Synthesis, crystal structure and effect of indeno[1,2-b]indole derivatives on prostate cancer in vitro. Potential effect against MMP-9.

A highly regiospecific synthesis of a series of indenoindoles is reported, together with X-ray studies and their activity against human prostate cancer cells PC-3 and LNCaP in vitro. The most effective compound 7,7-dimethyl-5-[(3,4-dichlorophenyl)]-(4bRS,9bRS)-dihydroxy-4b,5,6,7,8,9bhexahydro-indeno[1,2-b]indole-9,10-dione 7q reduced the viability in both cell lines in a time and dose-dependent manner. Inhibitory effects were also observed on the adhesion, migration, and invasion of the prostate cancer cells as well as on clonogenic possibly by inhibition of MMP-9 activity. Molecular docking of 7q and 6k into MMP-9 human active site was also performed to determine the probable binding mode.

N'-Formyl-2-(5-nitrothiophen-2-yl)benzothiazole-6-carbohydrazide as a potential anti-tumour agent for prostate cancer in experimental studies.

OBJECTIVES: Benzothiazoles (BZTs) represent organic compounds with different biological actions. In this study we aimed to investigate ten newly synthesized BZT derivatives as potential anti-tumour agents against prostate cancer in vitro and in vivo. METHODS: The cytotoxic effect of these compounds was screened on the human prostate cancer cell lines PC-3 and LNCaP. The most effective compound, N'-formyl-2-(5-nitrothiophen-2-yl)benzothiazole-6-carbohydrazide, was further characterized regarding its dose- and time-dependent effects on cell viability and proliferation (XTT test) as well as on adhesion and spreading (real-time cell analyzer xCelligence), migration (scratch-wound repair assay) and invasion (Boyden chamber) of the cells. This BZT derivative was also tested as an inhibitor of angiogenesis (chicken chorioallantoic membrane assay), clonogenic activity (soft agar) and matrix metalloproteinase 9 (gelatin zymography). KEY FINDINGS: N'-Formyl-2-(5-nitrothiophen-2-yl)benzothiazole-6-carbohydrazide significantly inhibited all tested properties of the prostate cancer cell lines and showed low toxic in vitro and in vivo effects. The in vitro anti-tumour activity of this compound was confirmed by the in vivo effects on PC-3 xenografts in nude mice. Tumour growth was decreased in treated compared with untreated mice. CONCLUSIONS: These results suggest the potential capacity of BZTs and in particular N'-formyl-2-(5-nitrothiophen-2-yl)benzothiazole-6-carbohydrazide as anti-tumour agents for the treatment of prostate cancer.

Human prostate cancer in a clinically relevant xenograft mouse model: identification of ß(1,6)-branched oligosaccharides as a marker of tumor progression.

PURPOSE: To establish xenograft mouse models of metastatic and nonmetastatic human prostate cancer and to apply these models to the search for aberrant glycosylation patterns associated with tumor progression in vivo and in patients. EXPERIMENTAL DESIGN: Prostate cancer cells (LNCaP, PC-3, LuCaP 23.1, and DU-145) were xenografted subcutaneously into immunodeficient pfp(-/-)/rag2(-/-) mice. Tumor growth and metastasis formation were quantified and as altered glycosylation patterns have been associated with metastasis formation in several other malignancies, prostate cancer cells were profiled by a quantitative real-time PCR (qRT-PCR) glycosylation array and compared with normal human prostate cells. The activity of upregulated glycosyltransferases was analyzed by their sugar residues end products using lectin histochemistry on primary tumors and metastases in the animal experiments and on 2,085 clinical samples. RESULTS: PC-3 cells produced the largest number of spontaneous lung metastases, followed by LNCaP and LuCaP 23.1, whereas DU-145 was nonmetastatic. qRT-PCR revealed an upregulation of ß1,6-N-acetylglucosaminyltransferase-5b (Mgat5b) in all prostate cancer cell lines. Mgat5b products [ß(1,6)-branched oligosaccharides] were predominantly detectable in metastatic xenografts as shown by increased binding of Phaseolus vulgaris leukoagglutinin (PHA-L). The percentage of prostate cancer patients who were PHA-L positive was 86.5. PHA-L intensity correlated with serum prostate-specific antigen and a cytoplasmic staining negatively affected disease-free survival. CONCLUSION: We show a novel xenograft mouse model for human prostate cancer respecting the complete metastatic cascade. Specific glycosylation patterns reveal Mgat5b products as relevant markers of both metastatic competence in mice and disease-free survival in patients. This is the first description of Mgat5b in prostate cancer indicating a significant biologic importance of ß(1,6)-branched oligosaccharides for prostate cancer progression.

