Peroxymonosulfate activation by CuMn-LDH for the degradation of bisphenol A: Effect, mechanism, and pathway.

The remediation of water contaminated with bisphenol A (BPA) has gained significant attention. In this study, a hydrothermal composite activator of Cu3Mn-LDH containing coexisting phases of cupric nitrate (Cu(NO3)2) and manganous nitrate (Mn(NO3)2) was synthesized. Advanced oxidation processes were employed as an effective approach for BPA degradation, utilizing Cu3Mn-LDH as the catalyst to activate peroxymonosulfate (PMS). The synthesis of the Cu3Mn-LDH material was characterized using X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and transmission electron microscopy (TEM). According to the characterization data and screening experiments, Cu3Mn-LDH was selected as the best experimental material. Cu3Mn-LDH exhibits remarkable catalytic ability with PMS, demonstrating good degradation efficiency of BPA under neutral and alkaline conditions. With a PMS dosage of 0.25 g·L-1 and Cu3Mn-LDH dosage of 0.10 g·L-1, 10 mg·L-1 BPA (approximately 17.5 µM) can be completely degraded within 40 min, of which the TOC removal reached 95%. The reactive oxygen species present in the reaction system were analyzed by quenching experiments and EPR. Results showed that sulfate free radicals (SO4•-), hydroxyl free radicals (•OH), superoxide free radicals (•O2-), and nonfree radical mono-oxygen were generated, while mono-oxygen played a key role in degrading BPA. Cu3Mn-LDH exhibits excellent reproducibility, as it can still completely degrade BPA even after four consecutive cycles. The degradation intermediates of BPA were detected by GCMS, and the possible degradation pathways were reasonably predicted. This experiment proposes a nonradical degradation mechanism for BPA and analyzes the degradation pathways. It provides a new perspective for the treatment of organic pollutants in water.

Multiplex Hybrid Antigen-Capture LC-MRM Quantification in Sera and Nasal Lining Fluid of AZD7442, a SARS-CoV-2-Targeting Antibody Combination.

AZD7442 (tixagevimab [AZD8895]/cilgavimab [AZD1061]) is a monoclonal antibody (mAb) combination in development for the prevention and treatment of coronavirus disease 2019. Traditionally, bioanalysis of mAbs is performed using ligand binding assays (LBAs), which offer sensitivity, robustness, and ease of implementation. However, LBAs frequently require generation of critical reagents that typically take several months. Instead, we developed a highly sensitive (5 ng/mL limit of quantification) method using a hybrid LBA-liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) approach for quantification of the two codosed antibodies in serum and nasal lining fluid (NLF), a rare matrix. The method was optimized by careful selection of multiple reaction monitoring, capture reagents, magnetic beads, chromatographic conditions, evaluations of selectivity, and matrix effect. The final assay used viral spike protein receptor-binding domain as capture reagent and signature proteotypic peptides from the complementarity-determining region of each mAb for detection. In contrast to other methods of similar/superior sensitivity, our approach did not require multidimensional separations and can be operated in an analytical flow regime, ensuring high throughput and robustness required for clinical analysis at scale. The sensitivity of this method significantly exceeds typical sensitivity of ∼100 ng/mL for analytical flow 1D LBA-LC-MS/MS methods for large macromolecules, such as antibodies. Furthermore, infection and vaccination status did not impact method performance, ensuring method robustness and applicability to a broad patient population. This report demonstrated the general applicability of the hybrid LBA-LC-MS/MS approach to platform quantification of antibodies with high sensitivity and reproducibility, with specialized extension to matrices of increasing interest, such as NLF.

Research Progress on the Protective Effect of Brown Algae-Derived Polysaccharides on Metabolic Diseases and Intestinal Barrier Injury.

In the human body, the intestine is the largest digestive and immune organ, where nutrients are digested and absorbed, and this organ plays a key role in host immunity. In recent years, intestinal health issues have gained attention and many studies have shown that oxidative stress, inflammation, intestinal barrier damage, and an imbalance of intestinal microbiota may cause a range of intestinal diseases, as well as other problems. Brown algae polysaccharides, mainly including alginate, fucoidan, and laminaran, are food-derived natural products that have received wide attention from scholars owing to their good biological activity and low toxic side effects. It has been found that brown algae polysaccharides can repair intestinal physical, chemical, immune and biological barrier damage. Principally, this review describes the protective effects and mechanisms of brown algae-derived polysaccharides on intestinal health, as indicated by the ability of polysaccharides to maintain intestinal barrier integrity, inhibit lipid peroxidation-associated damage, and suppress inflammatory cytokines. Furthermore, our review aims to provide new ideas on the prevention and treatment of intestinal diseases and act as a reference for the development of fucoidan as a functional product for intestinal protection.

