Midazolam suppresses interleukin-1β-induced interleukin-6 release from rat glial cells

<p>Abstract</p> <p>Background</p> <p>Peripheral-type benzodiazepine receptor (PBR) expression levels are low in normal human brain, but their levels increase in inflammation, brain injury, neurodegenerative states and gliomas. It has been reported that PBR functions as an immunomodulator. The mechanisms of action of midazolam, a benzodiazepine, in the immune system in the CNS remain to be fully elucidated. We previously reported that interleukin (IL)-1β stimulates IL-6 synthesis from rat C6 glioma cells and that IL-1β induces phosphorylation of inhibitory kappa B (IκB), p38 mitogen-activated protein (MAP) kinase, stress-activated protein kinase (SAPK)/c-<it>Jun </it>N-terminal kinase (JNK), extracellular signal-regulated kinase 1/2, and signal transducer and activator of transcription (STAT)3. It has been shown that p38 MAP kinase is involved in IL-1β-induced IL-6 release from these cells. In the present study, we investigated the effect of midazolam on IL-1β-induced IL-6 release from C6 cells, and the mechanisms of this effect.</p> <p>Methods</p> <p>Cultured C6 cells were stimulated by IL-1β. IL-6 release from C6 cells was measured using an enzyme-linked immunosorbent assay, and phosphorylation of IκB, the MAP kinase superfamily, and STAT3 was analyzed by Western blotting.</p> <p>Results</p> <p>Midazolam, but not propofol, inhibited IL-1β-stimulated IL-6 release from C6 cells. The IL-1β-stimulated levels of IL-6 were suppressed by wedelolactone (an inhibitor of IκB kinase), SP600125 (an inhibitor of SAPK/JNK), and JAK inhibitor I (an inhibitor of JAK 1, 2 and 3). However, IL-6 levels were not affected by PD98059 (an inhibitor of MEK1/2). Midazolam markedly suppressed IL-1β-stimulated STAT3 phosphorylation without affecting the phosphorylation of p38 MAP kinase, SAPK/JNK or IκB.</p> <p>Conclusion</p> <p>These results strongly suggest that midazolam inhibits IL-1β-induced IL-6 release in rat C6 glioma cells via suppression of STAT3 activation. Midazolam may affect immune system function in the CNS.</p

Effects of novelly synthesized nucleolipides on different tumor cell lines (HT29, HepG2, Panc-1, RenCa) with special respect to glioma cell lines (BT4Ca, GOS3, G28, G112, U251, U87) of human or other species

