leukemia (MLL)-rearranged (MLLr) infant B-cell acute lymphoblastic leukemia (iMLLr-B-ALL) has a dismal prognosis and is associated with a pro-B/mixed phenotype, therapy refractoriness and frequent central nervous system (CNS) disease/relapse. Neuron-glial antigen 2 (NG2) is specifically expressed in MLLr leukemias and is used in leukemia immunophenotyping because of its predictive value for MLLr acute leukemias. NG2 is involved in melanoma metastasis and brain development; however, its role in MLL-mediated leukemogenesis remains elusive. Here we evaluated whether NG2 distinguishes leukemia-initiating/propagating cells (L-ICs) and/or CNS-infiltrating cells (CNS-ICs) in iMLLr-B-ALL. Clinical data from the Interfant cohort of iMLLr-B-ALL demonstrated that high NG2 expression associates with lower event-free survival, higher number of circulating blasts and more frequent CNS disease/relapse. Serial xenotransplantation of primary MLL-AF4+ leukemias indicated that NG2 is a malleable marker that does not enrich for L-IC or CNS-IC in iMLLr-B-All. However, NG2 expression was highly upregulated in blasts infiltrating extramedullar hematopoietic sites and CNS, and specific blockage of NG2 resulted in almost complete loss of engraftment. Indeed, gene expression profiling of primary blasts and primografts revealed a migratory signature of NG2+ blasts. This study provides new insights on the biology of NG2 in iMLLr-B-ALL and suggests NG2 as a potential therapeutic target to reduce the risk of CNS disease/relapse and to provide safer CNS-directed therapies for iMLLr-B-ALL.

No abstract

BackgroundThe immunosuppressive and anti-inflammatory properties of mesenchymal stromal cells (MSC) have prompted their therapeutic application in several autoimmune diseases, including rheumatoid arthritis. Adult MSC are finite and their clinical use is restricted by the need for long-term expansion protocols that can lead to genomic instability. Inhibition of Smad2/3 signaling in human pluripotent stem cells (hPSC) provides an infinite source of MSC that match the phenotype and functional properties of adult MSC. Here, we test the therapeutic potential of hPSC-MSC of embryonic origin (embryonic stem cell-derived mesenchymal stromal cells, hESC-MSC) in the experimental model of collagen-induced arthritis (CIA).MethodsCIA was induced in DBA/1 mice by immunization with type II collagen (CII) in Complete Freund’s Adjuvant (CFA). Mice were treated with either a single dose (106 cells/mouse) of hESC-MSC on the day of immunization (prophylaxis) or with three doses of hESC-MSC every other day starting on the day of arthritis onset (therapy). Arthritis severity was evaluated daily for six weeks and ten days, respectively. Frequency of Treg (FoxP3+), Th1 (IFNγ+) and Th17 (IL17+) CD4+ T cells in inguinal lymph nodes (ILN) was quantified by flow cytometry. Serum levels of anti-CII antibodies were determined by ELISA. Detection of hESC-MSC and quantification of murine and human indoleamine 2,3 dioxygenase (IDO1) expression was performed by quantitative real-time PCR. Statistical differences were analyzed by ANOVA and the Mann-Whitney U test.ResultsAdministration of hESC-MSC to mice with established arthritis reduced disease severity compared to control-treated mice. Analysis of CD4 T cell populations in treated mice showed an increase in FoxP3+ Treg and IFNγ+ Th1 cells but not in Th17 cells in the ILN. Anti-CII antibody levels were not affected by treatment. Migration of hESC-MSC to the ILN in treated mice was associated with the induction of murine IDO1.ConclusionTreatment with hESC-MSC ameliorates CIA by inducing IFNγ+ Th1 cells and IDO1 in the host. Thus, hESC-MSC can provide an infinite cellular source for treatment of rheumatoid arthritis.

