pyrimidinones and Glioblastoma

Epithelioid glioblastoma multiforme (eGBM) is a rare and aggressive variant of glioblastoma multiforme (GBM) that predominantly affects younger patients and can be difficult to distinguish from other gliomas. Data on how patients with eGBM might be best treated are limited, although genomic analyses have shown that almost half of tumours harbour activating BRAF gene mutations. Here we present the case of a young female with BRAF V600E-mutant eGBM who had a prolonged response to targeted therapy with the BRAF and MEK1/2 inhibitors dabrafenib and trametinib. We review current knowledge about eGBM, including the emerging role for BRAF- ± MEK1/2- targeted therapy.

Topics: Antineoplastic Agents; Brain Neoplasms; Fatal Outcome; Female; Glioblastoma; Humans; Imidazoles; MAP Kinase Kinase 1; MAP Kinase Kinase 2; Oximes; Proto-Oncogene Proteins B-raf; Pyridones; Pyrimidinones; Spinal Cord Neoplasms; Young Adult

A rapid, sensitive, and robust aqueous normal-phase chromatography method coupled with tandem mass spectrometry was developed and validated for the quantitation of AZD1775, a Wee-1 inhibitor, in human plasma and brain tumor tissue. Sample preparation involved simple protein precipitation with acetonitrile. Chromatographic separation was achieved on ethylene bridged hybrid stationary phases (i.e., Waters XBridge Amide column) under an isocratic elution with the mobile phase consisting of acetonitrile/ammonium formate in water (10mM, pH 3.0) (85:15,v/v) at a flow rate of 0.8mL/min for 5min. The lower limit of quantitation (LLOQ) was 0.2ng/mL of AZD1775 in plasma and tissue homogenate. The calibration curve was linear over AZD1775 concentration range of 0.2-1000ng/mL in plasma and tissue homogenate. The intra- and inter-day precision and accuracy were within the generally accepted criteria for bioanalytical method (<15%). The method was successfully applied to assess the penetration of AZD1775 across the blood-brain tumor barrier, as assessed by the unbound brain-to-plasma ratio, in patients with glioblastoma.

Topics: Brain; Brain Neoplasms; Cell Cycle Proteins; Chromatography, High Pressure Liquid; Glioblastoma; Humans; Limit of Detection; Nuclear Proteins; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Tandem Mass Spectrometry

Glioblastoma (GB) is a highly aggressive, difficult to treat brain tumour. Successful treatment, consisting of maximal safe tumour de-bulking, followed by radiotherapy and treatment with the alkylating agent Temozolomide (TMZ), can extend patient survival to approximately 15 months. Combination treatments based on the inhibition of the PI3K pathway, which is the most frequently activated signalling cascade in GB, have so far only shown limited therapeutic success. Here, we use the clinically approved MEK inhibitor Trametinib to investigate its potential use in managing GB. Trametinib has a strong anti-proliferative effect on established GB cell lines, stem cell-like cells and their differentiated progeny and while it does not enhance anti-proliferative and cell death-inducing properties of the standard treatment, i.e. exposure to radiation or TMZ, neither does MEK inhibition block their effectiveness. However, upon MEK inhibition some cell populations appear to favour cell-substrate interactions in a sprouting assay and become more invasive in the Chorioallantoic Membrane assay, which assesses cell penetration into an organic membrane. While this increased invasion can be modulated by additional inhibition of the PI3K signalling cascade, there is no apparent benefit of blocking MEK compared to targeting PI3K.

Topics: Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Brain Neoplasms; Cell Adhesion; Cell Death; Cell Line, Tumor; Cell Proliferation; Chorioallantoic Membrane; Drug Screening Assays, Antitumor; Glioblastoma; Humans; MAP Kinase Kinase 1; Neoplasm Invasiveness; Neoplasm Metastasis; Phosphatidylinositol 3-Kinases; Phosphorylation; Pyridones; Pyrimidinones; Signal Transduction; Temozolomide; Translational Research, Biomedical