Characterization of Antibody-Drug Conjugate Pharmacokinetics and in Vivo Biotransformation Using Quantitative Intact LC-HRMS and Surrogate Analyte LC-MRM.

Antibody-drug conjugates (ADCs) pose challenges to bioanalysis because of their inherently intricate structures and potential for very complex catabolism. Common bioanalysis strategy is to measure the concentration of ADCs and Total Antibody (Ab) as well as deconjugated warhead in circulation. The ADCs and the Total Ab can be quantified with ligand binding assays (LBA) or with hybrid immunocapture-liquid chromatography coupled with multiple reaction monitoring mass spectrometry (LBA-LC-MRM). With the LBA-LC-MRM approach, a surrogate analyte, often the signature peptide, and released warhead can be used for the quantification of the Total Ab and ADCs, respectively. Recent advances in analytical instrumentation, especially the development of high resolution mass spectrometers (HRMS), have enabled characterization and quantification of intact macromolecules such as ADCs. The LBA-LC-HRMS approach employs immunocapture, followed by chromatographic separation at the macromolecule level and detection of the intact analyte. We developed an intact quantification method with 1-10 µg/mL linear dynamic range using 25 µL of plasma sample volume. This method was qualified for the measurement of naked monoclonal antibody (mAb), a site-specific cysteine-conjugated ADC with drug to antibody ratio ∼2 (DAR2) and a site-nonspecific cysteine-conjugated ADC (DAR8) in rat plasma. Samples from a rat pharmacokinetic (PK) study were analyzed with both methods. For the naked mAb, the results from both assays matched well. For ADCs, new species were observed from the LBA-HRMS method. The results demonstrated that potential biotransformation of the ADC was unveiled using the intact quantification approach while not being observed with traditional LBA-LC-MRM approach. Our work demonstrated an application of novel intact quantification by supporting animal PK studies. Moreover, our results suggest that the intact quantification method can provide novel perspectives on ADC in vivo characterization and quantification, which can benefit future drug candidate optimization as well as the immunogenicity impact evaluation and safety assessment.

Multifaceted Bioanalytical Methods for the Comprehensive Pharmacokinetic and Catabolic Assessment of MEDI3726, an Anti-Prostate-Specific Membrane Antigen Pyrrolobenzodiazepine Antibody-Drug Conjugate.

Complex biotherapeutic modalities, such as antibody-drug conjugates (ADC), present significant challenges for the comprehensive bioanalytical characterization of their pharmacokinetics (PK) and catabolism in both preclinical and clinical settings. Thus, the bioanalytical strategy for ADCs must be designed to address the specific structural elements of the protein scaffold, linker, and warhead. A typical bioanalytical strategy for ADCs involves quantification of the Total ADC, Total IgG, and Free Warhead concentrations. Herein, we present bioanalytical characterization of the PK and catabolism of a novel ADC. MEDI3726 targets prostate-specific membrane antigen (PMSA) and is comprised of a humanized IgG1 antibody site-specifically conjugated to tesirine (SG3249). The MEDI3726 protein scaffold lacks interchain disulfide bonds and has an average drug to antibody ratio (DAR) of 2. Based on the structural characteristics of MEDI3726, an array of 4 bioanalytical assays detecting 6 different surrogate analyte classes representing at least 14 unique species was developed, validated, and employed in support of a first-in-human clinical trial (NCT02991911). MEDI3726 requires the combination of heavy-light chain structure and conjugated warhead to selectively deliver the warhead to the target cells. Therefore, both heavy-light chain dissociation and the deconjugation of the warhead will affect the activity of MEDI3726. The concentration-time profiles of subjects dosed with MEDI3726 revealed catabolism of the protein scaffold manifested by the more rapid clearance of the Active ADC, while exhibiting minimal deconjugation of the pyrrolobenzodiazepine (PBD) warhead (SG3199).

Population pharmacokinetics, efficacy, and safety of moxetumomab pasudotox in patients with relapsed or refractory hairy cell leukaemia.