Today cancer is the second leading cause of death around the world. The World Health Organization predicts an increase from 9.6 million in 2018 up to 16.4 million cancer deaths in 2040. Although tumors of the brain and the nervous system are rather unusual among adults (2 %), they are the second most common diagnosed types of cancer in childhood. Gliomas and meningiomas are the most frequent types of brain tumors and account for 30 % of all tumors in the brain and central nervous system as well as for 80 % of all malignant intra-cranial tumors. Usually surgical excision is the first step in the treatment of cancer followed by radio- and chemotherapy. Due to the mainly intracranial location of gliomas, surgical interventions are extremely risky. Therefore, the use of radiation and chemotherapeutic agents become more important. 5-Fluorouracil (5-FU) is a well-known drug, usually used as chemotherapeutic agent in the standard treatment of fast proliferating cancer cells. Unfortunately, it carries along a wide range of unpleasant side effects in multiple organ systems. To improve the passage through the blood-brain-barrier and the cell membrane penetration thereby gaining a higher efficiency, our cooperation partners Prof. Dr. Rosemeyer and his colleagues from the Institute of Chemistry and Materials of the University of Osnabrück synthesized more than 120 novel nucleolipids. These molecules have been designed based on the 5-FU derivative 5-Fluorouridine (5-FUrd) or the nucleosides adenosine, cladribin, formycin, guanosine, inosine and others. The derivatives are modified at several positions and have different additional chemical side groups to mainly influence the hydrophilicity and lipophilicity and thus, to consequently improve the permeability across the blood-brain-barrier as well as the uptake into the cells. At first we screened the impact of 5-FUrd and the 120 novelly synthesized derivatives on the viability of rat BT4Ca and human GOS3 glioma cells as well as differentiated human macro-phages (THP1). Five derivatives (S.18, 19, 38, 98 and 101), which show low cytotoxic effects on the macrophages and are effective against both, the rat BT4Ca and the human GOS3 glioma cells, were selected to be further analyzed. Although we screened 120 derivatives consisting of purines and pyrimidines, it is remarkable that all five selected, most effective derivatives (S.18, 19, 38, 98 and 101) belong to the group of pyrimidines. Comparing the chemical structure of the selected derivatives, the position of the attached farnesyl chain seems to be of high importance for the effectiveness of the most efficient derivative (S.98) against rat as well as human glioma cell lines. We were able to show that the selected derivatives, derived out of the nucleosides adenosine (S.38), formycin (S.98) and inosine (S.18, 19 and 101) are able to keep up with or even overcome the cytostatic/cytotoxic effects of the well-known chemotherapeutic agent 5-FU on further human glioma cell lines (G28, G112, U251 and U87) and several other tumor entities (HT29, HepG2, Panc-1 and RenCa). Certainly, these effects depend on the applied derivative as well as its concentration and vary among the specific cell type analyzed. Furthermore, using the rat BT4Ca glioma cells as a model of fast proliferating glioblastomas, we studied several intracellular mechanisms, possibly responsible for the examined cytotoxic effects of the selected derivatives (S.18, 19, 38, 98 and 101). Our results indicate that the treatment of the rat BT4Ca glioma cells with each derivative lead to a concentration-dependent increase of the apoptotic cell rate. At concentrations 25 and 50 µM each of the selected derivatives led to a distinct activation of the caspase 3, confirming the induction of apoptosis. We also observed an increase of the necrotic cell rate with ascending concentration (12.5, 25 and 50 µM) of each derivative presumably due to secondary necrosis. Beside the loss of cell mass due to apoptosis, we detected lower levels of PCNA after the treatment with any selected derivative (12.5 and 25 µM) indicating a reduced cell proliferation of the rat BT4Ca glioma cells. In contrast, the treatment of the rat BT4Ca glioma cells with 5-FUrd did not affect the amount of PCNA. Thus, at our experimental conditions 5-FUrd showed no impact on the proliferation rate consistent with its cytostatic effects. Moreover, we determined a tremendous accumulation of ROS per cell after the treatment with each derivative (50 µM). Especially the treatment with the derivatives S.18, 38 and 98 caused high oxidative stress in the rat BT4Ca glioma cells. But regarding the ratio of rGSH/GSSG only the derivative S.98 showed an explicit effect on the glutathione antioxidant system by lowering the rGSH/GSSG ratio. Nevertheless, the activation of NFκB, through the translocation of the subunit p65 from the cytoplasm to the nucleus, was not triggered by the treatment with any selected derivative or 5-FUrd. For a validation of the effects of the derivatives on the migratory capacity of the rat BT4Ca glioma cells during the treatment, further experiments need to be done with a more accurate method. The results indicate that our five selected derivatives S.18, 19, 38, 98 and 101 bear fundamen-tal attributes, which are beneficial for the treatment of malignant cancer cells. These proper-ties and the broad range of cytotoxicity as well as the inhibition of the cell proliferation make the five derivatives, especially S.19 and S.98, highly interesting as prospects against several tumor entities for the use as possible novel drugs with a high potential as chemotherapeutic agent.Aktuell gehört Krebs zu den häufigsten Todesursachen weltweit. Die Welt-Gesundheits-Organisation prognostiziert einen Anstieg von 9,5 Millionen Krebs assoziierten Todesfällen im Jahre 2018 auf 16,4 Millionen im Jahre 2040. Obwohl Tumore des Gehirns und des zentralen Nervensys-tems bei Erwachsenen eher selten vorkommen (2 %), sind sie bei Kindern die am zweithäufigsten diagnostizierte Krebsart. Gliome und Meningiome stellen 30 % aller Tumore des Gehirns und Zent-ralen Nervensystems sowie 80 % aller bösartigen intrakranialen Tumore. Die Standardtherapie beginnt im Regelfall mit der operativen Entfernung, gefolgt von Radio- und Chemotherapie. Auf Grund der intrakranialen Lokalisation von Gliomen sind chirurgische Maßnahmen jedoch sehr ris-kant, wodurch Bestrahlungen und chemotherapeutische Behandlungen an Bedeutung gewinnen. 