ABSTRACT
In spite of the successful development of effective countermeasures against Covid-19, variants have and will continue to emerge that could compromise the efficacy of currently approved neutralizing antibodies and vaccines. Consequently, novel and more efficacious agents are urgently needed. We have developed a bispecific antibody, 2022, consisting of two antibodies, 2F8 and VHH18. 2F8 was isolated from our proprietary fully synthetic human IDEAL (Intelligently Designed and Engineered Antibody Library)-VH/VL library and VHH18 is a single domain antibody isolated from IDEAL-nanobody library. 2022 was constructed by attaching VHH18 to the C-terminal of Fc of 2F8. 2022 binds two non-overlapping epitopes simultaneously on the RBD of the SARS-CoV-2 spike protein and blocks the binding of RBD to human angiotensin-converting enzyme 2 (ACE2). 2022 potently neutralizes SARS-CoV-2 and all of the variants tested in both pseudovirus and live virus assays, including variants carrying mutations known to resist neutralizing antibodies approved under EUA and that reduce the protection efficiency of current effective vaccines. The half-maximum inhibitory concentration (IC50) of 2022 is 270 pM, 30 pM, 20 pM, and 1 pM, for wild-type, alpha, beta, and delta pseudovirus, respectively. In the live virus assay, 2022 has an IC50 of 26.4 pM, 13.3 pM, and 88.6 pM, for wild-type, beta, and delta live virus, respectively. In a mouse model of SARS-CoV-2, 2022 showed strong prophylactic and therapeutic effects. A single administration of 2022 intranasal (i.n.) or intraperitoneal (i.p.) 24 hours before virus challenge completely protected all mice from bodyweight loss, as compared with up to 20% loss of bodyweight in placebo treated mice. In addition, the lung viral titers were undetectable (FRNT assay) in all mice treated with 2022 either prophylactically or therapeutically, as compared with around 1x105 pfu/g lung tissue in placebo treated mice. In summary, bispecific antibody 2022 showed potent binding and neutralizing activity across a variety of SARS-CoV-2 variants and could be an attractive weapon to combat the ongoing waves of the COVID-19 pandemic propagated mainly by variants, especially, the much more contagious delta variant.
ABSTRACT
Objective To simulate the chemical microenvironment after traumatic brain injury (TBI) and to investigate the effect of this microenvironment on the survival and differentiation of neural stem cells (NSCs).Methods The brain tissue homogenate of TBI rat model was harvested to simulate the chemical microenvironment after TBI.The primary NSCs of rat model were isolated and extraction,and then identified the phenotype characteristics with immunofluorescence staining.The experiments were divided into control group,normal brain tissue extract group (BTE group) and traumatic brain injury tissue extract group (TBITE group).The cell growth and morphological changes of each group were observed dynamically.The expression of apoptosis related protein,which includes Bax,Bcl-2,caspase-3 and cleaved caspase-3,were detected by Western Blot 24 h after experiments.The proliferation of NSCs was detected by MTT assay and Western Blot after 3 days.The differentiation level of NSCs to neurons was detected by immunofluorescence staining after 7 days.Results The results of Western Blot showed that compared with the control group,there was no significant change of apoptosis in the BTE group,while the apoptosis in the BTE group was significantly increased,showed a increase of expression levels of Bax (F=18.06,P＜0.01) and Cleaved caspase-3 (F=23.86,P＜0.01),and a decrease of that of Bcl-2 (F=22.95,P＜0.01).The results of MTT assay showed that compared with the BTE group,the proliferation of NSCs in the TBITE group was decreased (F=41.99,P＜0.01).The immunofluorescence staining showed that compared with the control group,the neuronal differentiation rate was increased in the BTE group.Further,compared with the BTE group,the neuronal differentiation rate in the TBITE group was decreased (F=66.93,P＜0.01).Conclusion The injury microenvironment after TBI can significantly inhibit the survival and differentiation of NSCs,which provides a theoretical basis for clarifying the mechanism of endogenous nerve regeneration after TBI.
