Evaluation of a novel macromolecular cascade-polymer contrast medium for dynamic contrast-enhanced MRI monitoring of antiangiogenic bevacizumab therapy in a human melanoma model.

RATIONALE AND OBJECTIVES: To assess the applicability of a novel macromolecular polyethylene glycol (PEG)-core gadolinium contrast agent for monitoring early antiangiogenic effects of bevacizumab using dynamic contrast-enhanced (DCE) magnetic resonance imaging (MRI). MATERIALS AND METHODS: Athymic rats (n = 26) implanted with subcutaneous human melanoma xenografts underwent DCE-MRI at 2.0 T using two different macromolecular contrast agents. The PEG core cascade polymer PEG12,000-Gen4-(Gd-DOTA)16, designed for clinical development, was compared to the prototype, animal-only, macromolecular contrast medium (MMCM) albumin-(Gd-DTPA)35. The treatment (n = 13) and control (n = 13) group was imaged at baseline and 24 hours after a single dose of bevacizumab (1 mg) or saline to quantitatively assess the endothelial-surface permeability constant (K(PS), µL⋅min⋅100 cm(3)) and the fractional plasma volume (fPV,%), using a two-compartment kinetic model. RESULTS: Mean K(PS) values, assessed with PEG12,000-Gen4-(Gd-DOTA)16, declined significantly (P < .05) from 29.5 ± 10 µL⋅min⋅100 cm(3) to 10.4 ± 7.8 µL⋅min⋅100 cm(3) by 24 hours after a single dose of bevacizumab. In parallel, K(PS) values quantified using the prototype MMCM albumin-(Gd-DTPA)35 showed an analogous, significant decline (P < .05) in the therapy group. No significant effects were detected on tumor vascularity or on microcirculatory parameters in the control group between the baseline and the follow-up scan at 24 hours. CONCLUSION: DCE-MRI enhanced with the novel MMCM PEG12,000-Gen4-(Gd-DOTA)16 was able to monitor the effects of bevacizumab on melanoma xenografts within 24 hours of a single application, validated by the prototype, animal-only albumin-(Gd-DTPA)35. PEG12,000-Gen4-(Gd-DOTA)16 may be a promising candidate for further clinical development as a macromolecular blood pool contrast MRI agent.

Permeability to macromolecular contrast media quantified by dynamic MRI correlates with tumor tissue assays of vascular endothelial growth factor (VEGF).

PURPOSE: To correlate dynamic MRI assays of macromolecular endothelial permeability with microscopic area-density measurements of vascular endothelial growth factor (VEGF) in tumors. METHODS AND MATERIAL: This study compared tumor xenografts from two different human cancer cell lines, MDA-MB-231 tumors (n=5), and MDA-MB-435 (n=8), reported to express respectively higher and lower levels of VEGF. Dynamic MRI was enhanced by a prototype macromolecular contrast medium (MMCM), albumin-(Gd-DTPA)35. Quantitative estimates of tumor microvascular permeability (K(PS); µl/min × 100 cm(3)), obtained using a two-compartment kinetic model, were correlated with immunohistochemical measurements of VEGF in each tumor. RESULTS: Mean K(PS) was 2.4 times greater in MDA-MB-231 tumors (K(PS)=58 ± 30.9 µl/min × 100 cm(3)) than in MDA-MB-435 tumors (K(PS)=24 ± 8.4 µl/min × 100 cm(3)) (p<0.05). Correspondingly, the area-density of VEGF in MDA-MB-231 tumors was 2.6 times greater (27.3 ± 2.2%, p<0.05) than in MDA-MB-435 cancers (10.5 ± 0.5%, p<0.05). Considering all tumors without regard to cell type, a significant positive correlation (r=0.67, p<0.05) was observed between MRI-estimated endothelial permeability and VEGF immunoreactivity. CONCLUSION: Correlation of MRI assays of endothelial permeability to a MMCM and VEGF immunoreactivity of tumors support the hypothesis that VEGF is a major contributor to increased macromolecular permeability in cancers. When applied clinically, the MMCM-enhanced MRI approach could help to optimize the appropriate application of VEGF-inhibiting therapy on an individual patient basis.

Correlative dynamic contrast MRI and microscopic assessments of tumor vascularity in RIP-Tag2 transgenic mice.

