PEG-g-poly(GdDTPA-co-L-cystine): a biodegradable macromolecular blood pool contrast agent for MR imaging

Biodegradable PEGylated Gd-DTPA l-cystine copolymers, PEG-g-poly(GdDTPA-co-l-cystine), were prepared and tested as a blood pool contrast agent in mice. The biodegradable macromolecular agent was designed to be broken down into smaller Gd complexes by endogenous thiols via the disulfide-thiol exchange reaction to facilitate the clearance of Gd complexes after the contrast-enhanced MRI examination. Gd-DTPA l-cystine copolymers were synthesized by condensation polymerization of l-cystine and DTPA-dianhydride in water followed by chelating with Gd(OAc)(3). MPEG-NH(2) (MW = 2000) was then conjugated to the polymeric backbone in different ratios. The macromolecular contrast agent was readily degraded with the incubation of l-cysteine. It also demonstrated superior contrast enhancement in the heart and blood vessels as compared to a low molecular weight control agent, Gd-(DTPA-BMA). At 1 h postcontrast, the PEGylated macromolecular agent still showed prominent enhancement, while little contrast enhancement was detectable in the blood pool by the control agent. PEG-g-poly(GdDTPA-co-l-cystine) shows promise as an MR blood pool imaging agent.     

Polydisulfide Gd(III) chelates as biodegradable macromolecular magnetic resonance imaging contrast agents

Macromolecular gadolinium (Gd)(III) complexes have a prolonged blood circulation time and can preferentially accumulate in solid tumors, depending on the tumor blood vessel hyperpermeability, resulting in superior contrast enhancement in magnetic resonance (MR) cardiovascular imaging and cancer imaging as shown in animal models. Unfortunately, safety concerns related to these agents' slow elimination from the body impede their clinical development. Polydisulfide Gd(III) complexes have been designed and developed as biodegradable macromolecular magnetic resonance imaging (MRI) contrast agents to facilitate the clearance of Gd(III) complexes from the body after MRI examinations. These novel agents can act as macromolecular contrast agents for in vivo imaging and excrete rapidly as low-molecular-weight agents. The rationale and recent development of the novel biodegradable contrast agents are reviewed here. Polydisulfide Gd(III) complexes have relatively long blood circulation time and gradually degrade into small Gd(III) complexes, which are rapidly excreted via renal filtration. These agents result in effective and prolonged in vivo contrast enhancement in the blood pool and tumor tissue in animal models, yet demonstrate minimal Gd(III) tissue retention as the clinically used low-molecular-weight agents. Structural modification of the agents can readily alter the contrast-enhancement kinetics. Polydisulfide Gd(III) complexes are promising for further clinical development as safe, effective, biodegradable macromolecular MRI contrast agents for cardiovascular and cancer imaging, and for evaluation of therapeutic response.     

Gold nanoparticles functionalised by Gd-complex of DTPA-bis(amide) conjugate of glutathione as an MRI contrast agent

The work is directed toward the synthesis of gold nanoparticles (Au NPs) coated with paramagnetic Gd-complex of DTPA-bis(amide) conjugate of glutathione (GdL) for use as a highly efficient MRI contrast agent. Well-dispersed spherical Au NPs coated with gadolinium complexes, abbreviated as Au@GdL, have been obtained; the mean size of Au@GdL is 5-7 nm, and the numbers of GdL are 1.36x10(4) per Au NP. Au@GdL exhibits high longitudinal (r1) and transverse (r2) relaxivities of 1.87x10(5) and 3.02x10(5) mM(-1) s(-1), respectively.     

