Geraniin suppresses tumor cell growth and triggers apoptosis in human glioma via inhibition of STAT3 signaling

Natural phytochemicals are attracting increasing interest as anticancer agents. The aim of this study is to evaluate the therapeutic potential of geraniin, a major ellagitannin extracted from Geranium sibiricum L., in human glioma. Human U87 and LN229 glioma cells were treated with different concentrations of geraniin, and cell viability, apoptosis, and gene expression were assessed. The involvement of STAT3 signaling in the action of geraniin was examined. We found that geraniin treatment for 48 h significantly (P < 0.05) impaired the phosphorylation of STAT3 and reduced the expression of downstream target genes Bcl-xL, Mcl-1, Bcl-2, and cyclin D1. Exposure to geraniin led to a concentration-dependent decline in cell viability and increase in apoptosis in glioma cells, but had no significant impact on the viability of normal human astrocytes. Measurement of caspase-3 activity showed that geraniin-treated U87 and LN229 cells showed a 1.8–2.5-fold higher caspase-3 activity than control cells. Overexpression of constitutively active STAT3 significantly (P < 0.05) reversed geraniin-mediated growth suppression and apoptosis, which was accompanied by restoration of Bcl-xL, Mcl-1, Bcl-2, and cyclin D1 expression. In an xenograft tumor mouse model, geraniin treatment significantly retarded tumor growth and induced apoptosis. Western blot analysis confirmed the suppression of STAT3 phosphorylation in glioma xenograft tumors by geraniin. Taken together, these data suggest that geraniin exerts growth-suppressive and pro-apoptotic effects on glioma cells via inhibition of STAT3 signaling and may have therapeutic benefits in malignant gliomas.     

Decreased expression of microRNA-200b is an independent unfavorable prognostic factor for glioma patients

Background and aimAs a member of the microRNA (miR)-200 family, miR-200b has been recognized as one of the fundamental regulators of epithelial–mesenchymal transition, chemosensitivity, cell proliferation, and cell cycle. Especially in glioma, miR-200b targets the CREB1 gene and suppresses the tumor cell growth in vitro. However, its involvement in human glioma has not yet been determined. The aim of this study was to investigate the clinical significance of miR-200b expression in this disease.MethodsmiR-200b expression in 266 pairs of human gliomas and matched nonneoplastic brain tissues was measured by real-time quantitative RT-PCR assay.ResultsCompared with nonneoplastic brain tissues, the expression level of miR-200b was significantly decreased in glioma tissues (tumor vs. normal: 2.87 ± 2.05 vs. 8.78 ± 2.50, P < 0.001). Of 266 patients with gliomas, 166 (62.41%) were in low miR-200b expression group. In addition, we found that the glioma tissues from high-grade tumors (grade III and IV) had much lower miR-200b expression than glioma tissues from low grade tumors (grade I and II). Moreover, the expression level of miR-200b was positively correlated with Karnofsky performance status (KPS) scores of glioma tissues. The results of a 60-month follow-up in 266 glioma patients further demonstrated that lower miR-200b expression was correlated with worse progression-free survival and overall survival in the patients with grade III and IV gliomas. Both univariate and multivariate analyses revealed that miR-200b was an independent prognostic indicator for glioma.ConclusionThese findings prove that the decreased expression of miR-200b may be associated with malignant tumor progression and poor prognosis in patients with gliomas, suggesting the potential role of miR-200b in glioma management. miR-200b may be a novel and valuable signature for predicting the clinical outcome of patients with gliomas.     

Characterizing Photothrombotic Distal Middle Cerebral Artery Occlusion and YAG Laser-Induced Reperfusion Model in the Izumo Strain of Spontaneously Hypertensive Rats