Although B cells have been shown to be refractory to reprogramming into pluripotency, induced pluripotent stem cells (iPSCs) have been very recently generated, at very low efficiency, from human cord blood (CB)-and peripheral blood (PB)-derived CD191CD20 1 B cells using nonintegrative tetracistronic OSKM-expressing Sendai Virus (SeV). Here, we addressed whether cell ontogeny and hierarchy influence the reprogramming efficiency of the B-cell compartment. We demonstrate that human fetal liver (FL)-derived CD19 1 B cells are 110-fold easier to reprogram into iPSCs than those from CB/PB. Similarly, FL-derived CD341CD19 1 B progenitors are reprogrammed much easier than mature B cells (0.46% vs. 0.11%). All FL B-cell iPSCs carry complete VDJH rearrangements while 55% and 45% of the FL B-progenitor iPSCs carry incomplete and complete VDJH rearrangements, respectively, reflecting the reprogramming of developmentally different B progenitors (pro-B vs. pre-B) within a continuous differentiation process. Finally, our data suggest that successful B-cell reprogramming relies on active cell proliferation, and it is MYC-dependent as identical nonintegrative polycistronic SeV lacking MYC (OSKL or OSKLN) fail to reprogram B cells. The ability to efficiently reprogram human fetal primary B cells and B precursors offers an unprecedented opportunity for studying developmental B-lymphopoiesis and modeling B-cell malignances. STEM CELLS 2016;34:581-587 SIGNIFICANCE STATEMENTUsing tetracistronic OSKM-expressing nonintegrative Sendai vectors, we show that the developmental ontogeny and cellular hierarchy largely influences the reprogramming efficiency of human B cells into iPSCs. We also show that Ig/VDJH rearrangements do not constitute a reprogramming barrier and that B-cell reprogramming seems to rely on cell proliferation and it is cMYC-dependent. The ability to efficiently reprogram human fetal primary B cells and B precursors offers an unprecedented opportunity for studying developmental B-lymphopoiesis and modeling B-cell malignances.

B cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer, with cure rates of ∼80%. MLL-rearranged (MLLr) B-ALL (MLLr-B-ALL) has, however, an unfavorable prognosis with common therapy refractoriness and early relapse, and therefore new therapeutic targets are needed for relapsed/refractory MLLr-B-ALL. MLLr leukemias are characterized by the specific expression of chondroitin sulfate proteoglycan-4, also known as neuron-glial antigen-2 (NG2). NG2 was recently shown involved in leukemia invasiveness and central nervous system infiltration in MLLr-B-ALL, and correlated with lower event-free survival (EFS). We here hypothesized that blocking NG2 may synergize with established induction therapy for B-ALL based on vincristine, glucocorticoids, and l-asparaginase (VxL). Using robust patient-derived xenograft (PDX) models, we found that NG2 is crucial for MLLr-B-ALL engraftment upon intravenous (i.v.) transplantation. In vivo blockade of NG2 using either chondroitinase-ABC or an anti-NG2-specific monoclonal antibody (MoAb) resulted in a significant mobilization of MLLr-B-ALL blasts from bone marrow (BM) to peripheral blood (PB) as demonstrated by cytometric and 3D confocal imaging analysis. When combined with either NG2 antagonist, VxL treatment achieved higher rates of complete remission, and consequently higher EFS and delayed time to relapse. Mechanistically, anti-NG2 MoAb induces neither antibody-dependent cell-mediated not complement-dependent cytotoxicity. NG2 blockade rather overrides BM stroma-mediated chemoprotection through PB mobilization of MLLr-B-ALL blasts, thus becoming more accessible to chemotherapy. We provide a proof of concept for NG2 as a therapeutic target for MLLr-B-ALL.

It has been commonly found that in patients presenting Pancreatic Ductal Adenocarcinoma (PDAC), after a period of satisfactory response to standard treatments, the tumor becomes non-responsive and patient death quickly follows. This phenomenon is mainly due to the rapid and uncontrolled development of the residual tumor. The origin and biological characteristics of residual tumor cells in PDAC still remain unclear. In this work, using PDACs from patients, preserved as xenografts in nude mice, we demonstrated that a residual PDAC tumor originated from a small number of CD44+ cells present in the tumor. During PDAC relapse, proliferating CD44+ cells decrease expression of ZEB1, while overexpressing the MUC1 protein, and gain morphological and biological characteristics of differentiation. Also, we report that CD44+ cells, in primary and residual PDAC tumors, are part of a heterogeneous population, which includes variable numbers of CD133+ and EpCAM+ cells. We confirmed the propagation of CD44+ cells in samples from cases of human relapse, following standard PDAC treatment. Finally, using systemic administration of anti-CD44 antibodies in vivo, we demonstrated that CD44 is an efficient therapeutic target for treating tumor relapse, but not primary PDAC tumors. We conclude that CD44+ cells generate the relapsing tumor and, as such, are themselves promising therapeutic targets for treating patients with recurrent PDAC.