Design of (Gd-DO3A)n-polydiamidopropanoyl-peptide nucleic acid-D(Cys-Ser-Lys-Cys) magnetic resonance contrast agents

We hypothesized that chelating Gd(III) to 1,4,7-tris(carboxymethylaza)cyclododecane-10-azaacetylamide (DO3A) on peptide nucleic acid (PNA) hybridization probes would provide a magnetic resonance genetic imaging agent capable of hybridization to a specific mRNA. Because of the low sensitivity of Gd(III) as an magnetic resonance imaging (MRI) contrast agent, a single Gd-DO3A complex per PNA hybridization agent could not provide enough contrast for detection of cancer gene mRNAs, even at thousands of mRNA copies per cell. To increase the Gd(III) shift intensity of MRI genetic imaging agents, we extended a novel DO3An-polydiamidopropanoyl (PDAPm) dendrimer, up to n = 16, from the N-terminus of KRAS PNA hybridization agents by solid phase synthesis. A C-terminal D(Cys-Ser-Lys-Cys) cyclized peptide analog of insulin-like growth factor 1 (IGF1) was included to enable receptor-mediated cellular uptake. Molecular dynamic simulation of the (Gd-DO3A-AEEA)16-PDAP4-AEEA2-KRAS PNA-AEEA-D(Cys-Ser-Lys-Cys) genetic imaging nanoparticles in explicit water yielded a pair correlation function similar to that of PAMAM dendrimers, and a predicted structure in which the PDAP dendron did not sequester the PNA. Thermal melting measurements indicated that the size of the PDAP dendron included in the (DO3A-AEEA)n-PDAPm-AEEA2-KRAS PNA-AEEA-D(Cys-Ser-Lys-Cys) probes (up to 16 Gd(III) cations per PNA) did not depress the melting temperatures (Tm) of the complementary PNA/RNA hybrid duplexes. The Gd(III) dendrimer PNA genetic imaging agents in phantom solutions displayed significantly greater T1 relaxivity per probe (r1 = 30.64 +/- 2.68 mM(-1) s(-1) for n = 2, r1 = 153.84 +/- 11.28 mM(-1) s(-1) for n = 8) than Gd-DTPA (r1 = 10.35 +/- 0.37 mM(-1) s(-1)), but less than that of (Gd-DO3A)32-PAMAM dendrimer (r1 = 771.84 +/- 20.48 mM(-1) s(-1)) (P < 0.05). Higher generations of PDAP dendrimers with 32 or more Gd-DO3A residues attached to PNA-D(Cys-Ser-Lys-Cys) genetic imaging agents might provide greater contrast for more sensitive detection.     

Synthesis and characterization of poly(L-glutamic acid) gadolinium chelate: a new biodegradable MRI contrast agent

Most currently evaluated macromolecular contrast agents for magnetic resonance imaging (MRI) are not biodegradable. The goal of this study is to synthesize and characterize poly(l-glutamic acid) (PG) gadolinium chelates as biodegradable blood-pool MRI contrast agents. Two PG chelates of gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) were synthesized through the use of difunctional and monofunctional DTPA precursors. The conjugates were characterized with regard to molecular weight and molecular weight distribution, gadolinium content, relaxivity, and degradability. Distributions of the polymeric MRI contrast agents in various organs were determined by intravenous injection of (111)In-labeled polymers into mice bearing murine breast tumors. MRI scans were performed at 1.5 T in mice after bolus injection of the polymeric chelates. PG-Hex-DTPA-Gd, obtained from aminohexyl-substituted PG and DTPA-dianhydride, was partially cross-linked and was undegradable in the presence of cathepsin B. On the other hand, PG-Bz-DTPA-Gd synthesized directly from PG and monofunctional p-aminobenzyl-DTPA(acetic acid-tert-butyl ester) was a linear polymer and was degradable. The relaxivities of the polymers at 1.5 T were 3-8 times as great as that of Gd-DTPA. Both polymers had high blood concentrations and were primarily accumulated in the kidney. However, PG-Bz-DTPA-Gd was gradually cleared from the body and had significantly less retention in the blood, the spleen, and the kidney. MRI with PG-Bz-DTPA-Gd in mice showed enhanced vascular contrast at up to 2 h after the contrast agent injection. The ability of PG-Bz-DTPA-Gd to be degraded and cleared from the body makes it a favorable macromolecular MRI contrast agent.     

