gdc-0449 and Lung-Neoplasms

It has long been known that the hedgehog signaling pathway is crucial for embryonic development, in both Drosophila and vertebrate systems. During the past few years its implication in carcinogenesis has become clear and today it is acknowledged that this pathway plays a role in the malignant transformation of multiple cell types, either owing to the mutation of some of its components or to its erratic activation. New molecular targeted therapies that inhibit the pathway have shown their unquestionable efficiency in several tumours -among which basal cell carcinoma, medulloblastoma, or pancreatic adenocarcinoma. The assessment of these inhibitors in other types of tumours is currently underway with promising results, suggesting that the Sonic hedgehog signalling pathway may become one of the therapeutic targets of the future.

Topics: Anilides; Animals; Carcinoma, Basal Cell; Cerebellar Neoplasms; Embryonic Development; Embryonic Induction; Hedgehog Proteins; Humans; Lung Neoplasms; Medulloblastoma; Molecular Targeted Therapy; Mutation; Neoplasms; Neoplastic Stem Cells; Pancreatic Neoplasms; Patched Receptors; Patient Selection; Pyridines; Receptors, Cell Surface; Signal Transduction; Skin Neoplasms; Small Cell Lung Carcinoma

Preclinical targeting of the hedgehog pathway by vismodegib and of insulin-like growth factor 1 receptor by cixutumumab enhances the efficacy of chemotherapy and also demonstrates activity against the tumor cell fraction responsible for disease recurrence in small cell lung cancer.. Patients with newly diagnosed extensive-stage small cell lung cancer (SCLC-ED) were randomized to receive four 21-day cycles of cisplatin and etoposide alone (cisplatin at 75 mg/m(2) on day 1 and etoposide at 100 mg/m(2) on days 1-3; arm A) or in combination with either vismodegib (150 mg/d by mouth; arm B) or cixutumumab (6 mg/kg/wk intravenously on day 1; arm C). The primary endpoint was progression-free survival (PFS). Circulating tumor cells (CTCs) were isolated/enumerated with the Veridex CellSearch platform at the baseline.. One hundred fifty-two eligible patients were treated. Patient demographics and disease characteristics were well balanced between the 3 arms except for the higher rate with a performance status of 0 in arm B (P = .03). The median PFS times in arms A, B, and C were 4.4, 4.4, and 4.6 months, respectively; the median overall survival (OS) times were 8.8, 9.8, and 10.1 months, respectively; and the response rates were 48%, 56%, and 50%, respectively. None of the comparisons of these outcomes were statistically significant. The median OS was 10.5 months for those with low CTC counts (≤100/7.5 mL) at baseline and 7.2 months for those with high CTC counts (hazard ratio, 1.74; P = .006).. There was no significant improvement in PFS or OS with the addition of either vismodegib or cixutumumab to chemotherapy in patients with SCLC-ED. A low baseline CTC count was associated with a favorable prognosis. Cancer 2016;122:2371-2378. © 2016 American Cancer Society.

Topics: Adult; Aged; Aged, 80 and over; Anilides; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Biomarkers; Cisplatin; Etoposide; Female; Humans; Lung Neoplasms; Male; Middle Aged; Neoplasm Staging; Neoplastic Cells, Circulating; Pyridines; Small Cell Lung Carcinoma; Treatment Outcome

The use of 18-fluorodeoxyglucose (FDG) positron emission tomography with computed tomography (PET/CT) in subjects with advanced basal cell carcinoma (BCC) has not been fully explored due to the rarity of disease presentation. This study evaluated PET/CTs from subjects with advanced BCC participating in a phase I dose-escalation clinical trial of vismodegib. Fourteen subjects with BCC were imaged with 18-FDG PET/CT for lesion identification and response categorizing (European Organisation for Research and Treatment for Cancer [EORTC] and PET response criteria in solid tumors [PERCIST] 1.0). Several parameters including metabolic activity of target lesions, site of disease presentation and spread, treatment response, and prognostic significance of metabolic activity following therapy were evaluated. All subjects exhibited at least one hypermetabolic lesion. Most subjects had only four organ systems involved at study enrollment: skin-muscle (93%), lung (57%), lymph nodes (29%), and bone (21%). SUVmax measured across all lesions decreased (median 33%, SD ± 45%) following therapy with metabolic activity normalizing or disappearing in 42% of lesions. No significant difference was observed between EORTC and PERCIST 1.0. Subjects that demonstrated at least a 33% reduction in SUVmax from baseline had a significantly longer progression-free survival (PFS) (median 17 months, 95% confidence interval [CI] ±4 months vs. 9 months, 95% CI ±5 months, P = 0.038) and overall survival (OS) (median 24 months, 95% CI ±4 months vs. 17 months, 95% CI ±13 months, P = 0.019). BCC lesions are hypermetabolic on 18-FDG PET/CT. A decrease in SUVmax was associated with improved PFS and OS. These results further support the incorporation of 18-FDG PET/CT scans in advanced BCC management.