5-Fluorouracil (5-FU) ist ein seit vielen Jahren genutzter Wirkstoff, das als Chemotherapeutikum gegen schnell wachsende Krebszellen eingesetzt wird. Bedauerlicherweise bringt er auch ein breites Spektrum an unerwünschten Nebeneffekten für einige Organsysteme mit sich. Um die Passage durch die Blut-Hirn-Schranke und die Aufnahme in die Zelle und damit die Effektivität solcher Sub-stanzen zu verbessern, entwickelten und synthetisierten unsere Kooperationspartner Prof. Dr. Ro-semeyer und seine Kollegen am Institut für Chemie und Materialien der Universität Osnabrück über 120 neue Nukleolipidderivate. Die Struktur dieser Nukleolipide beruht auf dem 5-FU Derivat 5-Fluorouridine (5-FUrd) oder den Nukleosiden Adenosin, Cladribin, Formycin, Guanosin, Inosin und weiteren. Die Derivate dieser Grundsubstanzen sind an mehreren Positionen verändert und tragen zusätzlich verschiedene chemische Seitenketten um ihre Wasser- und Fettlöslichkeit zu beeinflus-sen. Durch diese Modifikationen soll der Übergang durch die Blut-Hirn-Schranke sowie die Aufnahme in die Zelle erleichtert werden. Zu Beginn der Studie untersuchten wir den Einfluss von 5-FUrd und den 120 neu synthetisierten Derivaten auf die Viabilität der aus Ratten stammenden BT4Ca und humanen GOS3 Gliom Zellen sowie differenzierten humanen Makrophagen (THP1). Fünf Derivate (S.18, 19, 38, 98 und 101), die keine oder nur eine geringe schädliche Wirkung für die Makrophagen zeigten, gleichzeitig jedoch effektiv gegen beide Gliom Zelllinien (BT4Ca und GOS3) wirkten, wurden ausgewählt und weiter analysiert. Bemerkenswert ist, dass trotz der Anzahl von 120 getesteten Substanzen, die sowohl Purine als auch Pyrimidine einschlossen, alle fünf ausgewählten Derivate zur Gruppe der Pyrimidine gehören. Vergleicht man die chemische Struktur der ausgewählten Derivate, spielt die Position der angehängten Farnesylkette offenbar eine wichtige Rolle für die Effektivität (S.98 versus S.18, 38 und 101). Wir zeigen, dass die ausgewählten Derivate, deren Grundlage Adenosin (S.38), Formycin (S.98) und Inosin (S.18, 19, 101) bilden, einen vergleichbaren oder sogar stärkeren zytostati-schen/zytotoxischen Effekt wie das bekannte chemotherapeutische Mittel 5-FU haben. Auch bei weiteren untersuchten humanen Gliom Zellen (G28, G112, U251 und U87) und anderen Tumorenti-täten (HT29, HepG2, Panc-1 und RenCa) können wir diese Effekte beobachten. Diese sind abhängig vom eingesetzten Derivat sowie dessen Konzentration und des verwendeten, spezifischen Zelltyps. Für weiterführende Versuche wurden die Ratten BT4Ca Gliom Zellen exemplarisch als Model für schnellwachsende Glioblastomzellen verwendet. Wir analysierten unterschiedliche intrazelluläre Mechanismen, die möglicherweise an den bereits beobachteten zytotoxischen Wirkungen der fünf ausgewählten Derivate (S.18, 19, 38, 98 und 101) beteiligt sind. Unsere Versuchsergebnisse dokumentieren, dass die Behandlung der Ratten BT4Ca Gliom Zellen mit einem dieser fünf Derivate zu einem konzentrationsabhängigen Anstieg der apoptotischen Zellzahl führt. In den Konzentrationen 25 und 50 µM löst jedes der ausgewählten Derivate eine deutliche Steigerung der Capspase 3 Aktivität aus, wodurch die beobachtete Induktion der Apoptose bestätigt wird. Mit steigenden Konzentrationen (12.5, 25 und 50 µM) sehen wir allerdings auch einen erhöhten Anteil an nekrotischen Zellen. Dieser Anstieg ist vermutlich auf den Prozess der sekundär-Nekrose zurückzuführen. Neben dem Zelltod durch Apoptose, detektieren wir nach der Behandlung mit den jeweiligen Derivaten auch ein verringertes Level des Proteins PCNA, wodurch die Zellteilung der Ratten BT4Ca Zellen zusätzlich vermindert wird. Im Gegensatz zu den Derivaten hat die Behandlung mit 5-FUrd keinen Einfluss auf die gemessene Menge von PCNA. Demnach hat 5-FUrd unter unseren experimentellen Bedingungen keinen Einfluss auf die Zellteilungsrate der Ratten BT4Ca Zellen, was den beobachteten zytostatischen Effekt von 5-FUrd erklärt. Darüber hinaus beobachten wir nach der Behandlung der Ratten BT4Ca Zellen mit den Derivaten (50 µM) eine enorme Anhäufung von Sauerstoff Radikalen (ROS) pro Zelle. Besonders der Einsatz der Derivate S.18, 38 und 98 verursacht hohen oxidativen Stress für die Ratten BT4Ca Zellen. Allerdings führt nur die Behandlung mit Derivat S.98 zu einer eindeutigen Verringerung des rGSH/GSSG Verhältnisses und beeinflusst damit die physiologisch zur Verfügung stehende Menge des Antioxidans Glutathion. Keines der fünf ausgewählten Derivate führt zur Translokation der NFκB-Untereinheit p65 vom Zytoplasma in den Zellkern und damit zur Aktivierung von NFκB in den Ratten BT4Ca Zellen. Der Einfluss der Derivate auf die Migration der Ratten BT4Ca Zellen muss durch weitere Untersu-chungen validiert werden. Die bisherigen Analysen zeigen, dass die Derivate S.18, 19, 38, 98 und 101 grundlegende Eigenschaften aufweisen, wodurch sie für die Behandlung von diversen bösarti-gen Krebszellen geeignet sind. Die fünf ausgewählten Derivate, besonders, S.19 und S.98, tragen durch ihr breites zytotoxisches Wirkungsspektrum und die Hemmung der Zellteilungsrate ein hohes Potential gegen mehrere, unterschiedliche Tumorentitäten als neue Wirkstoffe im Bereich der Chemotherapie Verwendung zu finden

