Comparative stability study and aggregate analysis of Bevacizumab marketed formulations using advanced analytical techniques.

Bevacizumab (Bvz) is the most preferred recombinant humanized monoclonal antibody in biosimilar development due to its prominence as a standard treatment in the oncology space. Therapeutic monoclonal antibodies are typically more complex and unlikely to produce a replica. As a result, regulatory agencies allow approval of biosimilars that differ structurally and functionally from their reference product, but these differences should not have any clinical significance. To identify these significant discrepancies, it is essential to perform a thorough characterization of critical product attributes both in real-time and after storage until the product's expiration. In the present study, two Bvz biosimilar brands (Bio-1 and Bio-2) marketed in India were evaluated and compared with the reference product Avastin® to assess their degree of similarity. A comprehensive physicochemical characterization of biosimilars and reference product was performed using orthogonal techniques including LC-ESI-QTOF, MALDI-TOF, FTIR-ATR, iCIEF, rCE, nrCE, UV280, and RP-HPLC. Furthermore, Bvz formulations under study were subjected to various stress conditions of thermal (elevated temperature 50 ± 2 °C), chemical (acidic pH 3.0 ± 0.2, neutral pH 7.0 ± 0.2, and basic pH 10.0 ± 0.2), and mechanical (agitation 200 rpm) for comparative stability evaluation. Any alteration in the secondary structure of the native protein was detected and quantified using far-UV circular dichroism (CD), indicating an average of 15% and 11% loss in native antiparallel ß-sheet conformation respectively in Bio-1 and Bio-2 upon exposure to elevated temperature and high pH. Additionally, covalent or non-covalent aggregates formed as a function of elevated temperature and agitation were quantified using SEC-MALS.

Recent updates in the clinical trials of therapeutic monoclonal antibodies targeting cytokine storm for the management of COVID-19.

Clinical studies have identified a cytokine storm in the third stage of disease progression in critical ill patients with coronavirus disease 2019 (COVID-19). Hence, effectively suppressing the uncontrolled immune response of the host towards the invaded viruses in a cytokine storm is a critical step to prevent the deterioration of patient conditions and decrease the rate of mortality. Therapeutic monoclonal antibodies (mAbs) are found to be effective for the management of acute respiratory distress syndrome in patients with COVID-19. In this review, we compiled all therapeutic mAbs targeting cytokine storm, which are in clinical trials for its repurposing in the management of COVID-19. Compilation of clinical trial data indicated that therapeutic monoclonal antibodies targeting interleukins (IL-6, IL-1ra, IL-8, IL-1ß, IL-17A, IL-33), interferon-gamma, tumor necrosis factor-alpha, P-selectin, connective tissue growth factor, plasma kallikrein, tumor necrosis factor superfamily 14, granulocyte macrophage colony stimulating factor, colony stimulating factor 1 receptor, C-C chemokine receptor type 5, cluster of differentiation 14 and 147, vascular endothelial growth factor, programmed cell death protein-1, Angiopoietin - 2, human factor XIIa, complementary protein 5, natural killer cell receptor G2A, human epidermal growth factor receptor 2, immunoglobulin-like transcript 7 receptor, complement component fragment 5a receptor and viral attachment to the human cell were under investigation for management of severely ill patients with COVID-19. Among these, about 65 clinical trials are targeting IL-6 inhibition as the most promising one and Tocilizumab, an IL-6 inhibitor is considered to be the potential candidate to treat cytokine storm associated with the COVID-19.

Thiopurine-induced mitotic catastrophe in Rad51d-deficient mammalian cells.

Thiopurines are part of a clinical regimen used for the treatment of autoimmune disorders and childhood acute lymphoblastic leukemia. However, despite these successes, there are also unintended consequences such as therapy-induced cancer in long-term survivors. Therefore, a better understanding of cellular responses to thiopurines will lead to improved and personalized treatment strategies. RAD51D is an important component of homologous recombination (HR), and our previous work established that mammalian cells defective for RAD51D are more sensitive to the thiopurine 6-thioguanine (6TG) and have dramatically increased numbers of multinucleated cells and chromosome instability. 6TG is capable of being incorporated into telomeres, and interestingly, RAD51D contributes to telomere maintenance, although the precise function of RAD51D at the telomeres remains unclear. We sought here to investigate: (1) the activity of RAD51D at telomeres, (2) the contribution of RAD51D to protect against 6TG-induced telomere damage, and (3) the fates of Rad51d-deficient cells following 6TG treatment. These results demonstrate that RAD51D is required for maintaining the telomeric 3' overhangs. As measured by γ-H2AX induction and foci formation, 6TG induced DNA damage in Rad51d-proficient and Rad51d-deficient cells. However, the extent of γ-H2AX telomere localization following 6TG treatment was higher in Rad51d-deficient cells than in Rad51d-proficient cells. Using live-cell imaging of 6TG-treated Rad51d-deficient cells, two predominant forms of mitotic catastrophe were found to contribute to the formation of multinucleated cells, failed division and restitution. Collectively, these findings provide a unique window into the role of the RAD51D HR protein during thiopurine induction of mitotic catastrophe. Environ. Mol. Mutagen. 59:38-48, 2018. © 2017 Wiley Periodicals, Inc.

HJP272, a novel endothelin receptor antagonist, attenuates lipopolysaccharide-induced acute lung injury in hamsters.

INTRODUCTION: Previous studies from this laboratory indicate that endothelin-1 (ET-1), a potent vasoconstrictor, may play an important role in lipopolysaccharide (LPS)-induced release of neutrophils from the pulmonary microvasculature. To further test this concept, Syrian hamsters were treated with a novel endothelin receptor A (ETA) antagonist (HJP272) prior to intratracheal instillation of LPS. METHODS: The effect of HJP272 on the LPS-induced inflammatory reaction was determined by measuring: (1) lung histopathological changes, (2) total neutrophils in bronchoalveolar lavage fluid (BALF), (3) expression of tumor necrosis factor receptor 1 (TNFR1) by BALF macrophages, and (4) alveolar septal cell apoptosis. RESULTS: Treatment with HJP272 significantly reduced each of these parameters during a 24-hr period following LPS instillation, supporting the concept that limiting the activity of ET-1 may reduce the extent of lung injury. This hypothesis was further tested by giving ET-1 prior to LPS instillation, which resulted in a marked enhancement of LPS-induced lung inflammation, as measured by BALF neutrophils and TNFR1-positive macrophages. Furthermore, the increase in neutrophils resulting from treatment with ET-1 was significantly reduced by HJP272, again demonstrating the ability of ETA receptor antagonists to limit the influx of these cells into the lung. CONCLUSIONS: These findings suggest a potential therapeutic role for these agents in diseases where neutrophils are a significant cause of lung injury, such as bronchopneumonia, respiratory distress syndrome, and chronic obstructive pulmonary disease.