No study has systematically studied the relevance of original Izumo strain of spontaneously hypertensive rats (SHR/Izm) as a stroke model. Furthermore, both SHR/Izm and stroke-prone SHR/Izm (SHRSP/Izm) are commercially available, and recent progress in genetic studies allowed us to use several congenic strains of rats constructed with SHR/Izm and SHRSP/Izm as the genetic background strains. A total of 166 male SHR/Izm and 17 male SHRSP/Izm were subjected to photothrombotic middle cerebral artery (MCA) occlusion with or without YAG laser-induced reperfusion. The pattern of distal MCA was recorded. Infarct volumes were determined with 2,3,5-triphenyltetrazolium chloride. At 24 or 48 h after MCA occlusion, infarct volumes in the permanent occlusion and 2-h occlusion groups (88 ± 22 [SD] and 87 ± 25 mm³, respectively) were significantly larger than that in the 1-h occlusion group (45 ± 14 mm³), indicating the presence of sizeable zone of penumbra. Infarct size in SHRSP/Izm determined at 24 h after MCA occlusion was fairly large (124.0 ± 34.8 mm³, n = 10). Infarct volume in SHR/Izm with simple distal MCA was 76 ± 19 mm³, which was significantly smaller than 95 ± 22 mm³ in the other SHR/Izm with more branching MCA. These data suggest that this stroke model in SHR/Izm is useful in the preclinical testing of stroke therapies and elucidating the pathophysiology of cerebral ischemia/reperfusion.     

Intra-arterial administration of tumor-targeting Salmonella typhimurium A1-R regresses a cisplatin-resistant relapsed osteosarcoma in a patient-derived orthotopic xenograft (PDOX) mouse model

Previously, a patient-derived orthotopic xenograft (PDOX) model was established with a lung metastasis from an osteosarcoma patient which developed after adjuvant cisplatinum (CDDP) treatment. In this model, we previously demonstrated the efficacy of trabectedin (TRAB) and temozolomide (TEM) compared with CDDP. In the present report, osteosarcoma tissue was implanted orthotopically in the distal femur of mice which were randomized into the following groups when tumor volume reached approximately 100 mm³; On day 14 after initiation of treatment, all but CDDP significantly inhibited tumor volume growth compared with untreated controls. Control (G1): 793.7 ± 215.0 mm³; CDDP (G2): 588.1 ± 176.9 mm³; Salmonella typhimurium A1-R (S. typhimurium A1-R) intravenous (i.v.) (G3): 269.7 ± 72.7 mm³; S. typhimurium A1-R intra-arterial (i.a.) (G4): 70.2 ± 18.9 mm³ (CDDP: p = 0.056; S. typhimurium A1-R i.v.: p = 0.0001; S. typhimurium A1-R i.a.: p = 0.00003, all vs. untreated controls). i.a. administration of S. typhimurium A1-R was significantly more effective than either CDDP (p = 0.00007), or i.v. administration of S. typhimurium A1-R (p = 0.00007) and significantly regressed the tumor volume compared with day 0 (p = 0.001). The new model of i.a. administration of S. typhimurium A1-R has great promise for the treatment of recalcitrant osteosarcoma.     

The anti-tumor effects of the recombinant toxin protein rLj-RGD3 from Lampetra japonica on pancreatic carcinoma Panc-1 cells in nude mice

Recombinant Lampetra japonica RGD peptide (rLj-RGD3) is a soluble toxin protein with three RGD (Arg-Gly-Asp) motifs and a molecular weight of 13.5 kDa. The aim of this study was to investigate the effects and mechanisms of rLj-RGD3 on tumor growth and survival in pancreatic carcinoma Panc-1 cell-bearing mice. A Panc-1 human pancreatic carcinoma-bearing nude mouse model was successfully generated, and the animals were treated with different doses of rLj-RGD3 for 3 weeks. The volume and weight of the subcutaneous tumors, the survival of the nude mice, histopathological changes, the intratumoral MVD, the number of apoptotic Panc-1 cells, and apoptosis-related proteins and gene expressions were determined. rLj-RGD3 significantly decreased the tumor volumes and weights, and the maximum tumor volume and weight IR values were 53.2% (p < 0.001) and 55.9% (p < 0.001), respectively. The life expectancy of Panc-1-bearing nude mice treated with rLj-RGD3 was increased by 56.3% (p < 0.001). Meanwhile, rLj-RGD3 promoted the expression of Bax, caspase-3, and caspase-9 and inhibited Bcl-2 and VEGF expression. In addition, rLj-RGD3 did not change FAK, PI3K and Akt expression, but p-FAK, p-PI3K and p-Akt, levels were down-regulated. These results show that rLj-RGD3 induced potent anti-tumor activity in vivo and suppressed the growth of transplanted Panc-1 cells in a nude mouse model, implying that rLj-RGD3 may serve as a potent clinical therapeutic agent for human pancreatic carcinoma.     

Up-regulation of vimentin expression in low-density malignant glioma cells as immediate and late effects under irradiation and temozolomide treatment

Nervous system tumors are one of the leading causes of cancer related death. Specific mechanisms facilitating the invasive behavior of gliomas remain obscure. Advanced simulation models of the in vivo response to therapy conditions should potentially improve malignant glioma treatment. Expressional profiling of vimentin––one of reliable pro-invasive tumor makers––in those simulation models was the goal of this study, in order to estimate a pro-invasive response of surviving malignant glioma cells under clinically relevant therapeutic conditions. Human U87-MG malignant glioma cells were used. These cells are characterized by the wild p53-phenotype, which is relevant for the majority of primary malignant glioblastomas. Experimental design foresaw the cells to undergo either irradiation or chemo-treatment with temozolomide alone, or combined treatment. Expression profiling of vimentin was performed by quantitative “Real-Time”-PCR under all treatment conditions simulating diverse tumor regions. Here we demonstrated that vimentin expression patterns in human malignant glioma cells strongly depend on cellular density, algorithms of drug delivery and chemo/radio treatment. Substantial differences were recognized between immediate and late therapy effects. Significant increase in vimentin expression levels was detected particularly in low-density cell cultures under durable treatment with constant concentration levels of temezolomide. Simulation of variable intratumoral regional conditions (central intratumoral regions vs. disseminated malignant cells in peripheral regions) demonstrated differential response of vimentin expression in malignant glioma cell cultures treated under clinically relevant conditions. Slight ebbing of expression levels as late effects of the treatment in confluent cultures may correspond to necrotic processes clinically observed in central intratumoral regions. Contrary, in disseminated malignant cells of peripheral regions therapy resulted in vimentin-inducing effects. This is in agreement with the clinical observations of an increased aggressiveness and malignancy grade of post-operatively chemo/radio-treated malignant gliomas.     

Spatiotemporal Optoacoustic Mapping of Tumor Hemodynamics in a Clinically Relevant Orthotopic Rabbit Model of Head and Neck Cancer

The purpose of this study was to investigate the usefulness of photoacoustic imaging (PAI) for spatiotemporal mapping of tumor hemodynamics in a rabbit model of head and neck carcinoma. Shope cottontail rabbit papilloma virus associated VX2 carcinomas were established in adult male New Zealand White rabbits (n = 9) by surgical transplantation of tumor tissue in the neck. Noninvasive PAI with co-registered ultrasound (US) was performed to longitudinally monitor tumor growth, oxygen saturation (%sO₂), and hemoglobin concentration (HbT). PAI findings were validated with Doppler sonography measures of percent vascularity (PV). Differences in tumor volumes, %sO₂, HbT, and PV values over time were analyzed using repeated-measures analysis of variance with multiple comparisons. Two-tailed Spearman correlation analysis was performed to determine the correlation coefficient (r) for comparisons between %sO₂, HbT, and tumor volume. US revealed a significant (P < .0001) increase in tumor volume over the 3-week period from 549 ± 260 mm³ on day 7 to 5055 ± 438 mm³ at 21 days postimplantation. Consistent with this aggressive tumor growth, PAI revealed a significant (P < .05) and progressive reduction in %sO₂ from day 7 (37.6 ± 7.4%) to day 21 (9.5 ± 2.1%). Corresponding Doppler images also showed a decrease in PV over time. PAI revealed considerable intratumoral spatial heterogeneity with the tumor rim showing two- to three-fold higher %sO₂ values compared to the core. Noninvasive PAI based on endogenous contrast provides a label-free method for longitudinal monitoring of temporal changes and spatial heterogeneity in thick head and neck tumors.     

Pilot feasibility study of in vivo intraoperative quantitative optical coherence tomography of human brain tissue during glioma resection

This study investigates the feasibility of in vivo quantitative optical coherence tomography (OCT) of human brain tissue during glioma resection surgery in six patients. High‐resolution detection of glioma tissue may allow precise and thorough tumor resection while preserving functional brain areas, and improving overall survival. In this study, in vivo 3D OCT datasets were collected during standard surgical procedure, before and after partial resection of the tumor, both from glioma tissue and normal parenchyma. Subsequently, the attenuation coefficient was extracted from the OCT datasets using an automated and validated algorithm. The cortical measurements yield a mean attenuation coefficient of 3.8 ± 1.2 mm^?1 for normal brain tissue and 3.6 ± 1.1 mm^?1 for glioma tissue. The subcortical measurements yield a mean attenuation coefficient of 5.7 ± 2.1 and 4.5 ± 1.6 mm^?1 for, respectively, normal brain tissue and glioma. Although the results are inconclusive with respect to trends in attenuation coefficient between normal and glioma tissue due to the small sample size, the results are in the range of previously reported values. Therefore, we conclude that the proposed method for quantitative in vivo OCT of human brain tissue is feasible during glioma resection surgery.     