A gadolinium(III) complex of a carboxylic-phosphorus acid derivative of diethylenetriamine covalently bound to inulin, a potential macromolecular MRI contrast agent

A novel conjugate of a polysaccharide and a Gd(III) chelate with potential as contrast agent for magnetic resonance imaging (MRI) was synthesized. The structure of the chelate was derived from H5DTPA by replacing the central pendant arm by a phosphinic acid functional group, which was covalently bound to the polysaccharide inulin. On the average, each monosaccharide unit of the inulin was attached to approximately one (0.9) chelate moiety. The average molecular weight is 23110 and the average number of Gd3+ ions per molecule is 24. The ligand binds the Gd3+ ion in an octadentate fashion via three nitrogen atoms, four carboxylate oxygen atoms, and one P-O oxygen atom, and its first coordination sphere is completed by a water molecule. This compound shows promising properties for application as a contrast agent for MRI thanks to a favorable residence lifetime of this water molecule (170 ns at 298 K), a relatively long rotational correlation time (866 ps at 298 K), and the presence of two water molecules in the second coordination sphere of the Gd3+ ion. Furthermore, its stability toward transmetalation with Zn(II) is as high as that of the clinically used [Gd(DTPA)(H2O)]2-.     

Polyvalent saccharide-functionalized generation 3 poly(amidoamine) dendrimer-methotrexate conjugate as a potential anticancer agent

A saccharide-terminated generation 3 (G3) polyamidoamine (PAMAM) dendrimer was synthesized as a drug carrier. Utilizing this dendritic platform, we have successfully synthesized polyvalent conjugates (G3-MTX) containing the drug methotrexate (MTX). Surface Plasmon Resonance (SPR) results showed that G3-MTX presented three orders of magnitude enhancement in binding avidity to folate-binding protein (FBP) as compared to the free folic acid (FA). Flow cytometric and confocal microscopic analysis showed that conjugate (G3-MTX-FI) containing imaging agent fluorescein-5(6)-carboxamidohexanoic acid (FI) was internalized into folate receptor (FR)-expressing KB cells in dose-dependent and receptor-mediated fashion. The G3-MTX induced a dose-dependent cytotoxicity in the KB cells. Therefore, the polyvalent G3-MTX may have potential as an anticancer nanodevice for the specific targeting and killing of FR-expressing tumor cells.     

Activated clearance of a biotinylated macromolecular MRI contrast agent from the blood pool using an avidin chase

The enhancement characteristics of a contrast agent are dependent on its pharmacokinetics within the body. In the case of macromolecular contrast agents, prolonged enhancement of the blood pool is seen after the first dose, limiting opportunities for repeated injection in the same session. If the enhancement within the blood pool could be intentionally switched off, the macromolecular contrast agents could be used both to define blood volume and vessel permeability, properties that could be useful in studying angiogenesis. In the current study, the avidin-biotin system was coupled to a dendrimer-based macromolecular MRI contrast agent to switch enhancement from the blood pool to the liver. Because avidin causes rapid trapping of the contrast agent in the liver, the blood pool cleared within 2 min of the injection of avidin. This system can be applied to all dendrimer-based macromolecular MRI contrast agents to investigate blood volume and vascular permeability. Moreover, it permits the repeated injection of the contrast agent and the "avidin switch" during a single MR experiment.     