Topics: Adult; Aged; Aged, 80 and over; Anilides; Antineoplastic Agents; Bone Neoplasms; Carcinoma, Basal Cell; Disease-Free Survival; Female; Fluorodeoxyglucose F18; Humans; Lung Neoplasms; Lymphatic Metastasis; Male; Middle Aged; Multimodal Imaging; Muscle Neoplasms; Positron-Emission Tomography; Pyridines; Skin Neoplasms; Tomography, X-Ray Computed; Treatment Outcome

Lung adenocarcinoma (LUAD), which comprises over 50% of all cases of non-small-cell lung cancer, has a poor prognosis and requires novel therapeutic approaches. The sonic Hedgehog (Shh) pathway, which plays a crucial role in differentiation, proliferation, and survival of cancer cells, is likely to be activated in LUADs, suggesting the Shh pathway as a potential therapeutic target for LUAD treatment. In this study, we reported that vismodegib, an inhibitor of the Shh pathway, only elicited minor antitumor efficacy in A549 and NCI-H1975 LUAD cells as well as in the xenograft tumors, with overexpressed GLI2 and increased autophagic activity. The aberrant autophagy in LUAD cells was further confirmed by the three main stages of autophagic flux, including the formation of autophagosomes, the fusion of autophagosomes with lysosomes, and degradation of autophagosomes in lysosomes. Furthermore, inhibition of autophagy by siRNA against ATG5 or ATG7 rescued the sensitivity of A549 and NCI-H1975 LUAD cells to vismodegib in vitro. Meanwhile, administration of the pharmaceutical inhibitor of autophagy, chloroquine, contributed to the enhanced anti-LUAD efficacy of vismodegib in vivo, probably through overproduction of ROS, acceleration of apoptosis, and suppression of GLI2 in LUAD tissues. In summary, our research revealed that downregulating autophagy facilitated the anti-LUAD efficacy of the Shh pathway suppression, thus highlighting a potential approach for LUAD therapy via simultaneously targeting the Shh signaling and autophagy pathway.

Topics: A549 Cells; Adenocarcinoma of Lung; Anilides; Animals; Autophagy; Cell Line, Tumor; Down-Regulation; Drug Resistance, Neoplasm; Hedgehog Proteins; Heterografts; Humans; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Nuclear Proteins; Pyridines; Reactive Oxygen Species; Signal Transduction; Xenograft Model Antitumor Assays; Zinc Finger Protein Gli2

We report a case of a 60-year-old man who was a former cigar smoker with a slow-growing, large exophytic left shoulder mass (15 cm in diameter) and later found to have left axillary lymphadenopathy. Fine needle aspirate biopsy of the left shoulder mass revealed squamous cell carcinoma (SCC). However, pathology of the enlarged left axillary lymph node was reported as metastatic adenocarcinoma. The patient underwent surgical resection of the shoulder mass which comprised of SCC (>95%) and adenoid basal cell carcinoma (BCC) as a second component of the tumour. The BCC had identical histology as the metastatic carcinoma in the left axillary lymph node. Therefore, diagnosis was revised as cutaneous collision tumour with metastatic BCC. Six months later following adjuvant radiation therapy, the patient was diagnosed with metastatic BCC in the right lung. Stereotactic body radiation therapy (SBRT) and a selective hedgehog pathway inhibitor vismodegib were given with only limited efficacy. Clinical trial registration number NCT03132636.