Gamma-Secretase Represents a Therapeutic Target for the Treatment of Invasive Glioma Mediated by the p75 Neurotrophin Receptor

The multifunctional signaling protein p75 neurotrophin receptor (p75^NTR) is a central regulator and major contributor to the highly invasive nature of malignant gliomas. Here, we show that neurotrophin-dependent regulated intramembrane proteolysis (RIP) of p75^NTR is required for p75^NTR-mediated glioma invasion, and identify a previously unnamed process for targeted glioma therapy. Expression of cleavage-resistant chimeras of p75^NTR or treatment of animals bearing p75^NTR-positive intracranial tumors with clinically applicable γ-secretase inhibitors resulted in dramatically decreased glioma invasion and prolonged survival. Importantly, proteolytic processing of p75^NTR was observed in p75^NTR-positive patient tumor specimens and brain tumor initiating cells. This work highlights the importance of p75^NTR as a therapeutic target, suggesting that γ-secretase inhibitors may have direct clinical application for the treatment of malignant glioma.     

Effect of human interferon β gene transfer upon human glioma, transplanted into nude mouse brain, involves induced natural killer cells

We investigated the immunological responses induced by human interferon β (IFNβ) gene transfer in human gliomas produced in the brains of nude mice. A suspension of human glioma U251-SP cells was injected into the brains of nude mice. The IFNβ gene was transferred by intratumoral injection with cationic liposomes or cationic liposomes associated with anti-glioma monoclonal antibody (immunoliposomes). When intratumoral injection of liposomes or immunoliposomes containing the human IFNβ gene was performed every second day for a total of six injections, starting 7 days after tumor transplantation, complete disappearance of the tumor was observed in six of seven mice that had received liposomes and in all seven mice receiving immunoliposomes. In addition, experimental gliomas injected with immunoliposomes were much smaller than those injected with ordinary liposomes following delayed injections beginning 14 days after transplantation. An immunohistochemical study of the treated nude mouse brains revealed a remarkable induction of natural killer (NK) cells expressing asialoGM1 antigen. To investigate the significance of NK cells in the antitumor effect, we injected liposomes or immunoliposomes containing the human IFNβ gene into tumors in nude mice depleted of NK cells by irradiation and anti-asialoGM1 antibody administration. The antitumor effect of the liposomes or immunoliposomes was abolished. Subsequent subcutaneous glioma challenge of the nude mice after intracerebral tumor implantation and gene transfer resulted in no subcutaneous tumor growth. These results suggest that the induction of NK cells is important in the cytocidal effect of liposomes or immunoliposomes containing the human IFNβ gene upon experimental gliomas. Received: 10 February 1998 / Accepted: 1 September 1998     

Synthetic glycolipids for glioma growth inhibition developed from neurostatin and NF115 compound

Neurostatin, a natural glycosphingolipid, and NF115, a synthetic glycolipid, are inhibitors of glioma growth. While neurostatin shows high inhibitory activity on gliomas its abundance is low in mammalian brain. On the contrary NF115 exhibits less inhibitory activity on gliomas, but could be prepared by chemical synthesis. In this study we describe synthetic compounds, structurally related to NF115, capable of inhibiting glioma growth at low micromolar range. We used DNA microarray technology to compare the genes inhibited in U373-MG human glioma cells after treatment with the natural or synthetic inhibitor. New synthetic compounds were developed to interact with the product of Rho GDP dissociation inhibitor alpha gene, which was repressed in both treatments. Compounds that were inhibitors of glioma cell growth in assays for [3H]-thymidine incorporation were then injected in C6 tumor bearing rats and the tumor size in each animal group were measured. The GC-17, GC-4 and IG-5 are new compounds derived from NF115 and showed high antiproliferative activity on tumor cell lines. The GC-17 compound inhibited U373-MG glioblastoma cells (3.2 μM), the effects was fifty times more potent than NF115, and caused a significant reduction of tumor volume (P < 0.05) when tested in Wistar rats allotransplanted with C6 glioma cells.     