AIMS: To characterize the pharmacokinetics (PK) of moxetumomab pasudotox, an anti-CD22 recombinant immunotoxin, in adults with relapsed or refractory hairy cell leukaemia, we examined data from a phase 1 study (Study 1001; n = 49) and from the pivotal clinical study (Study 1053; n = 74). METHODS: Data from both studies were pooled (n = 123) to develop a population PK model. Covariates included demographics, disease state, liver and kidney function, prior treatment, and antidrug antibodies (ADAs). Exposure-response and exposure-safety were analysed separately by study. A 1-compartment model with linear elimination from the central compartment and 2 clearance (CL) rates was developed. RESULTS: Moxetumomab pasudotox was cleared more rapidly after cycle 1, day 1 (CL1 = 24.7 L/h) than subsequently (CL2 = 3.76 L/h), with high interindividual variability (116 and 109%, respectively). In Study 1053, patients with ADA titres >10 240 showed ~4-fold increase in CL. Higher exposures (≥median) were related to higher response rates, capillary leak syndrome and increased creatinine (Study 1053 only), or grade ≥3 adverse events (Study 1001 only). Clinical benefits were still observed in patients with lower exposure or high ADA titres. CONCLUSION: Despite a high incidence of immunogenicity with increased clearance, moxetumomab pasudotox demonstrated efficacy in hairy cell leukaemia.

Results from an international phase 2 study of the anti-CD22 immunotoxin moxetumomab pasudotox in relapsed or refractory childhood B-lineage acute lymphoblastic leukemia.

BACKGROUND: In a multicenter phase 1 study of children with relapsed/refractory acute lymphoblastic leukemia (ALL), moxetumomab pasudotox, an anti-CD22 immunotoxin, demonstrated a manageable safety profile and preliminary evidence of clinical activity. A phase 2 study further evaluated efficacy. PROCEDURE: This international, multicenter, phase 2 study enrolled children with relapsed/refractory B-cell precursor ALL who received moxetumomab pasudotox 40 µg/kg intravenously every other day, for six doses per 21-day cycle. The primary objective was to evaluate the complete response (CR) rate. Secondary objectives included safety, pharmacokinetics, and immunogenicity evaluations. RESULTS: Thirty-two patients (median age, 10 years) were enrolled at 16 sites; 30 received study drug and were evaluable for safety; 28 were evaluable for response. The objective response rate was 28.6%, with three patients (10.7%) achieving morphologic CR, and five patients (17.9%) achieving partial response. Disease progression occurred in 11 patients (39.3%). Ten patients (33.3%) experienced at least one treatment-related serious adverse event, including capillary leak syndrome (CLS; n = 6), hemolytic uremic syndrome (HUS; n = 4), and treatment-related death (n = 1) from pulmonary edema. No differences were observed in inflammatory markers in patients who did or did not develop CLS or HUS. CONCLUSIONS: Despite a signal for clinical activity, this phase 2 study was terminated at interim analysis for a CR rate that did not reach the stage 1 target. Preclinical data suggest enhanced efficacy of moxetumomab pasudotox via continuous infusion or in combination regimens; thus, further studies designed to optimize the efficacy and safety of moxetumomab pasudotox in pediatric ALL may be warranted.

Phase 1 study of the anti-CD22 immunotoxin moxetumomab pasudotox for childhood acute lymphoblastic leukemia.

Novel therapies are needed to overcome chemotherapy resistance for children with relapsed/refractory acute lymphoblastic leukemia (ALL). Moxetumomab pasudotox is a recombinant anti-CD22 immunotoxin. A multicenter phase 1 study was conducted to determine the maximum-tolerated cumulative dose (MTCD) and evaluate safety, activity, pharmacokinetics, and immunogenicity of moxetumomab pasudotox in children, adolescents, and young adults with ALL (N = 55). Moxetumomab pasudotox was administered as a 30-minute IV infusion at doses of 5 to 50 µg/kg every other day for 6 (cohorts A and B) or 10 (cohort C) doses in 21-day cycles. Cohorts B and C received dexamethasone prophylaxis against capillary leak syndrome (CLS). The most common treatment-related adverse events were reversible weight gain, hepatic transaminase elevation, and hypoalbuminemia. Dose-limiting CLS occurred in 2 of 4 patients receiving 30 µg/kg of moxetumomab pasudotox every other day for 6 doses. Incorporation of dexamethasone prevented further dose-limiting CLS. Six of 14 patients receiving 50 µg/kg of moxetumomab pasudotox for 10 doses developed hemolytic uremic syndrome (HUS), thrombotic microangiopathy (TMA), or HUS-like events, exceeding the MTCD. Treatment expansion at 40 µg/kg for 10 doses (n = 11) exceeded the MTCD because of 2 HUS/TMA/HUS-like events. Dose level 6B (ie, 50 µg/kg × 6 doses) was the MTCD, selected as the recommended phase 2 dose. Among 47 evaluable patients, an objective response rate of 32% was observed, including 11 (23%) composite complete responses, 5 of which were minimal residual disease negative by flow cytometry. Moxetumomab pasudotox showed a manageable safety profile and evidence of activity in relapsed or refractory childhood ALL. This trial was registered at www.clinicaltrials.gov as #NCT00659425.