Gene therapy of malignant glioma with retroviral vectors and tumor-infiltrating progenitor cells

Gene therapy as a therapeutic strategy in the treatment of human gliomas is limited by the efficacy of gene transfer and intratumoral distribution of viral vectors. The major goals of this study were to enhance the gene transfer to glioma cells in vivo by using lentiviral vectors and to improve intratumoral distribution by selecting migratory progenitor cells that could function as packaging cells for the viral vectors. Therefore, the final goal was to establish tumor infiltrating packaging cells that release viral vectors within glioma in vivo. Lentiviral vectors were chosen to deliver genes into glioma cells. In contrast to currently used retroviral vectors, they transduce quiescent as well as mitotic cells. This is of major importance as within a defined treatment window, the majority of tumor cells are not mitotically active. However, lentiviral vectors can also infect normal brain cells. To define vectors with a specific tropism for glioma cells, lentiviral vectors pseudotyped with two different glycoproteins were used. Vectors pseudotyped with glycoproteins of the lymphocytic choriomeningitis virus (LCMV-GP) mediated efficient and specific transduction of rat glioma cells in vitro and in vivo, whereas vectors pseudotyped with the glycoproteins of the vesicular stomatatitis virus (VSV-G) preferentially transduced normal brain cells [Miletic et al., 2004]. Bone marrow derived progenitor cells were isolated to establish the tumor-infiltrating cells (BM-TICs) that could also serve as packaging cells. BM-TICs have a high passaging capacity in vitro, which is necessary for genetic modification and large scale production in the clinic. They were also found to show specific migration towards and into malignant glioma in vivo. In a therapeutic approach using BM-TICs stably expressing a suicide gene, an efficient therapeutic effect was demonstrated. The modified cells were also detected in vivo by non-invasive positron emission tomography (PET) and therapeutic outcome was followed-up by imaging methods and correlated with histopathology [Miletic et al., 2007]. To test packaging capabilities of BMTICs, the cells were modified with packaging constructs for retroviral LCMV-GP pseudotypes (BM-TIPCs). BM-TIPCs continuously produced retroviral vector particles for several weeks. Upon injection into experimental rat glioma, these cells migrated and were widely distributed within the tumor. Furthermore, released vector particles transduced glioma cells in solid as well as border areas [Fischer, Miletic et al., 2007]. In conclusion, the presented packaging system is highly attractive for future therapeutic applications in human glioblastoma especially in conjunction with an imaging-guided approach

