Glucose transporters and FDG uptake in untreated primary human non-small cell lung cancer.

PET imaging of malignant tumors with 2-[fluorine-18]-fluoro-2-deoxy-D-glucose (FDG) as a tracer is a noninvasive diagnostic and prognostic tool that measures tumor metabolism. In this study, we assessed the relationships between FDG uptake and the expression of facilitative glucose transporters, the sizes of populations of proliferating cells and infiltrating macrophages in patients with primary non-small cell lung cancers (NSCLC). METHODS: FDG uptake and the expression of five glucose transporters and the proportions of proliferating cell and macrophage populations were studied in paraffin sections from untreated primary lung cancers by immunohistochemistry. The patients were imaged with FDG PET before surgery. RESULTS: All tumors could be detected by FDG PET. Uptake was correlated with tumor size (P = 0.004). FDG uptake was lower in adenocarcinomas (ACs) than in squamous cell carcinomas (SQCs) (P = 0.03) or large cell carcinomas (P = 0.002) [standardized uptake value corrected for lean body mass (SUL) = 5.42 +/- 2.77, 8.04 +/-3.25 and 10.42 +/- 4.54, respectively]. Glut-1 expression was significantly higher than that of any other transporter. All tumors tested (n = 23) were Glut-1-positive (70.8% +/- 26.1% of tumor cell area was positive and staining intensity was 2.8 +/- 1.2). Glut-1 expression was higher in SQCs (78% +/- 17.8% and 3.5 +/-0.6) than in ACs (47.5% +/- 30.3% and 1.6 +/- 1.1; P = 0.044 for positive tumor cell area and P = 0.005 for staining intensity). Proliferating cells constituted 15.3% +/- 13.1% of the cancer cells, and the average number of macrophages was 7.8% +/- 6.3%; neither correlated with FDG uptake. CONCLUSION: In this population of patients with NSCLC, Glut-1 is the major glucose transporter expressed. Both FDG uptake and Glut-1 expression appear to be associated with tumor size. No association was found between FDG uptake and either macrophage or proliferative cell populations.     

Steady-state blood volume measurements in experimental tumors with different angiogenic burdens a study in mice

PURPOSE: To experimentally validate the effectiveness of magnetic resonance (MR) imaging enhanced with long-circulating iron oxide for measurement of vascular volume fractions (VVFs) as indicators of angiogenesis in different experimental tumor models. MATERIALS AND METHODS: Tumors with differing degrees of angiogenesis-9L rodent gliosarcoma, DU4475 human mammary adenocarcinoma, HT1080 human fibrosarcoma, and EOMA hemangioendothelioma--were implanted in nude mice. Tumoral VVFs were measured at submillimeter voxel resolutions by using 1.5-T MR imaging. A technetium-labeled intravascular radiotracer was injected into a subset of the animals to validate the MR imaging measurements. Microvessel density and vascular endothelial growth factor (VEGF) also were measured. Statistical analysis was performed with analysis of variance. RESULTS: High-resolution multisection MR maps of tumor blood volume were obtained in all tumor models. Mean tumoral VVF differed significantly among the different tumors: 2.1% +/- 0.3 (standard error of mean) for 9L gliosarcoma, 3.1% +/- 0.4 for DU4475 mammary adenocarcinoma, 5.5% +/- 0.8 for HT1080 fibrosarcoma, and 6.6% +/- 0.9 for EOMA hemangioendothelioma (P <.01). There was a strong correlation between the MR imaging and radiotracer measurements. There was considerable intra- and intertumoral heterogeneity among the VVFs. MR imaging measurements were in accordance with conventional measurements of angiogenesis, such as microvessel density count and VEGF. CONCLUSION: Measurements of tumoral VVF at high-resolution MR imaging with long-circulating iron oxide are feasible and correlate with angiogenic burden in experimental tumor models.     

L-type amino acid transporters LAT1 and LAT4 in cancer: uptake of 3-O-methyl-6-18F-fluoro-L-dopa in human adenocarcinoma and squamous cell carcinoma in vitro and in vivo.