Intratumoral Delivery of Paclitaxel in Solid Tumor from Biodegradable Hyaluronan Nanoparticle Formulations

In the current study, novel paclitaxel-loaded cross-linked hyaluronan nanoparticles were engineered for the local delivery of paclitaxel as a prototype drug for cancer therapy. The nanoparticles were prepared using a desolvation method with polymer cross-linking. In vitro cytotoxicity studies demonstrated that less than 75% of the MDA-MB-231 and ZR-75-1 breast cancer cells were viable after 2-day exposure to paclitaxel-loaded hyaluronan nanoparticles or free paclitaxel, regardless of the dose. These results suggest that hyaluronan nanoparticles maintain the pharmacological activity of paclitaxel and efficiently deliver it to the cells. Furthermore, in vivo administration of the drug-loaded nanoparticles via direct intratumoral injection to 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumor in female rats was studied. The paclitaxel-loaded nanoparticles treated group showed effective inhibition of tumor growth in all treated rats. Interestingly, there was one case of complete remission of tumor nodule and two cases of persistent reduction of tumor size that was observed on subsequent days. In the case of free paclitaxel-treated group, the mean tumor volume increased almost linearly (R ² = 0.93) with time to a size that was 4.9-fold larger than the baseline volume at 57 days post-drug administration. Intratumoral administration of paclitaxel-loaded hyaluronan nanoparticles could be a promising treatment modality for solid mammary tumors.     

Effects of Lanreotide Autogel on Growth Hormone,Insulinlike Growth Factor 1, and Tumor Size in Acromegaly: a 1-Year Prospective Multicenter Study

ObjectiveTo evaluate the safety and effectiveness of lanreotide Autogel on growth hormone and insulinlike growth factor 1 (IGF-1) concentrations and tumor size in patients with acromegaly.MethodsBetween September 2004 and March 2006, patients with active acromegaly who had not previously been treated with somatostatin analogues or received irradiation were enrolled in a 1-year, prospective, open, multicenter study. Lanreotide Autogel was injected subcutaneously starting with 90 mg every 4 weeks for 2 cycles and then individually titrated, aiming for safe growth hormone concentrations (< 2.5 ng/mL) and normal age-matched IGF-1 concentrations. Tumor shrinkage, clinical score, pituitary function, and safety parameters were evaluated.ResultsTwenty-seven patients (15 women, 12 men) were enrolled. One patient withdrew because of treatment intolerance, and 5 proceeded to neurosurgery 6 months into the study. Lanreotide Autogel was the primary treatment in 19 patients (4 with microadenoma, 15 with macroadenoma) and the adjuvant treatment in 8 patients in whom it followed a previous unsuccessful neurosurgery. In the 26 patients, safe growth hormone values were achieved in 11 (42%), normal IGF-1 values in 14 (54%), and both targets were achieved in 10 (38%). Tumors shrank in 16 of the 22 patients (73%) in whom tumor shrinkage could be evaluated. The maximal vertical diameter of the tumor decreased by a mean of 24% (range, 0% to 50%), from 14.4 ± 8.4 mm to 10.4 ± 7 mm, and tumor volume decreased by a mean of 44% (range, 0% to 76%), from 2536 mm³ (range, 115-7737 mm³) to 1461 mm³ (range, 63-6217 mm³) (both P < .015). Symptom scores and lipid levels significantly improved. In the 26 patients, glucose metabolism deteriorated in 3 (12%) and improved in 4 (15%). New biliary alterations appeared in 26%. Pituitary function and safety parameters did not change.ConclusionsLanreotide Autogel treatment, titrated for optimal hormonal control, effectively controls IGF-1 and growth hormone levels, shrinks tumors, reduces acromegalic symptoms, and is well tolerated. (Endocr Pract. 2008;14:846-855)     

Clonal Analysis in Recurrent Astrocytic, Oligoastrocytic and Oligodendroglial Tumors Implicates IDH1- Mutation as Common Tumor Initiating Event

Background

To investigate the dynamics of inter- and intratumoral molecular alterations during tumor progression in recurrent gliomas.

