Mitochondrial response of glioma cells to temozolomide.

Metabolic adaptations are central for carcinogenesis and response to therapy, but little is known about the contribution of mitochondrial dynamics to the response of glioma cells to the standard treatment with temozolomide (TMZ). Glioma cells responded to TMZ with mitochondrial mass increased and the production of round structures of dysfunctional mitochondria. At single-cell level, asymmetric mitosis contributed to the heterogeneity of mitochondrial levels. It affected the fitness of cells in control and treated condition, indicating that the mitochondrial levels are relevant for glioma cell fitness in the presence of TMZ.

MG-Pe: A Novel Galectin-3 Ligand with Antimelanoma Properties and Adjuvant Effects to Dacarbazine.

Melanoma is a highly metastatic and rapidly progressing cancer, a leading cause of mortality among skin cancers. The melanoma microenvironment, formed from the activity of malignant cells on the extracellular matrix and the recruitment of immune cells, plays an active role in the development of drug resistance and tumor recurrence, which are clinical challenges in cancer treatment. These tumoral metabolic processes are affected by proteins, including Galectin-3 (Gal-3), which is extensively involved in cancer development. Previously, we characterized a partially methylated mannogalactan (MG-Pe) with antimelanoma activities. In vivo models of melanoma were used to observe MG-Pe effects in survival, spontaneous, and experimental metastases and in tissue oxidative stress. Analytical assays for the molecular interaction of MG-Pe and Gal-3 were performed using a quartz crystal microbalance, atomic force microscopy, and contact angle tensiometer. MG-Pe exhibits an additive effect when administered together with the chemotherapeutic agent dacarbazine, leading to increased survival of treated mice, metastases reduction, and the modulation of oxidative stress. MG-Pe binds to galectin-3. Furthermore, MG-Pe antitumor effects were substantially reduced in Gal-3/KO mice. Our results showed that the novel Gal-3 ligand, MG-Pe, has both antitumor and antimetastatic effects, alone or in combination with chemotherapy.

Plasma and cerebrospinal fluid pharmacokinetics of select chemotherapeutic agents following intranasal delivery in a non-human primate model.

The blood-brain barrier (BBB) limits entry of most chemotherapeutic agents into the CNS, resulting in inadequate exposure within CNS tumor tissue. Intranasal administration is a proposed means of delivery that can bypass the BBB, potentially resulting in more effective chemotherapeutic exposure at the tumor site. The objective of this study was to evaluate the feasibility and pharmacokinetics (plasma and CSF) of intranasal delivery using select chemotherapeutic agents in a non-human primate (NHP) model. Three chemotherapeutic agents with known differences in CNS penetration were selected for intranasal administration in a NHP model to determine proof of principle of CNS delivery, assess tolerability and feasibility, and to evaluate whether certain drug characteristics were associated with increased CNS exposure. Intravenous (IV) temozolomide (TMZ), oral (PO) valproic acid, and PO perifosine were administered to adult male rhesus macaques. The animals received a single dose of each agent systemically and intranasally in separate experiments, with each animal acting as his own control. The dose of the agents administered systemically was the human equivalent of a clinically appropriate dose, while the intranasal dose was the maximum achievable dose based on the volume limitation of 1 mL. Multiple serial paired plasma and CSF samples were collected and quantified using a validated uHPLC/tandem mass spectrometry assay after each drug administration. Pharmacokinetic parameters were estimated using non-compartmental analysis. CSF penetration was calculated from the ratio of areas under the concentration-time curves for CSF and plasma (AUCCSF:plasma). Intranasal administration was feasible and tolerable for all agents with no significant toxicities observed. For TMZ, the degrees of CSF drug penetration after intranasal and IV administration were 36 (32-57) and 22 (20-41)%, respectively. Although maximum TMZ drug concentration in the CSF (Cmax) was lower after intranasal delivery compared to IV administration due to the lower dose administered, clinically significant exposure was achieved in the CSF after intranasal administration with the lower doses. This was associated with lower systemic exposure, suggesting increased efficiency and potentially lower toxicities of TMZ after intranasal delivery. For valproic acid and perifosine, CSF penetration after intranasal delivery was similar to systemic administration. Although this study demonstrates feasibility and safety of intranasal drug administration, further agent-specific studies are necessary to optimize agent selection and dosing to achieve clinically-relevant CSF exposures.