Expression of system L amino acid transporters (LAT) is strongly increased in many types of tumor cells. The purpose of this study was to demonstrate that (18)F-labeled amino acids, for example, 3-O-methyl-6-(18)F-fluoro-L-dopa ((18)F-OMFD), that accumulate in tumors via LAT represent an important class of imaging agents for visualization of tumors in vivo by PET. METHODS: (18)F-OMFD uptake kinetics, transport inhibition, and system L messenger RNA expression were studied in vitro in human adenocarcinoma (HT-29), squamous cell carcinoma (FaDu), macrophages (THP-1), and primary aortic endothelial cells (HAEC) and in vivo in the corresponding mouse tumor xenograft models. RESULTS: Uptake of (18)F-OMFD in all cell lines tested was mediated mainly by the sodium-independent high-capacity LAT. We found higher uptake in FaDu cells (V(max), 10.6 +/- 1.1 nmol/min x mg of cell protein) and in the corresponding FaDu tumor xenografts than in the other cells and corresponding xenograft models studied. Quantitative messenger RNA analysis revealed that tumor cells and xenografts have a higher expression of LAT1 than do HAEC and THP-1 macrophages. However, only in the FaDu tumor model did an increased (18)F-OMFD uptake seem to be explained by increased LAT expression. Furthermore, we demonstrated a high expression of LAT4, a recently identified LAT. CONCLUSION: Our findings support the hypothesis that (18)F-OMFD is a tracer for visualization of tumor cells. (18)F-OMFD particularly seems to be a suitable tracer for diagnostic imaging of amino acid transport in poorly differentiated squamous cell head and neck carcinoma with increased LAT1 and LAT4 expression.     

Detection of lymph node metastases by contrast-enhanced MRI in an experimental model.

Lymph node size, the accumulation of a nodal lymphotrophic contrast agent (LCDIO), and MRI were compared as methods for detecting nodal metastases in an experimental murine model. Lymph node metastases (B16-F1 melanoma expressing green fluorescent protein (GFP) and C57BL/6 mice) were generated to obtain a wide spectrum of nodes, including normal nodes and nodes bearing micrometastases, small metastases, or large metastases. Nodal uptake of LCDIO was measured using (111)Indium-labeled LCDIO and was found to be lower in micrometastastic nodes (4.20 +/- 1.4%ID/gm) than in normal nodes (8.60 +/- 0.22% ID dose/gram, P < 0.005). Nodal tumor burden was quantified from the amount of GFP present in nodes measured using the Western blot method, and was found to correlate with the decrease of LCDIO uptake. By MRI, nodes bearing small and large metastases contained regions of high signal intensity (SI) that corresponded to the visual pattern of tumor in nodes. Micrometastatic nodes were distinguishable from normal nodes based on a diffuse pattern of inhomogeneous SI. The signal-to-background ratio (SBR) of normal nodes (0.0112 +/- 0.0061) was different from micrometastatic nodes (0.179 +/- 0.080, P < 0.00046) and nodes bearing small metastases (0.723 +/- 0.269, P < 0.00013), with high degrees of significance.     

Evaluation of 99mTc-L-methionine brain SPECT for detection of recurrent brain tumor: a pilot study with radiological and pathological correlation

PURPOSE: To evaluate technetium labeled L-methionine for imaging recurrent brain tumors. MATERIAL AND METHODS: Brain SPECT with 99mTc-L-methionine was performed to evaluate tumor viability in 42 patients with primary brain tumor. Findings of SPECT were correlated with radiological and histopathological findings as reference. RESULTS: 99mTc-L-methionine showed localized increased uptake in 40 patients with tumor recurrence, whereas 2 patients with post-radiation gliosis did not show tracer accumulation. A low differential uptake rate (DUR) 2.43 +/- 0.74 and methionine retention (MR) index 0.93 +/- 0.03 was seen in cases of post-radiation gliosis. A high DUR (36.20 +/- 10.31) and MR index (4.87 +/- 2.37) was seen in cases of recurrent tumor. Mean DUR in high-grade tumors (44.01 +/- 8.46) was significantly higher (P<0.001) than in low-grade tumors (30.42 +/- 7.38), and mean MR index in high-grade tumors (7.03 +/- 2.05) was significantly higher than in low-grade tumors (3.27 +/- 0.82) (P<0.001). CONCLUSION: 99mTc-L-methionine can be used as a SPECT tracer to differentiate tumor recurrence from post-radiation gliosis.     

Applicability of 99mTc-HL91, a putative hypoxic tracer, to detection of tumor hypoxia.

