Mechanism of Action of Brodalumab May Correlate With Efficacy in Patients With Inflammatory Skin Diseases.

Biologic therapy is used for systemic treatment of multiple inflammatory conditions, including moderate-to-severe plaque psoriasis. Brodalumab is an interleukin-17 (IL-17) receptor A antagonist indicated for the treatment of moderate-to-severe plaque psoriasis in adult patients who are candidates for systemic therapy or phototherapy and have failed to respond or have lost response to other systemic therapies. The unique mechanism of action of brodalumab, which blocks signaling mediated by multiple IL-17 family members, may play a key role in the overall efficacy, including in patients whose disease did not respond to other biologics. In this narrative review, we discuss the mechanism of action of brodalumab in inflammatory skin conditions, exploring how it relates to clinical and real-world efficacy, rescued responses after IL-17A inhibitor failure, and improvements in mental health and quality of life. J Drugs Dermatol. 2023;22(10):994-1000 doi:10.36849/JDD.7701.

On-column entrapment of alpha1-acid glycoprotein for studies of drug-protein binding by high-performance affinity chromatography.

An on-column approach for protein entrapment was developed to immobilize alpha1-acid glycoprotein (AGP) for drug-protein binding studies based on high-performance affinity chromatography. Soluble AGP was physically entrapped by using microcolumns that contained hydrazide-activated porous silica and by employing mildly oxidized glycogen as a capping agent. Three on-column entrapment methods were evaluated and compared to a previous slurry-based entrapment method. The final selected method was used to prepare 1.0 cm × 2.1 mm I.D. affinity microcolumns that contained up to 21 (±4) µg AGP and that could be used over the course of more than 150 sample applications. Frontal analysis and zonal elution studies were performed on these affinity microcolumns to examine the binding of various drugs with the entrapped AGP. Site-selective competition studies were also conducted for these drugs. The results showed good agreement with previous observations for these drug-protein systems and with binding constants that have been reported in the literature. The entrapment method developed in this study should be useful for future work in the area of personalized medicine and in the high-throughput screening of drug interactions with AGP or other proteins. Graphical abstract On-column protein entrapment using a hydrazide-activated support and oxidized glycogen as a capping agent.

Entrapment of alpha1-acid glycoprotein in high-performance affinity columns for drug-protein binding studies.

A slurry-based method was developed for the entrapment of alpha1-acid glycoprotein (AGP) for use in high-performance affinity chromatography to study drug interactions with this serum protein. Entrapment was achieved based on the physical containment of AGP in hydrazide-activated porous silica supports and by using mildly oxidized glycogen as a capping agent. The conditions needed for this process were examined and optimized. When this type of AGP column was used in binding studies, the association equilibrium constant (Ka) measured by frontal analysis at pH 7.4 and 37°C for carbamazepine with AGP was found to be 1.0 (±0.5)×10(5)M(-1), which agreed with a previously reported value of 1.0 (±0.1)×10(5)M(-1). Binding studies based on zonal elution were conducted for several other drugs with such columns, giving equilibrium constants that were consistent with literature values. An entrapped AGP column was also used in combination with a column containing entrapped HSA in a screening assay format to compare the binding of various drugs to AGP and HSA. These results also agreed with previous data that have been reported in literature for both of these proteins. The same entrapment method could be extended to other proteins and to the investigation of additional types of drug-protein interactions. Potential applications include the rapid quantitative analysis of biological interactions and the high-throughput screening of drug candidates for their binding to a given protein.

Use of entrapment and high-performance affinity chromatography to compare the binding of drugs and site-specific probes with normal and glycated human serum albumin.

Protein entrapment and high-performance affinity chromatography were used with zonal elution to examine the changes in binding that occurred for site-specific probes and various sulfonylurea drugs with normal and glycated forms of human serum albumin (HSA). Samples of this protein in a soluble form were physically entrapped within porous silica particles by using glycogen-capped hydrazide-activated silica; these supports were then placed into 1.0 cm × 2.1 mm inner diameter columns. Initial zonal elution studies were performed using (R)-warfarin and L-tryptophan as probes for Sudlow sites I and II (i.e., the major drug binding sites of HSA), giving quantitative measures of binding affinities in good agreement with literature values. It was also found for solutes with multisite binding to the same proteins, such as many sulfonylurea drugs, that this method could be used to estimate the global affinity of the solute for the entrapped protein. This entrapment and zonal approach provided retention information with precisions of ±0.1-3.3% (± one standard deviation) and elution within 0.50-3.00 min for solutes with binding affinities of 1 × 10(4)-3 × 10(5) M(-1). Each entrapped-protein column was used for many binding studies, which decreased the cost and amount of protein needed per injection (e.g., the equivalent of only 125-145 pmol of immobilized HSA or glycated HSA per injection over 60 sample application cycles). This method can be adapted for use with other proteins and solutes and should be valuable in high-throughput screening or quantitative studies of drug-protein binding or related biointeractions.

Intermolecular Interactions between Eosin Y and Caffeine Using 1H-NMR Spectroscopy.

DETECHIP has been used in testing analytes including caffeine, cocaine, and tetrahydrocannabinol (THC) from marijuana, as well as date rape and club drugs such as flunitrazepam, gamma-hydroxybutyric acid (GHB), and methamphetamine. This study investigates the intermolecular interaction between DETECHIP sensor eosin Y (DC1) and the analyte (caffeine) that is responsible for the fluorescence and color changes observed in the actual array. Using 1H-NMR, 1H-COSY, and 1H-DOSY NMR methods, a proton exchange from C-8 of caffeine to eosin Y is proposed.

Wild Plum: novel blue fluorescent compounds capable of luminosity restoration in sun-exposed skin.

BACKGROUND: Novel, blue fluorescent solids referred to as Wild Plum compounds can camouflage skin imperfections when incorporated into cosmetic products. We evaluated the relationship between sun exposure and skin fluorescence and determined if the application of Wild Plum formulations could restore lost fluorescence without harming the skin. METHODS: The forehead skin of two groups of volunteers of mixed gender and age was examined for fluorescence and redness. In addition, subjects answered questions describing any adverse sensations they experienced after their skin was exposed to Wild Plum formulations for extended periods of time. RESULTS: Fluorescence measurements of both solar and non-solar skin indicated that repeated sun exposure causes a loss of skin fluorescence. Application of Wild Plum formulations caused an increase in skin fluorescence at all concentrations, restoring solar skin fluorescence to values well beyond that of non-solar skin. Photo analysis and interview questions indicated that these formulations did not cause any symptoms of irritancy. CONCLUSION: Wild Plum compounds have the ability to restore fluorescence of solar skin to a level significantly higher than that associated with non-solar skin. Skin appears more luminous and therefore more youthful. This fluorescence restoration is achieved at relatively low concentrations, without any harmful side effects.

Improved image analysis of DETECHIP® allows for increased specificity in drug discrimination.

DETECHIP® is a novel molecular sensing array being developed for the detection and identification of a variety of compounds including controlled substances. This easy to use technology has the ability to produce a unique identifying binary code for each substance tested. Original analysis methodology relied on human vision to classify color and fluorescence changes within the array. New digital color image analysis techniques using red-green-blue (RGB) color values provided a higher degree of specificity and greater consistency. This image analysis technique was able to detect more subtle changes in color and was therefore able to properly discriminate between substances that previously produced identical codes. This technique was also expanded to analyze changes in RGB color values individually, increasing the length of the code to 48 digits and therefore potentially providing a further increase in specificity. To show the applicability of this new method, a blind study was performed, correctly identifying two unknown analytes.

Digital Image Analysis for DETCHIP® Code Determination.

DETECHIP® is a molecular sensing array used for identification of a large variety of substances. Previous methodology for the analysis of DETECHIP® used human vision to distinguish color changes induced by the presence of the analyte of interest. This paper describes several analysis techniques using digital images of DETECHIP®. Both a digital camera and flatbed desktop photo scanner were used to obtain Jpeg images. Color information within these digital images was obtained through the measurement of red-green-blue (RGB) values using software such as GIMP, Photoshop and ImageJ. Several different techniques were used to evaluate these color changes. It was determined that the flatbed scanner produced in the clearest and more reproducible images. Furthermore, codes obtained using a macro written for use within ImageJ showed improved consistency versus pervious methods.

Characterization of drug interactions with serum proteins by using high-performance affinity chromatography.

The binding of drugs with serum proteins can affect the activity, distribution, rate of excretion, and toxicity of pharmaceutical agents in the body. One tool that can be used to quickly analyze and characterize these interactions is high-performance affinity chromatography (HPAC). This review shows how HPAC can be used to study drug-protein binding and describes the various applications of this approach when examining drug interactions with serum proteins. Methods for determining binding constants, characterizing binding sites, examining drug-drug interactions, and studying drug-protein dissociation rates will be discussed. Applications that illustrate the use of HPAC with serum binding agents such as human serum albumin, α(1)-acid glycoprotein, and lipoproteins will be presented. Recent developments will also be examined, such as new methods for immobilizing serum proteins in HPAC columns, the utilization of HPAC as a tool in personalized medicine, and HPAC methods for the high-throughput screening and characterization of drug-protein binding.

CHROMATOGRAPHIC ANALYSIS OF DRUG INTERACTIONS IN THE SERUM PROTEOME.