Temozolomide increases MHC-I expression via NF-κB signaling in glioma stem cells.

Gliomas are the most common and primary tumors of the central nervous system in adults. Temozolomide (TMZ) is the main drug used to treat glioma; however, prognosis remains poor for most patients. Glioma stem cells (GSCs) are thought to enable glioma initiation and evasion from immune surveillance; their immunogenicity can be determined by expression of major histocompatibility complex (MHC)-I. The present study investigated the effect of TMZ on MHC-I expression in GSCs. Glioma spheres were cultured in serum-free medium containing epidermal growth factor, basic fibroblast growth factor, and B27; MHC-I expression was detected by immunocytochemistry, quantitative real-time PCR, and flow cytometry. Nuclear factor (NF)-κB expression in glioma stem cells was detected by Western blot. TMZ enhanced MHC-I expression in GSCs, and NF-κB was activated. TMZ treatment increased MHC-I expression via modulation of NF-κB signaling in GSCs. In addition to being a chemotherapeutic agent, TMZ may also serve as an immunomodulatory agent in the treatment of glioma patients.

Brain Targeting of Temozolomide via the Intranasal Route Using Lipid-Based Nanoparticles: Brain Pharmacokinetic and Scintigraphic Analyses.

The aim of the present work was to investigate the efficacy of temozolomide nanostructured lipid carriers (TMZ-NLCs) to enhance brain targeting via nasal route administration. The formulation was optimized by applying a four-factor, three-level Box-Behnken design. The developed formulations and the functional relationships between their independent and dependent variables were observed. The independent variables used in the formulation were gelucire (X1), liquid lipid/total lipid (X2), Tween 80 (X3), and sonication time (X4), and their effects were observed with regard to size (Y1), % drug release (Y2), and drug loading (Y3). The optimized TMZ-NLC was further evaluated for its surface morphology as well as ex vivo permeation and in vivo studies. All TMZ-NLC formulations showed sizes in the nanometer range, with high drug loading and prolonged drug release. The optimized formulation (TMZ-NLCopt) showed an entrapment efficiency of 81.64 ± 3.71%, zeta potential of 15.21 ± 3.11 mV, and polydispersity index of less than 0.2. The enhancement ratio was found to be 2.32-fold that of the control formulation (TMZ-disp). In vivo studies in mice showed that the brain/blood ratio of TMZ-NLCopt was found to be significantly higher compared to that of TMZ-disp (intranasal, intravenous). Scintigraphy images of mouse brain showed the presence of a high concentration of TMZ. The AUC ratio of TMZ-NLCopt to TMZ-disp in the brain was the highest among the organs. The findings of this study substantiate the existence of a direct nose-to-brain delivery route for NLCs.

Problems of Glioblastoma Multiforme Drug Resistance.

Glioblastoma multiforme (GBL) is the most common and aggressive brain neoplasm. A standard therapeutic approach for GBL involves combination therapy consisting of surgery, radiotherapy, and chemotherapy. The latter is based on temozolomide (TMZ). However, even by applying such a radical treatment strategy, the mean patient survival time is only 14.6 months. Here we review the molecular mechanisms underlying the resistance of GBL cells to TMZ including genetic and epigenetic mechanisms. Present data regarding a role for genes and proteins MGMT, IDH1/2, YB-1, MELK, MVP/LRP, MDR1 (ABCB1), and genes encoding other ABC transporters as well as Akt3 kinase in developing resistance of GBL to TMZ are discussed. Some epigenetic regulators of resistance to TMZ such as microRNA and EZH2 are reviewed.