Effect of quinolinyl acrylate derivatives on prostate cancer in vitro and in vivo.

Quinolines and acrylates are chemical compounds which were previously described as potential antitumor agents. In this study, a series of seven new quinolinyl acrylate derivatives were synthesized and evaluated against human prostate cancer cells PC-3 and LNCaP in vitro and in vivo. The most effective compound (E)-methyl 2-(7-chloroquinolin-4-ylthio)-3-(4 hydroxyphenyl) acrylate reduced the viability in both cell lines in a time- and dose-dependent manner. Inhibitory effects were also observed on the adhesion, migration, and invasion of the prostate cancer cells as well as on the neoangiogenesis, clonogenic and MMP-9 activity. The effect in vivo was studied in PC-3 xenografts in nude mice. The results were concordant with the in vitro effects and showed decreased tumor growth in treated animals compared to controls. The study suggests the multi-target efficacy of the quinolinyl derivate against human prostate cancer cells and supports its potential therapeutic usefulness.

New 4-maleamic acid and 4-maleamide peptidyl chalcones as potential multitarget drugs for human prostate cancer.

PURPOSE: The objective of this study was to investigate the effect of new 4-maleamic acid and 4-maleamide peptidyl chalcone derivatives against human prostate cancer in vitro and in vivo. METHODS: From a series of 21 chalcones, the effects of the three best inhibitors of PC-3 and LNCaP cell viability on growth, including cell cycle changes, adhesion, migration, and cell invasion, as well as their ability to inhibit angiogenesis, clonogenic activity, and matrix metalloproteinases MMP-2 and MMP-9, were tested. The effects in vivo were studied in PC-3 and LNCaP xenografts. RESULTS: Three of the examined chalcones reduced cell viability in both cell lines in a strong dose- and time-dependent manner. An inhibition of the cell cycle progress was observed. These changes were accompanied with the inhibition of cell adhesion, migration, and invasion as well as with reduced neovascularization in chick embryos, tumor colony formation, and MMP-9 activity. The in vivo results demonstrated the strong activity of these structures as inhibitors of tumor development in nude mice compared to non-treated animals. CONCLUSION: The results suggest the multitarget efficacy of 4-maleamic acid and 4-maleamide peptidyl chalcones against human prostate cancer cells and emphasize the potential therapeutic relevance of these compounds.

Divergent effects of taurolidine as potential anti-neoplastic agent: inhibition of bladder carcinoma cells in vitro and promotion of bladder tumor in vivo.

We investigated taurolidine (TRD) against various human bladder cell lines and the AY-27 rat bladder carcinoma cells. In vitro we tested the effect of TRD in ascending concentrations depending on different incubation times on cell proliferation by the XTT-test. Taurolidine had an inhibitory effect on all tested cell lines. Increasing concentrations and longer incubation times decreased the proliferation depending on the primary quantities of cells. For in vivo studies, an orthotopic rat bladder carcinoma was used. The animals were treated intravenously or intravesically and the tumors were harvested and weighted after the study. In contrast to other authors we could not find any anti-proliferative effect, we actually showed that instillation into the rat urinary bladder enhanced tumor growth.

Enhanced inhibitory effect of the matrix metalloproteinase inhibitor Ro 28-2653 in combination with estramustine and etoposide on the prostate carcinoma in the rat Dunning orthotopic tumor model.