The purpose of this study was to define the feasibility of dynamic contrast-enhanced magnetic resonance imaging (MRI) to estimate the vascular density and leakiness of spontaneous islet cell tumors in RIP-Tag2 transgenic mice. Dynamic T(1)-weighted spoiled gradient echo (SPGR) imaging at 2.0 T was performed in 17 RIP-Tag2 mice using a prototype blood pool macromolecular contrast medium (MMCM), albumin-(Gd-DTPA)(35). Kinetic analysis of the dynamic enhancement responses based on a two-compartment model was used to estimate fractional plasma volume (fPV) and the coefficient of endothelial permeability (K(PS)) for each tumor. The MRI estimate of fPV was correlated on a tumor-by-tumor basis with corresponding microscopic measurements of vascular density. The fPV assays by MMCM-enhanced imaging ranged from 2.4%-14.1% of tissue volume. Individual tumor fPV values correlated significantly (r = 0.79, P < 0.001) with the corresponding microscopic estimates of vascularity consisting of the combined area densities of lectin-perfused microvessels plus erythrocyte-stained blood lakes. A biotinylated derivative of the albumin-based MMCM confirmed extravasation of the contrast agent from some tumor blood vessels and accumulation in 25% of blood lakes. The K(PS) values ranged from 0 (no detectable leak) to 0.356 mL/min/100 cm(3). Dynamic MMCM-enhanced MRI is feasible in RIP-Tag2 pancreatic tumors, yielding estimates of vascular permeability and microscopically validated measurements of vascular richness.

In vivo monitoring of angiogenesis inhibitory treatment effects by dynamic contrast-enhanced computed tomography in a xenograft tumor model.

RATIONALE AND OBJECTIVES: To evaluate the potential of dynamic CT enhanced by iohexol or a novel macromolecular contrast agent, PEG12000-Gen4-triiodo, to monitor microvascular changes in tumors treated with the angiogenesis inhibitor bevacizumab. MATERIALS AND METHODS: Ten female nude rats with MDA-MB 435 xenograft tumors were treated with 1 mg intraperitoneal bevacizumab when tumors reached 1 cm diameter and, for 4 rats, treated again 7 days later. Just before and 24 hours after the first injection of anti-VEGF antibody, the tumors were imaged by dynamic CT scans enhanced with PEG12000-Gen4-triiodo (n = 3 rats) or iohexol (n = 3 rats). The other 4 rats underwent dynamic CT scans enhanced with PEG12000-Gen4-triiodo just before and 24 hours after the second injection of anti-VEGF antibody. Microvascular leakiness (K(PS)) was calculated for the tumors using a 2-compartment tissue model. RESULTS: PEG12000-Gen4-triiodo-enhanced CT scans showed progressive reductions in K(PS) from day 1 to 2 to 9 (from 2.55 to 1.27 to 0.69 microL min(-1) cm(-3), respectively, P < 0.005 for each comparison of day 1-2 and day 2-9). No significant difference was seen in the K(PS) estimates derived from iohexol-enhanced CT scans obtained before or after treatment (276 vs. 223.8 microL min(-1) cm(-3), respectively, P = 0.54). The microvascular leak (K(PS)) was significantly larger for iohexol than for PEG12000-Gen4-triiodo-enhanced CT, P < 0.05. CONCLUSION: Dynamic macromolecular contrast-enhanced CT can be used to monitor serial decreases in tumor microvessel leakiness induced by repeated doses of an angiogenesis inhibitor drug.

Magnetic resonance imaging for monitoring the effects of thalidomide on experimental human breast cancers.

Thalidomide, which inhibits angiogenesis in certain tumor types, reduced extravasation of a macromolecular contrast medium (MMCM) in a human breast cancer model as assayed by MMCM-enhanced dynamic magnetic resonance imaging (MRI) and fluorescence microscopy in the same tumors. After a 1-week, three-dose course of thalidomide, the mean MRI-assayed endothelial transfer coefficient, K(PS), decreased significantly (p < 0.05) from 19.4 +/- 9.1 to 6.3 +/- 9.1 microl/min.100 cm(3). Correspondingly, microscopic measurements of extravasated MMCM, expressed as fractional area of streptavidin staining, were significantly (p < 0.05) lower in thalidomide-treated tumors (18.6 +/- 11.9%) than in control saline-treated tumors (50.2 +/- 2.3%). On a tumor-by-tumor basis, post-treatment K(PS) values correlated significantly (r(2) = 0.55, p < 0.05) with microscopic measures of MMCM extravasation. However, no significant differences were observed between saline- and thalidomide-treated tumors with respect to rate of growth, vascular richness, or amount of VEGF-containing cells. Because of its sensitivity to the detection of changes in vascular leakage in tumors, this MMCM-enhanced MRI assay could prove useful for monitoring the effects of thalidomide on an individual patient basis. The significant correlation between MRI and fluorescence microscopic measures of MMCM extravasation supports the utility of the non-invasive MRI approach for assessing the action of thalidomide on tumor blood vessels.