Synergistic cytotoxicity of the poly (ADP-ribose) polymerase inhibitor ABT-888 and temozolomide in dual-drug targeted magnetic nanoparticles.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) is associated with a poor prognosis because of a lack of effective treatment options. The objective of this study was to examine a new strategy for HCC treatment, namely the use of poly (ADP-ribose) polymerase 1 (PARP-1) inhibitor (ABT-888) together with Temozolomide (TMZ) incorporated onto magnetic nanoparticles. METHODS: Magnetic Fe3 O4 /Fe cores were encapsulated within a silica shell to facilitate the simultaneous incorporation of ABT-888 and TMZ. In vitro tests were performed with HepG2, Hep3B and PLC-PRF-5 liver tumoural cell lines and with WRL-68 liver non-tumoural cells. RESULTS: The magnetic nanocarriers were loaded simultaneously with ABT-888 and TMZ. High stability and extended release were achieved in culture medium. Confocal microscopy images showed that drug-loaded particles were uptaken and accumulated into the cytoplasm of liver tumoural cells, inducing the following effects: G2/M cell cycle arrest (P < 0.05), accumulation of DNA damage (P < 0.05), mitochondrial depolarization (P < 0.01), reduction in BCL-xL, FOS, JUND gene expression (P < 0.05), PARP-1 fragmentation, Caspase-3 activation and apoptotic cell death (P < 0.05). Interestingly, drugs loaded onto nanoparticles exhibited better efficiency than free drugs (cell death triggered by drug delivery nanosystem: 53.5% vs. 34.5% by free drugs, P = 0.01). CONCLUSIONS: These magnetic nanocompounds are able to incorporate both drugs simultaneously, enter the tumour cells and release them. ABT-888/TMZ/NPs decrease the transcription of key genes involved in tumour survival and induce apoptotic cell death in a more effective manner than is achieved by free drugs.

Transport of treosulfan and temozolomide across an in-vitro blood-brain barrier model.

In vitro, treosulfan (TREO) has shown high effectiveness against malignant gliomas. However, a first clinical trial for newly diagnosed glioblastoma did not show any positive effect. Even though dosing and timing might have been the reasons for this failure, it might also be that TREO does not reach the brain in sufficient amount. Surprisingly, there are no published data on TREO uptake into the brain of patients, despite extensive research on this compound. An in-vitro blood-brain barrier (BBB) model consisting of primary porcine brain capillary endothelial cells was used to determine the transport of TREO across the cell monolayer. Temozolomide (TMZ), the most widely used cytotoxic drug for malignant gliomas, served as a reference. An HPLC-ESI-MS/MS procedure was developed to detect TREO and TMZ in cell culture medium. Parallel to the experimental approach, the permeability of TREO and the reference substance across the in-vitro BBB was estimated on the basis of their physicochemical properties. The detection limit was 30 nmol/l for TREO and 10 nmol/l for TMZ. Drug transport was measured in two directions: influx, apical-to-basolateral (A-to-B), and efflux, basolateral-to-apical (B-to-A). For TREO, the A-to-B permeability was lower (1.6%) than the B-to-A permeability (3.0%). This was in contrast to TMZ, which had higher A-to-B (13.1%) than B-to-A (7.2%) permeability values. The in-vitro BBB model applied simulated the human BBB properly for TMZ. It is, therefore, reasonable to assume that the values for TREO are also meaningful. Considering the lack of noninvasive, significant alternative methods to study transport across the BBB, the porcine brain capillary endothelial cell model was efficient to collect first data for TREO that explain the disappointing clinical results for this drug against cerebral tumors.

Photochemical internalization of bleomycin and temozolomide--in vitro studies on the glioma cell line F98.

Here we evaluate the photosensitizer meso-tetraphenyl chlorin disulphonate (TPCS2a) in survival studies of rat glioma cancer cells in combination with the novel photochemical internalization (PCI) technique. The tested anticancer drugs were bleomycin (BLM) and temozolomide (TMZ). Glioma cells were incubated with TPCS2a (0.2 µg ml(-1), 18 h, 37 °C) before BLM or TMZ stimulation (4 h) prior to red light illumination (652 nm, 50 mW cm(-2)). The cell survival after BLM (0.5 µm)-PCI (40 s light) quantified using the MTT assay was reduced to about 25% after 24 h relative to controls, and to 31% after TMZ-PCI. The supplementing quantification by clonogenic assays, using BLM (0.1 µm), indicated a long-term cytotoxic effect: the surviving fraction of clonogenic cells was reduced to 5% after light exposure (80 s) with PCI, compared to 70% in the case of PDT. In parallel, structural and morphological changes within the cells upon light treatment were examined using fluorescence microscopy techniques. The present study demonstrates that PCI of BLM is an effective method for killing F98 glioma cells, but smaller effects were observed using TMZ following the "light after" strategy. The results are the basis for further in vivo studies on our rat glioma cancer model using PDT and PCI.

Treatment of poorly differentiated glioma using a combination of monoclonal antibodies to extracellular connexin-43 fragment, temozolomide, and radiotherapy.