Resistance is an obstacle of glioma therapy. Despite targeted interventions, tumors harbor primary resistance or become resistant over short course of treatment. This study examined the mouse double minute 2 (MDM2) inhibitor RG7388 together with radiotherapy and analyzed strategies to overcome acquired MDM2 inhibitor resistance in glioblastoma.. Effects of RG7388 and radiotherapy were analyzed in p53 wild-type glioblastoma cell lines and glioma-initiating cells. RG7388 resistant cells were generated by increasing RG7388 doses over 3 months. Regulated pathways were investigated by microarray, qRT-PCR, and immunoblot analysis and specifically inhibited to evaluate rational salvage therapies at RG7388 resistance. Effects of RG7388 and trametinib treatment were challenged in an orthotopical mouse model with RG7388 resistant U87MG glioblastoma cells.. MDM2 inhibition required functional p53 and showed synergistic activity with radiotherapy in first-line treatment. Long-term exposure to RG7388 induced resistance by activation of the extracellular signal-regulated kinases 1/2 (ERK1/2)-insulin growth factor binding protein 1 (IGFBP1) signaling cascade, which was specifically overcome by ERK1/2 pathway inhibition with trametinib and knockdown of IGFBP1. Combining trametinib with continued RG7388 treatment enhanced antitumor effects at RG7388 resistance. These data provide a rationale for combining RG7388 and radiotherapy as first-line therapy with a specific relevance for tumors insensitive to alkylating standard chemotherapy and for the addition of trametinib to continued RG7388 treatment as salvage therapy after acquired resistance against RG7388 for clinical practice.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Drug Resistance, Neoplasm; Glioblastoma; Heterografts; Humans; Insulin-Like Growth Factor Binding Protein 1; Mice; para-Aminobenzoates; Proto-Oncogene Proteins c-mdm2; Pyridones; Pyrimidinones; Pyrrolidines; Signal Transduction; Tumor Suppressor Protein p53

Glioblastoma is the most common type of primary brain tumour in adults, and its pathogenesis is particularly complicated. Among the many possible mechanisms underlying its pathogenesis, hyperactivation of the PI3K/Akt pathway is essential to the occurrence and development of glioma through the loss of PTEN or somatic activating mutations in PIK3CA. In the present study, we investigated the effect of the PI3Kβ inhibitor AZD6482 on glioma cells. The CCK-8 assay showed dose-dependent cytotoxicity in glioma cell lines treated with AZD6482. Additionally, AZD6482 treatment was found to significantly induce apoptosis and cell cycle arrest as detected using flow cytometry. Moreover, as shown using western blot analysis, the levels of p-AKT, p-GSK-3β, Bcl-2, and cyclin D1 were decreased after AZD6482 treatment. In addition, we found that AZD6482 inhibited the migration and invasion of glioma cells as detected by wound healing and Transwell invasion assays. Taken together, our findings indicate that AZD6482 exerts an antitumour effect by inhibiting proliferation and inducing apoptosis in human glioma cells. AZD6482 may be applied as an adjuvant therapy to improve the therapeutic efficacy of glioblastoma treatment.

Topics: Apoptosis; Brain Neoplasms; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Movement; Cell Proliferation; Class I Phosphatidylinositol 3-Kinases; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Glioblastoma; Humans; Mutation; ortho-Aminobenzoates; Protein Kinase Inhibitors; PTEN Phosphohydrolase; Pyrimidinones; Signal Transduction

Hyperactivation of the RAS-RAF-MEK-ERK signaling pathway is exploited by glioma cells to promote their growth and evade apoptosis. MEK activation in tumor cells can increase replication of ICP34.5-deleted herpes simplex virus type 1 (HSV-1), but paradoxically its activation in tumor-associated macrophages promotes a pro-inflammatory signaling that can inhibit virus replication and propagation. Here we investigated the effect of blocking MEK signaling in conjunction with oncolytic HSV-1 (oHSV) for brain tumors.. Infected glioma cells co-cultured with microglia or macrophages treated with or without trametinib were used to test trametinib effect on macrophages/microglia. Enzyme-linked immunosorbent assay, western blotting, and flow cytometry were utilized to evaluate the effect of the combination therapy. Pharmacokinetic (PK) analysis of mouse plasma and brain tissue was used to evaluate trametinib delivery to the CNS. Intracranial human and mouse glioma-bearing immune deficient and immune competent mice were used to evaluate the antitumor efficacy.. Oncolytic HSV treatment rescued trametinib-mediated feedback reactivation of the mitogen-activated protein kinase signaling pathway in glioma. In vivo, PK analysis revealed enhanced blood-brain barrier penetration of trametinib after oHSV treatment. Treatment by trametinib, a MEK kinase inhibitor, led to a significant reduction in microglia- and macrophage-derived tumor necrosis factor alpha (TNFα) secretion in response to oHSV treatment and increased survival of glioma-bearing mice. Despite the reduced TNFα production observed in vivo, the combination treatment activated CD8+ T-cell mediated immunity and increased survival in a glioma-bearing immune-competent mouse model.. This study provides a rationale for combining oHSV with trametinib for the treatment of brain tumors.

