Characterization and response of newly developed high-grade glioma cultures to the tyrosine kinase inhibitors, erlotinib, gefitinib and imatinib.

High-grade gliomas (HGG), are the most common aggressive brain tumours in adults. Inhibitors targeting growth factor signalling pathways in glioma have shown a low clinical response rate. To accurately evaluate response to targeted therapies further in vitro studies are necessary. Growth factor pathway expression using epidermal growth factor receptor (EGFR), mutant EGFR (EGFRvIII), platelet derived growth factor receptor (PDGFR), C-Kit and C-Abl together with phosphatase and tensin homolog (PTEN) expression and downstream activation of AKT and phosphorylated ribosomal protein S6 (P70S6K) was analysed in 26 primary glioma cultures treated with the tyrosine kinase inhibitors (TKIs) erlotinib, gefitinib and imatinib. Response to TKIs was assessed using 50% inhibitory concentrations (IC(50)). Response for each culture was compared with the EGFR/PDGFR immunocytochemical pathway profile using hierarchical cluster analysis (HCA) and principal component analysis (PCA). Erlotinib response was not strongly associated with high expression of the growth factor pathway components. PTEN expression did not correlate with response to any of the three TKIs. Increased EGFR expression was associated with gefitinib response; increased PDGFR-α expression was associated with imatinib response. The results of this in vitro study suggest gefitinib and imatinib may have therapeutic potential in HGG tumours with a corresponding growth factor receptor expression profile.

The effect of tricyclic antidepressants on cutaneous melanoma cell lines and primary cell cultures.

The tricyclic antidepressants have previously been shown to exert activity against glioma cells in vitro. Initial studies in cell lines suggested that this might extend to melanoma cells. We have therefore conducted a study in primary cell cultures from metastatic cutaneous melanoma deposits using a well established ATP-based tumour chemosensitivity assay to confirm and extend these findings. Two cell lines and eight primary cell cultures from metastatic melanoma deposits were exposed to three tricyclic drugs, amitriptyline, nortriptyline and clomipramine, at concentrations ranging from 200 to 6.25 µmol/l in the ATP-based tumour chemosensitivity assay. All three drugs showed activity, although nortriptyline was more active than clomipramine or amitriptyline in both cell lines and primary cell cultures, with an IC50 of 9, 27 and 33 µmol/l, respectively. Tricyclic agents show activity against melanoma in vitro. This could be related to the lysosomal effects based on their cationic amphiphilic properties, or effects at the mitochondrial membrane.

Imatinib and docetaxel in combination can effectively inhibit glioma invasion in an in vitro 3D invasion assay.

The main problem in the treatment of malignant astrocytomas is their invasive behaviour. Successful resection of the main tumour mass cannot prevent recurrence due to single cells invading the surrounding brain parenchyma at the time of diagnosis. The classical combination therapy, PCV (Procarbazine, CCNU and Vincristine) used for over 30 years; has shown its clinical effectiveness in the treatment of malignant astrocytomas and glioblastomas is still doubtful. Using an in vitro three dimensional invasion model, we tested the effect of the tyrosine kinase inhibitor imatinib and the microtubule inhibitor docetaxel on the invasion activity of a panel of astrocytic tumour cell lines, including two established glioma cell lines, IPSB-18 and SNB-19, and two primary cell lines, originating from glioblastomas, CLOM002 and UPHHJA, and in normal astrocytes. A dose response curve for each drug alone and in combination was determined. The half maximal inhibitory concentration (IC(50)) concentration of imatinib was between 15.7 and 18.7 µM, which did not affect invasion activity of the cell lines. The IC(50) concentration of docetaxel was between 0.7 and 19.8 nM, and at 14.9 nM docetaxel had a slight transient inhibitory effect on invasion activity of all tested cells. The combination of imatinib at 13.5 µM and docetaxel at 14.9 nM, however, synergistically inhibited cell growth and invasion activity and could not be reversed by drug removal. A combination treatment with tyrosine kinase inhibitors and cytotoxic drugs shows promise in tackling both glioma proliferation and invasion, and could present a new treatment regimen for malignant astrocytomas.

