Thermostability and binding properties of single-chained Fv fragments derived from therapeutic antibodies.

Small antibody fragments have recently been used as alternatives to full-length monoclonal antibodies in therapeutic applications. One of the most popular fragment antibodies is single-chain fragment variables (scFvs), consisting of variable heavy (VH) and variable light (VL) domains linked by a flexible peptide linker. scFvs have small molecular sizes, which enables good tissue penetration and low immunogenicity. Despite these advantages, the use of scFvs, especially for therapeutic purpose, is still limited because of the difficulty to regulate the binding activity and conformational stability. In this study, we constructed and analyzed 10 scFv fragments derived from 10 representatives of FDA-approved mAbs to evaluate their physicochemical properties. Differential scanning calorimetry analysis showed that scFvs exhibited relatively high but varied thermostability, from 50 to 70°C of melting temperatures, and different unfolding cooperativity. Surface plasmon resonance analysis revealed that scFvs fragments that exhibit high stability and cooperative unfolding likely tend to maintain antigen binding. This study demonstrated the comprehensive physicochemical properties of scFvs derived from FDA-approved antibodies, providing insights into antibody design and development.

Extracellular Water-to-total Body Water Ratio as an Objective Biomarker for Frailty in Lung Cancer Patients.

BACKGROUND/AIM: Our previous study revealed the association between extracellular water-to-total body water ratio (ECW/TBW) and the therapeutic durability of chemotherapy and/or immune checkpoint inhibitors in advanced lung cancer. We retrospectively examined the usefulness of ECW/TBW in detecting frailty compared to other bioelectrical impedance (BIA) parameters in a larger number of patients. PATIENTS AND METHODS: Lung cancer patients underwent BIA before anti-cancer therapy at our hospital between June 1, 2018 and July 31, 2020. RESULTS: Of 99 patients, 26 were assigned to ECW/TBW≥0.4 (higher group: HG) and 57 to ECW/TBW<0.4 (lower group: LG). ECW/TBW increased significantly with performance deterioration and ageing. HG patients had significantly shorter time-to-treatment failure (TTF) than LG patients. In patients with performance status 0-1, those in the HG had shorter TTF than those in the LG. ECW/TBW was the only independent predictor of TTF according to multivariate analysis. CONCLUSION: ECW/TBW is an objective biomarker for detecting frailty among lung cancer patients.

Kinetic Analysis of the Digestion of Bovine Type I Collagen Telopeptides with Porcine Pepsin.

Collagen is frequently digested using pepsin in industries to produce a triple helical collagen without the N- and C-terminal telopeptides. However, kinetic analysis of this reaction is difficult because several Lys residues in the N- and in the C-terminal telopeptides form covalent bonds, leading to multiple substrates species, and pepsin cleaves collagen at various sites in the N-terminal and in the C-terminal telopeptides, yielding different products. Here we performed kinetic analysis of the digestion of bovine type I collagen with porcine pepsin. The reaction could be monitored by SDS-PAGE by measuring the intensity of the protein bands corresponding to the variant ß11 chain. We obtained kinetic parameters relative to the decrease in the variant ß11 chain upon digestion. At pH 4.0, the Km and kcat values increased with increasing temperature (30 to 65 °C), although the kcat /Km values were stable. Additional cleavage at the helical region was detected at 45 to 65 °C. At 37 °C, the Km and kcat values increased with decreasing pH, and the kcat /Km values at pH 2.1 to 4.5 were stable and higher than those at pH 5.0 and 5.5. No additional cleavage was detected at the examined pH. Thus, the optimal pH and temperatures for selective digestion of collagen telopeptides with pepsin are 2.1 to 4.5 and 30 to 40 °C, respectively. These results suggest that the method might be useful for the kinetic analysis of the digestion of collagen telopeptides with pepsin.

A high-temperature in situ cell with a large solid angle for fluorescence X-ray absorption fine structure measurement.

We present the design and performance of a high-temperature in situ cell with a large solid angle for fluorescence X-ray absorption fine structure (XAFS) spectra. The cell has a large fluorescence XAFS window (116 mm(ϕ)) near the sample in the cell, realizing a large half-cone angle of 56°. We use a small heater (25 × 35 mm(2)) to heat the sample locally to 873 K. We measured a Pt-SnO2 thin layer on a Si substrate at reaction conditions having a high activity. In situ measurement enables the analysis of the difference XAFS spectra between before and during the reaction to reveal the structure change during the operation.

Culicoides Latreille (Diptera: Ceratopogonidae) da vila de Alter do Chão, Santarém, Pará, Brasil / Culicoides Latreille (Diptera: Ceratopogonidae) from Alter do Chão village, Santarém, Pará State, Brazil

O objetivo deste estudo é levantar a fauna de Culicoides Latreille e registrar sua distribuição sazonal em uma zona de praia da vila de Alter do Chão (Município de Santarém, Estado do Pará). As coletas foram feitas mensalmente, entre maio de 2006 e abril de 2007, utilizando atração humana e armadilha luminosa modelo CDC. Foram coletados 58 exemplares de maruins pertencentes a cinco espécies: C. crucifer Clastrier, C. insinuatus Ortiz & Leon, C. fusipalpis Wirth & Blanton, capturados com atração humana; C. insignis Lutz com armadilha CDC, e C. ruizi Forattini, coletados com as duas técnicas. C. crucifer foi a espécie mais abundante picando humanos, enquanto que C. insignis foi predominantemente coletada em armadilha CDC. A maior incidência de Culicoides ocorreu em outubro de 2006. Durante o levantamento, não foi evidenciada correlação entre os fatores climáticos e os meses de coleta por meio do coeficiente de Spearman.

The aim of this work was to survey Culicoides Latreille fauna and to record their seasonal abundance in a beach area from the village of Alter do Chão (Municipality of Santarém, Pará State, Brazil). Collections were made monthly between May 2006 and April 2007 using CDC light trap and human attraction. A total of 58 specimens were collected belonging to five species: C. crucifer Clastrier, C. insinuatus Ortiz & Leon, and C. fusipalpis Wirth & Blanton species were collected while biting humans; C. insignis Lutz was collected by CDC light trap. C. ruizi Forattini midges were collected utilizing both methods. The most abundant human-biting species collected was C. crucifer, while C. insignis was the predominant species collected using CDC light traps. The greatest incidence of Culicoides occurred in October 2006. During the survey, no correlation between climatic factors and the number of Culicoides collected each month was observed using the Spearman coefficient.