Affinity For Plasma Proteins Research Articles

Rational designing of an antidote nanoparticle decorated with abiotic polymer ligands for capturing and neutralizing target toxins

Many of macromolecular toxins induce cell death by directly interacting with cells or induction of inflammatory cytokines. Abiotic polymer ligands (PLs) composed of functional monomers are able to bind and neutralize toxins in vivo and are of great interest for efficient antidotes. However, little has been reported about recognition and neutralization of target molecules in the bloodstream because of readily elimination from the bloodstream. Here, we report a rational design of PLs-decorated lipid nanoparticles (PL-NPs) for neutralizing a target toxin in vivo. PL that decorated on the NPs would cooperatively interacts with target biomacromolecules since the lipid molecules in NPs have a high degree of freedom. In the present study, N-isopropylacrylamide based PLs interacting with histones, major mediators of sepsis, were synthesized. Affinity between PL-NPs and histones depends on monomer composition and polymer length. The optimized PL-NP showed little affinity for plasma proteins. The PL-NPs inhibited the toxicity of histones both in vitro and in vivo, suggesting that PLs on the NPs cooperatively bound to histones and neutralized their toxicity. In addition, circulation time of optimized PL was significantly prolonged by the modification onto NPs. These results provide a platform for designing antidote nanoparticles neutralizing toxic biomacromolecules.

Tuning the Protein Corona of Hydrogel Nanoparticles: The Synthesis of Abiotic Protein and Peptide Affinity Reagents.

Nanomaterials, when introduced into a complex, protein-rich environment, rapidly acquire a protein corona. The type and amount of proteins that constitute the corona depend significantly on the synthetic identity of the nanomaterial. For example, hydrogel nanoparticles (NPs) such as poly(N-isopropylacrylamide) (NIPAm) have little affinity for plasma proteins; in contrast, carboxylated poly(styrene) NPs acquire a dense protein corona. This range of protein adsorption suggests that the protein corona might be "tuned" by controlling the chemical composition of the NP. In this Account, we demonstrate that small libraries of synthetic polymer NPs incorporating a diverse pool of functional monomers can be screened for candidates with high affinity and selectivity to targeted biomacromolecules. Through directed synthetic evolution of NP compositions, one can tailor the protein corona to create synthetic organic hydrogel polymer NPs with high affinity and specificity to peptide toxins, enzymes, and other functional proteins, as well as to specific domains of large proteins. In addition, many NIPAm NPs undergo a change in morphology as a function of temperature. This transformation often correlates with a significant change in NP-biomacromolecule affinity, resulting in a temperature-dependent protein corona. This temperature dependence has been used to develop NP hydrogels with autonomous affinity switching for the protection of proteins from thermal stress and as a method of biomacromolecule purification through a selective thermally induced catch and release. In addition to temperature, changes in pH or buffer can also alter a NP protein corona composition, a property that has been exploited for protein purification. Finally, synthetic polymer nanoparticles with low nanomolar affinity for a peptide toxin were shown to capture and neutralize the toxin in the bloodstream of living mice. While the development of synthetic polymer alternatives to protein affinity reagents is in its early stages, these recent successes using only small libraries of functional monomers are most encouraging. It is likely that by expanding the chemical diversity of functional hydrogels and other polymers, a much broader range of NP-biomacromolecule affinity pairs will result. Since these robust, nontoxic polymers are readily synthesized in the chemistry laboratory, we believe the results presented in this Account offer a promising future for the development of low cost alternatives to more traditional protein affinity reagents such as antibodies.