To elucidate the applicability of 99mTc-HL91 (HL91) a putative hypoxic tracer, to the imaging of hypoxia in tumors, a biodistribution study of the tracer was performed. The intratumoral distribution of HL91 was compared with that of 14C-deoxyglucose (DG) and the expression of glucose transporter 1 (GLUT1) in an implanted tumor. METHODS: Biodistribution of HL91 after intravenous injection into Wistar rats with rat mammary tumor (Walker-256) was studied by determining blood and tissue levels of radioactivity from 15 min to 6 h after injection. Dual ex vivo autoradiography was performed on sections of the tumor using HL91 (74 MBq) and DG (185 kBq). The same sections were immunohistologically analyzed with anti-GLUT1 antibody. Tumor tissue was histologically divided into areas of viable cancer cells, necrosis and granulation tissue. The viable cancer cell area was further divided into normoxic and hypoxic areas. Uptake of both tracers in each area was measured quantitatively. The intensity of GLUT1 staining (relative optical density [ROD]) in each area was evaluated by densitometry. RESULTS: The uptake of HL91 in the tumor reached a maximal value (0.897 +/- 0.118% ID [injected dose], mean +/- SD, n = 5) at 120 min after intravenous injection of HL91, then gradually decreased. The tumor-to-muscle ratio continued to increase until 360 min (4.34 at 120 min, 7.01 at 240 min and 10.4 at 360 min). HL91 accumulated to significantly higher levels in the hypoxic area than those in the other tissues (P < 0.0001). Uptake of DG and expression of GLUT1 were significantly higher in the hypoxic area than in the normoxic area (P < 0.0001). In the viable cancer cell area, uptake of HL91 and expression of GLUT1 were strongly correlated (r = 0.624-0.868, mean r = 0.743, P < 0.0001), and DG uptake was moderately correlated with GLUT1 expression (r = 0.328-0.669, mean r = 0.505, P < 0.0001). CONCLUSION: These results indicate that HL91 can be used to detect tumor hypoxia.     

Iron oxide nanoparticles as magnetic resonance contrast agent for tumor imaging via folate receptor-targeted delivery

RATIONALE AND OBJECTIVE: Targeted delivery is a highly desirable strategy for diagnostic imaging because of enhanced efficacy and reduced dosage/toxicity. Receptor-targeting was used to deliver contrast-producing superparamagnetic iron oxide (IO) nanoparticle to receptor-expressing tumors for in vivo magnetic resonance (MR) imaging. MATERIALS AND METHODS: Nanometer-sized, dextran-coated (maghemite) IO particles were prepared by a precipitation method. They were tethered with N-hydroxysuccinimide-folate and fluorescence isothiocyanate (FITC). For in vitro study of delivery specificity and efficiency, KB cells, a human nasopharyngeal epidermal carcinoma cell line expressing surface receptors for folic acid, were used as positive targets, and A549 cells, a human lung carcinoma cell line which lacks folate receptors, were used as negative control targets. In vivo MR images were obtained using mouse models with subcutaneous tumor xenografts grown from implanted KB cells. RESULTS: Internalization of nanoparticles into targeted cells only occurred when IO was conjugated to folate and when the folate receptors are available and accessible on the cells. The endocytosis was efficient and rapid, as 97.5% KB cells cultured with folate-FITC-IO showed FITC uptake after 1 hour of incubation. In in vivo MR imaging, an average intensity decrease of 38% was observed from precontrast to postcontrast images of the tumor, which was about three times the intensity decrease observed at a non-tumor-bearing muscle. CONCLUSION: Successful in vivo MR imaging of folate receptor-expressing tumors targeted by IO nanoparticles was demonstrated for the first time.     

Ocular and cerebral metastases in the VX2 rabbit tumor model: contrast- enhanced MR imaging

Ocular and cerebral metastases developed after the inoculation of a VX2 tumor cell suspension into the internal carotid artery of 15 rabbits. The hematogenous spread of tumor cells resulted in ocular metastases in 13 of 15 animals (86.7%) and cerebral system metastases in 14 of 15 animals (93%). Magnetic resonance (MR) imaging with Gd-DTPA demonstrated early disruption of the blood-ocular barrier and blood-brain barrier 5-7 days after infusion of tumor cells. Quantitative assessment of contrast enhancement revealed a mean increase in signal intensity of 145% +/- 51% in the anterior chambers, 102% +/- 70% for choroidal metastases, and 51% +/- 29% for central nervous system (CNS) metastases. These results indicate that contrast-enhanced MR imaging can be used to demonstrate a loss of blood-ocular barrier integrity that is similar to the breakdown of the blood-brain barrier associated with metastatic tumors to the CNS and eye.     

Warthin tumor of the parotid gland: diagnostic value of MR imaging with histopathologic correlation