Antitumor efficiencies of monoclonal antibodies to connexin-43 second extracellular loop (MAbE2Cx43), temozolomide, and fractionated γ-irradiation in the monotherapy mode and in several optimized combinations were studied in Wistar rats with induced C6 glioma. The survival of animals with glioma and the dynamics of intracerebral tumor development were evaluated by MRT. Temozolomide monotherapy (200 mg/m(2)) and isolated radiotherapy in a total dose of 36 Gy shifted the survival median from 28 days (no therapy) to 34 and 38 days, respectively; 100% animals died under conditions of temozolomide monotherapy and radiotherapy. Monotherapy with MAbE2Cx43 in a dose of 5 mg/kg led to significant regression of the tumor (according to MRT data), cure of 19.23% animals with glioma, and prolongation of the survival median to 39.5 days after tumor implantation. Combined therapy with MAbE2Cx43 and temozolomide completely abolished the antitumor effect (survival median 29 days). Treatment with MAbE2Cx43 in combination with radiotherapy was associated with mutual boosting of the therapeutic efficiencies, leading to a significant inhibition of tumor development and prolongation of the survival median to 60 days. The mechanism of tumorsuppressive activity of the antibodies could be due to connexon blockade in Cx43-positive glioma cells in the peritumor invasion zone. Higher efficiency of combined therapy was presumably due to the increase in blood-brain barrier permeability for antibodies after irradiation of the brain and to additional inhibitory effect of antibodies towards radioresistant migrating glioma cells. The results suggested that MAbE2Cx43 could be effective as the first-line drug in combined therapy for poorly differentiated gliomas.

In situ electrochemical evaluation of anticancer drug temozolomide and its metabolites-DNA interaction.

Temozolomide (TMZ) is an antineoplastic alkylating agent with activity against serious and aggressive types of brain tumours. It has been postulated that TMZ exerts its antitumor activity via its spontaneous degradation at physiological pH. The in vitro evaluation of the interaction of TMZ and its final metabolites, 5-aminoimidazole-4-carboxamide (AIC) and methyldiazonium ion, with double-stranded DNA (dsDNA) was studied using differential pulse voltammetry at a glassy carbon electrode. The DNA damage was electrochemically detected following the changes in the oxidation peaks of guanosine and adenosine residues. The results obtained revealed the decrease of the dsDNA oxidation peaks with incubation time, showing that TMZ and AIC/methyldiazonium ion interact with dsDNA causing its condensation. Furthermore, the experiments of the in situ TMZ and AIC/methyldiazonium ion-dsDNA interaction using the multilayer dsDNA-electrochemical biosensor confirmed the condensation of dsDNA caused by these species and showed evidence for a specific interaction between the guanosine residues and TMZ metabolites, since free guanine oxidation peak was detected. The oxidative damage caused to DNA bases by TMZ metabolites was also detected electrochemically by monitoring the appearance of the 8-oxoguanine/2,8-dyhydroxyadenine oxidation peaks. Nondenaturing agarose gel electrophoresis of AIC/methyldiazonium ion-dsDNA samples confirmed the occurrence of dsDNA condensation and oxidative damage observed in the electrochemical results. The importance of the dsDNA-electrochemical biosensor in the in situ evaluation of TMZ-dsDNA interactions is clearly demonstrated.

Chromatographic method for dacarbazine quantification in skin permeation experiments.

Dacarbazine (DTIC) is a chemotherapeutic drug currently used for the systemic treatment of melanomas. Considering the easy access to these tumors, a topical route of drug administration could provide a more comfortable and less toxic treatment. However, DTIC quantification aiming at the design of topical formulations is challenging, pondering all the interferents present in the drug samples recovered from the skin. Hence, this work intended to validate a selective chromatographic method for DTIC determination in skin permeation studies. A reversed-phase C18 column was used as a stationary phase, and gradient elution of a mobile phase consisting of methanol and pH 6.5 sodium phosphate monohydrate buffer (0.01 mol/L) at a flow rate of 1.0 mL/min was implemented. DTIC was detected at 364 nm. The method was selective against skin interferents, linear (r = 0.9995) in a concentration range of 1.0-15.0 µg/mL, precise with an overall variation coefficient lower than 3.8%, accurate achieving recovery from the skin layers within 91-112%, and sensitive for the proposed application (detection limit = 0.10 µg/ mL, quantification limit = 0.30 µg/mL). Furthermore, the analytical method was successfully tested in in vitro skin permeation studies. In conclusion, the developed method is appropriate for DTIC analysis from the skin sample matrix.