Methodology/Principal Findings

To address intertumoral heterogeneity we investigated non- microdissected tumor tissue of 106 gliomas representing 51 recurrent tumors. To address intratumoral heterogeneity a set of 16 gliomas representing 7 tumor pairs with at least one recurrence, and 4 single mixed gliomas were investigated by microdissection of distinct oligodendroglial and astrocytic tumor components. All tumors and tumor components were analyzed for allelic loss of 1p/19q (LOH 1p/19q), for TP53- mutations and for R132 mutations in the IDH1 gene. The investigation of non- microdissected tumor tissue revealed clonality in 75% (38/51). Aberrant molecular alterations upon recurrence were detected in 25% (13/51). 64% (9/14) of these were novel and associated with tumor progression. Loss of previously detected alterations was observed in 36% (5/14). One tumor pair (1/14; 7%) was significant for both. Intratumoral clonality was detected in 57% (4/7) of the microdissected tumor pairs and in 75% (3/4) of single microdissected tumors. 43% (3/7) of tumor pairs and one single tumor (25%) revealed intratumoral heterogeneity. While intratumoral heterogeneity affected both the TP53- mutational status and the LOH1p/19q status, all tumors with intratumoral heterogeneity shared the R132 IDH1- mutation as a common feature in both their microdissected components.

Conclusions/Significance

The majority of recurrent gliomas are of monoclonal origin. However, the detection of divertive tumor cell clones in morphological distinct tumor components sharing IDH1- mutations as early event may provide insight into the tumorigenesis of true mixed gliomas.     

Oviduct binding ability of porcine spermatozoa develops in the epididymis and can be advanced by incubation with caudal fluid

The sperm reservoir is formed when spermatozoa bind to the epithelium of the uterotubal junction and caudal isthmus of the oviduct. It is an important mechanism that helps synchronize the meeting of gametes by regulating untimely capacitation and polyspermic fertilization. This study investigated the influence of epididymal maturation and caudal fluid on the ability of spermatozoa to bind to oviduct epithelium using a model porcine oviduct explant assay. Spermatozoa from the rete testis, middle caput (E2-E3), middle corpus (E6), and cauda (E8) of Large White or Large White × Landrace boars aged 10 to 14 months were diluted in modified Androhep solution and incubated with porcine oviduct explants. Results reported in this study support our hypothesis that testicular spermatozoa need to pass through the regions of the epididymis to acquire the ability to bind to the oviduct. There was a sequential increase in the number of spermatozoa that bound to oviduct explants from the rete testis to caudal epididymis. Binding of caudal spermatozoa to isthmic explants was the highest (15.0 ± 1.2 spermatozoa per 1.25 mm², mean ± standard error of the mean; P ≤ 0.05) and lowest by spermatozoa from the rete testis (2.0 ± 0.3 per 1.25 mm²), and higher to isthmus from sows compared to gilts (35.8 ± 6.7 per 1.25 mm² vs. 14.8 ± 3.0 per 1.25 mm²; P ≤ 0.05). Binding of ejaculated spermatozoa to porcine isthmus was higher than that for caudal spermatozoa (26.3 ± 1.4 per 1.25 mm² vs. 15.0 ± 0.8 per 1.25 mm²; P ≤ 0.05) and higher to porcine than to bovine isthmus (26.3 ± 2.3 per 1.25 mm² vs. 18.8 ± 1.9 per 1.25 mm²; P ≤ 0.05). Incubation of spermatozoa from the caput and corpus in caudal fluid increased the ability of spermatozoa to bind to the oviduct epithelium (P ≤ 0.05). In conclusion, the capacity of testicular spermatozoa to bind to the oviduct epithelium increases during their maturation in the epididymis and can be advanced by components of the caudal fluid.     

Preclinical TSPO Ligand PET to Visualize Human Glioma Xenotransplants: A Preliminary Study

Current positron emission tomography (PET) imaging biomarkers for detection of infiltrating gliomas are limited. Translocator protein (TSPO) is a novel and promising biomarker for glioma PET imaging. To validate TSPO as a potential target for molecular imaging of glioma, TSPO expression was assayed in a tumor microarray containing 37 high-grade (III, IV) gliomas. TSPO staining was detected in all tumor specimens. Subsequently, PET imaging was performed with an aryloxyanilide-based TSPO ligand, [¹⁸F]PBR06, in primary orthotopic xenograft models of WHO grade III and IV gliomas. Selective uptake of [¹⁸F]PBR06 in engrafted tumor was measured. Furthermore, PET imaging with [¹⁸F]PBR06 demonstrated infiltrative glioma growth that was undetectable by traditional magnetic resonance imaging (MRI). Preliminary PET with [¹⁸F]PBR06 demonstrated a preferential tumor-to-normal background ratio in comparison to 2-deoxy-2-[¹⁸F]fluoro-D-glucose ([¹⁸F]FDG). These results suggest that TSPO PET imaging with such high-affinity radiotracers may represent a novel strategy to characterize distinct molecular features of glioma growth, as well as better define the extent of glioma infiltration for therapeutic purposes.     

