Cancer stem cells are rare tumor cells characterized by their ability to self-renew and to induce tumorigenesis. They are present in gliomas and may be responsible for the lethality of these incurable brain tumors. In the most aggressive and invasive type, glioblastoma multiforme (GBM), an average of about one year spans the period between detection and death [1]. The resistence of gliomas to current therapies may be related to the existence of cancer stem cells [2-6]. We find that human gliomas display a stemness signature and demonstrate that HEDGEHOG (HH)-GLI signaling regulates the expression of stemness genes in and the self-renewal of CD133(+) glioma cancer stem cells. HH-GLI signaling is also required for sustained glioma growth and survival. It displays additive and synergistic effects with temozolomide (TMZ), the current chemotherapeutic agent of choice. TMZ, however, does not block glioma stem cell self-renewal. Finally, interference of HH-GLI signaling with cyclopamine or through lentiviral-mediated silencing demonstrates that the tumorigenicity of human gliomas in mice requires an active pathway. Our results reveal the essential role of HH-GLI signaling in controlling the behavior of human glioma cancer stem cells and offer new therapeutic possibilities.

Melanoma is one of the most aggressive cancers, and its incidence is increasing. These tumors derive from the melanocyte lineage and remain incurable after metastasis. Here we report that SONIC HEDGEHOG (SHH)-GLI signaling is active in the matrix of human hair follicles, and that it is required for the normal proliferation of human melanocytes in culture. SHH-GLI signaling also regulates the proliferation and survival of human melanomas: the growth, recurrence, and metastasis of melanoma xenografts in mice are prevented by local or systemic interference of HH-GLI function. Moreover, we show that oncogenic RAS-induced melanomas in transgenic mice express Gli1 and require Hh-Gli signaling in vitro and in vivo. Finally, we provide evidence that endogenous RAS-MEK and AKT signaling regulate the nuclear localization and transcriptional activity of GLI1 in melanoma and other cancer cells. Our data uncover an unsuspected role of HH-GLI signaling in melanocytes and melanomas, demonstrate a role for this pathway in RAS-induced tumors, suggest a general integration of the RAS/AKT and HH-GLI pathways, and open a therapeutic approach for human melanomas.cancer ͉ melanocyte ͉ hair follicle ͉ metastasis ͉ skin

Prostate cancer is the most common solid tumor in men, and it shares with all cancers the hallmark of elevated, nonhomeostatic cell proliferation. Here we have tested the hypothesis that the SONIC HEDGEHOG (SHH)-GLI signaling pathway is implicated in prostate cancer. We report expression of SHH-GLI pathway components in adult human prostate cancer, often with enhanced levels in tumors versus normal prostatic epithelia. Blocking the pathway with cyclopamine or anti-SHH antibodies inhibits the proliferation of GLI1 ؉ ͞PSA ؉ primary prostate tumor cultures. Inversely, SHH can potentiate tumor cell proliferation, suggesting that autocrine signaling may often sustain tumor growth. In addition, pathway blockade in three metastatic prostate cancer cell lines with cyclopamine or through GLI1 RNA interference leads to inhibition of cell proliferation, suggesting cell-autonomous pathway activation at different levels and showing an essential role for GLI1 in human cells. Our data demonstrate the dependence of prostate cancer on SHH-GLI function and suggest a novel therapeutic approach.S ONIC HEDGEHOG (SHH) signaling has been implicated in different aspects of animal development, acting through several components, including the transmembrane proteins PATCHED1 (PTCH1) and SMOOTHENED (SMOH), to activate the GLI zinc-finger transcription factors (1, 2). In addition, we and others have shown that SHH signaling is implicated in a number of tumors (reviewed in refs. 2 and 3), such as basal cell carcinomas (4-6), medulloblastomas (7,8), gliomas (7), sarcomas (9, 10), tumors of the digestive tract (11), small cell lung cancers (12,) and pancreatic carcinomas (13). To date there is no direct evidence linking SHH signaling to prostate cancer, the most common solid cancer in men (14), although we have found that sporadic prostate tumors express GLI1 (7), a reliable marker of SHH signaling (15,16). This observation allowed us to propose the hypothesis that the SHH-GLI pathway participates in prostate cancer (7). Consistently, Shh signaling has been found to be essential for prostate patterning and development (17)(18)(19)(20)(21)(22), and genetic mapping data has revealed that at least two key components of the SHH-GLI pathway [SMOH and SUPPRESSOR OF FUSED (SUFUH)] are located in chromosomal regions implicated in familial human prostate cancer (23,24). Here we have tested the involvement of SHH-GLI signaling in prostate cancer. MethodsCell Lines and Primary Cultures. The PC3, LNCaP, and DU145 cell lines (25-27) were purchased from American Type Culture Collection and grown as specified. All primary prostate tumors were obtained following approved protocols. Tumors in PBS were chopped with a razor blade and incubated with Papain for 1 h at 37°C, they were then dissociated by passing them through a fire-polished pipette and washed several times in serum containing media. All dissociated primary tumors were plated in polyornithinand laminin-treated p16 plates in DMEM-F12 with 10% FBS at Ϸ30,000 cells per p16 well. Primary cultures were ...