Dynamic Near-Infrared Optical Imaging of 2-Deoxyglucose Uptake by Intracranial Glioma of Athymic Mice

BACKGROUND: It is recognized that cancer cells exhibit highly elevated glucose metabolism compared to non-tumor cells. We have applied in vivo optical imaging to study dynamic uptake of a near-infrared dye-labeled glucose analogue, 2-deoxyglucose (2-DG) by orthotopic glioma in a mouse model. METHODOLOGY AND PRINCIPAL FINDINGS: The orthotopic glioma model was established by surgically implanting U87-luc glioma cells into the right caudal nuclear area of nude mice. Intracranial tumor growth was monitored longitudinally by bioluminescence imaging and MRI. When tumor size reached >4 mm diameter, dynamic fluorescence imaging was performed after an injection of the NIR labeled 2-DG, IRDye800CW 2-DG. Real-time whole body images acquired immediately after i.v. infusion clearly visualized the near-infrared dye circulating into various internal organs sequentially. Dynamic fluorescence imaging revealed significantly higher signal intensity in the tumor side of the brain than the contralateral normal brain 24 h after injection (tumor/normal ratio, TNR = 2.8+/-0.7). Even stronger contrast was achieved by removing the scalp (TNR = 3.7+/-1.1) and skull (TNR = 4.2+/-1.1) of the mice. In contrast, a control dye, IRDye800CW carboxylate, showed little difference (1.1+/-0.2). Ex vivo fluorescence imaging performed on ultrathin cryosections (20 microm) of tumor bearing whole brain revealed distinct tumor margins. Microscopic imaging identified cytoplasmic locations of the 2-DG dye in tumor cells. CONCLUSION AND SIGNIFICANCE: Our results suggest that the near-infrared dye labeled 2-DG may serve as a useful fluorescence imaging probe to noninvasively assess intracranial tumor burden in preclinical animal models

Analysis of Tumor Metabolism Reveals Mitochondrial Glucose Oxidation in Genetically Diverse Human Glioblastomas in the Mouse Brain In Vivo

SummaryDysregulated metabolism is a hallmark of cancer cell lines, but little is known about the fate of glucose and other nutrients in tumors growing in their native microenvironment. To study tumor metabolism in vivo, we used an orthotopic mouse model of primary human glioblastoma (GBM). We infused 13C-labeled nutrients into mice bearing three independent GBM lines, each with a distinct set of mutations. All three lines displayed glycolysis, as expected for aggressive tumors. They also displayed unexpected metabolic complexity, oxidizing glucose via pyruvate dehydrogenase and the citric acid cycle, and using glucose to supply anaplerosis and other biosynthetic activities. Comparing the tumors to surrounding brain revealed obvious metabolic differences, notably the accumulation of a large glutamine pool within the tumors. Many of these same activities were conserved in cells cultured ex vivo from the tumors. Thus GBM cells utilize mitochondrial glucose oxidation during aggressive tumor growth in vivo

Correlation of BCL-2, Pp53, and MIB-1 Expression with Ependymoma Grade and Subtype.

In this study, we report our results on the proliferative activity of ependymomas as determined by MIB-1 (also known as Ki-67) immunohistochemical analysis, and we compare our results with those obtained by immunolabeling with monoclonal antibodies to p53 and bcl-2 proteins to assess whether expression correlated with ependymoma subtype or tumor grade. The study included 4 myxopapillary ependymomas (Grade I of the World Health Organization [WHO] scale), 10 subependymomas (WHO Grade I), 17 ependymomas (WHO Grade II), 2 papillary ependymomas (WHO grade II), and 4 anaplastic ependymomas (WHO Grade III). The MIB-1 proliferation index was significantly higher in tumors diagnosed as anaplastic ependymoma (P \u3c .001), with a moderate level of correlation (Kendall\u27s tau-b = 0.557, asymptotic standard error = 108). In addition, one ependymoma (WHO Grade II) not considered overtly anaplastic by routine histologic criteria showed a high MIB-1 labeling index, suggesting that the MIB-1 proliferation index might be a more objective indicator of tumor grade. The remaining WHO Grade I and Grade II ependymomas showed low proliferative activity. bcl-2 oncoprotein expression was identified in all of the four myxopapillary and in both papillary ependymomas. An additional observation was the correlation of p53 expression with increasing WHO grade. These data suggest that high MIB-1 and p53 immunolabeling might be objective indicators of high grade in ependymomas that do not otherwise meet routine histologic criteria for high-grade ependymoma. Subsequent clinicopathologic analyses will be important in assessing whether these markers are useful as independent predictors of survival

<i>Ex vivo</i> near infrared fluorescence imaging of ultrathin cryosections of tumor bearing brain tissues.

<p>Unstained coronal brain sections (20 µm) containing tumor tissues were obtained immediately after the 24 h <i>in vivo</i> image for <i>ex vivo</i> imaging. For the tumors with 2-DG dye, representative images for a larger (A) and a small (D) intracranial tumor showed predominant accumulation of the dye within the tumor mass. Tumor mass was clearly delineated from the surrounding normal tissues. Even a track of migrated tumor cells was depicted in each case (arrows in A and D), which correlated well with cresyl violet staining of the corresponding regions enlarged from B and E (arrows in C and F). Arrow heads refer to the dentate gyrus. The control dye showed no significant difference between the tumor regions and the normal brain (G and H). Tumor/normal ratio (TNR) of the 2-DG dye was significantly higher than that of the control dye (3.7±1.2 vs. 1.4±0.1; p<0.05; I).</p

Dynamic <i>in vivo</i> fluorescence imaging of the 2-DG dye uptake by intracranial glioma.

<p><b>A.</b> After i.v. injection of IRDye800CW 2-DG repeated <i>in vivo</i> fluorescence imaging was performed. During the first four hours, unmixed images showing stronger signal of the dye in the brain area, but there was no obvious contrast between the tumor side of the brain and the normal brain. However, 24 h later, the light signal remained only in the tumor side of the intact brain. Even better contrast was seen after reflection of the scalp or further removal of the skull. <b>B.</b> Normalized emission spectra showing the near infrared 2-DG dye with a peak emission wavelength at 810 nm (red), while the background signal (white) was at ∼770 nm. <b>C.</b> As a control of the 2-DG dye, IRDye800CW carboxyl was injected into a mouse bearing orthotopic glioma. No significant difference in light intensity between the two sides of the brain was observed at any time points.</p

<i>In vivo</i> real-time near infrared imaging of a mouse.

<p>A series of whole body images was acquired before and after injection of near infrared dye labeled 2-DG, IRDye800CW 2-DG, via a mouse tail vein. Selected images captured the first pass perfusion of the dye into various internal organs: heart and lung area at 1.2 s, brain and upper limbs at 2.5 s, kidney at 4.5 s, etc. Principal Component Analysis of the kinetics was used to identify tissue regions based on DyCE software as shown in the color presentation (top left) and representative time courses (bottom right).</p