Search

Rhabdomyosarcoma (RMS) is the most common pediatric soft tissue sarcoma. In children, the 2 major RMS subtypes are alveolar and embryonal RMS. Aberrant Hedgehog/Patched1 (Hh/Ptch) signaling is a hallmark of embryonal RMS. We demonstrate that mice carrying a Ptch mutation in mesodermal Delta1-expressing cells develop embryonal-like RMS at a similar rate as mice harboring a Ptch mutation in the germline or the brachury-expressing mesoderm. The tumor incidence decreases dramatically when Ptch is mutated in Myf5- or Pax3-expressing cells. No RMS develop from Myogenin/Mef2c-expressing cells. This suggests that Hh/Ptch-associated RMS are derived from Delta1-positive, Myf5-negative, Myogenin-negative and Pax3-negative mesodermal progenitors that can undergo myogenic differentiation but lack stable lineage commitment. Additional preliminary genetic data and data on mesodermal progenitors further imply an interplay of Hh/Ptch and Delta/Notch signaling activity during RMS initiation. In contrast, Wnt signals supposedly suppress RMS formation because RMS multiplicity decreases after inactivation of the Wnt-inhibitor Wif1. Finally, our results strongly suggest that the tumor-initiating event determines the lineage of RMS origin.

Liposomes are frequently used in cancer therapy to encapsulate and apply anticancer drugs. Here, we show that a systemic treatment of mice bearing skin tumors with empty phosphatidylcholine liposomes (PCL) resulted in inhibition of tumor growth, which was similar to that observed with the synthetic bacterial lipoprotein and TLR1/2 agonist Pam(3)CSK(4) (BLP). Both compounds led to a substantial decrease of macrophages in spleen and in the tumor-bearing skin. Furthermore, both treatments induced the expression of typical macrophage markers in the tumor-bearing tissue. As expected, BLP induced the expression of the M1 marker genes Cxcl10 and iNOS, whereas PCL, besides inducing iNOS, also increased the M2 marker genes Arg1 and Trem2. In vitro experiments demonstrated that neither PCL nor BLP influenced proliferation or survival of tumor cells, whereas both compounds inhibited proliferation and survival and increased the migratory capacity of bone marrow-derived macrophages (BMDM). However, in contrast to BLP, PCL did not activate cytokine secretion and induced a different BMDM phenotype. Together, the data suggest that similar to BLP, PCL induce an antitumor response by influencing the tumor microenvironment, in particular by functional alterations of macrophages, however, in a distinct manner from those induced by BLP.

Rhabdomyosarcoma (RMS) is the most common soft tissue neoplasm of children, and those metastatic at presentation have a poor prognosis. RMS development is related to defective skeletal muscle differentiation, involving a variety of cell signaling and transcriptional control pathways, including aberrant hedgehog signaling. Here we evaluate Fem1a, a gene highly expressed in skeletal muscle, as a candidate for involvement in RMS. Fem1a is a homolog of fem-1, which controls cell fate decisions in the sex determination pathway of Caenorhabditis elegans, a pathway with homology to mammalian hedgehog signaling. We show that Fem1a expression is activated during myocyte differentiation of C2C12 myoblasts, and this expression is largely confined to the terminally differentiating pool, not to the satellite-cell-like quiescent reserve cell pool. We find that the human homolog, FEM1A, is downregulated in all of 8 different human RMS cell lines, including those derived from embryonal and alveolar RMS. Using mouse genetic models of RMS development, we further show that Fem1a is consistently downregulated in primary RMS from Ptch1+/– mice, from p53–/– mice, from p53+/–; Ptch1+/– mice, and from HGF/SF-Ink4a/Arf–/– mice. Therefore, Fem1a downregulation may be involved in, and/or a marker of, an early cell fate defect fundamental to RMS pathogenesis.

The identification of mutations in the human homolog of the Drosophila segment polarity gene Patched in basal cell carcinoma has sparked intense interest in the role of this gene in human disorders. The transmembrane protein Patched is a receptor for the morphogene Sonic Hedgehog. Sonic Hedgehog/Patched signaling involves another transmembrane protein, Smoothened, and its intracellular effectors, including the proto-oncogene GLI1. During the past 2 years it has become evident that mutations in Patched or in one of the components of its signaling pathway contribute to the formation of several common human tumors. It is now well established that Patched is a tumor suppressor gene. The Sonic Hedgehog/Patched/Smoothened signaling pathway is thus rapidly emerging as one of the most important regulators of oncogenic transformation. This pathway also plays an important role during mammalian embryonic development. This dual role is especially visible in humans with inherited Patched mutations. Such patients suffer from Gorlin, or nevoid basal cell carcinoma, syndrome and exhibit a variety of developmental defects accompanied by a predisposition to tumor formation. Activating mutations in Sonic Hedgehog and Smoothened lead to similar phenotypes as do loss-of function mutations in Patched. By means of transgenic and gene targeting technologies the respective mutations have been expressed in the mouse. Such mutant mouse strains exhibit many symptoms observed in humans. These strains are useful models to study the pathogenesis of several common human tumors and developmental defects. Furthermore they provide important tools to study the Sonic Hedgehog/Patched/Smoothened signaling at the molecular and biochemical level.

Gorlin (or nevoid basal cell carcinoma) syndrome is characterized by a variety of clinical problems including generalized overgrowth of the body, cysts, developmental abnormalities of the skeleton and a predisposition to benign and malignant tumors. The syndrome results from germline mutations of the human homolog of the drosophila segment polarity gene patched (ptc). Here we report that mice heterozygous for ptc develop many of the features characteristic of Gorlin syndrome and that they exhibit a high incidence of rhabdomyosarcomas (RMS), the most common soft-tissue sarcoma in children. The downstream signalling partner of ptc, gli1, was overexpressed in all RMSs analyzed, indicating that abnormal signalling of the ptc-gli1 pathway may be common for the various tumors associated with the syndrome. igf2, implicated in the formation of RMSs, was also overexpressed, suggesting cross-talk between the ptc and igf2 pathways in tumorigenesis. Developmental defects in Gorlin syndrome resemble those induced by ionizing radiation. We show that ptc heterozygous mice exhibit increased incidence of radiation-induced teratogenesis. This suggests a role for ptc in the response to ionizing radiation and provides a model for both the systemic (developmental) and stochastic (cancer) abnormalities observed in Gorlin syndrome.

Tyrosine kinase growth factor receptors and Ras/Raf/MEK/MAPK signalling have been implicated in the suppression1–3 as well as augmentation of programmed cell death4. In addition, a Ras-independent role for Raf as a suppressor of programmed cell death has been suggested by the recent finding that Craf1 interacts with members of the Bcl-2 family at mitochondrial membranes5. However, genetic studies of C elegans6 and Drosophila7, as well as the targeted mutagenesis of the murine Araf gene8, have failed to support such a role. Here we show that mice with a targeted disruption in the Braf gene die of vascular defects during mid-gestation. Braf−/− embryos, unlike Araf−/−8 or Craf1−/− embryos (L.W. et al., unpublished), show an increased number of endothelial precursor cells, dramatically enlarged blood vessels and apoptotic death of differentiated endothelial cells. These results establish Braf as a critical signalling factor in the formation of the vascular system and provide the first genetic evidence for an essential role of a Raf gene in the regulation of programmed cell death.