[C]choline as a potential PET marker for imaging of breast cancer athymic mice

[¹¹C]Choline has been evaluated as a potential positron emission tomography (PET) marker for imaging of breast cancer. The biodistribution of [¹¹C]choline was determined at 45 min post iv injection in MCF-7’s transfected with IL-1alpha implanted athymic mice and MDA-MB-435 implanted athymic mice. The results showed the uptake of [¹¹C]choline in these tumors was high, 2.0% dose/g in MCF-7’s transfected with IL-1alpha implanted mice and 1.8% dose/g in MDA-MB-435 implanted mice; the ratios of tumor/muscle (T/M) and tumor/blood (T/B) were 1.7 (T/M, MCF-7’s), 2.1 (T/M, MDA-MB-435) and 6.9 (T/B, MCF-7’s), 12.5 (T/B, MDA-MB-435), respectively; the tumor/muscle ratios are moderate, and the tumor/blood ratios are high. The micro-PET imaging of [¹¹C]choline in both breast cancer athymic mice was acquired for 15 min from a MCF-7’s transfected with IL-1alpha and/or MDA-MB-435 implanted mouse at 45 min post iv injection of 1 mCi of the tracer using a dedicated high resolution (<3 mm full-width at half-maximum) small FOV (field-of-view) PET imaging system, Indy-PET II scanner, developed in our laboratory, which showed the uptake of [¹¹C]choline in MCF-7’s transfected with IL-1alpha tumor or MDA-MB-435 tumor implanted in a nude athymic mouse. These results suggest that [¹¹C]choline may be a potential PET breast cancer imaging agent.     

Comparative studies of potential cancer biomarkers carbon-11 labeled MMP inhibitors (S)-2-(4'-[11C]methoxybiphenyl-4-sulfonylamino)-3-methylbutyric acid and N-hydroxy-(R)-2-[[(4'-[11C]methoxyphenyl)sulfonyl]benzylamino]-3-methylbutanamide

(S)-2-(4'-[11C]methoxybiphenyl-4-sulfonylamino)-3-methylbutyric acid ([11C]MSMA) and N-hydroxy-(R)-2-[[(4'-[11C]methoxyphenyl)sulfonyl]benzylamino]-3-methylbutanamide ([11C]CGS 25966), carbon-11 labeled matrix metalloproteinase (MMP) inhibitors, have been synthesized for evaluation as new potential positron emission tomography (PET) cancer biomarkers. [11C]MSMA was prepared by appropriate precursor (S)-2-(4'-hydroxybiphenyl-4-sulfonylamino)-3-methylbutyric acid tert-butyl ester, which was synthesized in eight steps from amino acid (L)-valine in 39.4% chemical yield. This precursor was labeled by [11C]methyl triflate through O-[11C]methylation method at the hydroxyl position of biphenol under basic conditions, followed by a quick acid hydrolysis and isolated by solid-phase extraction (SPE) purification to produce pure target compound [11C]MSMA in 35-55% radiochemical yield, based on 11CO2, decay corrected to end of bombardment (EOB), and 20-25 min synthesis time. [11C]CGS 25966 was prepared in our previous work starting from amino acid (D)-valine. The biodistribution of [11C]MSMA and [11C]CGS 25966 were determined at 45 min post iv injection in breast cancer animal models MCF-7's transfected with IL-1alpha implanted athymic mice and MDA-MB-435 implanted athymic mice. The results showed the uptakes of [11C]MSMA and [11C]CGS 25966 in these tumors were 0.95 and 0.42%dose/g in MCF-7's transfected with IL-1alpha implanted mice, 0.98 and 1.53%dose/g in MDA-MB-435 implanted mice, respectively; the ratios of tumor/muscle (T/M) and tumor/blood (T/B) were 1.21 and 1.09 (T/M, MCF-7's), 0.99 and 0.84 (T/B, MCF-7's), 1.38 and 1.27 (T/M, MDA-MB-435), 1.27 and 1.95 (T/B, MDA-MB-435), respectively. The micro-PET images of [11C]MSMA and [11C]CGS 25966 in both breast cancer athymic mice were acquired for 15 min from a MCF-7's transfected with IL-1alpha and/or MDA-MB-435 implanted mouse at 45 min post iv injection of 1 mCi of the tracer using a dedicated high resolution (<3 mm full-width at half-maximum) small FOV (field-of-view) PET imaging system, Indy-PET II scanner, developed in our laboratory, which showed both tumors were invisible with both tracers. The results were compared. From our results, we concluded that both [11C]MSMA and [11C]CGS 25966 might be unsuitable as PET tracers for cancer imaging.     

