Characterization of the blood brain barrier in pediatric central nervous system neoplasms

OBJECTIVE: The normal blood-brain barrier (BBB) is composed of tight junctions between endothelial cells and surrounding astrocyte foot processes. Breakdown of the physiological astrocyte-endothelial cell relationship occurs in adult metastatic and primary brain tumors. However, the astrocyte-endothelial cell relationship has not been studied in pediatric tumors. MATERIALS AND METHODS: Utilizing specimens from cases of pilocytic astrocytoma (n = 5), medulloblastoma (n = 5), and low-grade diffuse astrocytoma (n = 1), immunofluorescence were performed using primary antibodies against CD31, glial fibrillary acidic protein (GFAP), and aquaporin 4 (AQ4). Clinical, magnetic resonance imaging, operative, and histopathological findings were analyzed. RESULTS: Strongly-enhancing areas of medulloblastoma exhibited complete BBB breakdown with sparse GFAP and AQ4 staining around CD31-positive vessels. Moderately enhancing regions of pilocytic astrocytomas exhibited regions of intact BBB and vasculature surrounded by dense GFAP staining but reduced and disorganized AQ4 staining, suggesting tumor cells could not fulfill physiological BBB support. Non-enhancing low-grade diffuse astrocytoma demonstrated intact BBB with intense peri-microvasculature GFAP and AQ4 staining. AQ4 stained so strongly that AQ4 visualization alone delineated CD31-positive vessels. CONCLUSION: Taken together, BBB breakdown in pediatric tumors corresponds to a loss of normal endothelial cell-astrocyte foot process relationships. Further development of pharmaceutical agents capitalizing on this disrupted BBB is warranted in medulloblastoma and pilocytic astrocytoma. However, BBB integrity remains a challenge in treating low-grade diffuse astrocytoma before progression toward secondary glioblastoma

Cells isolated from residual intracranial tumors after treatment express iPSC genes and possess neural lineage differentiation plasticityResearch in context

Background: The goal of this study is to identify and characterize treatment resistant tumor initiating cells (TRTICs) using orthotopic xenografts. Methods: TRTICs were enriched from GBM cell lines using mouse xenografts treated with fractionated doses of radiation and temozolomide. TRTICs were characterized by neurosphere clonogenicity and self-renewal, serial xenotransplantation, differentiation potential, and mRNA & miRNA transcriptomic profiling. We use an unbiased approach to identify antigens encoding TRTIC and glioma stem cells (GSC) populations. Co-culture experiments of TRTIC and differentiated cells were conducted to evaluate the reliance of TRTIC differentiation on the secretome of differentiated cells. Findings: TRTICs acquire stem-like gene expression signatures and increased side population staining resulting from the activation of multi-drug resistance genes. Genetic and functional characterization of TRTICs shows a striking resemblance with GSCs. TRTICs can differentiate towards specific progeny in the neural stem cell lineage. TRTIC-derived tumors display all the histological hallmarks of glioblastoma (GBM) and exhibit a miRNA-transcript and mRNA-transcriptomic profile associated with aggressiveness. We report that CD24+/CD44+ antigens are expressed in TRTICs and patient-derived GSCs. Double positive CD24+/CD44+ exhibit treatment resistance and enhanced tumorigenicity. Interestingly, co-culture experiments with TRTICs and differentiated cells indicated that the regulation of TRTIC differentiation could rely on the secretome in the tumor niche. Interpretation: Radiation and temozolomide treatment enriches a population of cells that have increased iPSC gene expression. As few as 500 cells produced aggressive intracranial tumors resembling patient GBM. CD24+/CD44+ antigens are increased in TRTICs and patient-derived GSCs. The enrichment for TRTICs may result in part from the secretome of differentiated cells. Fund: NIH/NCI 1RC2CA148190, 1R01CA108633, 1R01CA188228, and The Ohio State University Comprehensive Cancer Center. Keywords: Treatment-resistance, Tumor-initiating, Glioma stem cell, CD24high/CD44high, Transcriptome, Neural lineag

Preparation, Biodistribution and Neurotoxicity of Liposomal Cisplatin following Convection Enhanced Delivery in Normal and F98 Glioma Bearing Rats

The purpose of this study was to evaluate two novel liposomal formulations of cisplatin as potential therapeutic agents for treatment of the F98 rat glioma. The first was a commercially produced agent, Lipoplatin TM, which currently is in a Phase III clinical trial for treatment of non-small cell lung cancer (NSCLC). The second, produced in our laboratory, was based on the ability of cisplatin to form coordination complexes with lipid cholesteryl hemisuccinate (CHEMS). The in vitro tumoricidal activity of the former previously has been described in detail by other investigators. The CHEMS liposomal formulation had a Pt loading efficiency of 25 % and showed more potent in vitro cytotoxicity against F98 glioma cells than free cisplatin at 24 h. In vivo CHEMS liposomes showed high retention at 24 h after intracerebral (i.c.) convection enhanced delivery (CED) to F98 glioma bearing rats. Neurotoxicologic studies were carried out in non-tumor bearing Fischer rats following i.c. CED of Lipoplatin TM or CHEMS liposomes or their ‘‘hollow’ ’ counterparts. Unexpectedly, Lipoplatin TM was highly neurotoxic when given i.c. by CED and resulted in death immediately following or within a few days after administration. Similarly ‘‘hollow’’ Lipoplatin TM liposomes showed similar neurotoxicity indicating that this was due to the liposomes themselves rather than the cisplatin. This was particularly surprising since Lipoplatin TM has been well tolerated when administered intravenously. In contrast, CHEMS liposomes and their ‘‘hollow’ ’ counterparts were clinically well tolerated. However, a variety of dos