Topics: Animals; Blood-Brain Barrier; Brain Neoplasms; CD8-Positive T-Lymphocytes; Cell Line, Tumor; Disease Models, Animal; Glioblastoma; Glioma; Herpesvirus 1, Human; Humans; Immunocompetence; Macrophages; Mice; Microglia; Mitogen-Activated Protein Kinase Kinases; Oncolytic Virotherapy; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; RAW 264.7 Cells; Survival Rate; Tumor Necrosis Factor-alpha; Xenograft Model Antitumor Assays

The strategy of using biologically targeted therapeutics for cancer has yet to translate into effective treatment of gliomas. The neuro-oncology community is beginning to recognize that phase 0 studies should be performed to account for the impact of the blood-brain barrier on the ability of a therapeutic to reach its target(s).

Topics: Brain Neoplasms; Glioblastoma; Humans; Pyrazoles; Pyrimidines; Pyrimidinones

It is generally known that glioblastoma is the most common primary malignant brain tumor and that it is highly aggressive and deadly. Although surgical and pharmacological therapies have made long‑term progress, glioblastoma remains extremely lethal and has an uncommonly low survival rate. Therefore, further elucidation of the molecular mechanisms of glioblastoma initiation and its pathological processes are urgent. Arsenic resistance protein 2 (Ars2) is a highly conserved gene, and it has been found to play an important role in microRNA biosynthesis and cell proliferation in recent years. Furthermore, absence of Ars2 results in developmental death in Drosophila, zebrafish and mice. However, there are few studies on the role of Ars2 in regulating tumor development, and the mechanism of its action is mostly unknown. In the present study, we revealed that Ars2 is involved in glioblastoma proliferation and we identified a potential mechanistic role for it in cell cycle control. Our data demonstrated that Ars2 knockdown significantly repressed the proliferation and tumorigenesis abilities of glioblastoma cells in vitro and in vivo. Further investigation clarified that Ars2 deficiency inhibited the activation of the MAPK/ERK pathway, leading to cell cycle arrest in the G1 phase, resulting in suppression of cell proliferation. These findings support the conclusion that Ars2 is a key regulator of glioblastoma progression.

Topics: Animals; Brain Neoplasms; Carcinogenesis; Cell Line, Tumor; Cell Proliferation; Disease Progression; Female; G1 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Glioblastoma; Humans; MAP Kinase Signaling System; Mice; Mice, Inbred NOD; Mice, SCID; Nuclear Proteins; Phosphorylation; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; Xenograft Model Antitumor Assays

Determining the penetration of drugs across the blood-brain barrier is a significant challenge in central nervous system drug development. The use of a mechanistic physiologically based pharmacokinetic model can predict drug exposures in the brain without needing

Topics: Adult; Blood-Brain Barrier; Glioblastoma; Humans; Pyrazoles; Pyrimidines; Pyrimidinones

Glioblastoma is the most common type of primary brain tumor in adults, with high mortality and morbidity rates. More effective therapeutic strategies are imperative. Previous studies have shown that the known p110-β-selective inhibitor TGX-221 blocks the activation of PKB/Akt in PTEN-deficient cells. We treated U87 and U251 glioblastoma cells with TGX-221 to determine the effect of TGX-221. We performed a Cell Counting Kit-8 (CCK-8) test, EDU staining and cell cycle distribution analysis and found that TGX-221 inhibited glioblastoma cell proliferation. Next, the effect of TGX-221 on cell apoptosis was investigated using flow cytometry. These results demonstrated that TGX-221 induced apoptosis in glioblastoma cells. Moreover, migration and invasion assays revealed that TGX-221 inhibited human glioblastoma cell migration and invasion. Collectively, our study revealed that TGX-221 could inhibit proliferation and induce apoptosis in glioblastoma cells.