Synthesis, biodistribution and micro-PET imaging of a potential cancer biomarker carbon-11 labeled MMP inhibitor (2R)-2-[[4-(6-fluorohex-1-ynyl)phenyl]sulfonylamino]-3-methylbutyric acid [11C]methyl ester

(2R)-2-[[4-(6-fluorohex-1-ynyl)phenyl]sulfonylamino]-3-methylbutyric acid [(11)C]methyl ester ([(11)C]FMAME), a novel carbon-11 labeled matrix metalloproteinase (MMP) inhibitor, has been synthesized for evaluation as new potential positron emission tomography (PET) cancer biomarker. [(11)C]FMAME was prepared by appropriate precursor (2R)-2-[[4-(6-fluorohex-1-ynyl)phenyl]sulfonylamino]-3-methylbutyric acid (FMA), which was synthesized in six steps from (D)-valine in 71% chemical yield. This acid precursor was labeled by [(11)C]methyl triflate through O-[(11)C]methylation method under basic conditions and isolated by solid-phase extraction (SPE) purification to produce pure target compound in 40-55% radiochemical yield, based on (11)CO(2), decay corrected to end of bombardment, and 15-20 min synthesis time. The biodistribution of [(11)C]FMAME was determined at 30 min post IV injection in breast cancer animal models MCF-7 transfected with IL-1 alpha implanted athymic mice and MDA-MB-435 implanted athymic mice. The results showed the uptakes of [(11)C]FMAME in these tumors were 1.13% dose/g in MCF-7 transfected with IL-1 alpha implanted mice and 1.37% dose/g in MDA-MB-435 implanted mice, respectively; the ratios of tumor/muscle (T/M) and tumor/blood (T/B) were 1.05 +/- 0.29 (T/M, MCF-7's), 0.77 +/- 0.20 (T/B, MCF-7's) and 0.99 +/- 0.35 (T/M, MDA-MB-435), 1.44 +/- 0.69 (T/B, MDA-MB-435), respectively. Pretreatment of MCF-7 transfected with IL-1 alpha tumor-bearing mice with MMP inhibitor FMA had no effect on [(11)C]FMAME biodistribution. Likewise, pretreatment of MDA-MB-435 tumor-bearing mice with FMA also showed no effect on [(11)C]FMAME biodistribution. The micro-PET images were acquired for 15 min from a MCF-7 transfected with IL-1 alpha tumor-bearing mouse or a MDA-MB-435 tumor-bearing mouse at 30 min post IV injection of 1 mCi of [(11)C]FMAME using a dedicated high resolution (<3 mm full-width at half-maximum) PET imaging system (Indy-PET II scanner). The initial dynamic micro-PET images of [(11)C]FMAME in a MCF-7 transfected with IL-1 alpha tumor-bearing mouse during different time periods of 0-15, 15-30, 30-45 and 45-60 min were performed by Indy-PET II. The PET images clearly showed both tumors were visible with [(11)C]FMAME. These results suggest that the localization of [(11)C]FMAME in the tumor is mediated by non-specific processes, and the visualization of [(11)C]FMAME on the tumor using the Indy-PET II scanner is related to non-specific binding.     

Effect of the EGFR density of breast cancer cells on nuclear importation, in vitro cytotoxicity, and tumor and normal-tissue uptake of [111In]DTPA-hEGF

INTRODUCTION: Our objective was to evaluate the effect of epidermal growth factor receptor(s) (EGFR) density on the importation and nuclear localization of 111In-labeled diethylenetriaminepentaacetic acid human epidermal growth factor ([111In]DTPA-hEGF) in breast cancer (BC) cells in vitro and in tumor xenografts and normal tissues in vivo in athymic mice, as well as on its cytotoxicity and tumor and normal-tissue distribution. METHODS: The internalization and nuclear importation of [111In]DTPA-hEGF were measured in MCF-7, MDA-MB-231, BT-474 and MDA-MB-468 BC cells (10(4), 2 x 10(5), 6 x 10(5) and 10(6) EGFR/cell, respectively). The molecular size (Mr) distribution and immunoreactivity of nuclear radioactivity were characterized. Tumor and normal-tissue uptake of [111In]DTPA-hEGF in athymic mice implanted subcutaneously with BC xenografts were compared. Nuclear radioactivity in the tumor, lungs, liver, kidneys, spleen and colon was measured. RESULTS: There was a direct association between EGFR density and the nuclear localization of [111In]DTPA-hEGF in BC cells; nuclear importation approached saturation at 6 x 10(5) EGFR/cell. Almost all nuclear radioactivities exhibited an Mr of >100 kDa; immunoreactivity with anti-hEGF, anti-EGFR and anti-importin beta 1 antibodies was detected. The efflux of nuclear radioactivity was slowest for MDA-MB-468 cells. Cytotoxicity was correlated with EGFR expression. Uptake was greater in MDA-MB-468 than in MCF-7 xenografts and improved with preinjection of a 100-fold excess of unlabeled DTPA-hEGF. Nuclear importation was higher in liver, kidney and spleen cells than in tumor cells. CONCLUSION: [111In]DTPA-hEGF is translocated to the nucleus of BC cells complexed with EGFR and importin beta1. Nuclear importation and cytotoxicity are effected by EGFR density. The absence of hepatic and renal toxicities in [111In]DTPA-hEGF cannot be explained by a low efficiency of nuclear importation.     