Prognostic Role of the Expression of Latent-Membrane Protein 1 of Epstein-Barr Virus in Classical Hodgkin Lymphoma.

The prognostic impact of the presence of Epstein-Barr virus (EBV) in classical Hodgkin lymphoma (cHL) is controversial. Previous studies reported heterogeneous results, rendering difficult the clinical validation of EBV as a prognostic biomarker in this lymphoma. The objective of this study was to evaluate the survival impact of the expression of EBV Latent-Membrane Protein 1 (EBV-LMP1) in tumoral Hodgkin-Reed-Sternberg (HRS) cells of primary diagnostic samples of cHL. Formalin-Fixed Paraffin-Embedded (FFPE) lymph node samples from 88 patients with cHL were analyzed. Patients were treated with the standard first-line chemotherapy (CT) with Adriamycin, Bleomycin, Vinblastine and Dacarbazine (ABVD) followed by radiotherapy. The Kaplan-Meier method and the Cox proportional hazards model were used for carrying out the survival analysis. In order to investigate whether the influence of EBV was age-dependent, analyses were performed both for patients of all ages and for age-stratified subgroups. In bivariate analysis, the expression of EBV was associated with older age (p = 0.011), mixed cellularity subtype cHL (p < 0.001) and high risk International Prognostic Score (IPS) (p = 0.023). Overall survival (OS) and progression-free survival (PFS) were associated with the presence of bulky disease (p = 0.009) and advanced disease at diagnosis (p = 0.016). EBV-positive cases did not present a significantly lower OS and PFS in comparison with EBV-negative cases, for all ages and when stratifying for age. When adjusted for covariates, absence of bulky disease at diagnosis (HR: 0.102, 95% CI: 0.02-0.48, p = 0.004) and limited disease stages (I-II) (HR: 0.074, 95% CI: 0.01-0.47, p = 0.006) were associated with a significant better OS. For PFS, limited-disease stages also retained prognostic impact in the multivariate Cox regression (HR: 0.145, 95% CI: 0.04-0.57, p = 0.006). These results are of importance as the early identification of prognostic biomarkers in cHL is critical for guiding and personalizing therapeutic decisions. The prognostic role of EBV in cHL could be modulated by the type of CT protocol employed and interact with the rest of presenting features.

Developing a clinically relevant radiosensitizer for temozolomide-resistant gliomas.

The prognosis for patients with glioblastoma (GB) remains grim. Concurrent temozolomide (TMZ) radiation-the cornerstone of glioma control-extends the overall median survival of GB patients by only a few months over radiotherapy alone. While these survival gains could be partly attributed to radiosensitization, this benefit is greatly minimized in tumors expressing O6-methylguanine DNA methyltransferase (MGMT), which specifically reverses O6-methylguanine lesions. Theoretically, non-O6-methylguanine lesions (i.e., the N-methylpurine adducts), which represent up to 90% of TMZ-generated DNA adducts, could also contribute to radiosensitization. Unfortunately, at concentrations attainable in clinical practice, the alkylation capacity of TMZ cannot overwhelm the repair of N-methylpurine adducts to efficiently exploit these lesions. The current therapeutic application of TMZ therefore faces two main obstacles: (i) the stochastic presence of MGMT and (ii) a blunted radiosensitization potential at physiologic concentrations. To circumvent these limitations, we are developing a novel molecule called NEO212-a derivatization of TMZ generated by coupling TMZ to perillyl alcohol. Based on gas chromatography/mass spectrometry and high-performance liquid chromatography analyses, we determined that NEO212 had greater tumor cell uptake than TMZ. In mouse models, NEO212 was more efficient than TMZ at crossing the blood-brain barrier, preferentially accumulating in tumoral over normal brain tissue. Moreover, in vitro analyses with GB cell lines, including TMZ-resistant isogenic variants, revealed more potent cytotoxic and radiosensitizing activities for NEO212 at physiologic concentrations. Mechanistically, these advantages of NEO212 over TMZ could be attributed to its enhanced tumor uptake presumably leading to more extensive DNA alkylation at equivalent dosages which, ultimately, allows for N-methylpurine lesions to be better exploited for radiosensitization. This effect cannot be achieved with TMZ at clinically relevant concentrations and is independent of MGMT. Our findings establish NEO212 as a superior radiosensitizer and a potentially better alternative to TMZ for newly diagnosed GB patients, irrespective of their MGMT status.