Topics: Brain Neoplasms; Cell Cycle Proteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Class I Phosphatidylinositol 3-Kinases; Gene Expression Regulation, Neoplastic; Glioblastoma; Humans; Morpholines; Proto-Oncogene Proteins c-akt; Pyrimidinones; Signal Transduction

Glioblastoma (GBM) is the most common malignant primary brain cancer. With a median survival of about a year, new approaches to treating this disease are necessary. To identify signaling molecules regulating GBM progression in a genetically engineered murine model of proneural GBM, we quantified phosphotyrosine-mediated signaling using mass spectrometry. Oncogenic signals, including phosphorylated ERK MAPK, PI3K, and PDGFR, were found to be increased in the murine tumors relative to brain. Phosphorylation of CDK1 pY15, associated with the G2 arrest checkpoint, was identified as the most differentially phosphorylated site, with a 14-fold increase in phosphorylation in the tumors. To assess the role of this checkpoint as a potential therapeutic target, syngeneic primary cell lines derived from these tumors were treated with MK-1775, an inhibitor of Wee1, the kinase responsible for CDK1 Y15 phosphorylation. MK-1775 treatment led to mitotic catastrophe, as defined by increased DNA damage and cell death by apoptosis. To assess the extensibility of targeting Wee1/CDK1 in GBM, patient-derived xenograft (PDX) cell lines were also treated with MK-1775. Although the response was more heterogeneous, on-target Wee1 inhibition led to decreased CDK1 Y15 phosphorylation and increased DNA damage and apoptosis in each line. These results were also validated in vivo, where single-agent MK-1775 demonstrated an antitumor effect on a flank PDX tumor model, increasing mouse survival by 1.74-fold. This study highlights the ability of unbiased quantitative phosphoproteomics to reveal therapeutic targets in tumor models, and the potential for Wee1 inhibition as a treatment approach in preclinical models of GBM. Mol Cancer Ther; 15(6); 1332-43. ©2016 AACR.

Topics: Animals; Biomarkers, Tumor; Brain Neoplasms; CDC2 Protein Kinase; Cell Cycle Proteins; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Glioblastoma; Humans; Mass Spectrometry; Mice; Molecular Targeted Therapy; Nuclear Proteins; Phosphorylation; Protein-Tyrosine Kinases; Proteomics; Pyrazoles; Pyrimidines; Pyrimidinones; Xenograft Model Antitumor Assays

Wee1 regulates key DNA damage checkpoints, and in this study, the efficacy of the Wee1 inhibitor MK-1775 was evaluated in glioblastoma multiforme (GBM) xenograft models alone and in combination with radiation and/or temozolomide.. In vitro MK-1775 efficacy alone and in combination with temozolomide, and the impact on DNA damage, was analyzed by Western blotting and γH2AX foci formation. In vivo efficacy was evaluated in orthotopic and heterotopic xenografts. Drug distribution was assessed by conventional mass spectrometry (MS) and matrix-assisted laser desorption/ionization (MALDI)-MS imaging.. GBM22 (IC50 = 68 nmol/L) was significantly more sensitive to MK-1775 compared with five other GBM xenograft lines, including GBM6 (IC50 >300 nmol/L), and this was associated with a significant difference in pan-nuclear γH2AX staining between treated GBM22 (81% cells positive) and GBM6 (20% cells positive) cells. However, there was no sensitizing effect of MK-1775 when combined with temozolomide in vitro. In an orthotopic GBM22 model, MK-1775 was ineffective when combined with temozolomide, whereas in a flank model of GBM22, MK-1775 exhibited both single-agent and combinatorial activity with temozolomide. Consistent with limited drug delivery into orthotopic tumors, the normal brain to whole blood ratio following a single MK-1775 dose was 5%, and MALDI-MS imaging demonstrated heterogeneous and markedly lower MK-1775 distribution in orthotopic as compared with heterotopic GBM22 tumors.. Limited distribution to brain tumors may limit the efficacy of MK-1775 in GBM.

Topics: Animals; Blood-Brain Barrier; Cell Cycle Proteins; Dacarbazine; Disease Models, Animal; DNA Damage; Glioblastoma; Humans; Mice; Nuclear Proteins; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Temozolomide; Tumor Burden; Xenograft Model Antitumor Assays

To identify therapeutic targets for glioblastoma (GBM), we performed genome-wide CRISPR-Cas9 knockout (KO) screens in patient-derived GBM stem-like cells (GSCs) and human neural stem/progenitors (NSCs), non-neoplastic stem cell controls, for genes required for their in vitro growth. Surprisingly, the vast majority GSC-lethal hits were found outside of molecular networks commonly altered in GBM and GSCs (e.g., oncogenic drivers). In vitro and in vivo validation of GSC-specific targets revealed several strong hits, including the wee1-like kinase, PKMYT1/Myt1. Mechanistic studies demonstrated that PKMYT1 acts redundantly with WEE1 to inhibit cyclin B-CDK1 activity via CDK1-Y15 phosphorylation and to promote timely completion of mitosis in NSCs. However, in GSCs, this redundancy is lost, most likely as a result of oncogenic signaling, causing GBM-specific lethality.