BACKGROUND AND PURPOSE: The purpose of our study was to describe the MR imaging appearance of Warthin tumors multiple MR imaging techniques and to interpret the difference in appearance from that of malignant parotid tumors. METHODS: T1-weighted, T2-weighted, short inversion time inversion recovery, diffusion-weighted, and contrast-enhanced dynamic MR images of 19 Warthin tumors and 17 malignant parotid tumors were reviewed. MR imaging results were compared with those of pathologic analysis. RESULTS: Epithelial stromata and lymphoid tissue with slitlike small cysts in Warthin tumors showed early enhancement and a high washout rate (> or =30%) on dynamic contrast-enhanced images, and accumulations of complicated cysts showed early enhancement and a low washout ratio (< 30%). The areas containing complicated cysts showed high signal intensity on T1-weighted images, whereas some foci in those areas showed low signal intensity on short tau inversion recovery images. The mean minimum signal intensity ratios (SIRmin) of Warthin tumor on short tau inversion recovery (0.29 +/- 0.22 SD) (P < .01) and T2-weighted images (0.28 +/- 0.09) (P < .05) were significantly lower than those of malignant parotid tumors (0.53 +/- 0.19, 0.48 +/- 0.19). The average washout ratio of Warthin tumors (44.0 +/- 20.4%) was higher than that of malignant parotid tumors (11.9 +/- 11.6%). The mean apparent diffusion coefficient of Warthin tumors (0.96 +/- 0.13 x 10(-3)mm2/s) was significantly lower (P < .01) than that of malignant tumors (1.19 +/- 0.19 x 10(-3)mm2/s). CONCLUSION: Detecting hypointense areas of short tau inversion recovery and T2-weighted images or low apparent diffusion coefficient values on diffusion-weighted images was useful for predicting whether salivary gland tumors were Warthin tumors. The findings of the dynamic contrast-enhanced study also were useful.     

Imaging of gliomas with Cis-4-[18F]fluoro-L-proline

Tumor imaging with cis-4-[18F]fluoro-L-proline (cis-FPro) was compared to that of L-[3H]proline and L-[3H]methionine in F98 rat gliomas by dual-tracer autoradiography. All tracers exhibited high accumulation in the tumors but in the normal brain significant uptake was observed for L-[3H]methionine only. Tumor extent on autoradiograms with L-[3H]proline and L-[3H]methionine was identical to that of histological staining while autoradiograms of cis-FPro showed diffuse uptake in the penumbra of some tumors. First PET studies in 7 patients with cerebral gliomas demonstrated accumulation of cis-FPro in tumor areas with enhancement of Gd-DTPA on MR scans. Uptake of cis-FPro in normal brain tissue was negligible. In one patient with a glioblastoma accumulation of cis-FPro was also found in two brain areas without enhancement of Gd-DTPA on MR scans. Control of MRI suggested tumor growth in these areas at further follow up. Our results indicate that in most gliomas increased cis-FPro uptake is restricted to areas with disruption of the BBB which limits its clinical utility.     

Radioiodinated VEGF to image tumor angiogenesis in a LS180 tumor xenograft model

INTRODUCTION: Angiogenesis is essential for tumor growth or metastasis. A method involving noninvasive detection of angiogenic activity in vivo would provide diagnostic information regarding antiangiogenic therapy targeting vascular endothelial cells as well as important insight into the role of vascular endothelial growth factor (VEGF) and its receptor (flt-1 and KDR) system in tumor biology. We evaluated radioiodinated VEGF(121), which displays high binding affinity for KDR, and VEGF(165), which possesses high binding affinity for flt-1 and low affinity for KDR, as angiogenesis imaging agents using the LS180 tumor xenograft model. METHODS: VEGF(121) and VEGF(165) were labeled with (125)I by the chloramine-T method. Biodistribution was observed in an LS180 human colon cancer xenograft model. Additionally, autoradiographic imaging and immunohistochemical staining of tumors were performed with (125)I-VEGF(121). RESULTS: (125)I-VEGF(121) and (125)I-VEGF(165) exhibited strong, continuous uptake by tumors and the uterus, an organ characterized by angiogenesis. (125)I-VEGF(121) uptake in tumors was twofold higher than that of (125)I-VEGF(165) (9.12+/-98 and 4.79+/-1.08 %ID/g at 2 h, respectively). (125)I-VEGF(121) displayed higher tumor to nontumor (T/N) ratios in most normal organs in comparison with (125)I-VEGF(165). (125)I-VEGF(121) accumulation in tumors decreased with increasing tumor volume. Autoradiographic and immunohistochemical analyses confirmed that the difference in (125)I-VEGF(121) tumor accumulation correlated with degree of tumor vascularity. CONCLUSION: Radioiodinated VEGF(121) is a promising tracer for noninvasive delineation of angiogenesis in vivo.     

In vitro and in vivo tracer characteristics of an established multidrug-resistant human colon cancer cell line.