Recent approaches to improve the antitumor efficacy of temozolomide.

Temozolomide (TMZ) is an oral anticancer agent approved for the treatment of newly diagnosed glioblastoma in combination with radiotherapy. Moreover, TMZ has shown comparable efficacy with respect to dacarbazine, the reference drug for metastatic melanoma. Due to its favorable toxicity and pharmacokinetic profile, TMZ is under clinical investigation for brain metastasis from solid tumors and refractory leukemias. TMZ interacts with DNA generating a wide spectrum of methyl adducts mainly represented by N-methylpurines. However, its antitumor activity has been mainly attributed to O(6)-methylguanine, since tumor cell sensitivity inversely correlates with the levels of O(6)-alkylguanine DNA alkyltransferase and requires an intact mismatch repair system. Therefore, an increasing number of studies have been performed in order to identify patients who will benefit from TMZ treatment on the basis of their molecular/genetic profile. Unfortunately, resistance to the methylating agent occurs relatively often and strongly affects the rate and durability of the clinical response in cancer patients. Thus, different approaches have been developed to abrogate resistance or to increase the efficacy of TMZ and for many of them investigation is still underway. Herein, we provide an overview on the recent findings of preclinical and clinical studies on TMZ in combination with inhibitors of DNA repair, chemotherapeutic drugs with different mechanisms of action or radiotherapy, anti-angiogenic agents and other biological modulators.

Clinical features, mechanisms, and management of pseudoprogression in malignant gliomas.

Since the introduction of chemoradiotherapy with temozolomide as the new standard of care for patients with glioblastoma, there has been an increasing awareness of progressive and enhancing lesions on MRI, noted immediately after the end of treatment, which are not related to tumour progression, but which are a treatment effect. This so-called pseudoprogression can occur in up to 20% of patients who have been treated with temozolomide chemoradiotherapy, and can explain about half of all cases of increasing lesions after the end of this treatment. These lesions decrease in size or stabilise without additional treatments and often remain clinically asymptomatic. Additionally, there is evidence that treatment-related necrosis occurs more frequently and earlier after temozolomide chemotherapy than after radiotherapy alone. The mechanisms behind these events have not yet been fully elucidated, but the likelihood is that chemoradiotherapy causes a higher degree of (desired) tumour-cell and endothelial-cell killing. This increased cell kill might lead to secondary reactions, such as oedema and abnormal vessel permeability in the tumour area, mimicking tumour progression, in addition to subsequent early treatment-related necrosis in some patients and milder subacute radiotherapy reactions in others. In patients managed with temozolomide chemoradiotherapy who have clinically asymptomatic progressive lesions at the end of treatment, adjuvant temozolomide should be continued; in clinically symptomatic patients, surgery should be considered. If mainly necrosis is noted during surgery, continuation of adjuvant temozolomide is logical. Trials on the treatment of recurrent malignant glioma should exclude patients with progression within the first 3 months after temozolomide chemoradiotherapy unless histological confirmation of tumour recurrence is available. Further research is needed to establish reliable imaging parameters that distinguish between true tumour progression and pseudoprogression or treatment-related necrosis.

Biophysical interaction of temozolomide and its active metabolite with biomembrane models: The relevance of drug-membrane interaction for Glioblastoma Multiforme therapy.

Temozolomide (TMZ) is the first-line treatment for Glioblastoma Multiforme (GBM). After administration, TMZ is rapidly converted into its active metabolite (MTIC). However, its pharmacological activity is reduced due MTIC low bioavailability in the brain. Since drugs' permeability through biological barriers and tumor cell membranes affects its bioavailability, the ability of MTIC to interact with the biological membranes presents a major contribution on its pharmacological properties and activity. Biomembrane models mimic the physiological conditions, allowing to predict the drug's behavior at biological membranes and its effects on drug biodistribution profiles. In this work, lipid bilayer models using liposomes were applied for the drug-membrane interaction studies. The zwitterionic phospholipid, 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), and cholesterol were chosen for the composition of the model, since they represent the major components of the membranes of GBM cells and brain capillary endothelial cell. Thus, the molecular interactions between MTIC and these models were studied by the evaluation of the partition of the drug into the phospholipid's membrane, its location within the bilayer and its effect on the fluidity of the membrane. The attained results suggest that the composition of membranes affects drugs partition, showing that drug biodistribution depends not only on its physicochemical features, but also depends on the characteristics of the membrane such as the packing of the lipid molecules. Also, MTIC exhibited low affinity to biological membranes, explaining its low bioavailability on the target cells.