Prolactin and its receptor as therapeutic targets in glioblastoma multiforme

Although prolactin (PRL) and its receptor (PRLR) have been detected in glioblastoma multiforme (GBM), their role in its pathogenesis remains unclear. Our aim was to explore their contribution in GBM pathogenesis. We detected PRL and PRLR in all GBM cell lines tested. PRLR activation or overexpression using plasmid transfection increased proliferation, viability, clonogenicity, chemoresistance and matrix metalloproteinase activity in GBM cells, while PRLR antagonist ∆1–9-G129R-hPRL reduced their proliferation, viability, chemoresistance and migration. Meta-analysis of transcriptomic data indicated that PRLR was expressed in all grade II-III glioma (GII-III) and GBM samples. PRL was upregulated in GBM biopsies when compared to GII-III. While in the general population tumour PRL/PRLR expression did not correlate with patient survival, biological sex-stratified analyses revealed that male patients with PRL+/PRLRHIGH GBM performed worse than PRL+/PRLRLOW GBM. In contrast, all male PRL+/PRLRHIGH GII-III patients were alive whereas only 30% of PRL+/PRLRLOW GII-III patients survived after 100 months. Our study suggests that PRLR may be involved in GBM pathogenesis and could constitute a therapeutic target for its treatment. Our findings also support the notion that sexual dimorphism should be taken into account to improve the care of GBM patients.Fil: Asad, Antonela Sofía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Nicola Candia, Alejandro Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: González, Nazareno. Laboratorio Max Planck de Biología Estructural, Química y Biofísica Molecular de Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Zuccato, Camila Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Abt, Araceli. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Orrillo, Santiago Jordi. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Yael, Lastra. Universidad de Buenos Aires. Facultad de Ciencias Veterinarias. Cátedra de Fisiología Animal; ArgentinaFil: de Simone, Emilio Adrian. Universidad de Buenos Aires. Facultad de Ciencias Veterinarias. Cátedra de Fisiología Animal; ArgentinaFil: Boutillon, Florence. Inserm; FranciaFil: Goffin, Vincent. Inserm; FranciaFil: Seilicovich, Adriana. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Pisera, Daniel Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Ferraris, Maria Jimena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Candolfi, Marianela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentin

Two Types of K⁺ Channel Subunit, Erg1 and KCNQ2/3, Contribute to the M-Like Current in a Mammalian Neuronal Cell

The potassium M current was originally identified in sympathetic ganglion cells, and analogous currents have been reported in some central neurons and also in some neural cell lines. It has recently been suggested that the M channel in sympathetic neurons comprises a heteromultimer of KCNQ2 and KCNQ3 (Wang et al., 1998) but it is unclear whether all other M-like currents are generated by these channels. Here we report that the M-like current previously described in NG108–15 mouse neuroblastoma x rat glioma cells has two components, “fast” and “slow”, that may be differentiated kinetically and pharmacologically. We provide evidence from PCR analysis and expression studies to indicate that these two components are mediated by two distinct molecular species of K+ channel: the fast component resembles that in sympathetic ganglia and is probably carried byKCNQ2/3 channels, whereas the slow component appears to be carried by merg1a channels. Thus, the channels generating M-like currents in different cells may be heterogeneous in molecular composition

Molecular and Genetic Determinants of Glioma Cell Invasion.

A diffusely invasive nature is a major obstacle in treating a malignant brain tumor, "diffuse glioma", which prevents neurooncologists from surgically removing the tumor cells even in combination with chemotherapy and radiation. Recently updated classification of diffuse gliomas based on distinct genetic and epigenetic features has culminated in a multilayered diagnostic approach to combine histologic phenotypes and molecular genotypes in an integrated diagnosis. However, it is still a work in progress to decipher how the genetic aberrations contribute to the aggressive nature of gliomas including their highly invasive capacity. Here we depict a set of recent discoveries involving molecular genetic determinants of the infiltrating nature of glioma cells, especially focusing on genetic mutations in receptor tyrosine kinase pathways and metabolic reprogramming downstream of common cancer mutations. The specific biology of glioma cell invasion provides an opportunity to explore the genotype-phenotype correlation in cancer and develop novel glioma-specific therapeutic strategies for this devastating disease

Protein kinase a distribution differentiates human glioblastoma from brain tissue

