The transcription factor Forkhead box M1 (FoxM1) is overexpressed in malignant glioma. However, the functional importance of this factor in human glioma is not known. In the present study, we found that FoxM1B was the predominant FoxM1 isoform expressed in human glioma but not in normal brain tissue. The level of FoxM1 protein expression in human glioma tissues was directly correlated with the glioma grade. The level of FoxM1 protein expression in human glioblastoma tissues was inversely correlated with patient survival. Enforced FoxM1B expression caused SW1783 and Hs683 glioma cells, which do not form tumor xenografts, to regain tumorigenicity in nude mouse model systems. Moreover, gliomas that arose from FoxM1B-transfected anaplastic astrocytoma SW1783 cells displayed glioblastoma multiforme phenotypes. Inhibition of FoxM1 expression in glioblastoma U-87MG cells suppressed their anchorage-independent growth in vitro and tumorigenicity in vivo. Furthermore, we found that FoxM1 regulates the expression of Skp2 protein, which is known to promote degradation of the cell cycle regulator p27 Kip1 . These results showed that FoxM1 is overexpressed in human glioblastomas and contributes to glioma tumorigenicity. Therefore, FoxM1 might be a new potential target of therapy for human malignant gliomas. (Cancer Res 2006; 66(7): 3593-602)

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Chronic myelogenous leukemia (CML) in IntroductionThe management of chronic myelogenous leukemia (CML) has been revolutionized by kinase inhibitors that were developed in response to cues from biologic studies of the BCR-ABL1 oncogene. However two challenging problems persist: the progression of the disease to blast crisis and resistance to kinase inhibition. 1 Continued investigation of BCR-ABL1 kinase signaling will provide insight into these problems. Members of the Src kinase family, which regulate proliferation, differentiation, and motility, 2 are known downstream targets of BCR-ABL1. In myeloid cell lines, BCR-ABL1 activates Lyn and Hck. 3,4 Several reports link growth, survival, and imatinib resistance of Philadelphia chromosome-positive (Ph ϩ ) leukemias to Lyn kinase expression and activation. 5,6 However, reports examining Fyn, a ubiquitously expressed Src family member, are sparse. Of note, phase-specific gene expression in CML using microarray analyses revealed that Fyn gene expression was linked to imatinib relapse. 7 In addition, a separate study using combined systems biology and gene expression approaches in Ph ϩ acute lymphoblastic leukemia (ALL) specimens identified Fyn as a hub for signaling. 8 Here we show that Fyn protein expression is increased in patients with blast-crisis CML compared with chronic-phase disease. By examining effects of silencing Fyn using shRNA, we find that Fyn transduces a mitogenic signal. Collectively, our results identify a novel effect of BCR-ABL1-up-regulation of Fyn-and delineate consequences of the observed up-regulation. MethodsPatient specimens were used for this study and were collected after informed consent was obtained in accordance with the Declaration of Antibodies, chemicals, and cell linesAntibodies were purchased from sources outlined in Document S1 (available on the Blood website; see the Supplemental Materials link at the top of the online article). Imatinib was kindly provided by Dr Elisabeth Buchdunger at Novartis Pharmaceuticals (Basel, Switzerland). Murine growth factor-dependent pro-B lymphoid BaF3 cell lines transformed with vector, wild-type BCR-ABL1, or imatinib-resistant mutant BCR-ABL1 were kindly provided by Dr Charles Sawyers 9 and were cultured as previously described. 9 K562 cells, TonB210 cells stably expressing a tetracyclineinducible BCR-ABL1 expression vector (kindly provided by Dr George Daley, Children's Hospital Boston, Harvard Medical School, MA), 10 and mouse 32D and 32Dp210 cells were maintained in RPMI1640 medium with 10% FBS supplemented. Mouse 32D cells were supplemented with 10% WEHI-cultured conditioned medium as a source of interleukin-3 (IL-3) in addition to 10% FBS. Design of shRNA to FynK562 cells were transfected with Fyn shRNA and control vectors (TranSilent human shRNA from Panomics, Redwood, CA) using the Nucleofector system kit V and transfection program T-16 (Amaxa Biosystems, Cologne, Germany). Lentiviral knockdown of Fyn and rescue design is detailed in Document S1. To generate the rescue construct, 4 nucleo...

Purpose Large diameter perineural prostate cancer is associated with poor outcomes. GDNF, with its co-receptor GFRα1, binds RET and activates downstream pro-oncogenic signaling. Since both GDNF and GFRα1 are secreted by nerves, we examined the role of RET signaling in prostate cancer. Experimental Design Expression of RET, GDNF and/or GFRα1 was assessed. The impact of RET signaling on proliferation, invasion and soft agar colony formation, perineural invasion and growth in vivo was determined. Cellular signaling downstream of RET was examined by Western blotting. Results RET is expressed in all prostate cancer cell lines. GFRα1 is only expressed in 22Rv1 cells, which is the only line that responds to exogenous GDNF. In contrast, all cell lines respond to GDNF plus GFRα1. Conditioned medium from dorsal root ganglia contains secreted GFRα1 and promotes transformation related phenotypes, which can be blocked by anti-GFRα1 antibody. Perineural invasion in the dorsal root ganglion assay is inhibited by anti-GFRα antibody and RET knockdown. In vivo, knockdown of RET inhibits tumor growth. RET signaling activates ERK or AKT signaling depending on context, but phosphorylation of p70S6 kinase is markedly increased in all cases. Knockdown of p70S6 kinase markedly decreases RET induced transformed phenotypes. Finally, RET is expressed in 18% of adenocarcinomas and all three small cell carcinomas examined. Conclusions RET promotes transformation associated phenotypes, including perineural invasion in prostate cancer via activation of p70S6 kinase. GFRα1, which is secreted by nerves, is a limiting factor for RET signaling, creating a perineural niche where RET signaling can occur.