ABSTRACT
The detection of severe acute respiratory syndrome coronavirus (SARS-CoV-1) was demonstrated using screened Fv-antibodies for SPR biosensor and impedance spectrometry. The Fv-antibody library was first prepared on the outer membrane of E. coli using autodisplay technology and the Fv-variants (clones) with a specific affinity toward the SARS-CoV-1 spike protein (SP) were screened using magnetic beads immobilized with the SP. Upon screening the Fv-antibody library, two target Fv-variants (clones) with a specific binding affinity toward the SARS-CoV-1 SP were determined and the Fv-antibodies on two clones were named "Anti-SP1" (with CDR3 amino acid sequence: 1GRTTG5NDRPD11Y) and "Anti-SP2" (with CDR3 amino acid sequence: 1CLRQA5GTADD11V). The binding affinities of the two screened Fv-variants (clones) were analyzed using flow cytometry and the binding constants (KD) were estimated to be 80.5 ± 3.6 nM for Anti-SP1 and 45.6 ± 8.9 nM for Anti-SP2 (n = 3). In addition, the Fv-antibody including three CDR regions (CDR1, CDR2, and CDR3) and frame regions (FRs) between the CDR regions was expressed as a fusion protein (Mw. 40.6 kDa) with a green fluorescent protein (GFP) and the KD values of the expressed Fv-antibodies toward the SP estimated to be 15.3 ± 1.5 nM for Anti-SP1 (n = 3) and 16.3 ± 1.7 nM for Anti-SP2 (n = 3). Finally, the expressed Fv-antibodies screened against SARS-CoV-1 SP (Anti-SP1 and Anti-SP2) were applied for the detection of SARS-CoV-1. Consequently, the detection of SARS-CoV-1 was demonstrated to be feasible using the SPR biosensor and impedance spectrometry utilizing the immobilized Fv-antibodies against the SARS-CoV-1 SP.
ABSTRACT
The BTK inhibitors ibrutinib and acalabrutinib are FDA-approved drugs for the treatment of B cell malignances. Both drugs have demonstrated clinical efficacy and safety profiles superior to chemoimmunotherapy regimens in patients with chronic lymphocytic leukemia. Mounting preclinical and clinical evidence indicates that both ibrutinib and acalabrutinib are versatile and have direct effects on many immune cell subsets as well as other cell types beyond B cells. The versatility and immunomodulatory effects of both drugs have been exploited to expand their therapeutic potential in a wide variety of human diseases. Over 470 clinical trials are currently registered at ClinicalTrials.gov to test the efficacy of ibrutinib or acalabrutinib not only in almost every type of B cell malignancies, but also in hematological malignancies of myeloid cells and T cells, solid tumors, chronic graft versus host disease (cGHVD), autoimmune diseases, allergy and COVID-19 (http:www.clinicaltrials.gov). In this review, we present brief discussions of the clinical trials and relevant key preclinical evidence of ibrutinib and acalabrutinib as monotherapies or as part of combination therapies for the treatment of human diseases beyond B cell malignancies. Adding to the proven efficacy of ibrutinib for cGVHD, preliminary results of clinical trials have shown promising efficacy of ibrutinib or acalabrutinib for certain T cell malignancies, allergies and severe COVID-19. However, both BTK inhibitors have no or limited efficacy for refractory or recurrent solid tumors. These clinical data together with additional pending results from ongoing trials will provide valuable information to guide the design and improvement of future trials, including optimization of combination regimens and dosing sequences as well as better patient stratification and more efficient delivery strategies. Such information will further advance the precise implementation of BTK inhibitors into the clinical toolbox for the treatment of different human diseases.
