Yu-Jen Chen

SONIC HEDGEHOG SIGNALING PROTECTS HUMAN HEPATOCELLULAR CARCINOMA CELLS AGAINST IONIZING RADIATION IN AN AUTOCRINE MANNER

PURPOSEnSonic hedgehog (Shh) signaling is critical to embryogenesis and resistance to chemotherapy. We aimed to examine the role of Shh signaling in the response to radiation of human hepatocellular carcinoma (HCC) cells.nnnMETHODS AND MATERIALSnResponse to ionizing radiation therapy (RT) was evaluated by clonogenic assay. Quantitative RT-polymerase chain reaction for patched-1 (PTCH-1) expression was performed. Cytosolic accumulation of Shh and nuclear translocation of Gli-1 were assessed by immunofluorescence. Gli-1 knockdown was done by RNA interference (RNAi). Immunoprecipitation was performed to detect Shh ligand in conditioned medium. Immunofluorescent stain for γ-H2AX was used as an index of DNA double strand breaks (DSB). Expression of proteins related to DNA damage repair was assessed by Western blotting.nnnRESULTSnWe found that Shh ligand could protect human HCC HA22T and Sk-Hep1 cells against RT. In HA22T cells, Shh ligand activated the Shh signaling with upregulation of Shh, PTCH-1, and Gli-1 expression. The nuclear translocation of Gli-1 further supports the activation of Gli-1. The radioprotection by Shh ligand was partly blocked by Shh antibody neutralization and was abolished by Gli-1 RNAi, suggesting a critical role of Shh signaling in radiation resistance. Furthermore, we noted that soluble factors secreted into conditioned medium, either constitutively or responding to radiation, by HA22T or Sk-Hep1 cells protected subsequent culturing cells against RT. Immunoprecipitation shows the presence of Shh peptide in conditioned medium. Intriguingly, antibody neutralization of Shh ligand or knockdown of Gli-1 reversed the radioprotective effect of conditioned medium. Furthermore, Shh ligand reduced the RT-induced phosphorylation of checkpoint kinase 1 and impaired the repair of DNA DSB.nnnCONCLUSIONSnActivation of Shh signaling protects HCC cells against ionizing radiation in an autocrine manner. Impairment of DNA damage repair might involve mechanism of Shh-induced radioresistance. Targeting Shh signaling pathway may be a novel strategy to enhance the radioresponse of human HCC cells.

Expression of hedgehog signaling molecules as a prognostic indicator of oral squamous cell carcinoma.

Recent studies have indicated hedgehog pathway plays a role in carcinogenesis of certain cancers. We investigated the clinical significance of its signaling components, including Sonic hedgehog (Shh), Patched (Ptch), and Gli‐1, in oral squamous cell carcinoma (OSCC).

Sonic hedgehog signaling regulates Bcr-Abl expression in human chronic myeloid leukemia cells.

PURPOSEnBcr-Abl fusion protein activates tyrosine kinase, resulting in the proliferation of leukemia cells, especially chronic myeloid leukemia (CML) cells. Imatinib (IM) effectively targets Bcr-Abl tyrosine kinase, but development of resistance to IM occurs with varying frequency.nnnMETHODSnElucidation of the common regulatory pathway upstream of Bcr-Abl in IM-sensitive and IM-resistant CML cells is important for developing novel therapeutics against CML.nnnRESULTSnThis study demonstrated that IM preferentially inhibited the viability and Bcr-Abl expression in IM-sensitive K562 (K562) cells, but not in Bcr-Abl overexpressing IM-resistant K562 (K562R) cells. Both K562 and K562R cells expressed Shh preproprotein, cleaved Shh C-terminal and N-terminal peptides, as well as mRNA level of major Shh signaling molecules, including sonic hedgehog (Shh), patched (PTCH), smoothened (Smo) and Gli-1. Moreover, Gli-1 translocation into nucleus was evident in these two cell lines, suggesting that both K562 and K562R cells possess activated and major components of the Shh signaling pathway. Silencing of Gli-1 by interference RNA was accompanied by inhibition of Bcr-Abl protein expression. Pharmacological suppression of Bcr-Abl expression was restored by the Smo agonist purmorpharmine. Treatment of Shh peptide in both K562 and K562R cells not only increased Shh and Gli-1 expression, but also up-regulated Bcr-Abl expression. Resveratrol, a known Bcr-Abl inhibitor, reduced Gli-1 activation and inhibited the viability of CML cells.nnnCONCLUSIONSnShh signaling may regulate Bcr-Abl expression in human chronic myeloid leukemia cells. Novel compounds inhibiting both Shh signaling and Bcr-Abl expression, such as resveratrol, may have potential to be effective agents against CML independent of IM resistance.