Polyaspartamide gadolinium complexes containing sulfadiazine groups as potential macromolecular MRI contrast agents

Both diethylenetriaminepentaacetic acid (DTPA) and sulfadiazine (SD) were incorporated into polyaspartamides with different side chains, including poly-alpha,beta-[N-(2-hydroxyethyl)-L-aspartamide] (PHEA), poly-alpha,beta-[N- (3-hydroxypropyl)-L-aspartamide] (PHPA), poly-alpha,beta-[N-(2-aminoethy1)-L-aspartamide] (PAEA), poly-alpha,beta-[N-(4-aminobuty1)-L-aspartamide] (PABA), and poly-alpha,beta-[N-(6-aminohexyl)-L-aspartamide] (PAHA). The polyaspartamide ligands containing DTPA and SD groups were further reacted with gadolinium chloride to give the corresponding macromolecular gadolinium complexes. Experimental data of 1H NMR, IR, UV, and elemental analysis exhibited the formation of the polyaspartamide ligands and gadolinium complexes. Relaxivity studies indicated that the macromolecular chelates possess higher relaxivities than that of the clinically used Gd-DTPA. MR imaging showed that the macromolecular chelate PAEA-Gd-DTPA-SD greatly enhanced the contrast of MR images of hepatoma in the lower limb of mice and provided prolonged intravascular duration. Thus the polyaspartamide gadolinium complex containing SD groups is expected to be used as the potential macromolecular MRI contrast agents for hepatoma in mice.     

RGD targeted poly(L-glutamic acid)-cystamine-(Gd-DO3A) conjugate for detecting angiogenesis biomarker alpha(v) beta3 integrin with MRT, mapping

Cyclic Arg-Gly-Asp-D-Phe-Lys [c(RGDfK)] targeted poly(L-glutamic acid) (PGA)-(Gd-DO3A) conjugate with a biodegradable cystamine spacer was prepared and evaluated for in vivo detection of an angiogenesis biomarker, alpha(v)beta3 integrin, in neoplastic tissues with T1 mapping, a quantitative magnetic resonance imaging (MRI) technique. The binding activity of the c(RGDfK) containing conjugate was investigated using in vitro vitronectin assay with human prostate carcinoma DU145 cell line and Kaposi's sarcoma SLK cell line. The peptide c(RGDfK) and PGA-cystamine-(Gd-DO3A) conjugate were used as controls. The binding affinity of polymer bound c(RGDfK) was slightly lower than free c(RGDfK) peptide. The RGD targeted conjugate had higher binding affinity to the DU145 cells than the SLK cells, which was consistent to free c(RGDfK). The imaging of alpha(v)beta3 integrin with targeted PGA-cystamine-(Gd-DO3A) was evaluated in nude mice bearing DU145 and SLK xenografts at a dose of 5 micromol-Gd/kg. The targeted conjugate demonstrated higher in vivo binding affinity to the DU145 xenografts than the SLK xenografts, resulting in a significant decrease of T1 values of water protons in the periphery of the DU145 tumors as shown in the MR T1 maps. No significant decrease of T1 values was observed in the SLK tumor with the targeted conjugate and in both tumors with the non-targeted conjugate. The targeted polymeric Gd(III) chelate conjugate with a degradable spacer has the potential to be a new paradigm for safe and effective probes in molecular imaging with quantitative MR T1 mapping.     

Synthesis and visualization of a membrane-permeable MRI contrast agent

The study of in vivo developmental events has undergone significant advances with the advent of biological molecular imaging techniques such as computer enhanced light microscopy imaging, positron emission tomography (PET), micro-CT, and magnetic resonance imaging (MRI). MRI has proven to be a particularly powerful tool in clinical and biological settings. Images can be acquired of opaque living animals, with the benefit of tracking events of extended periods of time on the same specimen. Contrast agents are routinely used to enhance regions, tissues, and cells that are magnetically similar but histologically distinct. A principal barrier to the development of MR contrast agents for investigating developmental biological questions is the ability to deliver the agent across cellular membranes. As part of our research, we are investigating a number of small molecules that facilitate transport of charged and uncharged species across cell membranes. Here we describe the synthesis and testing of a Gd(III)-based MR contrast agent conjugated to polyarginine that is able to permeate cell membranes. We confirmed cellular uptake of the agent using two-photon laser microscopy to visualize a Eu(III) derivative of the contrast agent in cell culture, and verified this uptake by T₁ analysis of the Gd(III) agent in cells.Abbreviations DOTA 1,4,7,10-tetraazacyclododecane-N,N,N,N-tetraacetic acid - DOTA(tris-t-Bu ester) 1,4,7,10-tetraazacyclododecane-1,4,7-tris(acetic acid-tert-butyl ester)-10-acetic acid - DO3A(tris-t-Bu ester) 1,4,7-tris(tert-butoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane - MRI magnetic resonance imaging - PET positron emission tomography - TPLM two-photon laser microscopy     