Magnetic resonance imaging assays for dimethyl sulfoxide effect on cancer vasculature.

OBJECTIVES: To evaluate the potential of quantitative assays of vascular characteristics based on dynamic contrast-enhanced magnetic resonance imaging (MRI) using a macromolecular contrast medium (MMCM) to search for and measure effects of dimethyl sulfoxide (DMSO) on cancer vasculature with microscopic correlations. MATERIAL AND METHODS: Saline-treated control (n = 8) and DMSO-treated (n = 7) human breast cancer xenografts (MDA-MB-435) in rats were imaged dynamically by MMCM-enhanced MRI using albumin-(Gd-DTPA)27-(biotin)11 (molecular weight approximately 90 kDa), before and after a 1-week, 3-dose treatment course. After the posttreatment MRI examinations, tumors were perfused with lectin and fixative and subsequently stained with RECA-1 and streptavidin for quantitative fluorescent microscopy. Quantitative MRI estimates of cancer microvessel permeability (KPS; microL/min.100 cm3) and fractional plasma volume (fPV; %) were based on a 2-compartment kinetic model. Fluorescent microscopy yielded estimates of MMCM extravasation and vascular density that were compared to the MRI results. RESULTS: DMSO decreased cancer vascular endothelial permeability significantly (P < 0.05) from tumor KPSday0 = 19.3 +/- 8.8 microL/min.100 cm3 to KPSday7 = 0 microL/min.100 cm3). K values in the saline-treated tumors did not change significantly. The amount of extravasated albumin-Gd-(DTPA)27-(biotin)11, as assayed by a fluorescently labeled streptavidin stain that strongly binds to the biotin tag on the MMCM, was significantly (P < 0.05) lower in the DMSO-treated cancers than in the control cancers (57.7% +/- 5.5% vs. 34.2% +/- 4.9%). Tumor vascular richness as reflected by the MRI-assayed fPV and by the RECA-1 and lectin-stained microscopy did not change significantly with DMSO or saline treatment. CONCLUSION: Reductions in cancer microvascular leakiness induced by a 7-day course of DMSO could be detected and measured by dynamic MMCM-enhanced MRI and were confirmed by microscopic measurements of the leaked macromolecular agents in the same cancers. Results support the robustness of an MMCM-enhanced MRI approach to the characterization of cancers and providing first evidence for an in vivo effect of DMSO on cancer blood vessels.

Vascular permeability during antiangiogenesis treatment: MR imaging assay results as biomarker for subsequent tumor growth in rats.

PURPOSE: To prospectively evaluate in rats the acute change in tumor vascular leakiness (K(PS)) assayed at magnetic resonance (MR) imaging after a single dose of the angiogenesis inhibitor bevacizumab as a predictive biomarker of tumor growth response after a prolonged treatment course. MATERIALS AND METHODS: Institutional animal care and use committee approval was obtained. Seventeen female rats with implanted human breast cancers underwent dynamic albumin-(Gd-DTPA)(30)-enhanced MR imaging followed by an initial dose of bevacizumab or saline (as a control). Treatment was continued every 3rd day, for a total of four doses at five possible dose levels: 0 mg bevacizumab (n = 4 [control rats]), 0.1 mg bevacizumab (n = 3), 0.25 mg bevacizumab (n = 2), 0.5 mg bevacizumab (n = 5), and 1.0 mg bevacizumab (n = 3). A second MR imaging examination was performed 24 hours after the initial dose to enable calculation of the acute change in MR imaging-assayed leakiness, or Delta K(PS). This acute change in K(PS) at MR imaging was correlated with tumor growth response for each cancer at the completion of the 11-day treatment course. For statistical analyses, an unpaired two-tailed t test, analysis of variance, and linear regression analyses were used. RESULTS: The MR imaging-assayed change in tumor microvascular leakiness, tested as a potential biomarker, correlated strongly with tumor growth rate (R(2) = 0.74, P < .001). K(PS) and tumor growth decreased significantly in all bevacizumab-treated cancers compared with these values in control group cancers (P < .05). CONCLUSION: The MR imaging-assayed acute change in vascular leakiness after a single dose of bevacizumab was an early, measurable predictive biomarker of tumor angiogenesis treatment response.

New macromolecular polymeric MRI contrast agents for application in the differentiation of cancer from benign soft tissues.