Andrographolide Sensitizes Ras-Transformed Cells to Radiation in vitro and in vivo

PURPOSEnIncreasing the sensitivity of tumor cells to radiation is a major goal of radiotherapy. The present study investigated the radiosensitizing effects of andrographolide and examined the molecular mechanisms of andrographolide-mediated radiosensitization.nnnMETHODS AND MATERIALSnAn H-ras-transformed rat kidney epithelial (RK3E) cell line was used to measure the radiosensitizing effects of andrographolide in clonogenic assays, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide assays, and a xenograft tumor growth model. The mechanism of andrographolide-sensitized cell death was analyzed using annexin V staining, caspase 3 activity assays, and terminal transferase uridyl nick end labeling assays. The roles of nuclear factor kappa B (NF-kappaB) and Akt in andrographolide-mediated sensitization were examined using reporter assays, electrophoretic mobility shift assays, and Western blotting.nnnRESULTSnConcurrent andrographolide treatment (10 microM, 3 h) sensitized Ras-transformed cells to radiation in vitro (sensitizer enhancement ratio, 1.73). Andrographolide plus radiation (one dose of 300 mg/kg peritumor andrographolide and one dose of 6 Gy radiation) resulted in significant tumor growth delay (27 +/- 2.5 days) compared with radiation alone (22 +/- 1.5 days; p <.05). Radiation induced apoptotic markers (e.g., caspase-3, membrane reversion, DNA fragmentation), and andrographolide treatment did not promote radiation-induced apoptosis. However, the protein level of activated Akt was significantly reduced by andrographolide. NF-kappaB activity was elevated in irradiated Ras-transformed cells, and andrographolide treatment significantly reduced radiation-induced NF-kappaB activity.nnnCONCLUSIONnAndrographolide sensitized Ras-transformed cells to radiation both in vitro and in vivo. Andrographolide-mediated radiosensitization was associated with downregulation of Akt and NF-kappaB activity. These observations indicate that andrographolide is a novel radiosensitizing agent with potential application in cancer radiotherapy.

Targeting Sonic Hedgehog Signaling by Compounds and Derivatives from Natural Products

Cancer stem cells (CSCs) are a major cause of cancer treatment failure, relapse, and drug resistance and are known to be responsible for cancer cell invasion and metastasis. The Sonic hedgehog (Shh) signaling pathway is crucial to embryonic development. Intriguingly, the aberrant activation of the Shh pathway plays critical roles in developing CSCs and leads to angiogenesis, migration, invasion, and metastasis. Natural compounds and chemical structure modified derivatives from complementary and alternative medicine have received increasing attention as cancer chemopreventives, and their antitumor effects have been demonstrated both in vitro and in vivo. However, reports for their bioactivity against CSCs and specifically targeting Shh signaling remain limited. In this review, we summarize investigations of the compounds cyclopamine, curcumin, epigallocatechin-3-gallate, genistein, resveratrol, zerumbone, norcantharidin, and arsenic trioxide, with a focus on Shh signaling blockade. Given that Shh signaling antagonism has been clinically proven as effective strategy against CSCs, this review may be exploitable for development of novel anticancer agents from complementary and alternative medicine.

Transdermal Fentanyl for Pain Caused by Radiotherapy in Head and Neck Cancer Patients Treated in an Outpatient Setting: A Multicenter Trial in Taiwan

