ace2 inhibitor mln 4760  (Merck & Co)

Journal: Journal of Biomedical Science

Article Title: The ACE2 decoy receptor can overcome immune escape by rapid mutating SARS-CoV-2 variants and reduce cytokine induction and clot formation

doi: 10.1186/s12929-025-01156-4

Figure Lengend Snippet: Varied properties of Spike protein variants. A The expression of different Spike variants in 293 T-ACE2 Cells. Two days after transfection, the presence of various Spike protein variants in cell lysates was confirmed by immunoblotting. β-actin was used as a loading control. B Cell Fusion in 293 T-ACE2 Cells. Images showing how different Spike protein variants induced cell fusion and syncytia formation. Scale bars represent 150 µm. C Quantitative syncytia analysis. The measured areas of syncytia formation to quantify the extent of cell fusion. D Levels of different Spike protein variants expressed in another cell line, H1650-ACE2. E Syncytia formation in H1650-ACE2 Cells. Similar to panel B, but demonstrating syncytia formation in H1650-ACE2 cells. Scale bars represent 150 µm. F Cytotoxicity post-Spike transfection. The cytotoxic effects observed 48 h after transfecting different Spike variants into H1650-ACE2 cells, with Triton-X100 treated cells serving as the 100% cytotoxicity control. G , H Impact on PBMCs. After treating PBMCs for 24 h with supernatants from H1650-ACE2 cells transfected with various Spike variants for 48 h, the levels of inflammation markers IL-6 and TNF-α were measured. Data are presented as mean ± SD from three replicates. Statistical analysis was performed by One-Way ANOVA with multiple comparisons *P < 0.05

Article Snippet: For the ACE2-Fc treatment groups, trimeric Spike protein was pre-incubated for 15 min with ACE2-Fc alone, ACE2-Fc plus Spike antibody (Genetex), or ACE2-Fc plus the ACE2 catalytic inhibitor MLN-4760 (MedChemExpress), prior to mixing with plasma and initiating clot formation.

Techniques: Expressing, Transfection, Western Blot, Control

Journal: Journal of Biomedical Science

Article Title: The ACE2 decoy receptor can overcome immune escape by rapid mutating SARS-CoV-2 variants and reduce cytokine induction and clot formation

doi: 10.1186/s12929-025-01156-4

Figure Lengend Snippet: Blocking of pseudovirus entry by ACE2-Fc across different Spike variants. A Recognition by ACE2-Fc. Flow cytometry analysis showing the binding of ACE2-Fc, tagged with a fluorescent marker (FITC), to different Spike protein variants expressed on 293 T cells. Mouse IgG-FITC was used as an isotype control to validate the specificity of ACE2-Fc binding. B , C Inhibition of Pseudovirus Entry. These panels demonstrate the effectiveness of ACE2-Fc in blocking the entry of pseudoviruses into two types of cells: 293 T-ACE2 ( B ) and H1650-ACE2 ( C ). The results indicate that ACE2-Fc can prevent pseudovirus infection by interfering with the interaction between the Spike protein and the ACE2 receptor on the surface of target cells. Data are presented as mean ± SD from three replicates. Statistical analysis was performed by unpaired two-tail t-test. *P < 0.05

Article Snippet: For the ACE2-Fc treatment groups, trimeric Spike protein was pre-incubated for 15 min with ACE2-Fc alone, ACE2-Fc plus Spike antibody (Genetex), or ACE2-Fc plus the ACE2 catalytic inhibitor MLN-4760 (MedChemExpress), prior to mixing with plasma and initiating clot formation.