Synthesis and physicochemical characterization of Gd-DTPA-B(sLex)A, a new MRI contrast agent targeted to inflammation

A new magnetic resonance imaging (MRI) contrast agent designed to mimic sialyl Lewis X (sLeX) and to target inflammation has been synthesized and characterized. The evolution of its proton longitudinal relaxivity as a function of the magnetic field (NMRD) and temperature has been studied. The exchange rate of the water coordinated to the metal has been assessed by oxygen-17 relaxometry. The transmetalation by zinc(II) ions and the noncovalent binding to human serum albumin have been evaluated. The results show no limitation by the residence time of the coordinated water molecule above room temperature, a higher stability of the complex versus transmetalation by zinc(II) ions than a parent complex, the clinically used Gd-DTPA-BMA, and negligible interaction with human serum albumin.     

In vitro and in vivo evaluation of a novel 99mTc(CO)3-pyrazolyl conjugate of cyclo-(Arg-Gly-Asp-d-Tyr-Lys)

Radiolabeled peptides containing the Arg-Gly-Asp amino acid sequence (single letter code = RGD) have been studied extensively to target integrin receptors upregulated on tumor cells and neovasculature. Integrins are cell surface transmembrane glycoproteins that exist as alphabeta heterodimers. The alphavbeta3 integrin is known to be overexpressed in many tumor types and is expressed at lower levels in normal tissues. Furthermore, alphavbeta3 and alphavbeta5 subtypes are expressed in neovasculature during angiogenesis. Thus, there is some impetus to image angiogenesis and tumor formation in vivo using RGD-based peptide targeting vectors. In this study, we report the design and development of a new cyclic RGD analogue cyclo-[Arg-Gly-Asp-d-Tyr-Lys(PZ)] (PZ = 3,5-Me2-pz(CH2)2N((CH2)3COOH)(CH2)2NH2) that can be radiolabeled with the [99mTc(CO)3(H2O)3]+ metal aquaion. Radiochemical evaluation of this new conjugate in vitro indicated a facile radiosynthesis of the new 99mTc-RGD conjugate with high radiolabeling yields (>or=95%) and high specific activities. In vitro internalization and blocking assays in alphavbeta3 receptor-positive, human M21 melanoma cancer cells showed the ability of this conjugate to target the integrin receptor with high specificity and selectivity. In vivo pharmacokinetic studies in normal CF-1 mice showed rapid clearance from blood with excretion primarily via/through the renal-urinary system. In vivo accumulation of radioactivity in mice bearing either alphavbeta3 receptor-positive or negative human melanoma tumors showed receptor specific uptake of tracer with accumulations of 2.50 +/- 0.29 and 0.71 +/- 0.08% ID/g in alphavbeta3 integrin positive (M21) and negative (M21L) tumors at 1 h postinjection (p.i.), respectively.     

In vivo MR detection of vascular endothelial injury using a new class of MRI contrast agent

A new class of dye-based MRI contrast agents, EB-DTPA-Gd, was designed and synthesized. The contrast agent was found to accumulate at the site of endothelial injury when the reagent was applied to isolated porcine blood vessels or in an ex vivo experiment using rat. In vivo MR detection of vascular endothelial injury was also successful in rat with its common carotid artery injured by balloon treatment. These results indicate that EB-DTPA-Gd is potentially useful for the diagnosis of vascular diseases.     

Synthesis and evaluation of nanoglobular macrocyclic Mn(II) chelate conjugates as non-gadolinium(III) MRI contrast agents

Because of the recent observation of the toxic side effects of Gd(III) based MRI contrast agents in patients with impaired renal function, there is strong interest on developing alternative contrast agents for MRI. In this study, macrocyclic Mn(II) chelates were conjugated to nanoglobular carriers, lysine dendrimers with a silsesquioxane core, to synthesize non-Gd(III) based MRI contrast agents. A generation 3 nanoglobular conjugate of Mn(II)-1,4,7-triaazacyclononane-1,4,7-triacetate-GA amide (G3-NOTA-Mn) was also synthesized and evaluated. The per ion T(1) and T(2) relaxivities of G2, G3, G4 nanoglobular Mn(II)-DOTA monoamide conjugates decreased with increasing generation of the carriers. The T(1) relaxivities of G2, G3, and G4 nanoglobular Mn(II)-DOTA conjugates were 3.3, 2.8, and 2.4 mM(-1) s(-1) per Mn(II) chelate at 3 T, respectively. The T(1) relaxivity of G3-NOTA-Mn was 3.80 mM(-1) s(-1) per Mn(II) chelate at 3 T. The nanoglobular macrocyclic Mn(II) chelate conjugates showed good in vivo stability and were readily excreted via renal filtration. The conjugates resulted in much less nonspecific liver enhancement than MnCl(2) and were effective for contrast-enhanced tumor imaging in nude mice bearing MDA-MB-231 breast tumor xenografts at a dose of 0.03 mmol Mn/kg. The nanoglobular macrocyclic Mn(II) chelate conjugates are promising nongadolinium based MRI contrast agents.     

Synthesis and characterization of novel natural product-Gd(III) MRI contrast agent conjugates

Several novel gadolinium chelates conjugated with paclitaxel, colchicine and thyroxine have been prepared as MRI contrast agents targeted to tubulin and thyroxine-binding globulin, respectively.     

A novel cholic acid-based contrast enhancement agent for targeted MRI

The novel Gd(III) complexes of heptadentate ligands NE3TA and NE3TA-Bn were prepared, and their relaxivities were measured and favorably compared to the commercially available MRI contrast enhancement agent Gd(DOTA). NE3TA was conjugated with cholic acid (CA) to produce CA-NE3TA. TEM images of Gd(CA-NE3TA) indicate that the complex self-assembles forming nano-sized micelles and displays an over threefold increased relaxivity compared to Gd(DOTA). The new cholic acid-conjugated nanoparticle MR contrast enhancement agent, Gd(CA-NE3TA) possesses great promise for use in targeted MRI.     

新型生物可降解塑料——多聚羟基烷酸研究进展

面对日益严重的白色污染 ,人们迫切需要一种能在自然界较快分解的新型塑料。多聚羟基烷酸是原核生物在不平衡代谢条件下形成的碳源和能源贮藏物质 ,这种贮藏物质如同淀粉、糖原一样 ,当生命活动需要时可以再分解利用。由于多聚羟基烷酸有着与石化塑料相似的理化性质 ,又能在一定条件下被微生物迅速而彻底地降解 ,因此是一种理想的传统石化塑料替代品。1　多聚羟基烷酸的理化及生物学特性1 1　多聚羟基烷酸的分子结构及理化性质多聚羟基烷酸是由羟基脂肪酸单体首尾相联构成的高分图 1　多聚羟基烷酸的分子结构子聚合物。又分为不同种类。如多…     

Synthesis,in vitro and in vivo studies of Gd-DTPA-XDA-D1-Glc(OH) complex as a new potential MRI contrast agent

A new type of dendritic molecules Gd-DTPA-XDA-D1-Glc(OH), which work as a functionalized ligand coordinating gadolinium(III) ion at the center of their frameworks with two glucose moieties on the molecular surfaces, were readily synthesized with high yield. The structures were established by IR, ¹H, ¹³C NMR, and mass spectral studies. Its bio-distribution patterns were evaluated on rats.