PURPOSE: Therapeutic efficacy of the novel matrix metalloproteinase (MMP) inhibitor Ro 28-2653 has been shown in various models of different tumor entities. We hypothesized that the inhibitor effect of Ro 28-2653 on the tumor growth could be improved by combination with chemotherapeutic drugs and examined therefore the effect of Ro 28-2653 alone and in combination with etoposide or estramustine in the MatLyLu Dunning R-3327 rat tumor model characteristic for the androgen-independent prostate cancer (PCa). METHODS: In vitro effects were estimated measuring the proliferation of MatLyLu cells incubated with the three agents alone or in combination using the XTT test. The in vivo effects of the agents alone or in combination were examined by measuring the tumor weight 18 days after tumor cell injection. RESULTS: The proliferation rate was only inhibited by etoposide while that effect was increased in combination with Ro 28-2653 and estramustine. Ro 28-2653 reduced the tumor weight by 86%. That effect was significantly increased in combination with etoposide to 92%. CONCLUSIONS: The inhibitory effect of the MMP inhibitor Ro 28-2653 on the tumor growth in the Dunning PCa model is enhanced by the standard chemotherapeutic drug etoposide. A combined application of both agents could be considered as potential tool to improve the therapy of patients with advanced PCa after failure of hormonal treatment.

Neointima inhibition: comparison of effectiveness of non-stent-based local drug delivery and a drug-eluting stent in porcine coronary arteries.

PURPOSE: To compare the inhibition of neointimal proliferation by using non-stent-based local drug delivery and a drug-eluting stent in porcine coronary arteries. MATERIALS AND METHODS: Experiments were conducted with permission of the animal protection committee of the local government. Paclitaxel was either dissolved in a nonionic contrast medium or coated on balloons. Stents were crimped on the coated balloons. Effectiveness was tested in 22 pigs. Two coronary stents were placed in each pig, and slight overstretch was applied. The animals were treated as follows: group A (control group), uncoated balloons, bare stents, and "plain" contrast medium; group B, same treatment as group A, but with paclitaxel in the contrast medium; group C, paclitaxel-coated balloons, with premounted bare stents and plain contrast medium; and group D, sirolimus-eluting stents, noncoated balloons, and plain contrast medium. Stenosis was assessed 4 weeks later at angiography and histomorphometry. For exploratory purposes, continuous variables of quantitative coronary angiography and histomorphometry were compared by using analysis of variance. RESULTS: Results at follow-up angiography indicated a mean of 1.00 mm +/- 0.18 (standard deviation) lumen diameter loss in the control group and 0.14 mm +/- 0.18 loss in the group treated with the paclitaxel-coated balloon (group C; P < .001). Findings at histomorphometry confirmed the effectiveness of drug delivery, with the most impressive inhibition of neointimal proliferation from coated balloons-the neointimal area was 2.4 mm2 +/- 0.3 (P < .01 vs all other groups), compared with 5.2 mm2 +/- 0.3 in group A (control group), 4.3 mm2 +/- 0.3 in group B, and 3.8 mm2 +/- 0.3 in group D. CONCLUSION: In spite of the short intima contact time, paclitaxel coated on the balloon inhibits neointimal formation in the porcine model of coronary stent placement.

Focal liver lesions: SPIO-, gadolinium-, and ferucarbotran-enhanced dynamic T1-weighted and delayed T2-weighted MR imaging in rabbits.

PURPOSE: To compare a superparamagnetic iron oxide (SPIO), VSOP-C184, with a gadopentetate dimeglumine with regard to signal-enhancing effects on T1-weighted dynamic magnetic resonance (MR) images and with another SPIO contrast medium with regard to signal-reducing effects on delayed T2-weighted MR images. MATERIALS AND METHODS: All experiments were approved by the responsible Animal Care Committee. Twenty rabbits (five for each contrast agent and dose) implanted with VX-2 carcinoma were imaged at 1.5 T. VSOP-C184 at 0.015 and 0.025 mmol Fe/kg was compared with gadopentetate dimeglumine at 0.15 mmol Gd/kg and ferucarbotran at 0.015 mmol Fe/kg. The imaging protocol comprised a T1-weighted dynamic gradient-echo (GRE) MR before injection and at 6-second intervals for up to 42 seconds after injection and a T2-weighted turbo spin-echo MR before and 5 minutes after injection. Images were evaluated quantitatively, and contrast media were compared by using nonparametric analysis of variance. RESULTS: At dynamic T1-weighted GRE MR imaging with 0.015-mmol Fe/kg VSOP-C184, 0.025-mmol Fe/kg VSOP-C184, gadopentetate dimeglumine, and ferucarbotran, the median peak contrast-to-noise ratio (CNR) was 20.7 (25th percentile, 16.3; 75th percentile, 22.6), 24.2 (25th percentile, 19.3; 75th percentile, 28.5), 16.4 (25th percentile, 13.7; 75th percentile, 20.3), and 14.0 (25th percentile, 11.4; 75th percentile, 16.8), respectively. Both doses of VSOP-C184 yielded significantly higher CNR (P < .05) than the other two agents. At T2-weighted turbo spin-echo imaging with 0.015-mmol Fe/kg VSOP-C184, 0.025-mmol Fe/kg VSOP-C184, gadopentetate dimeglumine, and ferucarbotran, the median CNR was 15.0 (25th percentile, 13.4; 75th percentile, 21.3), 15.7 (25th percentile, 14.5; 75th percentile, 19.8), 11.3 (25th percentile, 8.2; 75th percentile, 12.2), and 15.7 (25th percentile, 12.5; 75th percentile, 22.4), respectively. There was no significant difference between VSOP-C184 and ferucarbotran; both had a significantly higher CNR than did gadopentetate dimeglumine. CONCLUSION: VSOP-C184 produces higher liver-to-tumor contrast at dynamic T1-weighted imaging than does gadopentetate dimeglumine; at delayed T2-weighted imaging, the contrast is comparable to that achieved with ferucarbotran.