A comparison of EGF and MAb 528 labeled with 111In for imaging human breast cancer.

Our objective was to compare 111In-labeled human epidermal growth factor (hEGF), a 53-amino acid peptide with anti-epidermal growth factor receptor (EGFR) monoclonal antibody (MAb) 528 (IgG2a) for imaging EGFR-positive breast cancer. METHODS: hEGF and MAb 528 were derivatized with diethylenetriamine pentaacetic acid (DTPA) and labeled with 111In acetate. Receptor binding assays were conducted in vitro against MDA-MB-468 human breast cancer cells. Biodistribution and tumor imaging studies were conducted after intravenous injection of the radiopharmaceuticals in athymic mice bearing subcutaneous MCF-7, MDA-MB-231, or MDA-MB-468 human breast cancer xenografts or in severe combined immunodeficiency mice implanted with a breast cancer metastasis (JW-97 cells). MCF-7, MDA-MB-231, JW-97, and MDA-MB-468 cells expressed 1.5 x 10(4), 1.3 x 10(5), 2.7 x 10(5), and 1.3 x 106 EGFR/cell, respectively in vitro. RESULTS: 111In-DTPA-hEGF and 111In-DTPA-MAb 528 bound with high affinity to MDA-MB-468 cells (Ka of 7.5 x 10(8) and 1.2 x 10(8) L/mol, respectively). 111In-DTPA-hEGF was eliminated rapidly from the blood with < 0.2% injected dose/g (%ID/g) circulating at 72 h after injection, whereas 111In-DTPA-MAb 528 was cleared more slowly (3%ID/g in the blood at 72 h). Maximum localization of 111In-DTPA-hEGF in MDA-MB-468 tumors (2.2 %ID/g) was 10-fold lower than with 111In-DTPA-MAb 528 (21.6 %ID/g). There was high uptake in the liver and kidneys for both radiopharmaceuticals. Tumor-to-blood ratios were greater for 111In-labeled hEGF than for MAb 528 (12:1 versus 6:1), but all other tumor-to-normal tissue ratios were higher for MAb 528. MDA-MB-468 and JW-97 tumors were imaged successfully with both radiopharmaceuticals, but tumors were more easily visualized using 111In-labeled MAb 528. There was no direct quantitative relationship between EGFR expression on breast cancer cell lines in vitro, and tumor uptake of the radiopharmaceuticals in vivo, but control studies showed that tumor uptake was receptor mediated. CONCLUSION: Our results suggest that the tumor uptake in vivo of receptor-binding radiopharmaceuticals is controlled to a greater extent by their elimination rate from the blood than by the level of receptor expression on the cancer cells. Radiolabeled anti-EGFR MAbs would be more effective for tumor imaging in cancer patients than peptide-based radiopharmaceuticals such as hEGF, because they exhibit higher tumor uptake at only moderately lower tumor-to-blood ratios.     

[11C]Choline as pharmacodynamic marker for therapy response assessment in a prostate cancer xenograft model

Purpose

[¹¹C]Choline has been established as a PET tracer for imaging prostate cancer. The aim of this study was to determine whether [¹¹C]choline can be used for monitoring the effects of therapy in a prostate cancer mouse xenograft model.

Methods

The androgen-independent human prostate cancer cell line PC-3 was implanted subcutaneously into the flanks of 13 NMRI (nu/nu) mice. All mice were injected 4–6 weeks after xenograft implantation with 37 MBq [¹¹C]choline via a tail vein. Dynamic imaging was performed for 60 min with a small-animal PET/CT scanner (Siemens Medical Solutions). Six mice were subsequently injected intravenously with docetaxel twice (days 1 and 5) at a dose of 3 mg/kg body weight. Seven mice were treated with PBS as a control. [¹¹C]Choline imaging was performed prior to and 1, 2 and 3 weeks after treatment. To determine choline uptake the images were analysed in terms of tumour-to-muscle (T/M) ratios. Every week the size of the implanted tumour was determined with a sliding calliper.

Results

The PC-3 tumours could be visualized by [¹¹C]choline PET. Before treatment the T/M_mean ratio was 1.6±0.5 in the control group and 1.8±0.4 in the docetaxel-treated group (p=0.65). There was a reduction in the mean [¹¹C]choline uptake after docetaxel treatment as early as 1 week after initiation of therapy (T/M ratio 1.8±0.4 before treatment, 0.9±0.3 after 1 week, 1.1±0.3 after 2 weeks and 0.8±0.2 after 3 weeks). There were no decrease in [¹¹C]choline uptake in the control group following treatment (T/M ratio 1.6±0.5 before treatment, 1.7±0.4 after 1 week, 1.8±0.7 after 2 weeks and 1.7±0.4 after 3 weeks). For analysis of the dynamic data, a generalized estimation equation model revealed a significant decrease in the T/M_dyn ratios 1 week after docetaxel treatment, and the ratio remained at that level through week 3 (mean change ?0.93±0.24, p<0.001, after 1  week; ?0.78±0.21, p<0.001, after 2 weeks; ?1.08±0.26, p<0.001, after 3 weeks). In the control group there was no significant decrease in the T/M_dyn ratios (mean change 0.085±0.39, p=0.83, after 1 week; 0.31±0.48, p=0.52, after 2 weeks; 0.11±0.30, p=0.72, after 3 weeks). Metabolic changes occurred 1 week after therapy and preceded morphological changes of tumour size during therapy.