ABSTRACT
The clinical success of the two BTK inhibitors, ibrutinib and acalabrutinib, represents a major breakthrough in the treatment of chronic lymphocytic leukemia (CLL) and has also revolutionized the treatment options for other B cell malignancies. Increasing evidence indicates that in addition to their direct effects on B lymphocytes, both BTK inhibitors also directly impact the homeostasis, phenotype and function of many other cell subsets of the immune system, which contribute to their high efficacy as well as adverse effects observed in CLL patients. In this review, we attempt to provide an overview on the overlapping and differential effects of ibrutinib and acalabrutinib on specific receptor signaling pathways in different immune cell subsets other than B cells, including T cells, NK cells, monocytes, macrophages, granulocytes, myeloid-derived suppressor cells, dendritic cells, osteoclasts, mast cells and platelets. The shared and distinct effects of ibrutinib versus acalabrutinib are mediated through BTK-dependent and BTK-independent mechanisms, respectively. Such immunomodulatory effects of the two drugs have fueled myriad explorations of their repurposing opportunities for the treatment of a wide variety of other human diseases involving immune dysregulation.
ABSTRACT
Several endemic corona viruses (eCoVs) have been reported to be the most common etiologic agents for the seasonal common cold and also cause pneumonia. These eCoVs share extensive sequence homology with SARS-CoV-2, and immune responses to eCoVs can cross-react with SARS-CoV-2 antigens. Based on such cross-reactivity of antigens among eCoVs, the IgG antibodies against the spike protein (SP) of severe acute respiratory syndrome coronavirus (SARS-CoV) were isolated from pig serum using magnetic beads immobilized with SARS-CoV SP and a protein-A column. The selectivity of the isolated antibodies was tested using different types of antigens, such as SARS-CoV-2 nucleoprotein (NP), influenza A virus (Beijing type), influenza B virus (Tokio and Florida types), human hepatitis B virus surface antigen (HBsAg), and bovine serum albumin (BSA). From the selectivity test, the anti-SP antibodies isolated from pig serum had sufficient selectivity to other kinds of viral antigens, and the apparent binding constant of the isolated antibodies was approximately 1.5 × 10-8 M from the surface plasmon resonance (SPR) measurements. Finally, the isolated anti-SP antibodies were applied to the immunoassay of SP using competitive immunoassay configuration. The feasibility of the detection as well as the quantitative analysis of the SARS-CoV viral culture fluid was determined using four viral culture samples, namely, SARS-CoV, SARS-CoV-2, MERS-CoV, and CoV-229E.
ABSTRACT
Anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) nucleoprotein (NP) antibodies were isolated from pig sera using human SARS-CoV-2 NP-immobilized magnetic beads. The binding properties of the isolated antibodies against SARS-CoV-2 NP were tested via flow cytometry using SARS-CoV-2 NP-immobilized magnetic beads. A competitive immunoassay was developed for detecting SARS-CoV-2 NP as well as SARS-CoV-2 in the culture fluid using magnetic beads with immobilized anti-SARS-CoV-2 NP antibodies. Selectivity tests were carried out during the competitive immunoassay for SARS-CoV, MERS-CoV, and CoV strain 229E in the culture fluid.
ABSTRACT
Anti-coronavirusdisease-2019 (COVID-19; anti-severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2)) antibodies against nucleoprotein (NP) were purified from pig sera. Following the separation of the antibody fraction using a protein-A column, the final yield of the purified antibodies against SARS-CoV-2 NPs was estimated to be 0.26 ± 0.05 % (absolute amount of 143.4 ± 25.2 ng, n=5) from 1 mL of pig sera. The binding activities of the isolated antibodies were confirmed using immunoassay and immunostaining. Based on the specific binding activity to NPs, a quantitative assay was performed using a surface plasmon resonance (SPR) biosensor. From the doseresponse curve, the binding constant (Kd) was calculated to be 185 pM and the limit of detection was estimated to be 1.02 pM. The SPR biosensor with the isolated antibodies against SARS-CoV-2 NPs was applied for the detection of SARS-CoV-2, MERS-CoV, and CoV strain 229E in culture fluid.