Topics: Anilides; Axilla; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Hedgehog Proteins; Humans; Lung Neoplasms; Lymph Nodes; Male; Middle Aged; Pyridines; Radiosurgery; Radiotherapy, Adjuvant; Skin Neoplasms; Treatment Outcome

Tyrosine kinase inhibitors (TKIs) of the EGF receptor (EGFR) have provided a significant improvement in the disease outcome of nonsmall cell lung cancer (NSCLC). Unfortunately, resistance to these agents frequently occurs, and it is often related to the activation of the Hedgehog (Hh) and MET signaling cascades driving the epithelial-to-mesenchymal transition (EMT). Because the concomitant inhibition of both Hh and MET pathways restores the sensitivity to anti-EGFR drugs, here we aimed at discovering the first compounds that block simultaneously MET and SMO. By using an "in silico drug repurposing" approach and by validating our predictions both in vitro and in vivo, we identified a set of compounds with the desired dual inhibitory activity and enhanced antiproliferative activity on EGFR TKI-resistant NSCLC. The identification of the known MET TKIs, glesatinib and foretinib, as negative modulators of the Hh pathway, widens their application in the context of NSCLC.

Topics: Animals; Antineoplastic Agents; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Proliferation; Drug Repositioning; Drug Resistance, Neoplasm; ErbB Receptors; Female; HEK293 Cells; Humans; Lung; Lung Neoplasms; Mice, Inbred BALB C; Mice, Nude; Molecular Docking Simulation; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-met; Smoothened Receptor

Abnormal expression of chondroitin sulfate has been found in many types of cancer, while its biological functions in hepatocellular carcinoma (HCC) progression remain uninvestigated. Here, we report that chondroitin sulfate synthase 1 (CHSY1), the enzyme that mediates the polymerization step of chondroitin sulfate, is a critical mediator of malignant character in HCC that acts via modulating the activity of the hedgehog signaling. CHSY1 was up-regulated frequently in HCC where these events were associated with worse histologic grade and poor survival. Enforced expression of CHSY1 was sufficient to enhance cell growth, migration, invasion, and epithelial-mesenchymal transition, whereas silencing of CHSY1 suppressed these malignant phenotypes. Mechanistic investigations revealed that the increase of cell surface chondroitin sulfate by CHSY1 promoted sonic hedgehog binding and signaling. Inhibiting hedgehog pathway with vismodegib decreased CHSY1-induced migration, invasion, and lung metastasis of HCC cells, establishing the critical role of hedgehog signaling in mediating the effects of CHSY1 expression. Together, our results indicate that CHSY1 overexpression in HCC contributes to the malignant behaviors in cancer cells, we provide novel insights into the significance of chondroitin sulfate in hedgehog signaling and HCC pathogenesis.

Topics: Anilides; Animals; Carcinoma, Hepatocellular; Cell Movement; Cell Proliferation; Chondroitin Sulfates; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Glucuronosyltransferase; Hedgehog Proteins; Hep G2 Cells; Humans; Liver Neoplasms; Lung Neoplasms; Male; Mice, Inbred C57BL; Middle Aged; Multifunctional Enzymes; N-Acetylgalactosaminyltransferases; Neoplasm Grading; Neoplasm Invasiveness; Phenotype; Pyridines; RNA Interference; Signal Transduction; Transfection; Xenograft Model Antitumor Assays

An autocrine-driven upregulation of the Hedgehog (Hh) signaling pathway has been described in malignant pleural mesothelioma (MPM), in which the ligand, desert Hh (DHH), was produced from tumor cells. However, our investigation revealed that the Hh pathway is activated in both tumor and stroma of MPM tumor specimens and an orthotopic immunocompetent rat MPM model. This was demonstrated by positive immunohistochemical staining of Glioma-associated oncogene 1 (GLI1) and Patched1 (PTCH1) in both tumor and stromal fractions. DHH was predominantly expressed in the tumor fractions. To further investigate the role of the Hh pathway in MPM stroma, we antagonized Hh signaling in the rat model of MPM using a Hh antagonist, vismodegib, (100 mg/kg orally). Daily treatment with vismodegib efficiently downregulated Hh target genes Gli1, Hedgehog Interacting Protein (Hhip), and Ptch1, and caused a significant reduction of tumor volume and tumor growth delay. Immunohistochemical analyses revealed that vismodegib treatment primarily downregulated GLI1 and HHIP in the stromal compartment along with a reduced expression of previously described fibroblast Hh-responsive genes such as Fibronectin (Fn1) and Vegfa Primary cells isolated from the rat model cultured in 3% O2 continued to express Dhh but did not respond to vismodegib in vitro However, culture supernatant from these cells stimulated Gli1, Ptch1, and Fn1 expression in mouse embryonic fibroblasts, which was suppressed by vismodegib. Our study provides new evidence regarding the role of Hh signaling in MPM stroma in the maintenance of tumor growth, emphasizing Hh signaling as a treatment target for MPM. Mol Cancer Ther; 15(5); 1095-105. ©2016 AACR.