A CD80-Biased CTLA4-Ig Fusion Protein with Superior In Vivo Efficacy by Simultaneous Engineering of Affinity, Selectivity, Stability, and FcRn Binding.

Affinity- and stability-engineered variants of CTLA4-Ig fusion molecules with enhanced pharmacokinetic profiles could yield improved therapies with the potential of higher efficacy and greater convenience to patients. In this study, to our knowledge, we have, for the first time, used in vitro evolution to simultaneously optimize CTLA4 affinity and stability. We selected for improved binding to both ligands, CD80 and CD86, and screened as dimeric Fc fusions directly in functional assays to identify variants with stronger suppression of in vitro T cell activation. The majority of CTLA4 molecules showing the largest potency gains in primary in vitro and ex vivo human cell assays, using PBMCs from type 1 diabetes patients, had significant improvements in CD80, but only modest gains in CD86 binding. We furthermore observed different potency rankings between our lead molecule MEDI5265, abatacept, and belatacept, depending on which type of APC was used, with MEDI5265 consistently being the most potent. We then created fusions of both stability- and potency-optimized CTLA4 moieties with human Fc variants conferring extended plasma t1/2 In a cynomolgus model of T cell-dependent Ab response, the CTLA4-Ig variant MEDI5265 could be formulated at >100 mg/ml for s.c. administration and showed superior efficacy and significantly prolonged serum t1/2 The combination of higher stability and potency with prolonged pharmacokinetics could be compatible with very infrequent, s.c. dosing while maintaining a similar level of immune suppression to more frequently and i.v. administered licensed therapies.

Preparation and application of molecularly imprinted polymer solid-phase microextraction fiber for the selective analysis of auxins in tobacco.

As signal molecules, auxins play an important role in mediating plant growth. Due to serious interfering substances in plants, it is difficult to accurately detect auxins with traditional solid-phase extraction methods. To improve the selectivity of sample pretreatment, a novel molecularly imprinted polymer -coated solid-phase microextraction fiber, which could be coupled directly to high-performance liquid chromatography, was prepared with indole acetic acid as template molecule for the selective extraction of auxins. The factors influencing the polymer formation, such as polymerization solvent, cross-linker, and polymerization time, were investigated in detail to enhance the performance of indole acetic acid-molecularly imprinted polymer coating. The morphological and chemical stability of this molecularly imprinted polymer-coated fiber was characterized by scanning electron microscopy, infrared spectrometry, and thermal analysis. The extraction capacity of the molecularly imprinted polymer-coated solid-phase microextraction fiber was evaluated for the selective extraction of indole acetic acid and indole-3-pyruvic acid followed by high-performance liquid chromatography analysis. The linear range for indole acetic acid and indole-3-pyruvic acid was 1-100 µg/L and their detection limit was 0.5 µg/L. The method was applied to the simultaneous determination of two auxins in two kinds of tobacco (Nicotiana tabacum L and Nicotiana rustica L) samples, with recoveries range from 82.1 to 120.6%.

Characterization, dissolution and solubility of the hydroxypyromorphite-hydroxyapatite solid solution [(PbxCa1-x)5(PO4)3OH] at 25 °C and pH 2-9.