Conclusion

Our results demonstrate that [¹¹C]choline has the potential for use in the early monitoring of the therapeutic effect of docetaxel in a prostate cancer xenograft animal model. The results also indicate that PET with radioactively labelled choline derivatives might be a useful tool for monitoring responses to taxane-based chemotherapy in patients with advanced prostate cancer.     

Investigations into tumor accumulation and peroxisome proliferator activated receptor binding by F-18 and C-11 fatty acids

[11C]Acetate, a myocardial PET imaging agent for analysis of oxidative metabolism, has potential use in tumor imaging. Aromatic fatty acids display antitumor effects with phenylacetate currently in clinical trial. Tumor differentiation and cytostasis resulting from phenylacetate treatment may involve the peroxisome proliferator-activated receptor alpha (PPARalpha). To examine whether aromatic fatty acids are potential imaging agents for PPARalpha or tumors in general, [11C]phenylacetic acid (PAA) and [18F]fluorophenyl-acetic acid (FPAA) were synthesized and evaluated in EMT-6 tumor bearing mice and 9L-Glioma tumor bearing rats and compared to [11C]acetate. [11C]Acetate showed better tumor accumulation than PAA or FPAA. The aromatic fatty acids did not directly bind PPARalpha as confirmed by a biodistribution study of PAA in PPARalpha -/- mice.     

Carbon-11 labeled sigma2 receptor ligands for imaging breast cancer

Four conformationally flexible benzamide analogs having a high affinity and outstanding selectivity for sigma(2) versus sigma(1) receptors were synthesized and radiolabeled with carbon-11 by reaction with [(11)C]methyl iodide. The four (11)C-labeled radiotracers were evaluated for their potential to image the proliferative status of breast tumors with positron emission tomography (PET). In vivo studies in female BALB/C mice bearing EMT-6 breast tumors showed that one radiotracer, (2-methoxy-(11)C)-N-(4-(3,4-dihydro-6,7-dimethoxy-isoquinolin-2(1H)-yl)butyl)-5-methylbenzamide ([(11)C]2), had a high tumor uptake and suitable tumor/background ratio for imaging purposes. Blocking studies were consistent with the labeling of sigma(2) receptors in vivo. A study comparing the in vivo properties of [(11)C]2 and (18)F-3'-fluoro-3'-deoxy-L-thymidine ([(18)F]FLT) indicated that [(11)C]2 had either similar (lung, fat) or better (blood, muscle) tumor/organ ratios than [(18)F]FLT in the tissues that are important for breast tumor imaging. Consequently, [(11)C]2 is a potential radiotracer for imaging the proliferative status of breast tumors in vivo with PET.     

Synthesis and preclinical evaluation of the choline transport tracer deshydroxy-[18F]fluorocholine ([18F]dOC)

11C-labeled choline ([11C]CHO) and 18F-fluorinated choline analogues have been demonstrated to be valuable tracers for in vivo imaging of neoplasms by means of positron emission tomography (PET). The objective of the present study was to evaluate whether deshydroxy-[18F]fluorocholine, ([18F]dOC), a non-metabolizable [18F]fluorinated choline analogue, can serve as a surrogate for cholines that are able to be phosphorylated and thus allow PET-imaging solely by addressing the choline transport system. The specificity of uptake of [18F]dOC was compared with that of [11C]choline ([11C]CHO) in cultured rat pancreatic carcinoma and PC-3 human prostate cancer cells in vitro. In addition, biodistribution of [18F]dOC and [11C]CHO was compared in AR42J- and PC-3 tumor bearing mice. The in vitro studies revealed that membrane transport of both compounds can be inhibited in a concentration dependent manner by similar concentrations of cold choline (IC50 [18F]dOC= 11 microM; IC50 [11C]CHO=13 microM. In vitro studies with PC-3 and AR42J cells revealed that the internalized fraction of [18F]dOC after 5 min incubation time is comparable to that of [11C]CHO, whereas the uptake of [11C]CHO was superior after 20 min incubation time. As for [11C]CHO, kidney and liver were also the primary sites of uptake for [18F]dOC in vivo. Biodistribution data after simultaneous injection of both tracers into AR42J tumor bearing mice revealed slightly higher tumor uptake for [18F]dOC at 10 min post-injection, whereas [11C]CHO uptake was higher at later time points. In conclusion, [18F]dOC is taken up into AR42J rat pancreatic carcinoma and PC-3 human prostate cancer cells by a choline specific transport system. Similar transport rates of [18F]dOC and [11C]CHO result in comparable cellular uptake levels at early time points. In contrast to [18F]dOC, which is transported but not intracellularly trapped, the choline kinase substrate [11C]CHO is transported into tumor cells and retained. Thus, the signal obtained by imaging early after injection is mainly reflecting transport, whereas a valid quantification of choline kinase activity needs imaging at later time points. Further studies have to clarify whether quantification of the transport capacity or the choline kinase activity result in a better pathophysiological correlate and thus is the more useful process for tumor characterization.     