Brain tumor glioblastoma has no clear molecular signature and there is no effective therapy. In rodents, the intracellular distribution of the cyclic AMP (cAMP)-dependent protein kinase (Protein kinase A, PKA) R2Alpha subunit was previously shown to differentiate tumor cells from healthy brain cells. Now, we aim to validate this observation in human tumors. The distribution of regulatory (R1 and R2) and catalytic subunits of PKA was examined via immunohistochemistry and Western blot in primary cell cultures and biopsies from 11 glioblastoma patients. Data were compared with information obtained from 17 other different tumor samples. The R1 subunit was clearly detectable only in some samples. The catalytic subunit was variably distributed in the different tumors. Similar to rodent tumors, all human glioblastoma specimens showed perinuclear R2 distribution in the Golgi area, while it was undetectable outside the tumor. To test the effect of targeting PKA as a therapeutic strategy, the intracellular cyclic AMP concentration was modulated with different agents in four human glioblastoma cell lines. A significant increase in cell death was detected after increasing cAMP levels or modulating PKA activity. These data raise the possibility of targeting the PKA intracellular pathway for the development of diagnostic and/or therapeutic tools for human glioblastoma

Adjuvant therapeutic potential of tonabersat in the standard treatment of glioblastoma : a preclinical F98 glioblastoma rat model study

Purpose Even with an optimal treatment protocol, the median survival of glioblastoma (GB) patients is only 12-15 months. Hence, there is need for novel effective therapies that improve survival outcomes. Recent evidence suggests an important role for connexin (Cx) proteins (especially Cx43) in the microenvironment of malignant glioma. Cx43-mediated gap junctional communication has been observed between tumor cells, between astrocytes and between tumor cells and astrocytes. Therefore, gap junction directed therapy using a pharmacological suppressor or modulator, such as tonabersat, could be a promising target in the treatment of GB. In this preclinical study, we evaluated the possible therapeutic potential of tonabersat in the F98 model. Procedures Female Fischer rats were inoculated with +/- 25.000 F98 tumor cells in the right frontal lobe. Eight days post-inoculation contrast-enhanced T1-weighted (CE-T1w) magnetic resonance (MR) images were acquired to confirm tumor growth in the brain. After tumor confirmation, rats were randomized into a Control Group, a Connexin Modulation Group (CM), a Standard Medical Treatment Group (ST), and a Standard Medical Treatment with adjuvant Connexin Modulation Group (STCM). To evaluate therapy response, T2-weighted (T2w) and CE-T1w sequences were acquired at several time points. Tumor volume analysis was performed on CE-T1w images and statistical analysis was performed using a linear mixed model. Results Significant differences in estimated geometric mean tumor volumes were found between the ST Group and the Control Group and also between the STCM Group and the Control Group. In addition, significant differences in estimated geometric mean tumor volumes between the ST Group and the STCM Group were demonstrated. No significant differences in estimated geometric mean tumor volumes were found between the Control Group and the CM Group. Conclusion Our results demonstrate a therapeutic potential of tonabersat for the treatment of GB when used in combination with radiotherapy and temozolomide chemotherapy

Efficacy of Combined 5-Fluorouracil and Photodynamic Therapy in Glioma Spheroids

Standard treatment regimens consisting of surgery, radiation and chemotherapy have proven ineffective for the treatment of high-grade gliomas such as glioblastoma multiforme (GBM). An effective cure requires elimination of nests of tumor cells that have migrated from the resection margin and infiltrated normal brain. A number of localized therapies, including light-based approaches such as photodynamic therapy (PDT) and photochemical internalization (PCI) are currently under investigation for the management of GBM patients. Several studies have demonstrated a high degree of synergy between PDT and bleomycin, via the PCI mechanism, in a variety of in vitro and in vivo models, including glioma cell lines. The purpose of this study was to examine the efficacy of combined treatments consisting of PDT and the chemotherapeutic agent, 5-fluorouracil (5-FU) in a 3-dimensional spheroid model consisting of F98 rat glioma cells. Spheroids were incubated with a photosensitizer (aluminum phthalocyanine disulfonate; AlPcS2a) and irradiated with 670 nm laser light. Three different wash protocols (0, 4 and 24 h) were employed to determine whether any observed interactions between PDT and 5-FU could be attributed to the PCI mechanism, or were simply due to different cytotoxic pathways of the two treatment modalities. Although the combined PDT + 5-FU treatments resulted in greater suppression of spheroid growth compared to either treatment alone, no statistically significant differences in growth effects were observed between 0 and 4 h wash protocols suggesting that the combined treatment effects were due to different mechanisms of cytotoxicity, rather than a PCI effect