A peptide-modified solid lipid nanoparticle formulation of paclitaxel modulates immunity and outperforms dacarbazine in a murine melanoma model.

Melanoma is a highly aggressive skin cancer. A paclitaxel formulation of solid lipid nanoparticles modified with Tyr-3-octreotide (PSM) is employed to treat melanoma that highly expresses somatostatin receptors (SSTRs). PSM exerts more apoptotic and anti-invasive effects in B16F10 mice melanoma cells as compared to dacarbazine (DTIC), an approved chemotherapeutic drug for treating aggressive melanoma. Besides, PSM induces one of the biomarkers of immunogenic cell death in vitro and in vivo as confirmed by calreticulin exposure on the B16F10 cell surface. We observed a significant number of CD8 positive T cells in the tumor bed of the PSM treated group. As a result, PSM effectively reduces tumor volume in vivo as compared to DTIC. PSM also induces a favorable systemic immune response as determined in the spleen and sera of the treated animals. Importantly, PSM can reduce the number of nodule formations in the experimental lung metastasis model. Our experimentations indicate that the metronomic PSM exhibits remarkable anti-melanoma activities without any observable toxicity. This immune modulation behavior of PSM can be exploited for the therapy of melanoma and probably for other malignancies.

Receptor-mediated PLGA nanoparticles for glioblastoma multiforme treatment.

Glioblastoma multiforme is the most lethal type of brain tumor and the established therapy only extends patients survival to approximately one year. Its first-line treatment is based on of chemotherapy with the alkylating agent temozolomide (TMZ). As many other chemotherapeutic drugs, TMZ presents several limitations as high toxicity and low bioavailability. The delivery of TMZ using poly(lactic-co-glycolic acid) nanoparticles is proposed in this work. Stable nanoparticles functionalized with a OX26 type monoclonal antibody for transferrin receptor were developed, targeting the glioblastoma tumor cells, since these cells are known for overexpressing this receptor. The release profile of TMZ from the nanoparticles was studied mimicking physiological conditions, and targeted cellular internalization was also investigated. Two glioblastoma cell lines - U215 and U87 - were used to evaluate the in vitro cytotoxicity of the drug, showing that the prepared nanocarriers enhance the anticancer activity of TMZ. The functionalization with the monoclonal antibody for transferrin receptor proved to be advantageous in enhancing the cellular internalization in glioblastoma cells.

Experimental therapy of malignant gliomas using the inhibitor of histone deacetylase MS-275.

Inhibitors of histone deacetylases are promising compounds for the treatment of cancer but have not been systematically explored in malignant brain tumors. Here, we characterize the benzamide MS-275, a class I histone deacetylase inhibitor, as potent drug for experimental therapy of glioblastomas. Treatment of four glioma cell lines (U87MG, C6, F98, and SMA-560) with MS-275 significantly reduced cell growth in a concentration-dependent manner (IC(90), 3.75 micromol/L). Its antiproliferative effect was corroborated using a bromodeoxyuridine proliferation assay and was mediated by G(0)-G(1) cell cycle arrest (i.e., up-regulation of p21/WAF) and apoptotic cell death. Implantation of enhanced green fluorescent protein-transfected F98 glioma cells into slice cultures of rat brain confirmed the cytostatic effect of MS-275 without neurotoxic damage to the organotypic neuronal environment in a dose escalation up to 20 micromol/L. A single intratumoral injection of MS-275 7 days after orthotopic implantation of glioma cells in syngeneic rats confirmed the chemotherapeutic efficacy of MS-275 in vivo. Furthermore, its propensity to pass the blood-brain barrier and to increase the protein level of acetylated histone H3 in brain tissue identifies MS-275 as a promising candidate drug in the treatment of malignant gliomas.