99mTc-methoxyisobutylisonitrile (99mTc-MIBI) has been suggested as a tracer for the scintigraphic detection of multidrug resistance (MDR). The aim of this study was to compare MDR characteristics in vitro and in vivo by immunohistochemic and functional uptake assays in established tumor cell lines cultured and grown in severe combined immunodeficient (SCID) mice. METHODS: The presence of MDR was assessed in vitro in drug-resistant HT-29(mdr1) colon carcinoma cells and in nonresistant HT-29(par) cells by JSB-1 immunohistochemistry, uptake of the fluorescent dye Rhodamine 123, and quantitative measurement of 99mTc-MIBI accumulation. For in vivo imaging, SCID mice bearing subcutaneous xenografts of these cell lines were injected with 99mTc-MIBI and 18F-FDG for scintigraphic and PET examination. After imaging, tumors were analyzed by immunohistochemistry and electron microscopy. RESULTS: All HT-29(mdr1) cells cultured in vitro exhibited distinct JSB-1 immunoreactivity, although to a variable degree, whereas HT-29(par) cells were completely devoid of JSB-1 staining. Rhodamine 123 accumulated poorly in HT-29(mdr1) cells but strongly in HT-29(par) cells. Accumulation of 99mTc-MIBI was 0.05% +/- 0.01% of the activity of the external medium in HT-29(mdr1) cells, but about eight times higher in HT-29(par) cells (0.40% +/- 0.09%), a very low percentage compared with other tumor cell lines. No difference in 201TlCl accumulation was observed between both cell lines. In vivo, neither HT-29(par) nor HT-29(mdr1) tumors grown in SCID mice could be detected by 99mTc-MIBI scintigraphy. In FDG PET, both HT-29(mdr1) and HT-29(par) tumors were clearly visible. FDG uptake was, however, markedly higher in HT-29(par) than in HT-29(mdr1) tumors. Both tumor types were poorly vascularized, as shown histologically. JSB-1 immunoreactivity was absent in all HT-29(par) tumors examined, whereas the majority of HT-29(par) tumor cells were stained. Electron microscopy showed that HT-29(par) tumors contained significantly less mitochondria than hepatocytes of the SCID mouse liver, which displayed high 99mTc-MIBI uptake in our scintigraphy studies. CONCLUSION: Sufficient 99mTc-MIBI uptake is the major prerequisite for distinguishing successfully between drug-resistant and sensitive cells. Negative 99mTc-MIBI scintigrams are not necessarily associated with MDR expression. In some tumors, FDG may be an in vivo marker for MDR as suggested by PET.     

Renal angiomyolipoma with minimal fat: differentiation from other neoplasms at double-echo chemical shift FLASH MR imaging

PURPOSE: To prospectively evaluate the diagnostic performance of double-echo gradient-echo (GRE) chemical shift magnetic resonance (MR) imaging in the differentiation of angiomyolipoma (AML) with minimal fat from other renal neoplasms, with pathologic examination or follow-up data serving as the reference standard. MATERIALS AND METHODS: Institutional review board approval and informed consent were obtained. Double-echo GRE chemical shift MR imaging was performed in 55 patients (29 men and 26 women; mean age, 49 years +/- 14 [standard deviation]) with 55 renal tumors, including 37 (67%) pathologically proved tumors (23 renal cell carcinomas, nine AMLs, two oncocytomas, two lymphomas, and one reninoma) and 18 (33%) clinically diagnosed tumors (17 AMLs and one indeterminate malignancy). All tumors showed no intratumoral fat and had homogeneous enhancement and a prolonged or gradual enhancement pattern on biphasic helical computed tomographic scans. Signal intensity was measured in the renal tumor and spleen on in-phase and opposed-phase images. The signal intensity index and tumor-to-spleen ratio in AMLs and non-AMLs were calculated and compared with the Student t test. Receiver operating characteristic (ROC) analysis was performed to evaluate the diagnostic accuracy of the signal intensity index and tumor-to-spleen ratio and to extract the optimal cut-off values in the differentiation of AMLs and non-AMLs. RESULTS: The signal intensity index and tumor-to-spleen ratio were different between AMLs (42% +/- 11 and -43% +/- 17, respectively) and non-AMLs (5% +/- 14 and -4% +/- 16, respectively) (P < .001). The area under the ROC curve was 0.975 for the signal intensity index and 0.952 for the tumor-to-spleen ratio. For differentiation of AMLs from non-AMLs, sensitivity and specificity were (a) 96% and 93%, respectively, with a signal intensity index of 25% and (b) 88% and 97%, respectively, with a tumor-to-spleen ratio of -32%. CONCLUSION: Double-echo GRE chemical shift MR imaging can be used to differentiate AML with minimal fat from other renal neoplasms.     