The level of insulin growth factor-1 receptor expression is directly correlated with the tumor uptake of (111)In-IGF-1(E3R) in vivo and the clonogenic survival of breast cancer cells exposed in vitro to trastuzumab (Herceptin)

INTRODUCTION: Our objective was to define the relationships between tumor uptake of [(111)In]-IGF-1 and [(111)In]-IGF-1(E3R), an analogue which does not bind insulin growth factor-1 (IGF-1) binding proteins (i.e., IGFBP-3), and the level of IGF-1 receptor (IGF-1R) expression on human breast cancer (BC) xenografts in athymic mice, as well as the feasibility for tumor imaging. A second objective was to correlate IGF-1R (and HER2 density) with the cytotoxicity of trastuzumab in the absence/presence of IGFBP-3 or the IGF-1R tyrosine kinase inhibitor, AG1024. METHODS: The tumor and normal tissue uptake of [(111)In]-IGF-1 and [(111)In]-IGF-1(E3R) were determined at 4 h postinjection in mice implanted subcutaneously with MDA-MB-231, H2N, HR2 or MCF-7/HER2-18 human BC xenografts (8.5x10(4), 1.4x10(4), 4.0x10(4) and 1.0x10(5) IGF-1R/cell, respectively). The effect of co-injection of IGF-1 (50 microg) or IGFBP-3 (2 or 25 microg) was studied. The relationship between tumor uptake of [(111)In]-IGF-1(E3R) and IGF-1R density was examined. MicroSPECT/CT imaging was performed on mice with MCF-7/HER2-18 tumors injected with [(111)In]-IGF-1(E3R). The surviving fraction of BC cells exposed to trastuzumab (67.5 mug/ml) in the absence/presence of IGFBP-3 (1 microg/ml) or the IGF-1R kinase inhibitor, AG1024 (1 or 5 microg/ml), was determined. RESULTS: [(111)In]-IGF-1 was specifically taken up by MCF-7/HER2-18 xenografts; tumor uptake was decreased twofold when co-injected with IGF-1 (1.9+/-0.1 vs. 1.0+/-0.1 %ID/g). Co-injection of IGBP-3 decreased kidney uptake of [(111)In]-IGF-1 up to twofold and increased circulating radioactivity threefold. There was a strong linear correlation (r(2)=0.99) between the tumor uptake of (111)In-IGF-1(E3R) and IGF-1R density. Tumor uptake ranged from 0.4+/-0.05 %ID/g for H2N to 2.5+/-0.5 %ID/g for MCF-7/HER2-18 xenografts. MCF-7/HER2-18 tumors were visualized by microSPECT/CT. Resistance of BC cells to trastuzumab was directly associated with IGF-1R expression, despite co-expression of HER2. The resistance of HR2 cells could be partially reversed by IGFBP-3 or AG1024. CONCLUSION: Imaging of IGF-1R expression using [(111)In]-IGF-1(E3R) may be useful for identifying HER2-positive tumors in BC patients that are resistant to trastuzumab through this mechanism.     

Prostate carcinoma: diffusion-weighted imaging as potential alternative to conventional MR and 11C-choline PET/CT for detection of bone metastases

In a technical development study approved by the institutional ethics committee, the feasibility of fast diffusion-weighted imaging as a replacement for conventional magnetic resonance (MR) imaging sequences (short inversion time inversion recovery [STIR] and T1-weighted spin echo [SE]) and positron emission tomography (PET)/computed tomography (CT) in the detection of skeletal metastases from prostate cancer was evaluated. MR imaging and carbon 11 ((11)C) choline PET/CT data from 11 consecutive prostate cancer patients with bone metastases were analyzed. Diffusion-weighted imaging appears to be equal, if not superior, to STIR and T1-weighted SE sequences and equally as effective as (11)C-choline PET/CT in detection of bone metastases in these patients. Diffusion-weighted imaging should be considered for further evaluation and comparisons with PET/CT for comprehensive whole-body staging and restaging in prostate and other cancers.     

124I-labeled engineered anti-CEA minibodies and diabodies allow high-contrast, antigen-specific small-animal PET imaging of xenografts in athymic mice.