BACKGROUND: The interaction between Ca-HAP and Pb(2+) solution can result in the formation of a hydroxyapatite-hydroxypyromorphite solid solution [(PbxCa1-x)5(PO4)3(OH)], which can greatly affect the transport and distribution of toxic Pb in water, rock and soil. Therefore, it's necessary to know the physicochemical properties of (PbxCa1-x)5(PO4)3(OH), predominantly its thermodynamic solubility and stability in aqueous solution. Nevertheless, no experiment on the dissolution and related thermodynamic data has been reported. RESULTS: Dissolution of the hydroxypyromorphite-hydroxyapatite solid solution [(PbxCa1-x)5(PO4)3(OH)] in aqueous solution at 25 °C was experimentally studied. The aqueous concentrations were greatly affected by the Pb/(Pb + Ca) molar ratios (XPb) of the solids. For the solids with high XPb [(Pb0.89Ca0.11)5(PO4)3OH], the aqueous Pb(2+) concentrations increased rapidly with time and reached a peak value after 240-720 h dissolution, and then decreased gradually and reached a stable state after 5040 h dissolution. For the solids with low XPb (0.00-0.80), the aqueous Pb(2+) concentrations increased quickly with time and reached a peak value after 1-12 h dissolution, and then decreased gradually and attained a stable state after 720-2160 h dissolution. CONCLUSIONS: The dissolution process of the solids with high XPb (0.89-1.00) was different from that of the solids with low XPb (0.00-0.80). The average K sp values were estimated to be 10(-80.77±0.20) (10(-80.57)-10(-80.96)) for hydroxypyromorphite [Pb5(PO4)3OH] and 10(-58.38±0.07) (10(-58.31)-10(-58.46)) for calcium hydroxyapatite [Ca5(PO4)3OH]. The Gibbs free energies of formation (ΔG f (o) ) were determined to be -3796.71 and -6314.63 kJ/mol, respectively. The solubility decreased with the increasing Pb/(Pb + Ca) molar ratios (XPb) of (PbxCa1‒x)5(PO4)3(OH). For the dissolution at 25 °C with an initial pH of 2.00, the experimental data plotted on the Lippmann diagram showed that the solid solution (PbxCa1-x)5(PO4)3(OH) dissolved stoichiometrically at the early stage of dissolution and moved gradually up to the Lippmann solutus curve and the saturation curve for Pb5(PO4)3OH, and then the data points moved along the Lippmann solutus curve from right to left. The Pb-rich (PbxCa1-x)5(PO4)3(OH) was in equilibrium with the Ca-rich aqueous solution. Graphical abstractLippmann diagrams for dissolution of the hydroxypyromorphite-hydroxyapatite solid solution [(PbxCa1-x)5(PO4)3OH] at 25 ˚C and an initial pH of 2.00.

Quantitative measurement of the target-mediated internalization kinetics of biopharmaceuticals.

PURPOSE: Measurement of internalization of biopharmaceuticals targeting cell surface proteins can greatly facilitate drug development. The objective of this study was to develop a reliable method for determination of internalization rate constant (kint) and to demonstrate its utility. METHODS: This method utilized confocal imaging to record the internalization kinetics of fluorescence-tagged biopharmaceuticals in live-cells and a quantitative image-analysis algorithm for kint determination. Kint was incorporated into a pharmacokinetic-pharmacodynamic (PK-PD) model for simulation of the drug PK profiles, target occupancy and the displacement of endogenous ligand. RESULTS: The method was highly sensitive, allowing kint determination in cells expressing as low as 5,000 receptors/cell, and was amenable to adherent and suspension cells. Its feasibility in a mixed cell population, such as whole blood, was also demonstrated. Accurate assessment of the kint was largely attributed to continuous monitoring of internalization in live cells, rapid confocal image acquisition and quantitative image-analysis algorithm. Translational PK-PD simulations demonstrated that kint is a major determinant of the drug PK profiles, target occupancy, and the displacement of endogenous ligand. CONCLUSIONS: The developed method is robust for broad cell types. Reliable kint assessment can greatly expedite biopharmaceutical development by facilitating target evaluation, drug affinity goal setting, and clinical dose projection.

A review of the degradation of antibiotic contaminants using advanced oxidation processes: modification and application of layered double hydroxides based materials.

In recent years, the treatment of organic pollutants has become a global concern due to the threat to human health posed by emerging contaminants, especially antibiotic contamination. Advanced oxidation processes (AOPs) can solve the organic pollution problem well, which have been identified as a promising solution for the treatment of hard-to-handle organic compounds including antibiotic contaminants. Layered double hydroxides (LDHs) are excellent catalysts because of their flexible tunability, favorable thermal stability, abundant active sites, and facile exchangeability of intercalated anions. This paper conducted a systematic review of LDHs-based materials used for common antibiotic removal by three significant AOP technologies, such as photocatalysis, the Fenton-like processes, and peroxymonosulfate catalysis. The degradation effects studied in various studies were reviewed, and the mechanisms were discussed in detail based on the type of AOPs. Finally, the challenges and the application trends of AOPs that may arise were prospected. The aim of this study is to suggest ways to provide practical guidance for the screening and improvement of LDH materials and the rational selection of AOPs to achieve efficient antibiotic degradation. This could lead to the development of more efficient and environmentally friendly materials and processes for degrading antibiotics, with significant implications for our ecological conservation by addressing water pollution.