Techniques: Blocking Assay, Flow Cytometry, Binding Assay, Marker, Control, Inhibition, Infection

Journal: Journal of Biomedical Science

Article Title: The ACE2 decoy receptor can overcome immune escape by rapid mutating SARS-CoV-2 variants and reduce cytokine induction and clot formation

doi: 10.1186/s12929-025-01156-4

Figure Lengend Snippet: Inhibitory effects of ACE2-Fc on Spike-induced cell fusion and cytotoxicity. A Syncytia formation in 293 T-ACE2 Cells. ACE2-Fc inhibits the formation of syncytia induced by different Spike protein variants. The effectiveness of the inhibition is visually represented, with scale bars measuring 150 µm. B Quantitative analysis of GFP area. The quantitative results of the green fluorescent (GFP) area, which reflects the extent of syncytia formation. The calculations are based on a formula detailed in the Methods section of the study. C Syncytia reversal in H1650-ACE2 Cells. ACE2-Fc can reverse syncytia formation caused by different Spike variants in another cell type, H1650-ACE2. Scale bars represent 150 µm. D Reduction of cytotoxicity. ACE2-Fc reduces cytotoxicity observed 48 h after transfecting different Spike variants into H1650-ACE2 cells. E – F Reduction of cytokine induction. ACE2-Fc effectively reduces Delta and BQ.1 Spike meditated the induction of IL-6 and TNF-α in human PBMCs. Data are presented as mean ± SD from three replicates. Statistical analysis was performed by One-Way ANOVA with multiple comparisons ( B , D ) or unpaired two-tail t-test ( E , F ). *P < 0.05

Article Snippet: For the ACE2-Fc treatment groups, trimeric Spike protein was pre-incubated for 15 min with ACE2-Fc alone, ACE2-Fc plus Spike antibody (Genetex), or ACE2-Fc plus the ACE2 catalytic inhibitor MLN-4760 (MedChemExpress), prior to mixing with plasma and initiating clot formation.

Techniques: Inhibition

Journal: Journal of Biomedical Science

Article Title: The ACE2 decoy receptor can overcome immune escape by rapid mutating SARS-CoV-2 variants and reduce cytokine induction and clot formation

doi: 10.1186/s12929-025-01156-4

Figure Lengend Snippet: Inhibition of SARS-CoV-2 entry into host cells by ACE2-Fc. A Plaque assay inhibition. The ability of ACE2-Fc to inhibit infection by different SARS-CoV-2 variants using a plaque assay. The effectiveness of ACE2-Fc is compared to human IgG (hIgG), which serves as a baseline for inhibition. B Yield reduction assay. This assay was conducted to evaluate the inhibitory effects of ACE2-Fc on various coronavirus variants in H1650-ACE2 cells. The assay measures the reduction in the number of infectious virus particles as a result of ACE2-Fc treatment. C Plaque assay in Vero-E6 cells. This plaque assay quantifies the viral titer in the supernatant collected from the yield reduction assay. This method assesses the amount of virus that remains infectious after treatment with ACE2-Fc. Data are presented as mean ± SD from three replicates. Statistical analysis was conducted using an unpaired two-tail t-test. * P < 0.05, ** P < 0.01, *** P < 0.001. D Inhibition of nucleocapsid protein expression. The effect of ACE2-Fc on the expression of the Nucleocapsid protein across different variants. The results show a reduction in Nucleocapsid protein levels, indicating effective inhibition of virus replication by ACE2-Fc

Article Snippet: For the ACE2-Fc treatment groups, trimeric Spike protein was pre-incubated for 15 min with ACE2-Fc alone, ACE2-Fc plus Spike antibody (Genetex), or ACE2-Fc plus the ACE2 catalytic inhibitor MLN-4760 (MedChemExpress), prior to mixing with plasma and initiating clot formation.