PURPOSE: To compare three new macromolecular polyethylene glycol (PEG) -core dendrimeric gadolinium(Gd)-based MRI contrast agents for their applicability in quantitative assays of endothelial leakiness and tissue vascular density for the differentiation of cancer from normal soft tissues. MATERIALS AND METHODS: Thirty-two athymic rats with human breast cancer xenografts (MDA-MB-435) were imaged by dynamic MRI following enhancement with one of three new (Gd-DOTA)-conjugated PEG-core dendrimer contrast agents (effective molecular weights 161 to 323 kDa). Results were compared with a prototype macromolecular contrast agent, albumin (Gd-DTPA). Assays of permeabilities (K(PS); microL/min . 100 cm(3)) and tumor fractional plasma volumes (%) based on a two-compartment kinetic model were performed for skeletal muscle and tumors. RESULTS: The largest PEG-core contrast agent, PEG(20,000)-Gen4-(Gd-DOTA), leaked in breast tumors (K(PS) = 50 +/- 23 microL/min . 100 cm(3)), while exhibiting no measurable transendothelial leak (K(PS) = 0 microL/min . 100 cm(3)) in normal soft tissue microvessels allowing successful differentiation (P < 0.05) of cancers from normal muscle. PEG(12,000)-Gen4-(Gd-DOTA) leaked in tumors and in normal muscle (K(PS) = 51 +/- 26 and K(PS) = 21 +/- 18 microL/min . 100 cm(3), respectively). The smallest agent, PEG(12,000)-Gen3-(Gd-DOTA) also showed a measurable leak in both normal and malignant microvessels. CONCLUSION: MRI assays of vascular endothelial leakiness using new PEG-core, (Gd-DOTA)-conjugated macromolecular contrast agents proved applicable for the differentiation of human breast cancer from normal soft tissue. The apparent threshold in effective molecular weight for a clear differentiation of cancer from normal muscle with no measurable leak in the muscle is between 194 and 323 kDa.

Cascade polymeric MRI contrast media derived from poly(ethylene glycol) cores: initial syntheses and characterizations.

Diagnostic contrast media for magnetic resonance imaging (MRI) are often applied to enhance the signal of blood allowing for quantitative definition of vascular functional characteristics including tissue blood volume, flow, and leakiness. Well-tolerated and safe macromolecular formulations are currently being sought that remain in the blood for a relatively long period and that leak selectively from diseased vessels, particularly cancer vessels. We synthesized a new class of macromolecular, water-soluble MRI contrast media by introducing two diverging polylysine cascade amplifiers at each end of a poly(ethylene glycol) (PEG) backbone, followed by substitution of terminal lysine amino groups with Gd-DTPA chelates. Four candidate PEG cascade conjugates are reported here, PEG3400-Gen4-(Gd-DTPA)8, PEG6000-Gen4-(Gd-DTPA)8, PEG12000-Gen4-(Gd-DTPA)8, and PEG3400-Gen5-(Gd-DTPA)13 with descriptions of their basic physical, biological, and kinetic properties, including real and effective molecular sizes, proton T1 relaxivities in water and plasma, partition coefficients, osmolalities, chelate stability, stability in plasma, stability to autoclaving, certain in vivo pharmacokinetics (blood half-life, blood clearance, volume of distribution), and whole body elimination profiles in normal rodents. These candidate PEG-core cascade MRI contrast media showed a range of effective molecular sizes similar to proteins weighing 74-132 kDa, although their actual molecular weights were much smaller, 12-20 kDa. All compounds exhibited a narrow range of size dispersity and relatively high T1 relaxivities (approximately 3 times the value for unconjugated Gd-DTPA at 2 T and 37 degrees C). Representative compounds also showed a high degree of hydrophilicity, stability in solution buffer and plasma, and lack of binding to proteins. The two candidate compounds with the largest effective molecular sizes, PEG12000-Gen4-(Gd-DTPA)8 and PEG3400-Gen5-(Gd-DTPA)13, had longer blood half-lives, 36 and 73 min, respectively (monoexponential kinetics for both), and showed strong, prolonged MRI enhancement of vessels. Results also indicate that in vivo pharmacokinetics and bodily elimination profiles can be adjusted by the selection of molecular size for the PEG core and the selection of the amplification degree of the cascade polylysine clusters. The initially evaluated compounds from this new class of contrast media show acceptable, desirable characteristics in many, but not all, respects. Further efforts are directed toward candidate macromolecules having higher thermodynamic stability, higher degree of substitution by gadolinium chelates, and more rapid bodily elimination.

MR monitoring of cyclooxygenase-2 inhibition of angiogenesis in a human breast cancer model in rats.

PURPOSE: To prospectively evaluate the ability of macromolecular contrast medium (MMCM)-enhanced dynamic magnetic resonance (MR) imaging to depict vascular changes in response to cyclooxygenase-2 (COX-2) inhibition of angiogenesis in a human breast cancer model. MATERIALS AND METHODS: The institutional committee for animal research approved this study. A human breast cancer cell line, MDA-MB-231, was implanted in 30 female homozygotous athymic rats that were alternately assigned to either a drug treatment group that received celecoxib on a daily basis for 7 days or a control group that received saline. Each animal underwent MR imaging after intravenous administration of a high-molecular-weight contrast agent at baseline and again 24 hours and 7 days after administration. Eleven rats in each group successfully underwent all three studies and had data sets of sufficient technical quality. A bidirectional two-compartment tissue model was used to estimate transendothelial permeability (K(PS)) and fractional plasma volume (fPV) for each tumor. Microvessel density was also measured to enable histologic assessment of angiogenesis. Repeated-measures analysis of variance and unpaired two-tailed t tests were used to evaluate differences in mean values between MR examinations performed in the same rats and between baseline values in treated and control rats, respectively. RESULTS: MR imaging-assayed microvascular K(PS) decreased significantly after 7 days of treatment with celecoxib (P < .05), but it was not significantly changed after 7 days in the control group. Likewise, microvascular density, a histologic surrogate of angiogenesis, was significantly (P < .05) lower in the treatment group than in the control group. The fPV did not significantly change in either group. CONCLUSION: Dynamic MR imaging revealed microvascular permeability to a high-molecular-weight contrast agent was significantly reduced by treatment with celecoxib.

MRI tumor characterization using Gd-GlyMe-DOTA-perfluorooctyl-mannose-conjugate (Gadofluorine M), a protein-avid contrast agent.

The rationale and objectives were to define the MRI tumor-characterizing potential of a new protein-avid contrast agent, Gd-GlyMe-DOTA-perfluorooctyl-mannose-conjugate (Gadofluorine M; Schering AG, Berlin, Germany) in a chemically induced tumor model of varying malignancy. Because of the tendency for this agent to form large micelles in water and to bind strongly to hydrophobic sites on proteins, it was hypothesized that patterns of dynamic tumor enhancement could be used to differentiate benign from malignant lesions, to grade the severity of malignancies and to define areas of tumor necrosis. Gadofluorine M, 0.05 mmol Gd kg(-1), was administered intravenously to 28 anesthetized rats that had developed over 10 months mammary tumors of varying degrees of malignancy as a consequence of intraperitoneal administration of N-ethyl-N-nitrosourea (ENU), 45-250 mg kg(-1). These tumors ranged histologically from benign fibroadenomas to highly undifferentiated adenocarcinomas. Dynamic enhancement data were analyzed kinetically using a two-compartment tumor model to generate estimates of fractional plasma volume (fPV), apparent fractional extracellular volume (fEV*) and an endothelial transfer coefficient (K(PS)) for this contrast agent. Tumors were examined microscopically for tumor type, degree of malignancy (Scarff-Bloom-Richardson score) and location of necrosis. Eighteen tumor-bearing rats were successfully imaged. MRI data showed an immediate strong and gradually increasing tumor enhancement. K(PS) and fEV*, but not fPV obtained from tumors correlated significantly (p < 0.05) with the SBR tumor grade, r = 0.65 and 0.56, respectively. Estimates for K(PS) and fEV* but not fPV were significantly lower in a group consisting of benign and low-grade malignant tumors compared with the group of less-differentiated high-grade tumors (1.61 +/- 0.64 vs 3.37 +/- 1.49, p < 0.01; 0.45 +/- 0.17 vs 0.78 +/- 0.24, p < 0.01; and 0.076 +/- 0.048 vs 0.121 +/- 0.088, p = 0.24, respectively). It is concluded that the protein-avid MRI contrast agent Gadofluorine M enhances tumors of varying malignancy depending on the tumor grade, higher contrast agent accumulation for more malignant lesions. The results show potential utility for differentiating benign and low-grade malignant lesions from high-grade cancers.

Magnetic resonance imaging enhancement of normal tissues and tumors using macromolecular Gd-based cascade polymer contrast agents: preclinical evaluations.

