Journal: bioRxiv

Article Title: Circulating biomarkers in serum from aortic valve stenosis patients predict sex-specific drug responses in valve myofibroblasts

doi: 10.64898/2026.01.16.700010

Figure Lengend Snippet: Schematic overview of the approach to link candidate serum biomarkers with in vitro drug efficacy. B, αSMA gradient mean intensity for male and female VICs cultured on hydrogels with sex-matched AVS patient serum (N=16 male, N=15 female). Statistical significance is indicated as *** = P < 0.001 (unpaired two-tailed t-test with Welch’s correction). C, Percent αSMA reduction in male and female VICs cultured on hydrogels with optimized doses of Ataciguat, Colchicine, or Evogliptin. Statistical significance is indicated as ** = P < 0.01 (unpaired two-tailed t-test with Welch’s correction). D, Heatmaps showing the correlation between each candidate serum factor and antifibrotic drug efficacy for male or female VICs cultured with sex-matched AVS serum (N=16 male, N=15 female). Statistical significance is indicated as * = P < 0.05 (paired two-tailed t-test). E, Correlation between serum IGFBP2 abundance (relative fluorescence units (RFU)) and percent αSMA reduction in male and female VICs cultured on gels with sex-matched AVS serum and Evogliptin (N=16 male, N=15 female).

Article Snippet: Human IGFBP2 neutralizing antibody (R&D systems, Cat. No AF674) and normal IgG control antibody (R&D systems, Cat. No AB-108-C) were resuspended in sterile PBS at 200 μg/mL and 1,000 μg/mL respectively.

Techniques: In Vitro, Cell Culture, Two Tailed Test, Fluorescence

Journal: bioRxiv

Article Title: Circulating biomarkers in serum from aortic valve stenosis patients predict sex-specific drug responses in valve myofibroblasts

doi: 10.64898/2026.01.16.700010

Figure Lengend Snippet: IGFBP2 mediates Evogliptin resistance in female VICs cultured with female AVS serum . A, Serum IGFBP2 concentrations quantified using an ELISA kit (N=31 AVS, N=5 healthy). Statistical significance is indicated as **** = P < 0.0001 (unpaired two-tailed t-test with Welch’s correction). B through C, Percent αSMA reduction in (B) female and (C) male VICs cultured on hydrogels with FBS and treated with Evogliptin and/or recombinant human IGFBP2 protein (N=6 gels). Statistical significance is indicated as * = P < 0.05 (one sample t and Wilcoxon test against a theoretical mean of zero). D, Serum IGFBP2 levels quantified using an ELISA kit with and without IGFBP2 neutralizing antibody (IGFBP2 Ab), IgG control antibody, and recombinant human IGFBP2 protein (N=3 male, N=3 female). Statistical significance is indicated as *** = P < 0.001 (one-way ANOVA with Tukey posttests). E, Percent αSMA reduction in male and female VICs cultured on hydrogels with sex-matched AVS patient serum samples, Evogliptin, IgG, IGFBP2 Ab, or recombinant human IGFBP2 protein (N=3 male, N=3 female). Statistical significance is indicated as *** = P < 0.001 (one-way ANOVA with Tukey posttests). F, Representative immunofluorescent images of male and female VICs cultured on hydrogels (green: αSMA, blue: nuclei). Scale bar = 100 μm. G, Normalized IGFBP to IGF1 ratio for male and female AVS serum samples (N=23 male, N=16 female). Statistical significance is indicated as *** = P < 0.001 (unpaired two-tailed t-test with Welch’s correction). H, Correlation between normalized IGFBP to IGF1 ratios and percent αSMA reduction in male and female VICs cultured on hydrogels with sex-matched serum and Evogliptin.

Article Snippet: Human IGFBP2 neutralizing antibody (R&D systems, Cat. No AF674) and normal IgG control antibody (R&D systems, Cat. No AB-108-C) were resuspended in sterile PBS at 200 μg/mL and 1,000 μg/mL respectively.

Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Recombinant, Control

Journal: bioRxiv

Article Title: Circulating biomarkers in serum from aortic valve stenosis patients predict sex-specific drug responses in valve myofibroblasts

doi: 10.64898/2026.01.16.700010

Figure Lengend Snippet: Principal component analysis clustering samples based on RNA sequencing results (N=3). B through C, Volcano plot of differential gene expression from RNA sequencing between (B) Vehicle and Evogliptin groups or (C) Vehicle and Evogliptin plus IGFBP2 neutralizing antibody groups (N=3). Statistical significance was defined as FDR-adjusted P-value < 0.05 (unpaired two-tailed t-test with Welch’s correction) and fold change greater than 2. D, Venn diagram comparing differentially expressed genes in the Evogliptin and Evogliptin plus IGFBP2 neutralizing antibody groups relative to vehicle control (N=3). E, Volcano plot of differential gene expression from RNA sequencing between Evogliptin and Evogliptin plus IGFBP2 neutralizing antibody groups (N=3). Statistical significance was defined as FDR-adjusted P-value < 0.05 (unpaired t-test with Welch’s correction) and fold change greater than 2. F, Heatmap of the top 20 up and down regulated genes in the Evogliptin plus IGFBP2 neutralizing antibody group relative to vehicle control (N=3). G through H, Top 15 upregulated pathways in Evogliptin versus Evogliptin plus IGFBP2 neutralizing antibody group identified by (G) KEGG and (H) Ingenuity Pathway Analysis. Abbreviations: Evo: Evogliptin, Evo + Ab: Evogliptin with IGFBP2 neutralizing antibody, Veh: Vehicle control. Number indicates patient serum sample ID (26, 30, and 45).

Article Snippet: Human IGFBP2 neutralizing antibody (R&D systems, Cat. No AF674) and normal IgG control antibody (R&D systems, Cat. No AB-108-C) were resuspended in sterile PBS at 200 μg/mL and 1,000 μg/mL respectively.

Techniques: RNA Sequencing, Gene Expression, Two Tailed Test, Control

Journal: bioRxiv

Article Title: Circulating biomarkers in serum from aortic valve stenosis patients predict sex-specific drug responses in valve myofibroblasts

doi: 10.64898/2026.01.16.700010

Figure Lengend Snippet: IGFBP2 promotes Evogliptin resistance in female VICs cultured with female AVS serum through Rho/ROCK and focal adhesion kinase signaling. A, Schematic overview of the approach to validate the mechanism of IGFBP2-mediated Evogliptin resistance. B, Percent αSMA reduction in female VICs cultured on hydrogels with female AVS serum and treated with various doses of H-1152 with and without Evogliptin (N=3 gels). For H-1152: + = 0.1 μM; ++ = 0.5 μM; +++ = 5 μM. Statistical significance is indicated as * = P < 0.05 (one-way ANOVA with Tukey posttests). C, Percent αSMA reduction in female VICs cultured on hydrogels with AVS serum and treated with various doses of PF-573228 with and without Evogliptin (N=3 gels). For PF-573228: + = 0.1 μM; ++ = 1 μM; +++ = 10 μM. Statistical significance is indicated as * = P < 0.05 (one-way ANOVA with Tukey posttests). D through E, Percent αSMA reduction in (D) female or (E) male VICs cultured on hydrogels with sex-matched AVS serum and treated with various combinations of Evogliptin, IGFBP2 neutralizing antibody, H-1152, and PF-573228 (N=3 serum samples per sex). Statistical significance is indicated as * = P < 0.05, ** = P < 0.01 (one-way ANOVA with Tukey posttests). F, Representative immunofluorescent images of male and female VICs cultured on hydrogels with sex-matched serum and various drug combinations (green: αSMA, blue: nuclei). Scale bar = 100 μm. Abbreviations: Evo = Evogliptin, H11 = H-1152, PF5 = PF-573228, Ab = IGFBP2 neutralizing antibody.

Article Snippet: Human IGFBP2 neutralizing antibody (R&D systems, Cat. No AF674) and normal IgG control antibody (R&D systems, Cat. No AB-108-C) were resuspended in sterile PBS at 200 μg/mL and 1,000 μg/mL respectively.

Techniques: Cell Culture

Journal: bioRxiv

Article Title: Circulating biomarkers in serum from aortic valve stenosis patients predict sex-specific drug responses in valve myofibroblasts

doi: 10.64898/2026.01.16.700010

Figure Lengend Snippet: Summary mechanistic figure illustrating how elevated IGFBP2 in female serum modulates Evogliptin efficacy in female VICs.

Article Snippet: Human IGFBP2 neutralizing antibody (R&D systems, Cat. No AF674) and normal IgG control antibody (R&D systems, Cat. No AB-108-C) were resuspended in sterile PBS at 200 μg/mL and 1,000 μg/mL respectively.

Techniques: