MR Elastography Analysis of Glioma Stiffness and IDH1-Mutation Status.

BACKGROUND AND PURPOSE: Our aim was to noninvasively evaluate gliomas with MR elastography to characterize the relationship of tumor stiffness with tumor grade and mutations in the isocitrate dehydrogenase 1 (IDH1) gene. MATERIALS AND METHODS: Tumor stiffness properties were prospectively quantified in 18 patients (mean age, 42 years; 6 women) with histologically proved gliomas using MR elastography from 2014 to 2016. Images were acquired on a 3T MR imaging unit with a vibration frequency of 60 Hz. Tumor stiffness was compared with unaffected contralateral white matter, across tumor grade, and by IDH1-mutation status. The performance of the use of tumor stiffness to predict tumor grade and IDH1 mutation was evaluated with the Wilcoxon rank sum, 1-way ANOVA, and Tukey-Kramer tests. RESULTS: Gliomas were softer than healthy brain parenchyma, 2.2 kPa compared with 3.3 kPa (P < .001), with grade IV tumors softer than grade II. Tumors with an IDH1 mutation were significantly stiffer than those with wild type IDH1, 2.5 kPa versus 1.6 kPa, respectively (P = .007). CONCLUSIONS: MR elastography demonstrated that not only were gliomas softer than normal brain but the degree of softening was directly correlated with tumor grade and IDH1-mutation status. Noninvasive determination of tumor grade and IDH1 mutation may result in improved stratification of patients for different treatment options and the evaluation of novel therapeutics. This work reports on the emerging field of "mechanogenomics": the identification of genetic features such as IDH1 mutation using intrinsic biomechanical information.

Genetically Defined Oligodendroglioma Is Characterized by Indistinct Tumor Borders at MRI.

BACKGROUND AND PURPOSE: In 2016, the World Health Organization revised the brain tumor classification, making IDH mutation and 1p/19q codeletion the defining features of oligodendroglioma. To determine whether imaging characteristics previously associated with oligodendroglial tumors are still applicable, we evaluated the MR imaging features of genetically defined oligodendrogliomas. MATERIALS AND METHODS: One hundred forty-eight adult patients with untreated World Health Organization grade II and III infiltrating gliomas with histologic oligodendroglial morphology, known 1p/19q status, and at least 1 preoperative MR imaging were retrospectively identified. The association of 1p/19q codeletion with tumor imaging characteristics and ADC values was evaluated. RESULTS: Ninety of 148 (61%) patients had 1p/19q codeleted tumors, corresponding to genetically defined oligodendroglioma, and 58/148 (39%) did not show 1p/19q codeletion, corresponding to astrocytic tumors. Eighty-three of 90 (92%) genetically defined oligodendrogliomas had noncircumscribed borders, compared with 26/58 (45%) non-1p/19q codeleted tumors with at least partial histologic oligodendroglial morphology (P < .0001). Eighty-nine of 90 (99%) oligodendrogliomas were heterogeneous on T1- and/or T2-weighted imaging. In patients with available ADC values, a lower mean ADC value predicted 1p/19q codeletion (P = .0005). CONCLUSIONS: Imaging characteristics of World Health Organization 2016 genetically defined oligodendrogliomas differ from the previously considered characteristics of morphologically defined oligodendrogliomas. We found that genetically defined oligodendrogliomas were commonly poorly circumscribed and were almost always heterogeneous in signal intensity.