OBJECTIVES: We sought to compare magnetic resonance imaging (MRI) enhancement using 4 novel macromolecular polyethyleneglycol (PEG)-based cascade-polymer gadolinium contrast agents (macromolecular contrast media) in normal soft tissues and a breast cancer model. MATERIALS AND METHODS: Four candidate PEG cascade polymers with effective molecular weights of 74, 82, 106, and 132 kDa, respectively, and T1-relaxivities of 8.1, 9.1, 9.7, and 10.0, respectively (at 2 Tesla and 37 degrees C in HEPES buffer), initially were used to characterize liver and kidney MRI-enhancement patterns in normal Sprague-Dawley rats (n = 4-5 per contrast agent). Kinetic analysis of dynamic MRI enhancement was used in 8 nude rats bearing MDA-MB 435 breast cancers to estimate fractional plasma volume and apparent endothelial leakiness (K) in tumors and muscle. RESULTS: Soft-tissue enhancement patterns followed closely the blood enhancement over the course of 30-50 minutes with estimated blood half-lives between 23 and 73 minutes, which varied with effective molecular weights. The 2 smaller compounds yielded measurable leaks in normal muscle [K = 204 and 56 microL/(min.100 cm), respectively], whereas the 2 larger molecules did not leak in muscle [K = 0 microL/(min.100 cm)]; however, MRI-assayed leakiness of tumor vessels with respect to those 2 larger macromolecular contrast media was 68 +/- 27 and 16 +/- 8 microL/(min.100 cm), respectively. CONCLUSIONS: Two relatively large (effective molecular weight >82 kDa) PEG-based cascade polymer contrast agents were well-suited for MRI quantification of tissue plasma volume and for differentiating leaky cancer microvessels from nonleaky normal vessels.

Dendritic iodinated contrast agents with PEG-cores for CT imaging: synthesis and preliminary characterization.

The purpose of this study was to design, synthesize, and initially characterize a representative set of novel constructs for large-molecular radiographic/computed tomography (CT) contrast agents, intended for a primarily intravascular distribution. A new assembly of well-known and biocompatible components consists of paired, symmetrical dendritic polylysines initiated from both ends of a poly(ethylene glycol) (PEG) core, yielding an array of multiple free amino groups to which were conjugated highly soluble and stable triiodophthalamide ("triiodo") moieties. An array of six dendritic contrast agents was synthesized originally, using three different PEG cores (3, 6, 12 kDa) with t-Boc lysine-generated dendrimer "amplifiers" (from three to five generations) containing 16 to 64 amino groups for conjugation with reactive triiodo moieties. A clinically used, nonionic, small molecular CT contrast agent, iobitridol, was derivatized via a hydroxyl protection/deprotection strategy, introducing a new carboxyl group available for conjugation to the lysine amino groups of dendrimers. Final products were purified by size exclusion chromatography and characterized by NMR, UV, HPLC, and elemental analysis. Preliminary evaluations were conducted for physicochemical characterization and in vivo CT contrast enhancement in a rat model. All six iodinated PEG-core dendrimer conjugates were synthesized in good yields, with a high degree of size monodispersity, large apparent molecular weight, favored physicochemical properties. A representative compound, PEG12000-carbamate-Gen4-IOB conjugate, 27% (w%) rich in iodine, demonstrated a desirable strong and persistent intravascular enhancement with a monoexponential blood half-life of approximately 35 min assayed by dynamic CT imaging and also showed high water solubility (>550 mg/mL at 25 degrees C), large apparent molecular size (comparable to a 143-kDa protein), high hydrophilicity (butanol-water partition coefficient 0.015), and stability to autoclaving conditions. This study showed the synthetic feasibility, desired basic characteristics, and potential utility for CT contrast enhancement achieved with a new type of iodinated, large-molecular PEG-core dendritic construct. Further development of this class of macromolecular contrast agents will be required to define the optimal formulation, pharmacology, safety profile, and the full range of diagnostic applications including tumor microvascular quantitative characterization by CT imaging.

Mixture model approach to tumor classification based on pharmacokinetic measures of tumor permeability.

PURPOSE: To categorize the disease severity of mammary tumors in an animal model through the application of a novel tumor permeability mixture model within a hierarchical modeling framework. MATERIALS AND METHODS: Thirty-six rats with mammary tumors of varying grade were imaged via dynamic contrast-enhanced (CE) MRI using albumin-(Gd-DTPA)30. Time-dependent contrast agent concentration curves for blood and tumor tissue were obtained and a mathematical model of microvascular blood-tissue exchange was developed under the hypothesis that endothelial integrity is disrupted in a manner proportional to the degree of malignancy, with benign tumors showing no disruption of the vasculature endothelium. This permeability model was incorporated into a statistical model for the benign and malignant tumor subgroups that enabled automatic subject classification. The structural and statistical models were implemented using the software Nonlinear Mixed Effects Modeling (NONMEM) to statistically separate subjects into the two subgroups. RESULTS: Individual tumor classifications (as benign or malignant) were evaluated against the Scarff-Bloom-Richardson microscopic scoring method as applied to the tumor histology of each subject. The model-based classification resulted in 90.9% sensitivity, 92.9% specificity, and 91.7% accuracy. CONCLUSION: Mixture model analysis provides a robust method for subject classification without user intervention and bias. Although the present results are promising, additional research is needed to further evaluate this technique for diagnostic purposes.

