Engineered human FcγRIIa fusion: A novel strategy to extend serum half-life of therapeutic proteins.

The immunoglobulin G (IgG) molecule has a long circulating serum half-life (~3 weeks) through pH- dependent FcRn binding-mediated recycling. To hijack the intracellular trafficking and recycling mechanism of IgG as a way to extend serum persistence of non-antibody therapeutic proteins, we have evolved the ectodomain of a low-affinity human FcγRIIa for enhanced binding to the lower hinge and upper CH2 region of IgG, which is very far from the FcRn binding site (CH2-CH3 interface). High-throughput library screening enabled isolation of an FcγRIIa variant (2A45.1) with 32-fold increased binding affinity to human IgG1 Fc (equilibrium dissociation constant: 9.04 × 10-7 M for wild type FcγRIIa and 2.82 × 10-8 M for 2A45.1) and significantly improved affinity to mouse serum IgG compared to wild type human FcγRIIa. The in vivo pharmacokinetic profile of PD-L1 fused with engineered FcγRIIa (PD-L1-2A45.1) was compared with that of PD-L1 fused with wild type FcγRIIa (PD-L1-wild type FcγRIIa) and human PD-L1 in mice. PD-L1-2A45.1 showed 11.7- and 9.7-fold prolonged circulating half-life (t1/2 ) compared to PD-L1 when administered intravenously and intraperitoneally, respectively. In addition, the AUCinf of PD-L1-2A45.1 was two-fold higher compared to that of PD-L1-wild type FcγRIIa. These results demonstrate that engineered FcγRIIa fusion offers a novel and successful strategy for prolonging serum half-life of therapeutic proteins.

Amelioration of colitis in mice by Leuconostoc lactis EJ-1 by M1 to M2 macrophage polarization.

Dysregulation of immune responses to environmental antigens by the intestine leads to the chronic inflammatory disease, inflammatory bowel disease (IBD). Recent studies have thus sought to identify a dietary component that can inhibit lipopolysaccharide (LPS)-induced nuclear factor-kappa beta (NF-κB) signaling to ameliorate IBD. This study assessed if the lactic acid bacteria (LAB) from kimchi, suppresses the expression of tumor necrosis factor-alpha (TNF-α) in peritoneal macrophages induced by LPS. Leuconostoc lactis EJ-1, an isolate from LAB, reduced the expression of interleukin-6 (IL-6) and IL-1ß in peritoneal macrophages induced by LPS. The study further tested whether EJ-1 alleviates colitis induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS) in mice. TNBS significantly increased myeloperoxidase (MPO) expression, macroscopic colitis scores, and colon shortening. Oral administration of L. lactis EJ-1 resulted in an inhibited in TNBS-induced loss in body weight, colon shortening, MPO activity, and NF-κB and inducible nitric oxide synthase expression; it also led to a marked reduction in cyclooxygenase-2 expression. L. lactis EJ-1 also inhibited the TNBS-induced expression of TNF-α, IL-1ß, and IL-6; however, it induced the expression of IL-10. The M2 macrophage markers arginase I, IL-10, and CD206 were elevated by EJ-1. Collectively, these results suggest that EJ-1 inhibits the NF-κB signaling and polarizes M1- to M2-macrophage transition, which help in ameliorating colitis.

Effect of bevel direction on the success rate of ultrasound-guided radial arterial catheterization.

BACKGROUND: This study assessed the effect of bevel direction on the success rate of ultrasound guided radial artery catheterization. METHODS: A total of 204 patients requiring radial artery catheterization were randomly divided into bevel-up (n = 102) and bevel-down (n = 102) groups. Success rate, cannulation time, and number of attempts were compared groups. RESULTS: In the bevel-down group, an arterial line was placed on the first attempt in 86 of 102 (84.3 %; 95 % confidence interval [CI] = 76 % to 90 %) patients versus 73 of 102 (71.6 %; 95 % CI = 62.1 % to 79.4 %) in the bevel-up group (p = 0.028). In the bevel-down group, the mean time to a successful radial arterial cannulation was 33.3 ± 6.3 seconds (95 % CI = 32.1-34.6) versus 35.9 ± 7.6 seconds (95 % CI = 34.4-37.2) in the bevel-up group (p = 0.011). The median score was 33.2 and interquartile range [IQR] was 10.9 (30.3-41.2) for the mean cannulation time in the bevel-up group. In the bevel-down group, the mean score was 32.3 (IQR 3.90, 30-33.9) for mean cannulation time. In the bevel-down group, 11 of 102 (7 %; 95 % CI = 0 to 16 %) patients developed a posterior wall puncture versus 22 of 102 ((21.6 %; 95 % CI = 14.7 to 17.2 %) in the bevel-up group. CONCLUSION: The bevel-down approach during ultrasound-guided radial artery catheterization exhibited a higher success with fewer complications compared to the bevel-up approach. TRIAL REGISTRATION: Clinical Research Information Service is Korean Clinical Trials Registry ( KCT0001836 ). It was registered retrospectively 30th Nov 2015.