Induction and intracellular regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) mediated apotosis in human malignant glioma cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) preferentially triggers apoptosis in tumor cells versus normal cells, thus providing a therapeutic potential. In this study, we examined a large panel of human malignant glioma cell lines and primary cultures of normal human astrocytes for their sensitivity to TRAIL. Of 13 glioma cell lines, 3 were sensitive (80-100% death), 4 were partially resistant (30-79% death), and 6 were resistant (< 30% death). Normal astrocytes were also resistant. TRAIL-induced cell death was characterized by activation of caspase-8 and -3, poly(ADP-ribose) polymerase cleavage, and DNA fragmentation. Decoy receptor (DcR1 and DcR2) expression was limited in the glioma cell lines and did not correlate with TRAIL sensitivity. Both sensitive and resistant cell lines expressed TRAIL death receptor (DR5), adapter protein Fas-associated death domain (FADD), and caspase-8; but resistant cell lines expressed 2-fold higher levels of the apoptosis inhibitor phosphoprotein enriched in diabetes/phosphoprotein enriched in astrocytes-15 kDa (PED/PEA-15). In contrast, cellular FADD-like IL-1beta-converting enzyme-like inhibitory protein (cFLIP) expression was similar in sensitive and resistant cells. Transfection of sense PED/PEA-15 cDNA in sensitive cells resulted in cell resistance, whereas transfection of antisense in resistant cells rendered them sensitive. Inhibition of protein kinase C (PKC) activity restored TRAIL sensitivity in resistant cells, suggesting that PED/ PEA-15 function might be dependent on PKC-mediated phosphorylation. In summary, TRAIL induces apoptosis in > 50% of glioma cell lines, and this killing occurs through activation of the DR pathway. This caspase-8-induced apoptotic cascade is regulated by intracellular PED/PEA-15, but not by cFLIP or decoy receptors. This pathway may be exploitable for glioma and possibly for other cancer therapies.

Gene gun transfection of human glioma and melanoma cell lines with genes encoding human IL-12 and GM-CSF.

We used particle-mediated gene transfer by a custom-built gene gun to transfect two well-established human glioma (D54MG and U251) and melanoma (SK mel 28 and Ed 141) cell lines, as well as two glioma lines locally established from primary patient tumors (Ed 147 and Ed 149). Using beta-galactosidase as a reporter gene, D54MG, U251, Ed 141 and SK mel 28 showed an average transfection efficiency of 15-40%, whereas Ed 147 and Ed 149 had mean transfection efficiencies of 3% and 5% respectively. Twenty-four hours after transfection with the gene encoding human interleukin-12 (IL-12), ELISA was performed on cell supernatants (mean of n = 12 for each cell line). IL-12 expression was extremely variable between the different cell lines, ranging from 52 to 1,151 pg/10(6) cells/24 h. Results were very similar when cells were exposed to 20,000 rads of gamma irradiation 2 h after transfection. When the cell lines were transfected with human granulocyte-macrophage colony-stimulating factor, 24 h levels were: 13.0 (Ed 147), 17.8 (Ed 149), 18.6 (Ed 141), 27.4 (D54MG) and 27.7 ng/10(6) cells/24h (U251). SK mel 28 produced 88.1 ng/10(6) cells/24 h. We conclude that the gene gun can efficiently transfect a variety of immortalized, well-established and locally-established glioma and melanoma cell lines. High dose gamma irradiation does not adversely affect the expression of the foreign gene (IL-12) at 24 h. Significantly, transfected cell lines show different levels of expression depending on the particular gene/plasmid introduced. Therefore, each cell line has to be assessed individually for the level of expression of each introduced gene.

Human glioma immunobiology in vitro: implications for immunogene therapy.

