A gene signature specific for intestinal stem cells (ISCs) has recently been shown to predict relapse in colorectal cancer (CRC) but the tumorigenic role of individual signature genes remains poorly defined. A prominent ISC-signature gene is the cancer stem cell marker CD44, which encodes various splice variants comprising a diverse repertoire of adhesion and signaling molecules. Using Lgr5 as ISC marker, we have fluorescence-activated cell sorting-purified ISCs to define their CD44 repertoire. ISCs display a specific set of CD44 variant isoforms (CD44v), but remarkably lack the CD44 standard (CD44s) isoform. These CD44v also stand-out in transformed human ISCs isolated from microadenomas of familial adenomatous polyposis patients. By employing knock-in mice expressing either CD44v4-10 or CD44s, we demonstrate that the CD44v isoform, but not CD44s, promotes adenoma initiation in Apc(Min/+)mice. Our data identify CD44v as component of the ISCs program critical for tumor initiation, and as potential treatment target in CRC.

Aberrant activation of Wnt/β-catenin signaling plays a central role in the pathogenesis of a wide variety of malignancies and is typically caused by mutations in core Wnt pathway components driving constitutive, ligand-independent signaling. In multiple myelomas (MMs), however, these pathway intrinsic mutations are rare despite the fact that most tumors display aberrant Wnt pathway activity. Recent studies indicate that this activation is caused by genetic and epigenetic lesions of Wnt regulatory components, sensitizing MM cells to autocrine Wnt ligands and paracrine Wnts emanating from the bone marrow niche. These include deletion of the tumor suppressor CYLD, promotor methylation of the Wnt antagonists WIF1, DKK1, DKK3, and sFRP1, sFRP2, sFRP4, sFRP5, as well as overexpression of the co-transcriptional activator BCL9 and the R-spondin receptor LGR4. Furthermore, Wnt activity in MM is strongly promoted by interaction of both Wnts and R-spondins with syndecan-1 (CD138) on the MM cell-surface. Functionally, aberrant canonical Wnt signaling plays a dual role in the pathogenesis of MM: (I) it mediates proliferation, migration, and drug resistance of MM cells; (II) MM cells secrete Wnt antagonists that contribute to the development of osteolytic lesions by impairing osteoblast differentiation. As discussed in this review, these insights into the causes and consequences of aberrant Wnt signaling in MM will help to guide the development of targeting strategies. Importantly, since Wnt signaling in MM cells is largely ligand dependent, it can be targeted by drugs/antibodies that act upstream in the pathway, interfering with Wnt secretion, sequestering Wnts, or blocking Wnt (co)receptors.

NLRX1 is a mitochondrial innate immune receptor involved in viral immunity. Stokman et al. found that loss of NLRX1 increased cellular mitochondrial activity, production of reactive oxygen species, and apoptosis during oxidative stress in kidney injury.

Multiple myeloma (MM) is characterized by the expansion of malignant plasma cells in the bone marrow (BM). Most MMs display aberrant Wnt/β-catenin signaling, which drives proliferation; however, they lack oncogenic Wnt pathway mutations, suggesting activation by autocrine Wnt ligands and/or paracrine Wnts from the BM microenvironment. Expression of the heparan sulfate (HS) proteoglycan syndecan-1 is a hallmark of MM. Syndecan-1 is a critical player in the complex reciprocal interaction between MM cells and their BM niche, mediating growth factor/cytokine binding and signaling by its HS chains. Here, by means of CRISPR/Cas9-mediated knockout and doxycycline-inducible short hairpin RNA-mediated knockdown of EXT1, a critical enzyme for HS polymerization, we demonstrate that the HS chains decorating syndecan-1 mediate aberrant Wnt pathway activation in MM. HS-deficient MM cells exhibited strongly decreased autocrine Wnt/β-catenin pathway activity and reduced Wnt pathway-dependent proliferation. In addition, we demonstrate that Wnts bind to the HS side chains of syndecan-1 and that this binding contributes to paracrine Wnt pathway activation through the Wnt receptor Frizzled (Fzd). Furthermore, in an HS-dependent fashion, syndecan-1 also binds osteoblast-produced R-spondin, which represses Fzd degradation by activation of LGR4, an R-spondin receptor aberrantly expressed on MM cells. Costimulation with R-spondin and its binding to HS chains decorating syndecan-1 are indispensable for optimal stimulation of Wnt signaling in MM. Taken together, our results identify syndecan-1 as a crucial component of the Wnt signalosome in MM cells, binding Wnts and R-spondins to promote aberrant Wnt/β-catenin signaling and cell growth, and suggest HS and its biosynthetic enzymes as potential targets in the treatment of MM.

In studies of intestinal organoid cultures and mice with inducible deletion of MET, we found HGF receptor signaling to regulate intestinal homeostasis and regeneration, as well as adenoma formation. These activities of MET are promoted by the stem cell CD44 isoform CD44v4-10. Our findings provide rationale for targeting signaling via MET and CD44 during anti-EGF receptor therapy of patients with colorectal cancer or in patients resistant to EGF receptor inhibitors.