Prolonged clearance kinetics have hampered the development of intact antibodies as imaging agents, despite their ability to effectively deliver radionuclides to tumor targets in vivo. Genetically engineered antibody fragments display rapid, high-level tumor uptake coupled with rapid clearance from the circulation in the athymic mouse/LS174T xenograft model. The anticarcinoembryonic antigen (CEA) T84.66 minibody (single-chain Fv fragment [scFv]-C(H)3 dimer, 80 kDa) and T84.66 diabody (noncovalent dimer of scFv, 55 kDa) exhibit pharmacokinetics favorable for radioimmunoimaging. The present work evaluated the minibody or diabody labeled with (124)I, for imaging tumor-bearing mice using a high-resolution small-animal PET system. METHODS: Labeling was conducted with 0.2-0.3 mg of protein and 65-98 MBq (1.7-2.6 mCi) of (124)I using an iodination reagent. Radiolabeling efficiencies ranged from 33% to 88%, and immunoreactivity was 42% (diabody) or >90% (minibody). In vivo distribution was evaluated in athymic mice bearing paired LS174T human colon carcinoma (CEA-positive) and C6 rat glioma (CEA-negative) xenografts. Mice were injected via the tail vein with 1.9-3.1 MBq (53-85 microCi) of (124)I-minibody or with 3.1 MBq (85 microCi) of (124)I-diabody and imaged at 4 and 18 h by PET. Some mice were also imaged using (18)F-FDG 2 d before imaging with (124)I-minibody. RESULTS: PET images using (124)I-labeled minibody or diabody showed specific localization to the CEA-positive xenografts and relatively low activity elsewhere in the mice, particularly by 18 h. Target-to-background ratios for the LS174T tumors versus soft tissues using (124)I-minibody were 3.05 at 4 h and 11.03 at 18 h. Similar values were obtained for the (124)I-diabody (3.95 at 4 h and 10.93 at 18 h). These results were confirmed by direct counting of tissues after the final imaging. Marked reduction of normal tissue activity, especially in the abdominal region, resulted in high-contrast images at 18 h for the (124)I-anti-CEA diabody. CEA-positive tumors as small as 11 mg (<3 mm in diameter) could be imaged, and (124)I-anti-CEA minibodies, compared with (18)F-FDG, demonstrated highly specific localization. CONCLUSION: (124)I labeling of engineered antibody fragments provides a promising new class of tumor-specific probes for PET imaging of tumors and metastases.     

Automated synthesis of [11C]choline, a positron-emitting tracer for tumor imaging.

(beta-Hydroxyethyl)tri([11C]methyl)ammonium ([11C]choline) is a tracer very effective in imaging various human tumors using positron emission tomography (PET). We have constructed a computer-controlled [11C]choline synthetic apparatus which carries out the whole process of synthesis and product purification automatically. The setup is simple and the process quick. In 20 min, 11 GBq of [11C]choline (chloride) is obtainable from 26 GBq of [11C]CO2. The final product is a sterile and pyrogen-free [11C]choline "injection".     

Comparison of integrated whole-body [11C]choline PET/MR with PET/CT in patients with prostate cancer

Purpose

To evaluate the performance of conventional [¹¹C]choline PET/CT in comparison to that of simultaneous whole-body PET/MR.

Methods

The study population comprised 32 patients with prostate cancer who underwent a single-injection dual-imaging protocol with PET/CT and subsequent PET/MR. PET/CT scans were performed applying standard clinical protocols (5 min after injection of 793?±?69 MBq [¹¹C]choline, 3 min per bed position, intravenous contrast agent). Subsequently (52?±?15 min after injection) PET/MR was performed (4 min per bed position). PET images were reconstructed iteratively (OSEM 3D), scatter and attenuation correction of emission data and regional allocation of [¹¹C]choline foci were performed using CT data for PET/CT and segmented Dixon MR, T1 and T2 sequences for PET/MR. Image quality of the respective PET scans and PET alignment with the respective morphological imaging modality were compared using a four point scale (0–3). Furthermore, number, location and conspicuity of the detected lesions were evaluated. SUVs for suspicious lesions, lung, liver, spleen, vertebral bone and muscle were compared.

Results

Overall 80 lesions were scored visually in 29 of the 32 patients. There was no significant difference between the two PET scans concerning number or conspicuity of the detected lesions (p not significant). PET/MR with T1 and T2 sequences performed better than PET/CT in anatomical allocation of lesions (2.87?±?0.3 vs. 2.72?±?0.5; p?=?0.005). The quality of PET/CT images (2.97?±?0.2) was better than that of the respective PET scan of the PET/MR (2.69?±?0.5; p?=?0.007). Overall the maximum and mean lesional SUVs exhibited high correlations between PET/CT and PET/MR (ρ?=?0.87 and ρ?=?0.86, respectively; both p?<?0.001).

Conclusion

Despite a substantially later imaging time-point, the performance of simultaneous PET/MR was comparable to that of PET/CT in detecting lesions with increased [¹¹C]choline uptake in patients with prostate cancer. Anatomical allocation of lesions was better with simultaneous PET/MR than with PET/CT, especially in the bone and pelvis. These promising findings suggest that [¹¹C]choline PET/MR might have a diagnostic benefit compared to PET/CT in patients with prostate cancer, and now needs to be further evaluated in prospective trials.     