Ferumoxide-enhanced MR imaging of hepatocellular carcinoma: correlation with histologic tumor grade and tumor vascularity

PURPOSE: To evaluate ferumoxide-enhanced MR imaging findings of hepatocellular carcinomas (HCCs) in correlation with the histologic tumor grades and the tumor vascularity evaluated by CT hepatic arteriography (CTHA) and CT during arterial portography (CTAP) combined. MATERIALS AND METHODS: By searching the radiologic, surgical, and pathologic reports of our institution between January 1999 and February 2001, we identified 43 patients with 51 pathologically confirmed HCCs who underwent ferumoxide-enhanced MR imaging and combination CTHA and CTAP within two weeks. The HCCs consisted of 17 well-differentiated, 28 moderately differentiated, and six poorly differentiated tumors. The MR and CT were retrospectively reviewed by two radiologists in consensus for signal intensity on MR images and vascularity on CT. The Spearman's rank correlation coefficient was calculated to correlate the frequency of tumors with ferumoxide uptake with the histologic tumor grades and the tumor vascularity on CTHA and CTAP. RESULTS: A total of 45 tumors (88%) did not take up ferumoxide, and thus showed distinct, homogeneous hyperintensity. Six tumors (12%) ranging 5-16 mm in size (mean, 11 mm) took up ferumoxide, and thus showed isointensity, mixed intensity, or hypointensity, including five of 17 (29%) well-differentiated tumors and one of 28 (4%) moderately differentiated tumors. Five of the six tumors (83%) showed hyper- or hypovascularity on CTHA or hypovascularity on CTAP. The frequency of tumors with ferumoxide uptake showed weak correlation with tumor grades (coefficient = 0.26, P < 0.01) and vascularity on CTHA (-0.35, P < 0.05) and CTAP (0.39, P < 0.01). CONCLUSION: Although a small number of well-differentiated HCC take up ferumoxide and show iso-, mixed, or hypointensity, most such tumors show increased hepatic arterial or decreased portal venous perfusion. The present results suggest the limitation of reticuloendothelial contrast imaging, particularly in the diagnosis of small, well-differentiated HCC.     

Comparison of intratumoral distribution of 99mTc-MIBI and deoxyglucose in mouse breast cancer models.

The aims of this study were to evaluate the distribution of 99mTc-methoxyisobutylisonitrile (MIBI) in 3 animal models of breast cancer, the effect of radiotherapy on 99mTc-MIBI uptake, and the relationship between uptake and microvessel density. METHODS: We used syngeneic, subcutaneously transplanted FM3A, MM48, and Ehrlich mouse breast cancer. 99mTc-MIBI and FDG were injected intravenously, and tumor uptake was measured 30 min later. Double-tracer macroautoradiography (ARG) images were prepared with 99mTc-MIBI and 2-deoxy-D-[1-14C]-glucose (14C-DG), analyzed quantitatively, and compared with histology. The radiotherapeutic effects of 20 Gy x-ray irradiation were monitored by measuring tumor volume, tumor uptake, and ARG findings using 99mTc-MIBI and FDG in FM3A tumors. Microvessel density was quantified by immunohistochemical staining for CD34 and compared with ARG using 99mTc-MIBI in FM3A tumors. RESULTS: FM3A, MM48, and Ehrlich tumors showed different growth rates and radiosensitivities. Uptake of FDG, but not of 99mTc-MIBI, correlated significantly with growth rates. Compared with 14C-DG, 99mTc-MIBI accumulated more in cancer cells and less in infiltrating fibroblasts and macrophages in all tumor models. Irradiation significantly decreased 99mTc-MIBI uptake, but a rapid increase was noted at recurrence on day 7. Changes in FDG uptake were not significant at recurrence. Microvessel density in tumor tissue correlated significantly with 99mTc-MIBI uptake on ARG. CONCLUSION: Accumulation of 99mTC-MIBI in cancer cells is preferential and can be used as a sensitive marker to examine the response to radiotherapy. Angiogenesis seems to enhance accumulation of 99mTc-MIBI in tumors. These characteristics may be favorable for tumor imaging using 99mTC-MIBI.     

Clinically applicable labeling of mammalian and stem cells by combining superparamagnetic iron oxides and transfection agents

PURPOSE: To label mammalian and stem cells by combining commercially available transfection agents (TAs) with superparamagnetic iron oxide (SPIO) magnetic resonance (MR) imaging contrast agents. MATERIALS AND METHODS: Three TAs were incubated with ferumoxides and MION-46L in cell culture medium at various concentrations. Human mesenchymal stem cells, mouse lymphocytes, rat oligodendrocyte progenitor CG-4 cells, and human cervical carcinoma cells were incubated 2-48 hours with 25 microg of iron per milliliter of combined TAs and SPIO. Cellular labeling was evaluated with T2 relaxometry, MR imaging of labeled cell suspensions, and Prussian blue staining for iron assessment. Proliferation and viability of mesenchymal stem cells and human cervical carcinoma cells labeled with a combination of TAs and ferumoxides were evaluated. RESULTS: When ferumoxides-TA or MION-46L-TA was used, intracytoplasmic particles stained with Prussian blue stain were detected for all cell lines with a labeling efficiency of nearly 100%. Limited or no uptake was observed for cells incubated with ferumoxides or MION-46L alone. For TA-SPIO-labeled cells, MR images and relaxometry findings showed a 50%-90% decrease in signal intensity and a more than 40-fold increase in T2s. Cell viability varied from 103.7% +/- 9 to 123.0% +/- 9 compared with control cell viability at 9 days, and cell proliferation was not affected by endosomal incorporation of SPIO nanoparticles. Iron concentrations varied with ferumoxides-TA combinations and cells with a maximum of 30.1 pg +/- 3.7 of iron per cell for labeled mesenchymal stem cells. CONCLUSION: Magnetic labeling of mammalian cells with use of ferumoxides and TAs is possible and may enable cellular MR imaging and tracking in experimental and clinical settings.     