Initial computed tomography imaging experience using a new macromolecular iodinated contrast medium in experimental breast cancer.

OBJECTIVE: The objective of this study was to evaluate computed tomography (CT) enhancement characteristics for a new iodinated macromolecular contrast medium (MMCM), PEG12000-Gen4-triiodo, for angiographic effect and for assessment of abnormal vascular permeability in cancer. MATERIALS AND METHODS: Time persistence of angiographic effect was evaluated on rat CT images acquired over 30 minutes using the iodinated polyethyleneglycol- (PEG) based macromolecule. Dynamic CT imaging after PEG12000-Gen4-triiodo-enhancement in tumor-bearing rats was used to quantitatively estimate plasma volume and microvascular transendothelial permeability for both tumor and normal soft tissue. Using identical doses of iodine, 300 mg iodine/kg, blood curves for this MMCM and iohexol were compared. RESULTS: Serial whole-body CT angiograms using PEG12000-Gen4-triiodo showed diagnostic vascular detail through 20 minutes, and the blood enhancement curve was higher and more persistent than with small-molecular iohexol. Permeability estimates were significantly (P<0.02; paired t test) higher in tumors (48.2+/-18.1 microL/min-1 100 mL) than in muscle (2.5+/-5.7 microL/min-1 100 mL). CONCLUSIONS: Use of PEG-based MMCM for experimental CT allowed for a persistent angiographic enhancement and for quantitative estimation of tumor microvascular characteristics.

Magnetic resonance characterization of tumor microvessels in experimental breast tumors using a slow clearance blood pool contrast agent (carboxymethyldextran-A2-Gd-DOTA) with histopathological correlation.

Carboxymethyldextran (CMD)-A2-Gd-DOTA, a slow clearance blood pool contrast agent with a molecular weight of 52.1 kDa, designed to have intravascular residence for more than 1 h, was evaluated for its potential to characterize and differentiate the microvessels of malignant and benign breast tumors. Precontrast single-slice inversion-recovery snapshot FLASH and dynamic contrast-enhanced MRI using an axial T1-weighted three-dimensional spoiled gradient recalled sequence was performed in 30 Sprague-Dawley rats with chemically induced breast tumors. Endothelial transfer coefficient and fractional plasma volume of the breast tumors were estimated from MRI data acquired with CMD-A2-Gd-DOTA enhancement injected at a dose of 0.1 mmol Gd/kg body weight using a two-compartment bidirectional model of the tumor tissue. The correlation between MRI microvessel characteristics and histopathological tumor grade was determined using the Scarff-Bloom-Richardson method. Using CMD-A2-Gd-DOTA, no significant correlations were found between the MR-estimated endothelial transfer coefficient or plasma volumes with histological tumor grade. Analysis of CMD-A2-Gd-DOTA-enhanced MR kinetic data failed to demonstrate feasibility for the differentiation of benign from malignant tumors or for image-based tumor grading.

Zinc absorption and kinetics during pregnancy and lactation in Brazilian women.

BACKGROUND: Adjustments in zinc absorption and endogenous excretion maintain zinc homeostasis in nonpregnant adults fed low-zinc diets. The effects on zinc homeostasis of a low zinc intake during pregnancy and lactation have not been described in a longitudinal study. OBJECTIVE: We examined longitudinal changes in fractional zinc absorption (FZA) and zinc kinetics in 10 healthy Brazilian women who habitually consumed a marginal zinc diet ( approximately 9 mg Zn/d). DESIGN: Zinc status was measured at 10-12 (early pregnancy; EP) and 34-36 (late pregnancy; LP) wk of pregnancy and at 7-8 wk after delivery (early lactation; EL). Zinc kinetics and FZA were studied by using stable isotopic tracers. RESULTS: Zinc intake averaged 9 +/- 3 mg/d throughout the study. FZA increased from 29 +/- 6% at EP to 43 +/- 10% at LP and to 39 +/- 13% at EL (P < 0.05). FZA was inversely related to plasma zinc at EL (r = -0.73, P = 0.02) and LP (r = -0.72, P = 0.07). Plasma zinc mass was 23% greater at LP than at EP or EL (P < 0.05). The amount of zinc (mg/d) that fluxed between plasma and the most-rapidly-turning-over extravascular pool was 53% greater at LP than at EP or EL (P < 0.05). The zinc flux between plasma and the less-rapidly-turning-over zinc pool at EL was 27% greater than that at EP or LP, but this difference was not significant. CONCLUSIONS: FZA increased significantly in women with marginal zinc intakes during pregnancy and lactation; the increase was higher in women with low plasma zinc. Plasma zinc was distributed into a different exchangeable pool at LP than at EL.