Prognostic value of the 6-gene OncoMasTR test in hormone receptor-positive HER2-negative early-stage breast cancer: Comparative analysis with standard clinicopathological factors.

AIM: The aim of the study was to assess the prognostic performance of a 6-gene molecular score (OncoMasTR Molecular Score [OMm]) and a composite risk score (OncoMasTR Risk Score [OM]) and to conduct a within-patient comparison against four routinely used molecular and clinicopathological risk assessment tools: Oncotype DX Recurrence Score, Ki67, Nottingham Prognostic Index and Clinical Risk Category, based on the modified Adjuvant! Online definition and three risk factors: patient age, tumour size and grade. METHODS: Biospecimens and clinicopathological information for 404 Irish women also previously enrolled in the Trial Assigning Individualized Options for Treatment [Rx] were provided by 11 participating hospitals, as the primary objective of an independent translational study. Gene expression measured via RT-qPCR was used to calculate OMm and OM. The prognostic value for distant recurrence-free survival (DRFS) and invasive disease-free survival (IDFS) was assessed using Cox proportional hazards models and Kaplan-Meier analysis. All statistical tests were two-sided ones. RESULTS: OMm and OM (both with likelihood ratio statistic [LRS] P < 0.001; C indexes = 0.84 and 0.85, respectively) were more prognostic for DRFS and provided significant additional prognostic information to all other assessment tools/factors assessed (all LRS P ≤ 0.002). In addition, the OM correctly classified more patients with distant recurrences (DRs) into the high-risk category than other risk classification tools. Similar results were observed for IDFS. DISCUSSION: Both OncoMasTR scores were significantly prognostic for DRFS and IDFS and provided additional prognostic information to the molecular and clinicopathological risk factors/tools assessed. OM was also the most accurate risk classification tool for identifying DR. A concise 6-gene signature with superior risk stratification was shown to increase prognosis reliability, which may help clinicians optimise treatment decisions.

Investigation of the role of p53 in chemotherapy resistance of lung cancer cell lines.

BACKGROUND: p53 is a tumour suppressor gene, which is mutated in more than half of all tumours. Most chemotherapeutic drugs cause DNA damage, which is sensed by p53; the cell can then try to repair the damage or induce cell suicide. If the p53 machinery is defective, effective chemotherapy is made more difficult. MATERIALS AND METHODS: Wild-type p53 was transfected into lung cancer cell lines with different p53 status. The transfected cells were tested for changes in sensitivity to a range of chemotherapeutic agents. RESULTS: We observed only modest changes in the sensitivity to the chemotherapeutic agents adriamycin, taxol and carboplatin in the transfected cells lines. p53 protein was detected in a transfected clone of the cell line H1299, whose parent cells are p53 null. However, the protein did not accumulate after DNA damage, suggesting that this cell line utilises alternative pathways for responding to stress, and no longer has a functional p53 pathway. CONCLUSION: The results suggest that introduction of wild-type p53 alone is not sufficient to substantially alter the sensitivity of a cell line to a given chemotherapeutic agent.

Hypoxia helps glioma to fight therapy.

Despite major improvements in the surgical management the prognosis for patients bearing malignant gliomas is still dismal. Malignant gliomas are notoriously resistant to treatment and the survival time of patients is between 3-8 years for low-grade and anaplastic gliomas and 6 - 12 month for glioblastoma. Increasing malignancy of gliomas correlates with an increase in cellularity and a poorly organized tumor vasculature leading to insufficient blood supply, hypoxic areas and ultimately to the formation of necrosis, a characteristic of glioblastoma. Hypoxic/necrotic tumors are more resistant to chemotherapy and radiation. Hypoxia induces either directly or indirectly (through the activation of transcription factors) changes in the biology of a tumor and its microenvironment leading to increased aggressiveness and tumor resistance to chemotherapy and radiation. This review is focused on hypoxia-induced molecular changes affecting glioma biology and therapy.