The influence of displacement compounds on the binding of ochratoxin A to human serum albumin examined with fluorescence anisotropy methods

The subject of the research is the application of the methods of fluorescence anisotropy measurements to study the interaction between human serum albumin (HSA) and toxins and selected medicines (ibuprofen, warfarin, flurbiprofen). Optical spectroscopic methods are useful tools for the study of biologically active compounds. Determining binding intensity of ochratoxin A (OTA) to albumin may be helpful in explaining the effects of toxic influence of OTA. The main factor influencing the distribution of OTA is its affinity for plasma proteins. It was shown that ochratoxin binds strongly to albumin. By the use of the method of fluorescence anisotropy it was proved that the unbound fraction of OTA is higher due to competing interactions with drugs. As a result of separating ochratoxin from protein by competitive compounds, a decrease in the fluorescence anisotropy of the HSA-OTA complex was observed. The largest increase in free fraction of OTA is caused by flurbiprofen, then ibuprofen and warfarin. It will accelerate OTA transport to target organs and shortening its half-life period, leading consequently to a decrease in chronic toxic effects.

Reversed- and normal-phase liquid chromatography in quantitative structure retention–property relationships of newly synthesized seco-androstene derivatives

The rational preselection of drug candidates includes also correlation between physico-chemical properties (lipophilicity, as the key one) and pharmacokinetic properties, as well as pharmacodynamic activity. Lipophilicity can be determined alternatively by chromatographic methods. Chromatographic behavior of nineteen newly synthesized derivatives of 16-cyano-16,17-seco-5-androstene has been studied by reversed-phase and normal-phase thin-layer chromatography (RP- and NP-TLC). Commercial plates RP-C18-HPTLC and water-dioxane and water-acetonitrile, as well as Lux® silica gel plates and toluene-dioxane and toluene-acetonitrile mixtures with different volume fractions of the solvents were used. Retention constants RM0 and C0 for each compound were determined and correlated with (i) theoretical log P values and (ii) pharmacokinetic predictors determined in silico. Significant linear relationship was found between RP TLC retention constants, RM0, and computational logP values as well as between NP TLC retention constants, C0, and logP. Lipophilicity values for the analytes, determined by RP TLC and NP TLC, were also correlated with computer calculated absorption constants, affinity for plasma proteins, volume of distribution and logarithm of blood–brain permeation. Significant linear relationships were obtained. These relations were further improved by introducing other regressors, as molecular size descriptors (molecular mass and/or volume) and a molecular polarity descriptor (total polar surface area). Retention parameters, RM0 and C0, are recommended for lipophilicity expression of analyzed compounds. In silico pharmacokinetic descriptors for the analytes can be expressed as function of the lipophilicity determined by chromatographic methods, the size and the polarity of the molecules expressed as molecular mass/volume and total polar surface area. The analyzed seco-androstene derivatives have adequate lipophilicity which should provide druglikeness and good pharmacokinetic profiles and they can be recommended for further studies in which their biological activity would be examined.

Biomimetic chromatographic analysis of selenium species: Application for the estimation of their pharmacokinetic properties

The retention behavior of selenites, selenates, seleno-DL-methionine, selenocystine, selenocystamine, selenourea, dimethyl selenide, and dimethyl diselenide was investigated by means of biomimetic liquid chromatography. For this purpose, two immobilized artificial membrane (IAM) columns, namely, IAM.PC.DD2 and IAM.PC.MG, and two immobilized plasma protein columns, human serum albumin (HSA) and alpha(1)-acid glycoprotein (AGP) columns, were employed using different mobile phase conditions in respect to pH and buffer composition. In general, satisfactory interrelations between retention factors obtained with the two IAM stationary phases and HSA/AGP columns were obtained. Large differences were observed between biomimetic retention factors and octanol-water logD values, since the latter fail to describe electrostatic interactions. In contrast, despite the column diversity, the net retention outcome on all four biomimetic columns was quite similar, especially in the presence of phosphate-buffered saline, which by its effective shielding alleviates the differences between the stationary phases. Of the two IAM columns, IAM.PC.DD2 showed better performance when compared with HSA and AGP columns as well as to octanol-water partitioning. Biomimetic chromatographic indices were further used to estimate the percentage of human oral absorption and plasma protein binding of the eight selenium species investigated, according to equations previously reported in the literature. The estimated values of human oral absorption imply moderate absorption only for dimethyl diselenide, which also may exhibit considerable plasma protein binding. Moderate affinity for plasma proteins should also be expected for dimethyl selenide and selenocystamine.