Endothelial cell pseudopods and angiogenesis of breast cancer tumors

Background  

A neoplastic tumor cannot grow beyond a millimeter or so in diameter without recruitment of endothelial cells and new blood vessels to supply nutrition and oxygen for tumor cell survival. This study was designed to investigate formation of new blood vessels within a human growing breast cancer tumor model (MDA MB231 in mammary fat pad of nude female mouse). Once the tumor grew to 35 mm³, it developed a well-vascularized capsule. Histological sections of tumors greater than 35 mm³ were stained with PAS, with CD-31 antibody (an endothelial cell maker), or with hypoxia inducible factor 1α antibody (HIF). The extent of blood vessel and endothelial cell pseudopod volume density was measured by ocular grid intercept counting in the PAS stained slides.     

Topotecan enhances immune clearance of gliomas

Despite aggressive surgery, radiation therapy, and chemotherapy, glioblastoma multiforme (GBM) is refractory to therapy, recurs quickly, and results in a median survival time of only 14 months. The modulation of the apoptotic receptor Fas with cytotoxic agents could potentiate the response to therapy. However, Fas ligand (FasL) is not expressed in the brain and therefore this Fas-inducing cell death mechanism cannot be utilized. Vaccination of patients with gliomas has shown promising responses. In animal studies, brain tumors of vaccinated mice were infiltrated with activated T cells. Since activated immune cells express FasL, we hypothesized that combination of immunotherapy with chemotherapy can activate Fas signaling, which could be responsible for a synergistic or additive effect of the combination. When we treated the human glioma cell line U-87 and GBM tumor cells isolated from patients with TPT, Fas was up regulated. Subsequent administration of soluble Fas ligand (sFasL) to treated cells significantly increased their cell death indicating that these Fas receptors were functional. Similar effect was observed when CD3⁺ T cells were used as a source of the FasL, indicating that the up regulated Fas expression on glioma cells increases their susceptibility to cytotoxic T cell killing. This additive effect was not observed when glioma cells were pre-treated with temozolomide, which was unable to increase Fas expression in tumor. Inhibition of FasL activity with the antagonistic antibody Nok-1 mitigated these effects confirming that these responses were specifically mediated by the Fas-FasL interaction. Furthermore, the CD3⁺ T cells co-cultured with topotecan treated U-87 and autologous GBM tumor cells showed a significant increase in expression in IFN-γ, a key cytokine produced by activated T cells, and accordingly enhanced tumor cytotoxicity. Based on our data we conclude that drugs, such as topotecan, which cause up regulation of Fas on glioma cells can be potentially exploited with immunotherapy to enhance immune clearance of tumors via Fas signaling. Jun Wei and Guillermo DeAngulo are Co-lead authors.     

Preparation of cross-linked carboxymethyl chitosan for repairing sciatic nerve injury in rats

A successful nerve regeneration process was achieved with nerve repair tubes made up of 1-ethyl-3(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) cross-linked carboxymethyl chitosan (CM-chitosan) with improved biodegradability. Chitosan has a very slow degradation rate, while the EDC cross-linked CM-chitosan tubes degraded to 30% of original weight during 8 weeks of incubation in lysozyme solution. In vitro cell culture indicated that the CM-chitosan films presented no cytotoxicity to Schwann cells. From in vivo studies using a 10 mm rat sciatic nerve defect model investigated by histomorphometry analysis, the average diameter of the fibers and the average thickness of myelin sheath in the CM-chitosan tubes were 3.7 ± 0.33 and 0.33 ± 0.04 μm, respectively, which demonstrated equivalence to nerve autografts (the current “gold” standard); furthermore, the average fiber density in the CM-chitosan tubes was 20.5 × 10³/mm², which was similar to that of autografts (21 × 10³/mm²) and significantly higher than that of common chitosan tubes (15.3 × 10³/mm²).