Human transferrin receptor gene as a marker gene for MR imaging

PURPOSE: To quantitate and characterize the expression of an engineered human transferrin receptor (ETR) as a marker gene by using magnetic resonance (MR) imaging. MATERIALS AND METHODS: Rat gliosarcoma 9L cells stably expressing ETR (ETR+) were used, with nontransfected (ETR-) cells serving as controls. A conjugate of transferrin and monocrystalline iron oxide (Tf-MION) nanoparticles was synthesized to probe for the activity of ETR. Accumulation of Tf-MION was examined by using cell internalization in culture and MR (n = 6) and nuclear (n = 4) imaging in a mouse model with ETR+ and ETR- tumors implanted in the opposite flanks. Autoradiographic and histopathologic results were correlated with MR findings. RESULTS: Tf-MION was internalized by ETR+ cells at 37 degrees C but not at 4 degrees C. Rhodamine-labeled Tf-MION and fluorescein-labeled antibody to ETR colocalized in small vesicle-like structures in the cytoplasm. Both findings were consistent with accumulation by the receptor-mediated endocytosis mechanism of ETR. Compared with ETR- tumors, ETR+ tumors accumulated more Tf-MION and had higher signal intensity on T1-weighted MR images and lower signal intensity on T2-weighted images. Autoradiographic findings showed a spatial correlation between MR signal intensity and TF-MION accumulation. CONCLUSION: ETR+ tumors internalize the MR imaging probe through the action of transferrin receptor in amounts that can be detected with MR imaging.     

64Cu-labeled folate-conjugated shell cross-linked nanoparticles for tumor imaging and radiotherapy: synthesis, radiolabeling, and biologic evaluation.

Long-circulating nanoparticles functionalized with ligands for receptors overexpressed by tumor cells have promising applications for active and passive tumor targeting. The purpose of this study was to evaluate 64Cu-radiolabeled folate-conjugated shell cross-linked nanoparticles (SCKs) as candidate agents to shuttle radionuclides and drugs into tumors overexpressing the folate receptor (FR). METHODS: SCKs were obtained by cross-linking the shell of micelles obtained from amphiphilic diblock copolymers. SCKs were then functionalized with folate, fluorescein thiosemicarbazide (FTSC), and 1,4,8,11-tetraazacyclotetradecane-N,N',N',N'-tetraacetic acid (TETA). The specific interaction of SCK-folate with the FR was investigated on KB cells. The biodistributions of 64Cu-TETA-SCK and 64Cu-TETA-SCK-folate were evaluated in athymic mice bearing small-size KB cell xenografts (10-100 mg), whereas the intratumor distributions were investigated by autoradiography in 0.3- to 0.6-g KB cell xenografts. RESULTS: A global solution-state functionalization strategy has been introduced for attaching optimum numbers of targeting and imaging agents onto the SCKs for increasing the efficiency of interaction with cell-surface receptors. Epifluorescence microscopy confirmed the specific interaction of FTSC-SCK-folate with the FR in vitro. 64Cu labeling of TETA-SCKs led to the radiolabeled compounds with 15%-20% yield and >95% radiochemical purity. The biodistribution results demonstrated high accumulation of 64Cu-labeled SCKs in organs of the reticuloendothelial system (RES) (56.0 +/- 7.1 %ID/g and 45.7 +/- 3.5 %ID/g [percentage injected dose per gram] in liver at 10 min after injection for folated and nonfolated SCKs, respectively) and a prolonged blood circulation. No increase of SCK tumor uptake deriving from folate conjugation was observed (5.9 +/- 2.8 %ID/g and 6.0 +/- 1.9 %ID/g at 4 h after injection for folated and nonfolated SCKs, respectively). However, tumor accumulation was higher in small-size tumors, where competitive block of SCK-folate uptake with excess folate was observed. Autoradiography results confirmed the extravasation of radiolabeled SCKs in vascularized areas of the tumor, whereas no diffusion was observed in necrotic regions. CONCLUSION: Despite high RES uptake, the evaluated 64Cu-labeled SCKs exhibited long circulation in blood and were able to passively accumulate in tumors. Furthermore, SCK-folate uptake was competitively blocked by excess folate in small-size solid tumors, suggesting interaction with the FR. For these reasons, functionalized SCKs are promising drug-delivery agents for imaging and therapy of early-stage solid tumors.     