A multicenter clinical trial on the use of (-5, -7) pro prostate specific antigen.

PURPOSE: The determination of pro prostate specific antigen (proPSA) forms has been suggested to be promising in prostate cancer diagnosis. In this multicenter trial we evaluated the diagnostic usefulness of (-5, -7) proPSA. MATERIALS AND METHODS: A total of 2,055 white men, including 1,046 with and 1,009 without prostate cancer, with total PSA (tPSA) between 0.28 and 81 ng/ml were retrospectively analyzed. Of these men 2,026 and 1,727 had tPSA less than 20 and less than 10 ng/ml, respectively. All subjects were untreated for prostatic disease and underwent multisector needle biopsy of the prostate. An Elecsys 2010 analyzer was used to determine tPSA, free PSA (fPSA) and (-5, -7) proPSA in the 2,055 serum samples. ROC analyses were performed to discriminate men with biopsy positive and negative results in the entire and in select tPSA ranges. RESULTS: In the select tPSA range 2 to 4 ng/ml the area under the ROC curve for proPSA (0.53) and proPSA/fPSA (0.59) was not significantly larger than that for tPSA (0.60) or the fPSA/tPSA (f/tPSA) ratio (0.64). In the tPSA range 4 to 10 ng/ml the area under the curve for the ratio proPSA/fPSA (0.67) was larger than for tPSA (0.53) but not larger than for f/tPSA (0.69). The f/tPSA ratio demonstrated the best discriminatory power in this tPSA range of 4 to 10 ng/ml. CONCLUSIONS: In this multicenter study no improvement in diagnostic accuracy was shown when comparing (-5, -7) proPSA and the corresponding ratios with tPSA or f/tPSA. Further studies using other proPSA forms or tumor associated proteins should be done.

Matrix metalloproteinase inhibitor Ro 28-2653 in combination with estramustine: tumor-reducing effects on hormone-sensitive prostate cancer in rats.

Therapeutic efficacy of the novel matrix metalloproteinase (MMP) inhibitor, Ro 28-2653 (5-biphenyl-4-yl-5-[4-(-nitro-phenyl)-piperazin-1-yl]-pyrimidine-2,4,6-trione), has been shown in various models of different tumor entities. The tumor growth-reducing effect has been demonstrated in the orthotopic rat prostate Dunning model (subline MatLyLu). Based on these results we investigated Ro 28-2653 in combination with estramustine on the G subline of the Dunning tumor. This subline is characterized by a low metastatic ability and androgen sensitivity. Efficacy was determined by recording tumor growth in vivo by magnetic resonance imaging (MRI). Tumor cells were injected into the prostates of 81 Copenhagen rats. MRI was performed at day 100 and at day 126 after tumor cell injection. The duration of therapy was 17 days with daily oral application of Ro 28-2653 (100 mg/kg) and four i.p. injections of estramustine (7.5 mg/kg). Histological evaluations were conducted to provide further information about the effects on tumor morphology. Orthotopic tumor induction was successful in 100% of the animals. Tumor volume calculations with MRI showed a significant difference between the control groups, the animals treated with Ro 28-2653, and the animals treated with the combination of Ro 28-2653 and estramustine. The new MMP inhibitor Ro 28-2653 reduces tumor growth and provides a compatible therapeutic alternative for patients with prostate cancer.