Topics: Anilides; Animals; Biomarkers; Cell Line, Tumor; Cell Proliferation; Cell Survival; Disease Models, Animal; Gene Expression Regulation, Neoplastic; Hedgehog Proteins; Humans; Lung Neoplasms; Magnetic Resonance Imaging; Mesothelioma; Mesothelioma, Malignant; Mice; NIH 3T3 Cells; Pleural Neoplasms; Pyridines; Rats; Signal Transduction; Stromal Cells; Tumor Burden; Xenograft Model Antitumor Assays

The present study sought to determine whether the Hedgehog (Hh) pathway is active and regulates the cell growth of cultured malignant pleural mesothelioma (MPM) cells and to evaluate the efficacy of pathway blockade using smoothened (SMO) antagonists (SMO inhibitor GDC-0449 or the antifungal drug itraconazole [ITRA]) or Gli inhibitors (GANT61 or the antileukemia drug arsenic trioxide [ATO]) in suppressing MPM viability.. Selective knockdown of SMO to inhibit Hh signaling was achieved by small interfering RNA in 3 representative MPM cells. The growth inhibitory effect of GDC-0449, ITRA, GANT61, and ATO was evaluated in 8 MPM lines, with cell viability quantified using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell death was determined by annexinV/propidium iodide staining and flow cytometry.. SMO small interfering RNA mediated a two- to more than fivefold reduction of SMO and Gli1 gene expression as determined by real-time quantitative reverse-transcriptase polymerase chain reaction, indicating significant Hh pathway blockade. This was associated with significantly reduced cell viability (34% ± 7% to 61% ± 14% of nontarget small interfering RNA controls; P = .0024 to P = .043). Treating MPM cells with Hh inhibitors resulted in a 1.5- to 4-fold reduction of Gli1 expression. These 4 Hh antagonists strongly suppressed MPM cell viability. More importantly, ITRA, ATO, GANT61 induced significant apoptosis in the representative MPM cells.. Hh signaling is active in MPM and regulates cell viability. ATO and ITRA were as effective as the prototypic SMO inhibitor GDC-0449 and the Gli inhibitor GANT61 in suppressing Hh signaling in MPM cells. Pharmaceutical agents Food and Drug Administration-approved for other indications but recently found to have anti-Hh activity, such as ATO or ITRA, could be repurposed to treat MPM.

Topics: Anilides; Antineoplastic Agents; Apoptosis; Arsenic Trioxide; Arsenicals; Cell Line, Tumor; Cell Proliferation; Cell Survival; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; Hedgehog Proteins; Humans; Itraconazole; Lung Neoplasms; Mesothelioma; Mesothelioma, Malignant; Molecular Targeted Therapy; Oxides; Pleural Neoplasms; Pyridines; Pyrimidines; Receptors, G-Protein-Coupled; RNA Interference; Signal Transduction; Smoothened Receptor; Time Factors; Transcription Factors; Transfection; Zinc Finger Protein GLI1

Basal cell carcinomas (BCCs) in patients with Gorlin syndrome have been reported to be extremely sensitive to Smoothened (SMO) inhibitors, a novel targeted therapy against the Hedgehog pathway, because of characteristic mutations in these patients. A few cases of disease refractory to oral therapy with SMO inhibitors have been reported in patients with Gorlin syndrome and nonmetastatic BCCs, but refractory disease in distantly metastatic tumors has not been documented in this high-risk group.. A man with Gorlin syndrome and innumerable cutaneous BCCs presented with biopsy-proven BCC in his lungs. After SMO inhibitor therapy, almost all of his cutaneous tumors shrank, but his lung metastases did not. These lung metastases remained refractory to treatment despite institution of a second SMO inhibitor.. We report a case of Gorlin syndrome in a patient with metastatic BCC refractory to SMO inhibitors. Furthermore, clinical responses in this patient's cutaneous tumors did not parallel the responses in the distant site. However, serial imaging after diagnosis of metastatic disease can be critical to monitor for response to therapy.