OBJECTIVEnThis study evaluated the efficacy and safety of transdermal fentanyl in the outpatient treatment of head and neck cancer patients with pain caused by radiotherapy.nnnMETHODSnPatients with a visual analogue scale score >or=4 were invited to participate in the study. The following variables were collected: visual analogue scale, the Brief Pain Inventory, concomitant pain medications and adverse effects. A total of 163 head and neck cancer patients were enrolled (148 males and 15 females; median age, 53 years; age range, 21-72 years). Seventy-two (44%) patients had a visual analogue scale score >6 at enrollment, despite the use of non-steroidal anti-inflammatory drugs or weak opioids. Ninety-four (57.7%) patients received concurrent chemotherapy.nnnRESULTSnA total of 88 patients completed the study, whereas 55 underwent a drop-out by side effects. The most frequently reported adverse events were vomiting (23.9%) and nausea (16.6%). Treatment with transdermal fentanyl resulted in a significant decrease in visual analogue scale and Brief Pain Inventory scores that persisted during treatment. In the overall efficacy evaluation, the pain-alleviating effect, the easiness of application and the overall impression of transdermal fentanyl were rated as good by 54.5%, 65.9% and 59.1% of the completers, respectively. Effects of transdermal fentanyl were rated as good by 64.8% of the investigators.nnnCONCLUSIONSnOur data provide evidence that transdermal fentanyl is effective and relatively easy to use for outpatient treatment of pain control in head and neck cancer patients following radiotherapy in selected patients. Reduction of side effects and effective pain management need to be paramount in the management of head and neck cancer patients undergoing radiotherapy.

Midostaurin (PKC412) modulates differentiation and maturation of human myeloid dendritic cells

Midostaurin, a tyrosine kinase inhibitor, has been shown efficacy against acute myeloid leukemia and various other malignancies in clinical trials. Prior studies indicate midostaurin affects the function of immune cells such as lymphocytes and macrophages. To understand the effect of midostaurin on human myeloid dendritic cells (DCs), we conducted an ex vivo study using immature DCs differentiated from CD14(+) monocytes and further maturated using lipopolysaccharide. Addition of midostaurin to a culture of starting CD14(+) monocytes markedly and dose-dependently reduced DC recovery. Mature DCs differentiating in the presence of midostaurin had fewer, shorter cell projections than those differentiating in the absence of midostaurin. Changes in morphological features characteristic of apoptotic cells were also evident. Moreover, midostaurin affected DC differentiation and maturation patterns; CD83 expression levels decreased, whereas CD14 and CD80 expressions increased. Additionally, DCs derived in the presence of midostaurin possessed a lower endocytotic capacity and less allostimulatory activity on naive CD4(+)CD45(+)RA(+) T cell proliferation than those derived in its absence, suggesting that midostaurin redirects DC differentiation toward a less mature stage and that this effect is not solely due to its cytotoxicity. Whether this effect underlies immune suppression or tolerance to disease treatments with unwanted immune reactions needs further evaluation.

Cancer Stem Cells and Sonic Hedgehog Signaling in Head and Neck Cancer: Potential Targets for Overcoming Chemoradiation Resistance

Cancer stem cells (CSC) are a small and distinct population of cancer cells that possess self-renewal and differentiation ability and are relatively resistant to treatment including chemotherapy and radiotherapy. Sonic hedgehog (SHH) and related signaling molecules are critical to embryonic development and regulate both proliferation and differentiation of various types of stem cells, including CSC. This article provides a brief overview of the SHH signaling pathway, summarizes the correlation between SHH signaling and treatment resistance of cancer cells and discusses the recent advances in targeting this signaling cascade to overcome treatment resistance. We proposed that CSC and their related the SHH signaling pathway might be potential targets for overcoming chemoradiation resistance of head and neck cancer cells. This has been under investigation by our comprehensive team treating head and neck cancers.

Complementary and Alternative Medicine in Cancer Stem Cells

Cancer stem cells (CSCs), a small subpopulation of cancer cells prone to form tumorigenesis, resemble normal stem cells with self-renewal and differentiation characteristics. CSCs have recently been identified in leukemia and various types of solid tumors. There is growing evidence indicating that CSCs might be the principal cause for the development of resistance to cancer treatments, recurrence, and metastasis. Emerging data indicates that CSCs and normal stem cells may share cellular pathways regulating proliferation and differentiation. These pathways include, but not limited to, polycomb group transcriptional repressor Bmi-1, Notch, Sonic hedgehog, and Wnt pathways. Therefore, identification of CSC markers, selection and development of unique therapeutics targeting CSCs and related signaling pathways have been extensively investigated. In this special issue, we publish articles that explore aspects of complementary and alternative medicine (CAM) associated with CSCs in basic research, in translational medicine, and in clinical application for cancer treatment. The topic will highlight current research, encourage additional research, draw attention to the subject, and ultimately help advance the field of CAM application in CSCs.