Research Progress on Biological Accumulation, Detection and Inactivation Technologies of Norovirus in Oysters.

Noroviruses (NoVs) are major foodborne pathogens that cause acute gastroenteritis. Oysters are significant carriers of this pathogen, and disease transmission from the consumption of NoVs-infected oysters occurs worldwide. The review discusses the mechanism of NoVs bioaccumulation in oysters, particularly the binding of histo-blood group antigen-like (HBGA-like) molecules to NoVs in oysters. The review explores the factors that influence NoVs bioaccumulation in oysters, including temperature, precipitation and water contamination. The review also discusses the detection methods of NoVs in live oysters and analyzes the inactivation effects of high hydrostatic pressure, irradiation treatment and plasma treatment on NoVs. These non-thermal processing treatments can remove NoVs efficiently while retaining the original flavor of oysters. However, further research is needed to reduce the cost of these technologies to achieve large-scale commercial applications. The review aims to provide novel insights to reduce the bioaccumulation of NoVs in oysters and serve as a reference for the development of new, rapid and effective methods for detecting and inactivating NoVs in live oysters.

[Application of Fe3[KG-*2/5]O4/Mulberry Stem Biochar Effects on Soil Arsenic Species and Rice Arsenic Content].

The passivation effect of Fe3O4/mulberry pole biochar (Fe-MBC) prepared at different carbonization temperatures on soil available arsenic content was studied through soil culture experiments, and Fe-MBC-800 (prepared by carbonization at 800℃) with good passivation effect was selected and characterized. The effects of 1%-7% (mass fraction of biochar to soil) Fe-MBC-800, MBC-800, and Fe3O4 on soil pH value, soil electrical conductivity, soil arsenic form, rice biomass, and total arsenic (As) content in rice were studied using a pot experiment. The results showed that:①Fe-MBC-800 successfully loaded Fe3O4, and its main functional groups were C=O double bond, O-H bond, C-O bond, and Fe-O bond. The specific surface areas of Fe-MBC-800, MBC-800, and Fe3O4 were 209.659 m2·g-1, 517.714 m2·g-1, and 68.025 m2·g-1, respectively. ②The addition of Fe-MBC-800 could increase the soil pH value, decrease the soil EC value, increase the content of residual arsenic in soil, and reduce the content of water-soluble arsenic and available arsenic in the soil. Under the treatment using 7% Fe-MBC-800 (ω) amendments, the content of water-soluble arsenic and available arsenic in the soil decreased by 81.6% and 56.33%, respectively. ③When the addition ratio of Fe-MBC-800 in the soil was 5%-7%, it could promote the growth of rice plants, increase rice biomass, and reduce the bioaccumulation of arsenic by between 62.5% and 68.75%.

Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody-Drug Conjugate.

Deamidation, a common post-translational modification, may impact multiple physiochemical properties of a therapeutic protein. MEDI7247, a pyrrolobenzodiazepine (PBD) antibody-drug conjugate (ADC), contains a unique deamidation site, N102, located within the complementarity-determining region (CDR), impacting the affinity of MEDI7247 to its target. Therefore, it was necessary to monitor MEDI7247 deamidation status in vivo. Due to the low dose, a sensitive absolute quantification method using immunocapture coupled with liquid chromatography-tandem mass spectrometry (LBA-LC-MS/MS) was developed and qualified. We characterized the isomerization via Electron-Activated Dissociation (EAD), revealing that deamidation resulted in iso-aspartic acid. The absolute quantification of deamidation requires careful assay optimization in order not to perturb the balance of the deamidated and nondeamidated forms. Moreover, the selection of capture reagents essential for the correct quantitative assessment of deamidation was evaluated. The final assay was qualified with 50 ng/mL LLOQ for ADC for total and nondeamidated antibody quantification, with qualitative monitoring of the deamidated antibody. The impact of deamidation on the pharmacokinetic characteristics of MEDI7247 from clinical trial NCT03106428 was analyzed, revealing a gradual reduction in the nondeamidated form of MEDI7247 in vivo. Careful quantitative biotransformation analyses of complex biotherapeutic conjugates help us understand changes in product PTMs after administration, thus providing a more complete view of in vivo pharmacology.