Topics: Anilides; Antineoplastic Agents; Basal Cell Nevus Syndrome; Carcinoma, Basal Cell; Drug Resistance, Neoplasm; Humans; Lung Neoplasms; Male; Middle Aged; Pyridines; Receptors, G-Protein-Coupled; Skin Neoplasms; Smoothened Receptor; Treatment Failure; Veratrum Alkaloids

Vismodegib (GDC-0449, Genentech, USA), a small molecule inhibitor of the Hedgehog signalling pathway, has potent anti-tumour activity in advanced basal cell carcinoma (BCC). We report a case of a 67-year-old Australian man with metastatic BCC including pulmonary disease with malignant effusion who showed a dramatic complete response to vismodegib but subsequently experienced a recurrence of pulmonary disease, indicative of chemoresistance to vismodegib. This case is the first to illustrate chemoresistance in a patient with metastatic BCC, and demonstrates the need for closely monitoring metastatic BCC patients even after an apparently complete response.

Topics: Aged; Anilides; Antineoplastic Agents; Bone Neoplasms; Carcinoma, Basal Cell; Drug Resistance, Neoplasm; Hedgehog Proteins; Humans; Lung Neoplasms; Lymphatic Metastasis; Male; Pyridines; Ribs; Signal Transduction; Skin Neoplasms

Epidermal growth factor receptor- tyrosine kinase inhibitors (EGFR-TKIs) benefit Non-small cell lung cancer (NSCLC) patients, and an EGFR-TKIi erlotinib, is approved for patients with recurrent NSCLC. However, resistance to erlotinib is a major clinical problem. Earlier we have demonstrated the role of Hedgehog (Hh) signaling in Epithelial-to-Mesenchymal transition (EMT) of NSCLC cells, leading to increased proliferation and invasion. Here, we investigated the role of Hh signaling in erlotinib resistance of TGF-β1-induced NSCLC cells that are reminiscent of EMT cells.. Hh signaling was inhibited by specific siRNA and by GDC-0449, a small molecule antagonist of G protein coupled receptor smoothened in the Hh pathway. Not all NSCLC patients are likely to benefit from EGFR-TKIs and, therefore, cisplatin was used to further demonstrate a role of inhibition of Hh signaling in sensitization of resistant EMT cells. Specific pre- and anti-miRNA preparations were used to study the mechanistic involvement of miRNAs in drug resistance mechanism.. siRNA-mediated inhibition as well as pharmacological inhibition of Hh signaling abrogated resistance of NSCLC cells to erlotinib and cisplatin. It also resulted in re-sensitization of TGF-β1-induced A549 (A549M) cells as well the mesenchymal phenotypic H1299 cells to erlotinib and cisplatin treatment with concomitant up-regulation of cancer stem cell (CSC) markers (Sox2, Nanog and EpCAM) and down-regulation of miR-200 and let-7 family miRNAs. Ectopic up-regulation of miRNAs, especially miR-200b and let-7c, significantly diminished the erlotinib resistance of A549M cells. Inhibition of Hh signaling by GDC-0449 in EMT cells resulted in the attenuation of CSC markers and up-regulation of miR-200b and let-7c, leading to sensitization of EMT cells to drug treatment, thus, confirming a connection between Hh signaling, miRNAs and drug resistance.. We demonstrate that Hh pathway, through EMT-induction, leads to reduced sensitivity to EGFR-TKIs in NSCLCs. Therefore, targeting Hh pathway may lead to the reversal of EMT phenotype and improve the therapeutic efficacy of EGFR-TKIs in NSCLC patients.

Topics: Anilides; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cisplatin; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; ErbB Receptors; Erlotinib Hydrochloride; Gene Knockdown Techniques; Hedgehog Proteins; Humans; Lung Neoplasms; MicroRNAs; Pyridines; Quinazolines; RNA, Small Interfering; Signal Transduction; Transfection