Potential Utility of FLAIR in MRI-negative Cushing\u27s Disease.

OBJECTIVE Accurate presurgical localization of microadenomas in Cushings disease (CD) leads to improved remission rates and decreased adverse events. Volumetric gradient recalled echo (3D-GRE) MRI detects pituitary microadenomas in CD in up to 50%-80% cases as a focus of hypointensity due to delayed contrast wash-in. The authors have previously reported that postcontrast FLAIR imaging may be useful in detecting otherwise MRI-negative pituitary microadenomas as foci of hyperintensity. This reflects theoretically complementary imaging of microadenomas due to delayed contrast washout. The authors report on the diagnostic accuracy and clinical utility of FLAIR imaging in the detection of microadenomas in patients with CD. METHODS The authors prospectively analyzed imaging findings in 23 patients (24 tumors) with biochemically proven CD who underwent transsphenoidal surgery for CD. Preoperatively, the patients underwent pituitary MRI with postcontrast FLAIR and postcontrast 3D-GRE sequences. RESULTS Postcontrast FLAIR hyperintensity was detected in macroadenomas, and in 3D-GRE-positive or -negative microadenomas. Overall, 3D-GRE was superior in detecting surgically and histopathologically confirmed, location-concordant microadenomas. Of 24 pituitary adenomas, 18 (75%; sensitivity 82%, positive predictive value 95%) were found on 3D-GRE, and 13 (50% [1 was false positive]; sensitivity 55%, positive predictive value 92%) were correctly identified on FLAIR. The stand-alone specificity of 3D-GRE and FLAIR was similar (50%). These results confirm the superiority of 3D-GRE as a stand-alone imaging modality. The authors then tested the utility of FLAIR as a complementary tool to 3D-GRE imaging. All 5 patients with negative 3D-GRE MRI displayed a distinct focus of FLAIR enhancement. Four of those 5 cases (80%) had location-concordant positive histopathological results and achieved postsurgical biochemical remission. The remaining patient was not cured, because resection did not include the region of FLAIR hyperintensity. CONCLUSIONS This study suggests that delayed microadenoma contrast washout may be detected as FLAIR hyperintensity in otherwise MRI-negative CD cases. The authors propose adding postcontrast FLAIR sequences to complement 3D-GRE for surgical planning in patients with CD. Clinical trial registration no.: NIH protocol 03-N-0164, NCT00060541 (clinicaltrials.gov)

The antitumor effects of IFN-α are abrogated in a STAT1-deficient mouse

IFN-α activates the signal transducer and activator of transcription (STAT) family of proteins; however, it is unknown whether IFN-α exerts its antitumor actions primarily through a direct effect on malignant cells or by stimulating the immune system. To investigate the contribution of STAT1 signaling within the tumor, we generated a STAT1-deficient melanoma cell line, AGS-1. We reconstituted STAT1 into AGS-1 cells by retroviral gene transfer. The resulting cell line (AGS-1(STAT1)) showed normal regulation of IFN-α–stimulated genes (e.g., H2k, ISG-54) as compared with AGS-1 cells infected with the empty vector (AGS-1(MSCV)). However, mice challenged with the AGS-1, AGS-1(STAT1), and AGS-1(MSCV) cell lines exhibited nearly identical survival in response to IFN-α treatment, indicating that restored STAT1 signaling within the tumor did not augment the antitumor activity of IFN-α. In contrast, STAT1(–/–) mice could not utilize exogenous IFN-α to inhibit the growth of STAT1(+/+) melanoma cells in either an intraperitoneal tumor model or in the adjuvant setting. The survival of tumor-bearing STAT1(–/–) mice was identical regardless of treatment (IFN-α or PBS). Additional cell depletion studies demonstrated that NK cells mediated the antitumor effects of IFN-α. Thus, STAT1-mediated gene regulation within immune effectors was necessary for mediating the antitumor effects of IFN-α in this experimental system

Clonogenic survival of F98 glioma cells following treatment with either free cisplatin or liposomal cisplatin for 4 or 24 h.

<p>Surviving fractions (S.Fs) were determined for the F98 glioma cells treated with (A) CHEMS lipsomes, (B) free cisplatin following a 4 h (•) or 24 h (○) exposure. Each data point represents the mean of 3 replicates ± the standard deviation.</p