The binding of drugs with serum proteins and binding agents such as human serum albumin, α1-acid glycoprotein, and lipoproteins is an important process in determining the activity and fate of many pharmaceuticals in the body. A variety of techniques have been used to study drug interactions with serum proteins, but there is still a need for faster or better methods for such work. High-performance liquid chromatography (HPLC) is one tool that has been utilized in many formats for these types of measurements. Advantages of using HPLC for this application include its speed and precision, its ability to be automated, its good limits of detection, and its compatibility with a wide range of assay formats and detectors. This review will discuss various approaches in which HPLC can be employed for the study of drug-protein interactions. These techniques include the use of soluble proteins in zonal elution and frontal analysis methods or vacancy techniques such as the Hummel-Dreyer method. Zonal elution and frontal analysis methods that make use of immobilized proteins and high-performance affinity chromatography will also be presented. A variety of applications will be examined, ranging from the determination of free drug fractions to the measurement of the strength or rate of a drug-protein interaction. Newer developments that will be discussed include recent work in the creation of novel mathematical approaches for HPLC studies of drug-protein binding, the use of HPLC methods for the high-throughput screening of drug-protein binding, and the creation and use of affinity monoliths or affinity microcolumns for examining drug-protein systems.

Chromatographic analysis of acetohexamide binding to glycated human serum albumin.

Acetohexamide is a drug used to treat type II diabetes and is tightly bound to the protein human serum albumin (HSA) in the circulation. It has been proposed that the binding of some drugs with HSA can be affected by the non-enzymatic glycation of this protein. This study used high-performance affinity chromatography to examine the changes in acetohexamide-HSA binding that take place as the glycation of HSA is increased. It was found in frontal analysis experiments that the binding of acetohexamide to glycated HSA could be described by a two-site model involving both strong and weak affinity interactions. The average association equilibrium constant (K(a)) for the high affinity interactions was in the range of 1.2-2.0×10(5)M(-1) and increased in moving from normal HSA to HSA with glycation levels that might be found in advanced diabetes. It was found through competition studies that acetohexamide was binding at both Sudlow sites I and II on the glycated HSA. The K(a) for acetohexamide at Sudlow site I increased by 40% in going from normal HSA to minimally glycated HSA but then decreased back to near-normal values in going to more highly glycated HSA. At Sudlow site II, the K(a) for acetohexamide first decreased by about 40% and then increased in going from normal HSA to minimally glycated HSA and more highly glycated HSA. This information demonstrates the importance of conducting both frontal analysis and site-specific binding studies in examining the effects of glycation on the interactions of a drug with HSA.

Preparation of high-capacity supports containing protein G immobilized to porous silica.

This study examined the preparation of high-capacity silica supports containing immobilized protein G. A maximum content of 39 mg protein G/g silica was obtained when using 100 A pore size silica, followed by 33 mg/g for 50 A silica and 9.3-24 mg/g for 300-4000 A silica. The surface coverage of protein G increased with pore size, with a maximum level of 0.037 micromol/m(2) being obtained for 4000 A silica. These supports gave comparable apparent activities (i.e., 30-47% binding to rabbit immunoglobulin G [IgG]), with the highest binding capacities (71-77 mg IgG/g silica) being obtained for 50-100 A silica.

Entrapment of proteins in glycogen-capped and hydrazide-activated supports.

A method is described for the entrapment of proteins in hydrazide-activated supports using oxidized glycogen as a capping agent. This approach is demonstrated using human serum albumin (HSA) as a model binding agent. After optimization of this method, a protein content of 43 (+/-1)mg of HSA/g support was obtained for porous silica. The entrapped HSA supports could retain a low-mass drug (S-warfarin) and had activities and equilibrium constants comparable to those for soluble HSA. It was also found that this approach could be used with other proteins and binding agents that had masses between 5.8 and 150kDa.

Characterization of drug-protein interactions in blood using high-performance affinity chromatography.

The binding of drugs with proteins in blood, serum, or plasma is an important process in determining the activity, distribution, rate of excretion, and toxicity of drugs in the body. High-performance affinity chromatography (HPAC) has received a great deal of interest as a means for studying these interactions. This review examines the various techniques that have been used in HPAC to examine drug-protein binding and discusses the types of information that can be obtained through this approach. A comparison of these techniques with traditional methods for binding studies (e.g., equilibrium dialysis and ultrafiltration) will also be presented. The use of HPAC with specific serum proteins and binding agents will then be discussed, including HSA and alpha(1)-acid glycoprotein (AGP). Several examples from the literature are provided to illustrate the applications of such research. Recent developments in this field are also described, such as the use of improved immobilization techniques, new data analysis methods, techniques for working directly with complex biological samples, and work with immobilized lipoproteins. The relative advantages and limitations of the methods that are described will be considered and the possible use of these techniques in the high-throughput screening or characterization of drug-protein binding will be discussed.