Despite improvements in chemoradiation, local control remains a major clinical problem in locally advanced non-small cell lung cancer. The Hedgehog pathway has been implicated in tumor recurrence by promoting survival of tumorigenic precursors and through effects on tumor-associated stroma. Whether Hedgehog inhibition can affect radiation efficacy in vivo has not been reported.. We evaluated the effects of a targeted Hedgehog inhibitor (HhAntag) and radiation on clonogenic survival of human non-small cell lung cancer lines in vitro. Using an A549 cell line xenograft model, we examined tumor growth, proliferation, apoptosis, and gene expression changes after concomitant HhAntag and radiation. In a transgenic mouse model of Kras(G12D)-induced and Twist1-induced lung adenocarcinoma, we assessed tumor response to radiation and HhAntag by serial micro-computed tomography (CT) scanning.. In 4 human lung cancer lines in vitro, HhAntag showed little or no effect on radiosensitivity. By contrast, in both the human tumor xenograft and murine inducible transgenic models, HhAntag enhanced radiation efficacy and delayed tumor growth. By use of the human xenograft model to differentiate tumor and stromal effects, mouse stromal cells, but not human tumor cells, showed significant and consistent downregulation of Hedgehog pathway gene expression. This was associated with increased tumor cell apoptosis.. Targeted Hedgehog pathway inhibition can increase in vivo radiation efficacy in lung cancer preclinical models. This effect is associated with pathway suppression in tumor-associated stroma. These data support clinical testing of Hedgehog inhibitors as a component of multimodality therapy for locally advanced non-small cell lung cancer.

Topics: Adenocarcinoma; Anilides; Animals; Apoptosis; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Survival; Female; Hedgehog Proteins; Humans; Lung Neoplasms; Mice; Mice, Nude; Mice, Transgenic; Neoplasm Proteins; Pyridines; Radiation Tolerance; Transplantation, Heterologous

The hedgehog (Hh) signaling pathway has been shown to be activated in the cancer stem cells of several tumor entities. The Hh inhibitor GDC-0449 has been proven to be effective in some cancers but not yet in lung cancer. We aimed at investigating whether GDC-0449 is effective in the lung cancer cell lines HCC (adenocarcinoma) and H1339 (small-cell-lung carcinoma), whether in these cell lines stem cell-like side populations (SPs) can be identified, and whether possible effects of GDC-0449 are mediated via SPs. SPs were identified by spectrum shift and decreased fluorescence after staining with 2.5 μg/ml Hoechst 33342. Expression of proteins was quantified by immunofluorescence. GDC-0449 (25 and 50 μM) inhibited concentration-dependent cell growth in HCC and H1339 cells. Further, the inhibitory effects of cisplatin on cell growth were augmented. In HCC and H1339 cell lines, SPs of 0.57 and 0.46% could be identified, respectively. SP, but not non-SP, cells were able to repopulate the original tumor population. The Hh receptor smoothened was detectable in SP but not in non-SP cells, showing the activation of the Hh pathway only in SPs. GDC-0449 considerably reduced SPs in HCC and H1339 cells. We demonstrate for the first time that GDC-0449 effectively reduces cell growth in lung cancer cell lines. This effect is mediated by the inhibition of stem cell-like SPs.

Topics: Anilides; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Benzimidazoles; Cell Line, Tumor; Cisplatin; Flow Cytometry; Fluorescence; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Hedgehog Proteins; Humans; Lung Neoplasms; Pyridines; Signal Transduction; Small Cell Lung Carcinoma; Time Factors

Cisplatin resistance is an important issue in lung cancer. We aimed at investigating if the Hedgehog pathway inhibitor GDC-0449 is effective in cisplatin-resistant cells and if it alters intracellular Ca(2+)-homeostasis.. The cytoplasmatic ([Ca(2+)](cyto)) and endoplasmatic ([Ca(2+)])(ER) Ca(2+) concentration of HCC (adeno carcinoma of the lung) and H1339 (small cell lung carcinoma) cells were measured with the calcium indicator dye Fura-2 AM. The expression of the inositol-1,4,5-trisphosphate receptor (IP(3)R) and sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) were analyzed using western blot analysis.. GDC-0449 inhibited cell growth in cisplatin-naïve and -resistant cells. In both cell types, GDC-0449 increased [Ca(2+)](cyto) and reduced endoplasmatic [Ca(2+)](ER). Cisplatin failed to considerably alter Ca(2+) homeostasis in resistant cells. The effects of GDC-0449 on intracellular Ca(2+) homeostasis were not mediated by an altered expression of IP(3)R or SERCA.. GDC-0449 alters intracellular Ca(2+) homeostasis and inhibits cell growth in cisplatin-resistant lung cancer cells.