Favorable biokinetic and tumor-targeting properties of 99mTc-labeled glucosamino RGD and effect of paclitaxel therapy.

Compared with the recent advancements in radiohalogenated Arg-Gly-Asp (RGD) peptides for alpha(v)beta(3)-targeted imaging, there has been limited success with (99m)Tc-labeled RGD compounds. In this article, we describe the favorable in vivo kinetics and tumor-imaging properties of a novel (99m)Tc-RGD compound that contains a glucosamine moiety. METHODS: Glucosamino (99m)Tc-d-c(RGDfK) was prepared by incorporating (99m)Tc(CO)(3) to the glucosamino peptide precursor in high radiochemical yield. Cell-binding characteristics were tested on human endothelial cells. Mice bearing RR1022 fibrosarcoma and Lewis lung carcinoma (LLC) tumors were used for in vivo biodistribution and blocking experiments and for imaging studies. Separate LLC-bearing mice underwent antiangiogenic therapy with 0, 20, or 40 mg of paclitaxel per kilogram of body weight every 2 d. Tumor volume was serially monitored, and tumor glucosamino (99m)Tc-d-c(RGDfK) uptake and Western blots of alpha(v) integrin expression were analyzed at day 14. RESULTS: Glucosamino (99m)Tc-d-c(RGDfK) binding to endothelial cells was dose-dependently inhibited by excess RGD. Biodistribution in mice showed rapid blood clearance of glucosamino (99m)Tc-d-c(RGDfK), with substantially lower liver uptake and higher tumor uptake compared with (125)I-c(RGD(I)yV). Tumor uptake was 1.03 +/- 0.21 and 1.18 +/- 0.26 %ID/g at 1 h and 0.85 +/- 0.05 and 0.89 +/- 0.28 %ID/g at 4 h for sarcomas and carcinomas, respectively. Excess RGD blocked uptake by 76.5% and 70.2% for the respective tumors. gamma-Camera imaging allowed clear tumor visualization, with an increase of sarcoma-to-thigh count ratios from 5.5 +/- 0.7 at 1 h to 10.1 +/- 2.2 at 4 h and sustained carcinoma-to-thigh count ratios from 4 to 17 h. Pretreatment with excess cRGDyV significantly reduced tumor contrast on images. Paclitaxel therapy in LLC tumor-bearing mice significantly retarded tumor growth. This was accompanied by a corresponding reduction of tumor glucosamino (99m)Tc-d-c(RGDfK) uptake, which correlated significantly with tumor alpha(v) integrin expression levels. CONCLUSION: Glucosamino (99m)Tc-d-c(RGDfK) has favorable in vivo biokinetics and tumor-imaging properties and may be useful for noninvasive evaluation of tumor integrin expression and response to antiangiogenic therapeutics. Because of the wide accessibility of gamma-cameras and high availability and excellent imaging characteristics of (99m)Tc, glucosamino (99m)Tc-d-c(RGDfK) may be an attractive alternative to radiohalogenated RGD peptides for angiogenesis-imaging research.     

The added value of the apparent diffusion coefficient calculation to magnetic resonance imaging in the differentiation and grading of malignant brain tumors

OBJECTIVE: ADC calculation can improve the diagnostic efficacy of MR imaging in brain tumor grading and differentiation. METHODS: Apparent diffusion coefficient (ADC) values and ratios of 33 low-grade (23 astrocytomas, 10 oligodendrogliomas) and 40 high-grade (25 metastases and 15 high-grade astrocytomas) malignant tumors were prospectively evaluated. RESULTS: Tumoral ADC values (r=-0.738, P <0.000) and ratios (r=-0.746, P < 0.000) were well correlated with higher degree of malignancy and quite effective in grading of malignant brain tumors (P < 0.000). By using cutoff values of 0.99 for tumoral ADC value and 1.22 for normalized ADC ratio, the sensitivity of MR imaging could be increased from 72.22% to 93.75% and 90.63%, the specificity from 81.08% to 92.68% and 90.24%, PPV from 78.79% to 90.91% and 87.88%, and NPV from 75.00% to 95.00% and 92.50%, respectively. CONCLUSION: ADC calculation was quite effective in grading of malignant brain tumors but not in differentiation of them and added more information to conventional contrast-enhanced MR imaging.