Comparison of the iron oxide-based blood-pool contrast medium VSOP-C184 with gadopentetate dimeglumine for first-pass magnetic resonance angiography of the aorta and renal arteries in pigs.

RATIONALE AND OBJECTIVES: VSOP-C184 at a dose of 0.045 mmol Fe/kg has been shown to be an efficient blood pool contrast medium for equilibrium magnetic resonance angiography (MRA) that can be administered as a bolus. The present study was performed to determine whether VSOP-C184 is also suitable for first-pass MRA. MATERIALS AND METHODS: Fifteen MRA examinations at 1.5 T were performed in minipigs using a fast 3D fast low-angle shot (FLASH) sequence (repetition time = 4.5 ms, echo time = 1.7 ms, excitation angle = 25 degrees, matrix 256, body phased-array coil). The citrate-stabilized iron oxide preparation VSOP-C184 was investigated (total particle diameter: 7.0 +/- 0.15 nm; core size: 4 nm) and compared with gadopentetate dimeglumine (Gd-DTPA). The following doses were tested: VSOP-C184: 0.015, 0.025, and 0.035 mmol Fe/kg; Gd-DTPA: 0.1 and 0.2 mmol Gd/kg; n = 3 examinations/dose. Data were analyzed quantitatively (signal enhancement (ENH) and vessel edge definition (VED)) and qualitatively. RESULTS: First-pass MRA using the 3 doses of VSOP-C184 yielded the following ENH: aorta: 9.4 +/- 2.6; 12.31 +/- 1.2; 16.53 +/- 1.7; renal arteries: 7.6 +/- 2.2; 9.9 +/- 1.0; 13.2 +/- 0.5. The values for the 2 doses of Gd-DTPA were aorta: 12.9 +/- 1.0; 16.8 +/- 2.2; renal arteries: 11.2 +/- 1.23; 11.3 +/- 1.7. VED for the 3 doses of VSOP-C184 was aorta: 106.3 +/- 31.0; 135.3 +/- 58.8; 141.3 +/- 71.0; renal arteries: 102.2 +/- 24.3; 146.8 +/- 63.0; 126.9 +/- 37.6 and for the 2 doses of Gd-DTPA, aorta: 157.2 +/- 47.8; 164.2 +/- 36.8; renal arteries: 165.9 +/- 30.4; 170.3 +/- 38.2 respectively. The differences between VSOP-C184 and Gd-DTPA are clinically not relevant and statistically not significant (p > or = .05). Qualitative evaluation of image quality, contrast, and delineation of vessels showed the results obtained with VSOP-C184 at doses of 0.025 and 0.035 mmol Fe/kg to be similar to those of Gd-DTPA at 0.1 and 0.2 mmol Gd/kg. CONCLUSION: VSOP-C184 is suitable for first-pass MRA at doses of 0.025 and 0.035 mmol Fe/kg and thus, in addition to its blood pool characteristics, allows for selective visualization of the arteries without interfering venous signal.

Paclitaxel balloon coating, a novel method for prevention and therapy of restenosis.

BACKGROUND: Drug-eluting stents have shown promising antirestenotic effects in clinical trials. Non-stent-based local delivery of antiproliferative drugs may offer additional flexibility and also reach vessel areas beyond the immediate stent coverage. The aim of the present study was to evaluate a novel method of local drug delivery based on angioplasty balloons. METHODS AND RESULTS: Stainless steel stents (n=40; diameter, 3.0 to 3.5 mm; length, 18 mm) were implanted in the left anterior descending and circumflex coronary arteries of domestic pigs. Both conventional uncoated and 3 different types of paclitaxel-coated, percutaneous transluminal coronary angioplasty balloons (contact with vessel wall for 1 minute) were used. No difference in short-term tolerance between coated and uncoated balloons and no signs of thrombotic events were observed. Quantitative angiography and histomorphometry of the stented arteries asserted the statistical equality of the baseline parameters between the control and the 3 treatment groups. Paclitaxel balloon coating led to a marked, dose-dependent reduction of parameters characterizing in-stent restenosis (reduction of neointimal area up to 63%). Despite the marked reduction in neointimal proliferation, endothelialization of stent struts was present in all samples. There was no evidence of a significant inflammatory response in the neighborhood of the stent struts. CONCLUSIONS: Paclitaxel balloon coating is safe, and it effectively inhibits restenosis after coronary angioplasty with stent implantation in the porcine model. The degree of reduction in neointimal formation was comparable to that achieved with drug-eluting stents.