Topics: Adenocarcinoma; Anilides; Antineoplastic Agents; Blotting, Western; Calcium; Cell Proliferation; Cisplatin; Cytoplasm; Drug Resistance, Neoplasm; Fura-2; Hedgehog Proteins; Homeostasis; Humans; Inositol 1,4,5-Trisphosphate Receptors; Lung Neoplasms; Pyridines; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Small Cell Lung Carcinoma; Tumor Cells, Cultured

The aim of this study was to assess the activity of hedgehog signaling pathway in malignant pleural mesothelioma (MPM).. The expression of hedgehog signaling components was assessed by quantitative PCR and in situ hybridization in 45 clinical samples. Primary MPM cultures were developed in serum-free condition in 3% oxygen and were used to investigate the effects of smoothened (SMO) inhibitors or GLI1 silencing on cell growth and hedgehog signaling. In vivo effects of SMO antagonists were determined in an MPM xenograft growing in nude mice.. A significant increase in GLI1, sonic hedgehog, and human hedgehog interacting protein gene expression was observed in MPM tumors compared with nontumoral pleural tissue. SMO antagonists inhibited GLI1 expression and cell growth in sensitive primary cultures. This effect was mimicked by GLI1 silencing. Reduced survivin and YAP protein levels were also observed. Survivin protein levels were rescued by overexpression of GLI1 or constitutively active YAP1. Treatment of tumor-bearing mice with the SMO inhibitor HhAntag led to a significant inhibition of tumor growth in vivo accompanied by decreased Ki-67 and nuclear YAP immunostaining and a significant difference in selected gene expression profile in tumors.. An aberrant hedgehog signaling is present in MPM, and inhibition of hedgehog signaling decreases tumor growth indicating potential new therapeutic approach.

Topics: Adaptor Proteins, Signal Transducing; Adult; Aged; Anilides; Animals; Cell Line, Tumor; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Humans; Inhibitor of Apoptosis Proteins; Lung Neoplasms; Male; Mesothelioma; Mice; Middle Aged; NIH 3T3 Cells; Phosphoproteins; Pleural Effusion, Malignant; Pyridines; Receptors, G-Protein-Coupled; RNA, Small Interfering; Signal Transduction; Smoothened Receptor; Survivin; Tomatine; Transcription Factors; Transplantation, Heterologous; Veratrum Alkaloids; YAP-Signaling Proteins; Zinc Finger Protein GLI1

HhAntag691 (GDC-0449), a low-molecular weight inhibitor of the tumor-promoting hedgehog (Hh) signaling pathway, has been used to treat medulloblastoma in animal models and has recently entered clinical trials for a variety of solid tumors. Here, we show that HhAntag691 inhibits multiple ATP-binding cassette (ABC) transporters. ATP-binding cassette transporters are within a family of membrane proteins, the overexpression of which is associated with multidrug resistance, a major impediment to successful cancer treatment. HhAntag691 is a potent inhibitor of two ABC transporters, ABCG2/BCRP and ABCB1/Pgp, and is a mild inhibitor of ABCC1/MRP1. In ABCG2-overexpressing HEK293 cells, HhAntag691 increased retention of the fluorescent ABCG2 substrate BODIPY-prazosin and resensitized these cells to mitoxantrone, an antineoplastic ABCG2 substrate. In Madin-Darby canine kidney II cells engineered to overexpress Pgp or MRP1, HhAntag691 increased the retention of calcein-AM and resensitized them to colchicine. HhAntag691 also resensitized human non-small cell lung carcinoma cells NCI-H460/par and NCI-H460/MX20, which overexpress ABCG2 in response to mitoxantrone, to mitoxantrone, and to topotecan or SN-38. The IC(50) values of HhAntag691 for inhibition of ABCG2 and Pgp were approximately 1.4 and approximately 3.0 microM, respectively. Because ABC transporters are highly expressed at the blood-brain barrier and on many tumor cells, they contribute significantly to treatment failure of many types of cancer, particularly of those within the neuraxis. In addition to its effect on Hh signaling, the ability of HhAntag691 and related compounds to inhibit two key ABC transporters could contribute to their effectiveness in treating malignancies.

Topics: Anilides; Animals; Antineoplastic Agents; ATP Binding Cassette Transporter, Subfamily B; ATP Binding Cassette Transporter, Subfamily B, Member 1; ATP Binding Cassette Transporter, Subfamily G, Member 2; ATP-Binding Cassette Transporters; Benzimidazoles; Carcinoma, Non-Small-Cell Lung; Cells, Cultured; Colchicine; Dogs; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Drug Resistance, Neoplasm; Hedgehog Proteins; Humans; Lung Neoplasms; Multidrug Resistance-Associated Proteins; Neoplasm Proteins; Pyridines; Signal Transduction