The unrestrained growth of tumor cells is generally attributed to mutations in essential growth control genes, but tumor cells are also affected by, or even addicted to, signals from the microenvironment. As therapeutic targets, these extrinsic signals may be equally significant as mutated oncogenes. In multiple myeloma (MM), a plasma cell malignancy, most tumors display hallmarks of active Wnt signaling but lack activating Wnt-pathway mutations, suggesting activation by autocrine Wnt ligands and/or paracrine Wnts emanating from the bone marrow (BM) niche. Here, we report a pivotal role for the R-spondin/leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) axis in driving aberrant Wnt/ β-catenin signaling in MM. We show that LGR4 is expressed by MM plasma cells, but not by normal plasma cells or B cells. This aberrant LGR4 expression is driven by IL-6/STAT3 signaling and allows MM cells to hijack R-spondins produced by (pre)osteoblasts in the BM niche, resulting in Wnt (co)receptor stabilization and a dramatically increased sensitivity to auto-and paracrine Wnts. Our study identifies aberrant R-spondin/LGR4 signaling with consequent deregulation of Wnt (co)receptor turnover as a driver of oncogenic Wnt/ β-catenin signaling in MM cells. These results advocate targeting of the LGR4/R-spondin interaction as a therapeutic strategy in MM.M ultiple myeloma (MM) in most patients is an incurable hematologic malignancy characterized by the accumulation of clonal plasma cells in the bone marrow (BM). Despite the wide variety of underlying structural and numerical genomic abnormalities (1-3), virtually all MMs are highly dependent on a protective BM microenvironment, or "niche," for growth and survival (4). Understanding the complex reciprocal interaction between MM cells and the BM microenvironment is critical for the development of new targeted therapies.In MM, aberrant activation of the canonical Wnt pathway drives proliferation and is associated with disease progression, dissemination, and drug resistance (5-9). Because MMs with hallmarks of active Wnt signaling do not harbor mutations that typically underlie constitutive Wnt pathway activation, this oncogenic Wnt pathway activity was proposed to involve autocrine and/or paracrine Wnt ligands (6,8,10). Wnts are lipid-modified glycoproteins that function as typical niche factors because they are relatively unstable and insoluble due to their hydrophobic nature, which constrains long-range signaling (11). Binding of a Wnt ligand to its receptor Frizzled (Fzd) initiates a signaling cascade that ultimately results in stabilization and nuclear translocation of the Wnt effector β-catenin. In cooperation with TCF/LEF family transcription factors this orchestrates a transcriptional program (12, 13), comprising targets such as MYC and CCND1 (Cyclin D1) that play crucial roles in the pathogenesis of MM (14-16). Aberrant Wnt signaling in cancer typically results from mutations in APC, β-catenin (CTNNB1), or AXIN that drive constitutive, ligand-independent pathway ac...

In multiple myeloma (MM), angiogenesis is strongly correlated to disease progression and unfavorable outcome, and may be promoted by bone marrow hypoxia. Employing gene-expression profiling, we here identified the pro-angiogenic factor adrenomedullin (AM) as the most highly upregulated gene in MM cells exposed to hypoxia. Malignant plasma cells from the majority of MM patients, belonging to distinct genetic subgroups, aberrantly express AM. Already under normoxic conditions, a subset of MM highly expressed and secreted AM, which could not be further enhanced by hypoxia or cobalt chloride-induced stabilization of hypoxia-inducible factor (HIF)1α. In line with this, expression of AM did not correlate with expression of a panel of established hypoxia-/HIF1α-target genes in MM patients. We demonstrate that MM-driven promotion of endothelial cell proliferation and tube formation is augmented by inducible expression of AM and strongly repressed by inhibition of endogenous and hypoxia-induced AM activity. Together, our results demonstrate that MM cells, both in a hypoxia-dependent and -independent fashion, aberrantly express and secrete AM, which can mediate MM-induced angiogenesis. Thus, AM secretion can be a major driving force for the angiogenic switch observed during MM evolution, which renders AM a putative target for MM therapy.

The Wnt/β-catenin pathway plays a crucial role in the pathogenesis of various human cancers. In multiple myeloma (MM), aberrant auto-and/or paracrine activation of canonical Wnt signaling promotes proliferation and dissemination, while overexpression of the Wnt inhibitor Dickkopf1 (DKK1) by MM cells contributes to osteolytic bone disease by inhibiting osteoblast differentiation. Since DKK1 itself is a target of TCF/β-catenin mediated transcription, these findings suggest that DKK1 is part of a negative feedback loop in MM and may act as a tumor suppressor. In line with this hypothesis, we show here that DKK1 expression is low or undetectable in a subset of patients with advanced MM as well as in MM cell lines. This absence of DKK1 is correlated with enhanced Wnt pathway activation, evidenced by nuclear accumulation of β-catenin, which in turn can be antagonized by restoring DKK1 expression. Analysis of the DKK1 promoter revealed CpG island methylation in several MM cell lines as well as in MM cells from patients with advanced MM. Moreover, demethylation of the DKK1 promoter restores DKK1 expression, which results in inhibition of β-catenin/TCF-mediated gene transcription in MM lines. Taken together, our data identify aberrant methylation of the DKK1 promoter as a cause of DKK1 silencing in advanced stage MM, which may play an important role in the progression of MM by unleashing Wnt signaling.