OBJECTIVE: Human gliomas are known to be immunosuppressive. Recent reports have suggested novel strategies to overcome this immunosuppression, including immunogene therapy. We examined expression of 10 immunologically important molecules by human gliomas in vitro, and we discuss the implications for immunogene therapy. METHODS: Early passage human glioma cultures and established human glioma cell lines were analyzed by flow cytometry for expression of Class I and II major histocompatibility complex (MHC), B7-2 (CD86), and Fas (CD95). Culture supernatants were assayed by enzyme-linked immunosorbent assay for interleukin (IL)-6, IL-10, IL-12, transforming growth factor beta2, prostaglandin E2, and granulocyte-macrophage colony-stimulating factor levels. RESULTS: All cultures (16 of 16 samples) expressed Class I MHC and Fas, but few expressed Class II MHC (1 of 16 samples) or B7-2 (0 of 16 samples). Nearly all expressed high levels of IL-6 (19 of 21 samples; mean, 36.5 +/- 10.8 ng/10(6) cells/d) and prostaglandin E2 (21 of 21 samples; mean, 15.6 +/- 4.5 ng/10(6) cells/d) levels, and many expressed transforming growth factor beta2 (13 of 21 samples; mean, 8.6 +/- 3.7 ng/10(6) cells/d). Although several cultures (6 of 14 samples) expressed granulocyte-macrophage colony-stimulating factor, expression levels were very low (mean, 0.2 +/- 0.1 ng/10(6) cells/d). Few cultures (4 of 21 samples) expressed measurable IL-10, and none (0 of 22 samples) expressed IL-12. CONCLUSION: Class I MHC and Fas expression suggests that human glioma cells may be susceptible to Class I MHC-dependent cytotoxic T cell recognition and Fas-mediated killing. Unfortunately, transforming growth factor beta2 and prostaglandin E2 probably impair T cell activation, and IL-6 may shift immunity to less effective humoral (T helper 2) responses. Proinflammatory gene expression (B7-2, granulocyte-macrophage colony-stimulating factor, and/or IL-12) is lacking. Together, these results suggest that modifying glioma cells via proinflammatory gene transfer or immunoinhibitory gene suppression might stimulate immune responses that are effective against unmodified tumors.

Glioma immunology and immunotherapy.

OBJECTIVE: Despite advances in conventional therapy, the prognosis for most glioma patients remains dismal. This has prompted an intensive search for effective treatment alternatives. Immunotherapy, one such alternative, has long been recognized as a potentially potent cancer treatment but has been limited by an inadequate understanding of the immune system. Now, increased insight into immunology is suggesting more rational approaches to immunotherapy. In this article, we explore key aspects of modern immunology and discuss their implications for glioma therapy. METHODS: A thorough literature review of glioma immunology and immunotherapy was undertaken to inquire into the basic immunology, central nervous system immunology, glioma immunobiology, standard glioma immunotherapy, and recent immunotherapeutic advances in glioma treatment. RESULTS: Although gliomas express tumor-associated antigens and appear potentially sensitive to immune responses, many factors work together to inhibit antiglioma immunity. Not surprisingly, most clinical attempts at glioma immunotherapy have met with little success to date. However, novel immunostimulatory strategies, such as immunogene therapy, directed cytokine delivery, and dendritic cell manipulation, have recently yielded dramatic preclinical results in glioma models. This suggests that glioma-derived immunosuppression can be overcome. CONCLUSION: Modern molecular biology and immunology techniques have yielded a wealth of new data about glioma immunobiology. Armed with this information, many investigators have proposed novel means to stimulate antiglioma immune responses. Although definitive clinical results remain to be seen, the current renaissance in glioma immunology and immunotherapy shows great promise for the future.

Improved technique for establishing short term human brain tumor cultures.

Culturing human central nervous system tumors has been difficult compared to other neoplasms. We report improved success rates for establishing short term human brain tumor cultures using a modified tissue processing technique. Eighty-seven brain tumor specimens (56 glioblastomas, 8 mid grade astrocytomas, 8 oligodendrogliomas, 15 other) were obtained from June 1988 to March 1997. The first twenty-three samples were processed by dissection, partial enzyme dissociation, and filtration through a tissue culture sieve. Subsequent samples were processed identically except tumor cells were centrifuged on a density gradient prior to plating. Successful cultures were defined as those surviving greater than three passages in tissue culture and growing to sufficient numbers (>10(6) cells) to allow freezing. Success rate was 42% (10/23) using standard processing methods and 86% (55/64) with the addition of density gradient centrifugation. Glial fibrillary acidic protein (GFAP) and vimentin staining, karyotypes, and growth curves were obtained for representative glioma cultures. All cultures tested were positive for vimentin (29/29) while 62% (18/29) were positive for GFAP. Of four cultures karyotyped (two glioblastomas, two oligodendrogliomas), all but one oligodendroglioma culture exhibited clonal cytogenetic abnormalities. These immunohistochemical and karyotypic results are consistent with the malignant glial origin of these cells. Of note, low passage human glioma cultures grew slower and exhibited more contact inhibition than immortalized human glioblastoma cell lines. Nevertheless, this simple method for establishing short term human brain tumor cultures should aid in further developing human brain tumor pre-clinical models as well as enhancing clinical applications dependent on in vitro human brain tumor cell growth adjust.