Topics: CDC2 Protein Kinase; Cell Cycle Proteins; Cell Survival; CRISPR-Cas Systems; Cyclin B; ErbB Receptors; Gene Library; Genome, Human; Glioblastoma; Humans; Membrane Proteins; Microscopy, Video; Mitosis; Neoplastic Stem Cells; Nuclear Proteins; Phosphorylation; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-akt; Pyrazoles; Pyrimidines; Pyrimidinones; RNA Interference; Time-Lapse Imaging; Tumor Suppressor Protein p53

We investigated the efficacy of the Wee1 inhibitor MK-1775 in combination with radiation for the treatment of pediatric high-grade gliomas (HGGs), including diffuse intrinsic pontine gliomas (DIPGs).. Gene expression analysis was performed for 38 primary pediatric gliomas (3 grade I, 10 grade II, 11 grade III, 14 grade IV) and 8 normal brain samples using the Agilent 4 × 44 K array. Clonogenic survival assays were carried out in pediatric and adult HGG cell lines (n = 6) to assess radiosensitizing effects of MK-1775. DNA repair capacity was evaluated by measuring protein levels of γ-H2AX, a marker of double strand DNA breaks. In vivo activity of MK-1775 with radiation was assessed in 2 distinct orthotopic engraftment models of pediatric HGG, including 1 derived from a genetically engineered mouse carrying a BRAF(V600E) mutation, and 1 xenograft model in which tumor cells were derived from a patient's DIPG.. Wee1 is overexpressed in pediatric HGGs, with increasing expression positively correlated with malignancy (P = .007 for grade III + IV vs I + II) and markedly high expression in DIPG. Combination treatment of MK-1775 and radiation reduced clonogenic survival and increased expression of γ-H2AX to a greater extent than achieved by radiation alone. Finally, combined MK-1775 and radiation conferred greater survival benefit to mice bearing engrafted, orthotopic HGG and DIPG tumors, compared with treatment with radiation alone (BRAF(V600E) model P = .0061 and DIPG brainstem model P = .0163).. Our results highlight MK-1775 as a promising new therapeutic agent for use in combination with radiation for the treatment of pediatric HGGs, including DIPG.

Topics: Animals; Brain Neoplasms; Cell Cycle Proteins; Cell Death; Cell Line, Tumor; Chemoradiotherapy; Glioblastoma; Humans; Mice; Mice, Transgenic; Nuclear Proteins; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones

Topics: Animals; Brain Neoplasms; Cell Cycle Proteins; Glioblastoma; Humans; Nuclear Proteins; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones

Although diacylglycerol kinase α (DGKα) has been linked to several signaling pathways related to cancer cell biology, it has been neglected as a target for cancer therapy. The attenuation of DGKα activity via DGKα-targeting siRNA and small-molecule inhibitors R59022 and R59949 induced caspase-mediated apoptosis in glioblastoma cells and in other cancers, but lacked toxicity in noncancerous cells. We determined that mTOR and hypoxia-inducible factor-1α (HIF-1α) are key targets of DGKα inhibition, in addition to its regulation of other oncogenes. DGKα regulates mTOR transcription via a unique pathway involving cyclic AMP. Finally, we showed the efficacy of DGKα inhibition with short hairpin RNA or a small-molecule agent in glioblastoma and melanoma xenograft treatment models, with growth delay and decreased vascularity. This study establishes DGKα as a central signaling hub and a promising therapeutic target in the treatment of cancer.