Effective clearance of rituximab-resistant tumor cells by breaking the mirror-symmetry of immunoglobulin G and simultaneous binding to CD55 and CD20.

Complement-dependent cytotoxicity (CDC), which eliminates aberrant target cells through the assembly and complex formation of serum complement molecules, is one of the major effector functions of anticancer therapeutic antibodies. In this study, we discovered that breaking the symmetry of natural immunoglobulin G (IgG) antibodies significantly increased the CDC activity of anti-CD20 antibodies. In addition, the expression of CD55 (a checkpoint inhibitor in the CDC cascade) was significantly increased in a rituximab-resistant cell line generated in-house, suggesting that CD55 overexpression might be a mechanism by which cancer cells acquire rituximab resistance. Based on these findings, we developed an asymmetric bispecific antibody (SBU-CD55 × CD20) that simultaneously targets both CD55 and CD20 to effectively eliminate rituximab-resistant cancer cells. In various cancer cell lines, including rituximab-resistant lymphoma cells, the SBU-CD55 × CD20 antibody showed significantly higher CDC activity than either anti-CD20 IgG antibody alone or a combination of anti-CD20 IgG antibody and anti-CD55 IgG antibody. Furthermore, the asymmetric bispecific antibody (SBU-CD55 × CD20) exhibited significantly higher CDC activity against rituximab-resistant cancer cells compared to other bispecific antibodies with symmetric features. These results demonstrate that enhancing CDC with an asymmetric CD55-binding bispecific antibody could be a new strategy for developing therapeutics to treat patients with relapsed or refractory cancers.

Dealing with new members: Team members' reactions to newcomer's attractiveness and sex.

We examine how team members respond to the inclusion of new members' physical attractiveness and sex. Drawing on Social Exchange Theory, we argue and show that incumbent team members engage in three behaviors (mimicry, ingratiation, and challenging) in response to the inclusion of more or less attractive male or female members in their team. Using a multilevel experimental design, we show that existing team members mimic newcomers who are higher on physical attractiveness and that the effect is more pronounced when there is a sex match (i.e., existing males mimic new males more). Furthermore, they ingratiate toward the physically attractive newcomers who are also committed to the task. In addition, we find that existing team members challenge physically attractive females who are committed to the task. Our findings suggest that the basic combinations of primary cues of newcomers' characteristics affect intrateam behaviors and produce different outcomes across sexes for attractiveness. By shifting the attention to the effect that newcomers have on team behaviors, the study provides novel insights for scholars that help move the discussion of team membership changes beyond the traditional accounts of new member socialization and team effectiveness. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Development and Characterization of Phage-Display-Derived Novel Human Monoclonal Antibodies against the Receptor Binding Domain of SARS-CoV-2.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has resulted in an ongoing global pandemic crisis, caused by the life-threatening illness coronavirus disease 2019 (COVID-19). Thus, the rapid development of monoclonal antibodies (mAbs) to cope with COVID-19 is urgently necessary. In this study, we used phage display to develop four human mAbs specific to the receptor-binding domain (RBD) of SARS-CoV-2. Our intensive in vitro functional analyses demonstrated that K102.1, an anti-SARS-CoV-2 RBD-specific mAb, exerted potent neutralizing activity against pseudoviral and live viral infection and the interaction between SARS-CoV-2 RBD and human angiotensin-converting enzyme 2. Monotherapy with K102.1 also revealed the therapeutic potential against SARS-CoV-2 infection in vivo. Further, this study developed a sandwich enzyme-linked immunosorbent assay with a non-competing mAb pair, K102.1 and K102.2, that accurately detected the RBDs of SARS-CoV-2 wild-type and variants with high sensitivity in the picomolar range. These findings suggest that the phage-display-based mAb selection from an established antibody library may be an effective strategy for the rapid development of mAbs against the constantly evolving SARS-CoV-2.

Anti-inflammatory mechanism of ginsenoside Rh1 in lipopolysaccharide-stimulated microglia: critical role of the protein kinase A pathway and hemeoxygenase-1 expression.