The choice of region of interest measures in contrast-enhanced magnetic resonance image characterization of experimental breast tumors.

OBJECTIVES: The objectives of this study were to determine if magnetic resonance (MR) estimates of quantitative tissue microvascular characteristics from regions of interest (ROI) limited to the tumor periphery provided a better correlation with tumor histologic grade than ROI defined for the whole tumor in cross-section. METHODS: A metaanalysis was based on 98 quantitative MR image breast tumor characterizations acquired in 3 separate experimental studies using identical methods for tumor induction and contrast enhancement. RESULTS: The endothelial transfer coefficient (K) of albumin (Gd-DTPA)30 from the tumor periphery correlated (r = 0.784) significantly more strongly (P < 0.001) with the pathologic tumor grade than K derived from the whole tumor (r = 0.604). K estimates, either from the tumor periphery or from the whole tumor, correlated significantly more strongly with histologic grade (P < 0.01) than MR image estimates of fractional plasma volume (fPV) from either tumor periphery (r = 0.368) or whole tumor (r = 0.323). CONCLUSIONS: K estimates from the tumor periphery were the best of these measurable MR image microvascular characteristics for predicting the histologic grade.

MRI monitoring of Avastin antiangiogenesis therapy using B22956/1, a new blood pool contrast agent, in an experimental model of human cancer.

PURPOSE: To evaluate the diagnostic and prognostic potential of a new protein-binding contrast medium, B22956/1, for quantitatively characterizing tumor microvessels by MRI and monitoring response to antiangiogenic therapy. MATERIALS AND METHODS: Dynamic contrast-enhanced MRI (DCE-MRI) was performed in an experimental cancer model with the use of the novel protein-binding agent B22956/1, a low molecular contrast agent (ProHance), and a macromolecular contrast medium, albumin-(Gd-DTPA). MDA-MB-435, a human cancer cell line, was implanted in 22 athymic rats. Animals were assigned randomly to a control (saline) or drug-treated (Avastin) group. MRI was performed at baseline and after nine days of treatment. The transendothelial permeability (KPS) and the fractional blood volume (fBV) were estimated from the kinetic analysis of dynamic MR data using a two-compartment model. Tumor growth was also measured from volumetric MRI. RESULTS: Tumors grew more slowly, although not significantly (P=0.07), in the drug-treated group. The KPS determined for B22956/1 decreased significantly in the drug-treated group compared to baseline (P <0.05), and progressed significantly in the control group. However, no significant changes were resolved with the use of ProHance or albumin-(Gd-DTPA). CONCLUSION: With the use of appropriate contrast media, the therapeutic effects of an anti-VEGF antibody on tumor microvessels can be monitored by dynamic MRI. The dynamic range of permeability to B22956/1, and the sensitivity to change of this parameter suggest a potential application in the clinical setting.

Kinetic parameters and plasma zinc concentration correlate well with net loss and gain of zinc from men.

The search for a reliable, convenient indicator of Zn status was the focus of research for several decades. Plasma Zn concentration is still the most widely used clinical measurement, despite the known problems of interpretation. More recently, researchers suggested that isotopically determined kinetic parameters, such as the exchangeable Zn pool (EZP), may more accurately and reliably reflect body Zn status. The objective of this study was to examine the relationship between net body Zn loss and gain during acute changes in dietary Zn intake with biochemical and kinetic indices of Zn status. Five men participated in an 85-d Zn depletion/repletion study. Net body Zn loss and gain were determined from the difference between dietary plus intravenously administered Zn and Zn excretion. Biochemical indicators of Zn status included plasma Zn, plasma alkaline phosphatase activity, and plasma retinol binding protein concentration. Following intravenous administration of (70)Zn or (67)Zn, a compartmental model was used to determine EZP mass, fractional Zn absorption, endogenous zinc excretion (EZE), and plasma Zn flux. The changes in total body zinc correlated best with changes in plasma Zn (r(2) = 0.826, P < 0.001), EZE (r(2) = 0.773, P < 0.001), and plasma Zn flux (r(2) = 0.766, P < 0.001). This study confirms that plasma Zn concentration is a valid indicator of whole-body Zn status in the absence of confounding factors; however, further research is needed to determine how kinetic parameters respond to conditions where plasma Zn concentration is known to be unreliable.