Topics: Apoptosis; Brain Neoplasms; Cell Line, Tumor; Diacylglycerol Kinase; Gene Expression Regulation, Neoplastic; Glioblastoma; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Molecular Targeted Therapy; Piperidines; Pyrimidinones; Quinazolinones; RNA, Small Interfering; Thiazoles

The purpose of this study was to determine the capacity of MK-1775, a potent Wee-1 inhibitor, to abrogate the radiation-induced G(2) checkpoint arrest and modulate radiosensitivity in glioblastoma cell models and normal human astrocytes. The radiation-induced checkpoint response of established glioblastoma cell lines, glioblastoma neural stem (GNS) cells, and astrocytes were determined in vitro by flow cytometry and in vivo by mitosis-specific staining using immunohistochemistry. Mechanisms underlying MK-1775 radiosensitization were determined by mitotic catastrophe and γH2AX expression. Radiosensitivity was determined in vitro by the clonogenic assay and in vivo by tumor growth delay. MK-1775 abrogated the radiation-induced G(2) checkpoint and enhanced radiosensitivity in established glioblastoma cell lines in vitro and in vivo, without modulating radiation response in normal human astrocytes. MK-1775 appeared to attenuate the early-phase of the G(2) checkpoint arrest in GNS cell lines, although the arrest was not sustained and did not lead to increased radiosensitivity. These results show that MK-1775 can selectively enhance radiosensitivity in established glioblastoma cell lines. Further work is required to determine the role Wee-1 plays in checkpoint activation of GNS cells.

Topics: Animals; Brain Neoplasms; Cell Cycle Proteins; Cell Line, Tumor; G2 Phase Cell Cycle Checkpoints; Glioblastoma; Humans; Mice; Mice, Nude; Molecular Targeted Therapy; Nuclear Proteins; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Radiation Tolerance; Radiation-Sensitizing Agents; Up-Regulation; Xenograft Model Antitumor Assays

Photodynamic action of merocyanine 540 (MC540) on the plasma membrane of human glioblastoma(U-87MG) cells has been investigated. Plasma membrane was labeled with lipid specific probe 1,(4-trimethylammonium),6-diphenyl-1,3,5-hexatriene. Steady-state anisotropy, decay time and time-dependent anisotropy of TMA-DPH in U-87MG cells have been measured as a function of light dose. A decrease in the steady-state anisotropy and decay time of TMA-DPH in MC540-treated cells was observed upon light irradiation. The time-dependent anisotropy measurements showed a decrease in the limiting anisotropy (r infinity) and an increase in the rotational relaxation time (phi) of the probe upon photosensitization of cells. Analysis of these data using wobbling in cone model for probe rotation in the membrane indicated an increase in the cone angle (theta c) and a decrease in the order parameter (S). Protein specific probe N-(1-pyrene)-maleimide was used to study the effect of photosensitization on the plasma membrane proteins. An increase in the rotational relaxation time and a decrease in the ratio of excimer to monomer fluorescence intensity of PM was observed on photosensitization. Photodynamic action of MC540 also caused an inhibition of protein SH groups and Na(+)-K(+)-ATPase activity of plasma membrane. Our results demonstrate that the photodynamic action of MC540 decreases the order of the lipid bilayer and reduces the mobility of the proteins in the plasma membrane of cells.

Topics: Cell Membrane; Diphenylhexatriene; Dithionitrobenzoic Acid; Fluorescence Polarization; Fluorescent Dyes; Glioblastoma; Humans; Light; Maleimides; Photosensitizing Agents; Pyrimidinones; Sodium-Potassium-Exchanging ATPase; Sulfhydryl Reagents; Tumor Cells, Cultured

Mechanism of merocyanine 540 (MC540) mediated photosensitization in glioblastoma (U-87MG) and neuroblastoma (Neuro 2a) cells was investigated. Photoinduced lipid peroxidation was measured in the presence of mechanistic probes-deuterium oxide (D2O), sodium azide, superoxide dismutase (SOD), mannitol and sodium benzoate. In both the types of cells, the photoinduced lipid peroxidation was enhanced in D2O whereas it showed inhibition in the presence of sodium azide. SOD also inhibited the lipid peroxidation while sodium benzoate and mannitol had no effect. These results suggest that photosensitization of U-87MG and Neuro 2a cells by MC 540 involves both type I (free radical mediated) and type II (singlet oxygen mediated) mechanisms.

Topics: Animals; Drug Screening Assays, Antitumor; Glioblastoma; Humans; Lipid Peroxidation; Mice; Neuroblastoma; Photosensitizing Agents; Pyrimidinones; Tumor Cells, Cultured

Topics: Aminohydrolases; Astrocytoma; Central Nervous System; Glioblastoma; Humans; Meningeal Neoplasms; Meningioma; Neoplasms; Phosphates; Pyrimidinones