Techniques: Inhibition, Plaque Assay, Infection, Virus, Expressing

Journal: Journal of Biomedical Science

Article Title: The ACE2 decoy receptor can overcome immune escape by rapid mutating SARS-CoV-2 variants and reduce cytokine induction and clot formation

doi: 10.1186/s12929-025-01156-4

Figure Lengend Snippet: Blocking of SARS-CoV-2 induced cytotoxicity, cytokine release and clot formation by ACE2-Fc. A The cytotoxicity in H1650-ACE2 cells. The cytotoxic effects observed in cells infected with different SARS-CoV-2 variants over 24 and 48 h. After the infection period, the supernatant was collected to measure cell damage, using Triton-X100 treated cells serving as the 100% cytotoxicity control. B Cytokine levels in PBMCs post-infection. After 48 h of infection in H1650-ACE2 cells, the supernatants were used to treat PBMCs, and the levels of IL-6 and TNF-α were measured. *Significant differences compared to the MOCK group. P < 0.05. C Inhibition of cytotoxicity by ACE2-Fc. Pre-treatment with ACE2-Fc significantly reduces the cytotoxic effects in H1650-ACE2 cells infected with various coronavirus variants 48 h post-infection. D Reduction of cytokine release by ACE2-Fc. ACE2-Fc treatment effectively decreases the release of IL-6 and TNF-α by PBMCs that were exposed to supernatants from infected H1650-ACE2 cells. Data are presented as mean ± SD from three replicates. Statistical analysis was performed by One-Way ANOVA with multiple comparisons *P < 0.05. E The turbidity of human plasma clot formation with D614G, Delta, and BA.5 spike. F – H The effect of ACE2-Fc treatment on plasma clot formation induced by the D614G ( F ), Delta ( G ), and BA.5 H Spike proteins. I Effect of ACE2-Fc co-treatment with Spike antibody on plasma clot formation. ( J ) Effect of ACE2-Fc co-treatment with the ACE2 catalytic inhibitor MLN-4760 on plasma clot formation. E – J show representative results, with similar trends observed in three independent experiments

Article Snippet: For the ACE2-Fc treatment groups, trimeric Spike protein was pre-incubated for 15 min with ACE2-Fc alone, ACE2-Fc plus Spike antibody (Genetex), or ACE2-Fc plus the ACE2 catalytic inhibitor MLN-4760 (MedChemExpress), prior to mixing with plasma and initiating clot formation.

Techniques: Blocking Assay, Infection, Control, Inhibition, Clinical Proteomics

Journal: Journal of Biomedical Science

Article Title: The ACE2 decoy receptor can overcome immune escape by rapid mutating SARS-CoV-2 variants and reduce cytokine induction and clot formation

doi: 10.1186/s12929-025-01156-4

Figure Lengend Snippet: Schematic diagram of the impact of SARS-CoV-2 variants and ACE2-Fc inhibition. This diagram illustrates the process of cell fusion induced by different SARS-CoV-2 variants, which leads to cytotoxicity and cytokine induction in host cells. It also shows how ACE2-Fc treatment can inhibit these effects. ACE2-Fc is highlighted as a therapeutic agent that effectively blocks the disease progression at multiple stages: preventing the virus from entering cells, reducing cell fusion, mitigating cell damage, and decreasing the release of inflammatory cytokines

Article Snippet: For the ACE2-Fc treatment groups, trimeric Spike protein was pre-incubated for 15 min with ACE2-Fc alone, ACE2-Fc plus Spike antibody (Genetex), or ACE2-Fc plus the ACE2 catalytic inhibitor MLN-4760 (MedChemExpress), prior to mixing with plasma and initiating clot formation.

Techniques: Inhibition, Biomarker Discovery, Virus

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Participant demographics and health characteristics.

Article Snippet: Samples were assayed in duplicates without ACE2 inhibitor and in singles with 10 −5 M ACE2 inhibitor MLN‐4760 (Calbiochem, San Diego, CA, cat. 530616).

Techniques: Infection, Biomarker Discovery

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Overview of ACE2 serology results and establishment of seropositivity thresholds. (A) Overall view of anti‐ACE2 IgG, IgA, and IgM antibodies of the full cohort, including longitudinal sampling. (B) Quantitative Venn diagram representation of the distribution between IgM, IgA, and IgG ACE2 autoantibodies in the overall cohort of ACE2‐positive individuals. (C) IgG, IgA, and IgM anti‐ACE2 antibodies and associated threshold. IgG and IgA threshold was set at two cycles of 2 standard deviations (SD) from the mean of the presumed negative distribution. IgM threshold was set by a first cycle at 2SD and a second at 1SD from the mean of the presumed negative distribution.