"Idiopathic" cranial hypertrophic pachymeningitis responsive to antituberculous therapy: case report.

OBJECTIVE AND IMPORTANCE: We present a case similar to previously described cases of idiopathic hypertrophic cranial pachymeningitis. However, our patient responded to antituberculous therapy. This raises the possibility that some cases of "idiopathic" hypertrophic cranial pachymeningitis may represent occult tuberculous disease. CLINICAL PRESENTATION: A 55-year-old woman presented with a right fourth nerve palsy and a 5-month history of headaches. Magnetic resonance imaging with gadolinium revealed thick enhancing dura on the right half of the tentorium cerebelli, with edema of the adjacent midbrain, pons, and cerebral peduncle. INTERVENTION: Open biopsy of the tentorial lesion revealed only dense fibrosis with histiocytic infiltration. An exhaustive search failed to demonstrate an underlying cause. In particular, mycobacterial stains/cultures were negative, there was no granuloma formation, and the chest x-ray was unremarkable. However, because of a strongly positive purified protein-derivative skin test and residence in an area endemic for tuberculosis, the patient was placed on antituberculous medications. CONCLUSION: The patient's symptoms and signs resolved with antituberculous therapy. Resolution of the tentorial lesion was confirmed by gadolinium-enhanced magnetic resonance imaging. We conclude that this case represented occult tuberculous disease. An empiric trial of antituberculous therapy may be warranted in other cases of apparently idiopathic hypertrophic cranial pachymeningitis.

Granulocyte-macrophage colony-stimulating factor and B7-2 combination immunogene therapy in an allogeneic Hu-PBL-SCID/beige mouse-human glioblastoma multiforme model.

Glioblastoma multiforme is the most common primary central nervous system neoplasm. Its dismal prognosis has led to investigation of new treatment strategies such as immunogene therapy. We transduced the human glioblastoma cell line D54MG in vitro with genes encoding the proinflammatory cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF), the T cell co-stimulatory molecule B7-2, or both (in a bicistronic vector) via retroviral vectors. Therapeutic gene expression by D54MG was high after transduction and selection (30 ng/10(6) cells/day for GM-CSF and > 2 orders of magnitude fluorescence shift on flow cytometry for B7-2). The effect of GM-CSF and/or B7-2 transduction on D54MG tumor growth in vivo was monitored in a novel allogeneic human peripheral blood lymphocyte-severe combined immunodeficiency mouse (Hu-PBL-SCID) model. GM-CSF- or B7-2-transduced tumors showed growth suppression in hu-PBL-reconstituted mice compared to untransduced and/or unreconstituted controls. Growth suppression was greatest for B7-2. Furthermore, vaccination with irradiated GM-CSF/B7-2-transduced tumor cells markedly inhibited growth of wild-type tumors at distant sites. Thus, this study illustrates a potential gene therapy strategy for glioblastoma multiforme patients using GM-CSF and/or B7-2 transduced tumor vaccines. Although extension of these allogeneic studies to an autologous system is critical, this is the first demonstration of in vivo efficacy of combination GM-CSF and B7-2 immunogene therapy for human glioblastoma multiforme.