Differential routing of choline in implanted breast cancer and normal organs.

Choline is an essential nutrient participating as the initial substrate in major metabolic pathways. The differential metabolic routing of choline was investigated in MCF7 human breast cancer implanted in nude mice and in the kidney, liver, and brain of these mice. The distribution of metabolites following infusion of [1,2-(13)C]-choline was monitored by (13)C magnetic resonance spectroscopy. This infusion led to an 18-fold increase in plasma choline and to concomitant changes in the content and distribution of choline metabolites. In vivo kinetic studies of the tumor during the infusion demonstrated accumulation of choline in the interstitium and intracellular synthesis of phosphocholine. The amount of unlabeled choline metabolites was 7.1, 4.1, 3.5, and 1.4 micromol/g in the kidney, liver, tumor, and brain, respectively. The variations in the labeled metabolites were more pronounced with high amounts in the kidney and liver (8.0 and 4.3 micromol/g, respectively) and very low amounts in the tumor and brain (0.33 and 0.12 micromol/g, respectively). In the kidney and liver, betaine (unlabeled and labeled) was the predominant choline metabolite. The dominant unlabeled metabolite in breast cancer was phosphocholine and in the brain glycerophosphocholine. Magn Reson Med 46:31-38, 2001.     

Evaluation of biodistribution and anti-tumor effect of a dimeric RGD peptide–paclitaxel conjugate in mice with breast cancer

PURPOSE: Targeting drugs to receptors involved in tumor angiogenesis has been demonstrated as a novel and promising approach to improve cancer treatment. In this study, we evaluated the anti-tumor efficacy of a dimeric RGD peptide-paclitaxel conjugate (RGD2-PTX) in an orthotopic MDA-MB-435 breast cancer model. METHODS: To assess the effect of conjugation and the presence of drug moiety on the MDA-MB-435 tumor and normal tissue uptake, the biodistribution of (3)H-RGD2-PTX was compared with that of (3)H-PTX. The treatment effect of RGD2-PTX and RGD2+PTX was measured by tumor size, (18)F-FDG/PET, (18)F-FLT/PET, and postmortem histopathology. RESULTS: By comparing the biodistribution of (3)H-RGD2-PTX and (3)H-PTX, we found that (3)H-RGD2-PTX had higher initial tumor exposure dose and prolonged tumor retention than (3)H-PTX. Metronomic low-dose treatment of breast cancer indicated that RGD2-PTX is significantly more effective than PTX+RGD2 combination and solvent control. Although in vivo (18)F-FLT/PET imaging and ex vivo Ki67 staining indicated little effect of the PTX-based drug on cell proliferation, (18)F-FDG/PET imaging showed significantly reduced tumor metabolism in the RGD2-PTX-treated mice versus those treated with RGD2+PTX and solvent control. Terminal uridine deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining also showed that RGD2-PTX treatment also had significantly higher cell apoptosis ratio than the other two groups. Moreover, the microvessel density was significantly reduced after RGD2-PTX treatment as determined by CD31 staining. CONCLUSION: Our results demonstrate that integrin-targeted delivery of paclitaxel allows preferential cytotoxicity to integrin-expressing tumor cells and tumor vasculature. The targeted delivery strategies developed in this study may also be applied to other chemotherapeutics for selective tumor killing.     

In vitro and in vivo investigation of matrix metalloproteinase expression in metastatic tumor models

INTRODUCTION: Overexpression of matrix metalloproteinases (MMPs), particularly MMP-2 and MMP-9, has been correlated with poor prognosis in several cancer types including lung, colon and breast. Noninvasive detection of MMP expression might allow physicians to better determine when more aggressive cancer therapy is appropriate. The peptide CTT (CTTHWGFTLC) was identified as a selective inhibitor of MMP-2/9 that inhibits the growth of MDA-MB-435 human breast cancer xenografts. METHODS: CTT was conjugated with the bifunctional chelator DOTA (1,4,7,10-tetraazacyclotetradecane-N,N',N',N'-tetraacetic acid) for radiolabeling with (64)Cu (t(1/2)=12.7 h, 17.4% beta(+), 39% beta(-)), a radionuclide suitable for positron emission tomography (PET). In vitro affinity was determined in a fluorogenic substrate assay. Tumor gelatinase targeting was evaluated in both biodistribution and microPET imaging studies. RESULTS: Cu(II)-DOTA-CTT inhibited hMMP-2 (EC(50)=8.7 microM) and mMMP-9 (EC(50)=18.2 microM) with similar affinity to CTT (hMMP-2 EC(50)=13.2 microM; mMMP-9 EC(50)=11.0 microM). In biodistribution and microPET imaging studies, (64)Cu-DOTA-CTT was taken up by MMP-2/9-positive B16F10 murine melanoma tumors. Subsequently, imaging studies using (64)Cu-DOTA-CTT were performed on MDA-MB-435 tumor-bearing mice. With zymography, tumor MMP-2/9 expression in this model was shown to be inconsistent, resulting in microPET detection of the MDA-MB-435 tumor in only 1 of 24 imaged mice. Following limited imaging success, (64)Cu-DOTA-CTT was shown to have poor in vivo stability. CONCLUSIONS: Despite some evidence for selective uptake of (64)Cu-DOTA-CTT by gelatinase-expressing tumors, the low affinity for MMP-2 and MMP-9 and in vivo instability make this an inadequate radioligand for in vivo tumor evaluation.     

Syk表达对乳腺肿瘤细胞侵袭迁移作用的影响

目的:探讨脾源性酪氨酸激酶(spleen tyrosine kinase, Syk)在人乳腺癌细胞中的表达及其对肿瘤细胞侵袭迁移能力的影响.方法:RT-PCR法、免疫荧光法检测不同侵袭能力乳腺癌细胞株--MCF-7(低侵袭性)和MDA-MB-231(高侵袭性) Syk mRNA及蛋白的表达;采用脂质体介导的方法,将Syk基因导入Syk(-)乳腺癌细胞株,利用Transwell小室法检测Syk基因对肿瘤细胞侵袭迁移能力的影响.结果:Syk mRNA及蛋白在MCF-7细胞中呈中等程度表达,在MDA-MB-231中表达缺失;转染Syk基因的MDA-MB-231细胞与转染空载体及未转染MDA-MB-231细胞相比,侵袭及迁移实验中的穿膜细胞数(总迁移细胞数/5HPF)均明显减少(P＜0.05).结论:Syk表达可降低乳腺肿瘤细胞体外侵袭迁移能力.     

Breast cancer models to study the expression of estrogen receptors with small animal PET imaging

Different animal models of estrogen positive tumors (ER+) were evaluated for their suitability to follow tumor response after various treatment protocols, using small animal positron emission tomography (PET). ER+ human breast cancer cell lines MCF-7 and T-47D, using MDA-MB-231 as ER-; control, and murine mammary ductal carcinomas MC4-L2, MC4-L3, and MC7-L1, were compared for their in vivo growth rate and retention of ER+ status. Tumor metabolic activity was estimated from the relative uptake (% injected dose/g) of [18F]fluorodeoxyglucose (FDG) uptake, whereas ER content was determined from 16alpha-[18F]fluoroestradiol (FES) retention. F-18 activity values were obtained by small animal PET imaging and confirmed by tissue sampling and radioactivity counting. Reliable uptake measurements could be obtained for tumors of 200 microl or over. The human cell lines grew at a slower rate in vivo and failed to accumulate FES; in contrast, the Balb/c MC7-L1 and MC4-L2 grew well and showed good uptake of both FDG and FES. Chemotherapy and hormone therapy delayed the growth of MC7-L1 and MC4-L2 tumors, confirming their suitability as an ER+ model for therapeutic interventions. MC4-L3 tumors also showed promising results but required the presence of progestative pellets to grow. These data demonstrate that murine MC7-L1 and MC4-L2 tumors are suitable models for the monitoring of ER+ breast cancer therapy using small animal PET imaging.     

Value of integrated PET/CT for lesion localisation in cancer patients: a comparative study

The aim of this study was to retrospectively compare the value of integrated PET/CT and separate PET plus morphological imaging studies for lesion localisation in cancer patients. Two different series of consecutive patients who had previously been treated for neoplastic disease were considered. One series consisted of 105 patients who had undergone [¹⁸F]fluorodeoxyglucose (FDG) PET/CT (n=70) or [¹¹C]choline PET/CT (n=35) studies (PET/CT group). The other series comprised 105 patients who had undergone FDG PET scan (n=70) or [¹¹C]choline PET scan (n=35) alone; in this series, PET findings were correlated with the results of morphological imaging (MI) studies, i.e. CT (n=92) or MR imaging (n=13) (PET+MI group). Regions of abnormal tracer uptake at PET scanning were classified as ambiguous or unambiguous depending on their precise anatomical localisation. A total of 207 and 196 lesions were found in the PET/CT and PET+MI groups, respectively. The difference in terms of number of lesions per patient detected with the two imaging protocols was not statistically significant (P=0.718). When analysis of lesion localisation was performed, there were 7/207 (3.4%) and 30/196 (15.3%) ambiguous lesions in the PET/CT and PET+MI groups, respectively. The number of ambiguous lesions was significantly higher in the PET+MI group than in the PET/CT group (²=15.768, P<0.0001). Comparison of the effect of use of the different tracers on reporting of PET/CT versus PET+MI revealed that the improvement in the final report in [¹¹C]choline PET/CT studies was similar to that observed in [¹⁸F]FDG studies. In cancer patients, PET/CT shows higher diagnostic accuracy for lesion localisation than PET plus morphological imaging studies performed independently. This result does not seem to be affected by the type of tracer used.