Article Snippet: Samples were assayed in duplicates without ACE2 inhibitor and in singles with 10 −5 M ACE2 inhibitor MLN‐4760 (Calbiochem, San Diego, CA, cat. 530616).

Techniques: Sampling

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Influence of clinical features and demographic characteristics on ACE2 autoantibodies prevalence. Odd ratios (OR) and 95% confidence interval (CI) were calculated for demographic characteristics and clinical features for individuals in relation to (A) IgG, (B) IgA and (C) IgM ACE2 autoantibodies. A table with a complete list of OR and 95% CI is available in the Table . The OR (CI95) are shown here on a log10 formatted axis.

Article Snippet: Samples were assayed in duplicates without ACE2 inhibitor and in singles with 10 −5 M ACE2 inhibitor MLN‐4760 (Calbiochem, San Diego, CA, cat. 530616).

Techniques:

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Persistence of autoantibodies in serum over time and association with previous SARS‐CoV‐2 infection. (A) Some individuals had one or two follow‐up samples. ACE2 IgG, IgA, IgM is shown longitudinally (Follow up 1 ( n = 120), median time between draws (SD) = 35 days (10), Follow up 2 ( n = 35), median time between draws (SD) = 37 days (6)). (B) IgG, IgA, and IgM ACE2 autoantibodies levels are shown for individuals who never had SARS‐CoV‐2 and for individuals who had serological markers of SARS‐CoV‐2 infection and/or self‐reported a SARS‐CoV‐2 infection. Unpaired two‐sided Wilcoxon test was used to establish statistical significance ( n = 464), p ≤ 0.05 (*), p ≤ 0.01 (**), p ≤ 0.001 (***), p ≤ 0.0001 (****).

Article Snippet: Samples were assayed in duplicates without ACE2 inhibitor and in singles with 10 −5 M ACE2 inhibitor MLN‐4760 (Calbiochem, San Diego, CA, cat. 530616).

Techniques: Infection

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Effect of ACE2 autoantibodies on ACE2 enzymatic activity and SARS‐CoV‐2 spike‐ACE2 binding. (A) ACE2 autoantibodies levels (IgG, IgA, IgM) in a subset of samples for which ACE2 enzymatic activity was assessed. (B) Levels of ACE2 enzymatic activity in individuals who had IgG and/or IgA and/or IgM ACE2 autoantibodies in contrast to the individuals who had no detectable ACE2 autoantibodies. (C) Levels of ACE2 enzymatic activity in individuals who self‐reported a SARS‐CoV‐2 infection and/or had serological markers of previous infection. (D) ACE2 autoantibodies levels in a subset of samples in which ACE2 and spike interaction was assessed. (E) Anti‐Spike IgG antibodies levels pre‐ and post‐depletion in sera. (F) % inhibition representing the ability of the sera to inhibit spike and ACE2 interaction pre‐ and post‐depletion. (G) % inhibition representing the ability of the sera to inhibit spike and ACE2 interaction in individuals who had IgG and/or IgA and/or IgM ACE2 autoantibodies above threshold in contrast to the individuals who had no detectable ACE2 autoantibodies. (H) Linear correlation between the ability of sera to inhibit Spike and ACE2 binding and the levels of Spike IgG antibodies in depleted sera. Unpaired two‐sided Wilcoxon test was used to establish statistical significance where applicable, p ≤ 0.05 (*), p ≤ 0.01 (**), p ≤ 0.001 (***), p ≤ 0.0001 (****).

Article Snippet: Samples were assayed in duplicates without ACE2 inhibitor and in singles with 10 −5 M ACE2 inhibitor MLN‐4760 (Calbiochem, San Diego, CA, cat. 530616).

Techniques: Activity Assay, Binding Assay, Infection, Inhibition