Iosimenol, a low-viscosity nonionic dimer: preclinical physicochemistry, pharmacology, and pharmacokinetics.

RATIONALE AND OBJECTIVES: Newer radiologic techniques require fast bolus injections and thus low-viscosity, high-concentration, well-tolerated contrast media (CM), especially in vulnerable patients. To this end, we designed and developed iosimenol, a novel isotonic nonionic dimer, and have conducted tests to enable its clinical evaluation. METHODS: Standard physicochemical methods were used. Effects on erythrocyte morphology and coagulation were investigated in human and rat blood. Neural tolerance was assessed by behavioral tests in rats after intracisternal injection. Immunosensitizing potential was evaluated by the skin sensitization test in guinea pigs and by the popliteal lymph node assay in rats. Pharmacokinetics and biotransformation were investigated in rats and dogs. RESULTS: Iosimenol is extremely hydrophilic, it is less viscous than any other isotonic CM, has little effect on erythrocytes and blood coagulation, and has good neural tolerance. No immunosensitizing effect was found in validated animal models. Pharmacokinetics are identical with other angio- and urographic CM. CONCLUSIONS: Iosimenol is the only CM which, although isotonic, affords, unlike current nonionic dimers, at the same iodine concentration the low viscosity of monomeric, nonionic agents, which are all hypertonic. Iosimenol's pharmacologic characteristics closely resemble those of iotrolan and iodixanol.

Inhibition of restenosis in stented porcine coronary arteries: uptake of Paclitaxel from angiographic contrast media.

RATIONALE AND OBJECTIVES: Paclitaxel added to angiographic contrast medium (CM) has been shown to inhibit restenosis in the porcine coronary overstretch model. This study determined early local tissue concentrations after the administration of different paclitaxel doses and preparations. MATERIALS AND METHODS: Fifteen pigs received 2 stents each in the left coronary artery. During and/or after the intervention, paclitaxel-containing CM or diluted Taxol was injected. Fifteen minutes after the last intracoronary injection, paclitaxel concentrations in the arterial wall and myocardium were measured by high-performance liquid chromatography. RESULTS: Mean paclitaxel concentrations in the left coronary arteries reached 3-10 microM. Higher volumes and higher paclitaxel concentrations resulted in higher tissue concentrations. Paclitaxel in CM was better tolerated and led to higher local concentrations than diluted Taxol. Low paclitaxel concentrations in the uninjected right coronary artery and in plasma indicate selectivity. CONCLUSION: When admixed to CM, paclitaxel results in local tissue concentrations proportional to the amount of the drug injected.

Coronary MR angiography: experimental results with a monomer-stabilized blood pool contrast medium.

PURPOSE: To evaluate the signal-enhancing characteristics of monomer-coated very small superparamagnetic iron oxide (SPIO) particles used as a blood pool contrast medium for magnetic resonance (MR) angiography in the coronary arteries. MATERIALS AND METHODS: The particles used in this study were coated with citrate as the monomer (VSOP-C91). The particles have a total diameter of 7 nm and show the following relaxivities at 0.47 T: T1, 19 L/mmol. sec(-1); T2, 29 L/mmol. sec(-1). Fifteen cardiac MR examinations were performed at 1.5 T in five pigs. Images were acquired from immediately to 35 minutes (equilibrium phase) after intravenous injection of gadopentetate dimeglumine, gadobenate dimeglumine, and the very small SPIO particles (n = 5 for each substance). RESULTS: Immediately after administration of gadopentetate dimeglumine, gadobenate dimeglumine, and the very small SPIO particles, respectively, increases in the signal-to-noise ratio in blood were 94%, 103%, and 102% and in myocardium were 83%, 83%, and 29% (P <.05, very small SPIO particles versus the low-molecular-weight gadolinium-based compounds). Differences in the blood-to-myocardium contrast-to-noise ratio and visualization of the coronary arteries and their branches were also significant. CONCLUSION: VSOP-C91 significantly improves visualization of the coronary arteries at MR angiography from immediately to 35 minutes after injection.