Immobilization of Biomass Materials for Removal of Refractory Organic Pollutants from Wastewater.

In the field of environmental science and engineering, microorganisms, enzymes and algae are promising biomass materials that can effectively degrade pollutants. However, problems such as poor environmental adaptability, recycling difficulties, and secondary pollution exist in the practical application of non-immobilized biomass materials. Biomass immobilization is a novel environmental remediation technology that can effectively solve these problems. Compared with non-immobilized biomass, immobilized biomass materials have the advantages of reusability and stability in terms of pH, temperature, handling, and storage. Many researchers have studied immobilization technology (i.e., methods, carriers, and biomass types) and its applications for removing refractory organic pollutants. Based on this, this paper reviews biomass immobilization technology, outlines the mechanisms and factors affecting the removal of refractory organic pollutants, and introduces the application of immobilized biomass materials as fillers for reactors in water purification. This review provides some practical references for the preparation and application of immobilized biomass materials and promotes further research and development to expand the application range of this material for water purification.

Bioanalytical Methods and Strategic Perspectives Addressing the Rising Complexity of Novel Bioconjugates and Delivery Routes for Biotherapeutics.

In recent years, an increase in the discovery and development of biotherapeutics employing new modalities, such as bioconjugates or novel routes of delivery, has created bioanalytical challenges. The inherent complexity of conjugated molecular structures means that quantification of the bioconjugate and its multiple components is critical for preclinical/clinical studies to inform drug discovery and development. Moreover, bioconjugates involve additional multifactorial complexity because of the potential for in vivo catabolism and biotransformation, which may require thorough investigations in multiple biological matrices. Furthermore, excipients that enhance absorption are frequently evaluated and employed for the development of oral and inhaled biotherapeutics. Risk-benefit assessments are required for novel or existing excipients that utilize dosages above previously approved levels. Bioanalytical methods that can measure both excipients and potential drug metabolites in biological matrices are highly relevant to these emerging bioanalysis challenges. We discuss the bioanalytical strategies for analyzing bioconjugates such as antibody-drug conjugates and antibody-oligonucleotide conjugates and review recent advances in bioanalytical methods for the quantification and characterization of novel bioconjugates. We also discuss bioanalytical considerations for both biotherapeutics and excipients through novel administration routes and review analyses in various biological matrices, from the extensively studied serum or plasma to tissue biopsy in the context of preclinical and clinical studies from both technical and regulatory perspectives.

High-sensitivity quantification of antisense oligonucleotides for pharmacokinetic characterization.

Aim: Antisense oligonucleotides (ASOs) are a fast-growing drug modality. Pharmacokinetic characterization and accurate quantification of ASOs is critical for drug development. LC-MS and hybridization immunoassays are common methods to quantify ASOs but may lack sensitivity. In this study we aimed to develop an ASO quantification method with improved sensitivity. Methods: We developed a branched DNA approach for ASO quantification and compared it with hybridization immunoassays. Results: The branched DNA assay showed significantly improved sensitivity, with LLOQ 31.25 pg/ml in plasma, 6.4-and 16-fold higher than dual-probe hybridization electrochemiluminescence and single-probe hybridization ELISA, respectively, with adequate precision, accuracy, selectivity and specificity and acceptable matrix interference. Conclusion: Branched DNA for ASO quantification has significantly higher sensitivity and lower hemolysis interference.

Disease can be caused by genetic mutations that lead to overproduction or underproduction of an aberrant protein. Antisense oligonucleotides (ASOs) are a relatively new class of drugs. While most current drugs act at the protein level, ASOs work at the RNA level and minimize synthesis of the aberrant protein. ASOs are small synthetic nucleotides that specifically bind and modify the target RNA. Quantification of ASOs is important in drug development to understand how much of the drug is in circulation or in the body after a certain period of time. While there are methods available to quantify ASOs, they lack sensitivity. We developed a method called 'branched DNA' to quantify ASOs, and compared it with known ASO quantification methods. We found that the branched DNA method showed improved sensitivity compared with other existing methods and is a reliable method to quantify ASOs. This method may be used in clinical trials when improved sensitivity quantification is needed and thus facilitate the ASO drug development field.