Microglia activation plays a pivotal role in neurodegenerative diseases, and thus controlling microglial activation has been suggested as a promising therapeutic strategy for neurodegenerative diseases. In the present study, we showed that ginsenoside Rh1 inhibited inducible nitric oxide synthase, cyclooxygenase-2, and pro-inflammatory cytokine expression in lipopolysaccharide (LPS)-stimulated microglia, while Rh1 increased anti-inflammatory IL-10 and hemeoxygenase-1 (HO-1) expression. Suppression of microglial activation by Rh1 was also observed in the mouse brain following treatment with LPS. Subsequent mechanistic studies revealed that Rh1 inhibited LPS-induced MAPK phosphorylation and nuclear factor-κB (NF-κB)-mediated transcription without affecting NF-κB DNA binding. As the increase of pCREB (cAMP responsive element-binding protein) is known to result in suppression of NF-κB-mediated transcription, we examined whether Rh1 increased pCREB levels. As expected, Rh1 increased pCREB, which was shown to be related to the anti-inflammatory effect of Rh1 because pre-treatment with protein kinase A inhibitors attenuated the Rh1-mediated inhibition of nitric oxide production and the up-regulation of IL-10 and HO-1. Furthermore, treatment of HO-1 shRNA attenuated Rh1-mediated inhibition of nitric oxide and reactive oxygen species production. Through this study, we have demonstrated that protein kinase A and its downstream effector, HO-1, play a critical role in the anti-inflammatory mechanism of Rh1 by modulating pro- and anti-inflammatory molecules in activated microglia.

Lactobacillus sakei S1 Improves Colitis Induced by 2,4,6-Trinitrobenzene Sulfonic Acid by the Inhibition of NF-κB Signaling in Mice.

Lactobacillus sakei S1 strongly inhibits the expression of interleukin (IL)-6 and IL-1ß in lipopolysaccharide-induced peritoneal macrophages by a mechanism for which lactic acid bacteria from kimchi that inhibit tumor necrosis factor-alpha (TNF-κ) were isolated. Therefore, we further evaluated the protective effect of this strain on the colitis mouse model induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS). TNBS significantly elevated myeloperoxidase (MPO) expression, macroscopic scores, and colon shortening. Oral L. sakei S1 administration resulted in reduction of TNBS-induced loss in body weight, colon shortening, MPO activity, expression of cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS) and nuclear factor-kappa B (NF-κB). L. sakei S1 inhibited the expression of inflammatory cytokines IL-1ß, IL-6 and TNF-κ, induced by TNBS, but enhanced IL-10 expression. L. sakei S1 showed resistance to artificial digestive juices and adherence to intestinal epithelial Caco-2 cells. Thus, L. sakei S1 may inhibit the NF-κB pathway and be used in functional food to treat colitis.

Inhibition of matrix metalloproteinase-9 gene expression by an isoflavone metabolite, irisolidone in U87MG human astroglioma cells.

Matrix metalloproteinase-9 (MMP-9) plays an important role in mediating the invasion and angiogenic process of malignant gliomas. This study was undertaken to determine if an isoflavone metabolite, irisolidone, inhibits MMP-9 expression in human astroglioma cells. Irisolidone was found to inhibit the secretion and protein expression of MMP-9 induced by PMA in U87 MG glioma cells, accompanied by the inhibition of MMP-9 mRNA expression and promoter activity. Further mechanistic studies revealed that irisolidone inhibits the binding of NF-kappaB and AP-1 to the MMP-9 promoter and suppresses the PMA-induced phosphorylation of ERK and JNK, which are upstream signaling molecules in MMP-9 expression. The Matrigel-invasion assay showed that irisolidone suppresses the in vitro invasiveness of glioma cells. Therefore, the strong inhibition of MMP-9 expression by irisolidone might be a potential therapeutic modality for controlling the growth and invasiveness of gliomas.

Development of a radionuclide-labeled monoclonal anti-CD55 antibody with theranostic potential in pleural metastatic lung cancer.

Decay-accelerating factor (CD55 or DAF) inhibits complement-dependent cytotoxicity. We determined that CD55 is overexpressed in 76.47% of human non-small cell lung cancer tissue specimens. We therefore developed a lutetium-177-labeled chimeric monoclonal antibody against CD55. CD55-specific single-chain variable fragment (scFv) was selected from a naïve chicken scFv phage-display library, converted to IgG, and radiolabeled with lutetium-177 to generate a 177Lu-anti-CD55 antibody. We then charaterized the biodistribution of this antibody in a mouse model of pleural metastatic lung cancer. The 177Lu-anti-CD55 antibody was primarily retained in tumor tissue rather than normal tissue. Treatment of the mice with 177Lu-anti-CD55 reduced the growth of lung tumors and improved median survival in vivo by two-fold compared to controls. Finally, 177Lu-anti-CD55 also enhanced the antitumor activity of cisplatin both in vitro and in vivo. These data suggest 177Lu-anti-CD55 antibody is a promising theranostic agent for pleural metastatic lung cancer.