99mTc-labeling and in vitro characterization of N4- and N3S-RGDS-derivative peptides

The aim of this work was to characterize the in vitro behavior of N4- and N3S-RGDS-derivative peptides labeled with 99mTc. Peptides AGGG-Abu-GRGDSPK-NH2 (F22) and C(acm)-GGG-Abu-GRGDSPK-NH2 (SMA1) were synthesized by solid phase. The stability of 99mTc-labeled peptides was assessed in a 30-fold molar excess of cysteine and in plasma. The affinity for plasma proteins was also evaluated. Labeling yield was >95% for both peptides. 99mTc-F22 was not stable in presence of cysteine, but 63% of 99mTc remained chelated to SMA1 up to 24 hours. Both peptides showed low affinity to plasma proteins. N3S-RGDS-derivative peptide (SMA1) showed more stable coordination binding with 99mTc and a higher stability in plasma with regard to N4-RGDS-derivative peptide (F22).

New Antithrombotic Drugs

For many decades, heparin and oral anticoagulants have been the antithrombotic drugs of choice in the prevention and treatment of both venous and arterial thromboembolism. The efficacy and the relative safety of these compounds have been consistently proven by a large number of clinical studies that have also progressively contributed to improve their management. However, these drugs have limitations. Unfractionated heparin (UFH) acts by catalyzing the effect of antithrombin III, a natural anticoagulant able to inactivate factors IIa, IXa, and Xa, and also by inhibiting the activation of factors V and VIII by thrombin. The anticoagulant effect of UFH is unpredictable and varies considerably among patients, depending upon age, gender, body weight, smoking status, and renal function. This wide variability is caused by heparin binding to phase reactant proteins; the level of which varies among normal individuals and among disease states. Before therapeutic plasma levels are achieved, the binding to plasma proteins as well as to receptor sites on endothelium must be saturated. Unfractionated heparin must be given parenterally and it requires careful laboratory monitoring to ensure the anticoagulant response is therapeutic.1 Low molecular weight heparin (LMWH) also acts by catalyzing antithrombin III, but because of the reduced number of saccharide units, its antithrombotic activity is more selective and it is mainly based on inactivation of factor Xa because of a reduced ability to inactivate factor IIa. Low molecular weight heparin does not bind to the endothelium and have a lower affinity for plasma proteins. This results in a more predictable bioavailability, in a substantially longer half-life, and in a stable dose response when injected subcutaneously. Therefore, LMWH has a more predictable anticoagulant response that has overcome the need for laboratory monitoring. However, it also needs to be administered parenterally and thus has limited indications for long-term use.2 Characteristics and pharmacokinetics of UFH and LMWH are summarized in T1 and T2. In the last years, new antithrombotic agents have been developed to overcome some of heparin and warfarin limitations. One of the main goals was the search for a greater selectivity in the pharmacologic action of the new compounds. Direct factor Xa inhibitors (DTI) and direct thrombin inhibitors recently underwent extensive evaluation in the prevention and treatment of both venous and arterial thromboembolism. science [coagulation and hematology] New Antithrombotic Drugs Walter Ageno,1 Alexander G.G. Turpie2 1Department of Internal Medicine, University of Insubria, Varese, Italy, 2Department of Internal Medicine, McMaster University, Hamilton, Canada

Localisation of [3H] clonidine binding in rat neurohypophysis by means of electron-microscopic autoradiography.

Electron-microscopic autoradiography of rat neurohypophyses treated with [3H] clonidine, an alpha 2-agonist, showed that binding apparently occurred preferentially at the neurosecretory endings and blood vessels rather than on the pituicytes. Since it is known that clonidine has a high affinity for plasma proteins, the distribution over the neurosecretory nerve endings would suggest the existence of presynaptic alpha 2-binding sites on neurosecretory neurones, which could indicate a regulatory function for catecholamines in neurohypophysial hormone release.