Fig 1: Inhibitory effects of mAb-2E3 on the lung metastasis and tumor growth in mouse model. A-E. Effect of mAb-2E3 on lung metastasis in SCID/Beige mice. (A) The mice were intraperitoneally injected with antibody (10 mg/kg, 40 mg/kg) or 40mg/kg control non-immune IgG every three days for ten times at 1 day after intravenous injection of 2×106 KYSE30lm3 cells, and metastasis was monitored by IVIS imaging system for 4 weeks. (B) Effect of mAb-2E3 on lung metastases in the mice by IVIS. Removing extreme values (maximum and minima) for each group were shown and compared. (C) HE staining of mice lung tissues. Bars, 100µm. (D) Detection of PAI-1 level in plasma of lung metastasis mice model by ELISA. (E) Comparison of the mice weight (P=0.803). F-I. Effect of mAb-2E3 on KYSE30lm3 tumor growth in SCID/Beige mice. (F) 1×106 KYSE30lm3 cells were inoculated into the flank of SCID/Beige mice. The tumors were harvest after a month of treatment with mAb or control. (G) The tumor volume (mm3) in the mAb group decreased significantly compared with the control. (H) Comparison of the tumor weight. (I) There was no difference in the weight of the mice. *P<0.05, **P<0.01, ****P<0.0001, ns =0.05.
Fig 2: MAb-2E3 blocking the binding of LRP1-Cluster II-Fc and PAI-1. A. The binding activity of PAI-1 and LRP1-Cluster II-Fc protein was determined and EC50 value was calculated. B. mAb-2E3 was able to block the binding between LRP1-Cluster II-Fc and PAI-1. mIgG was used as the control. IC50 values were calculated.
Fig 3: PAI-1 expression in patients with ESCC and the clinicopathological characteristics of ESCC. A. The level of PAI-1 in serum of ESCC patients was higher than in that of healthy donors (HD group = 8.61 ± 0.4909, n=180; ESCC group = 21.07 ± 1.17, n=180). B. Serum PAI-1 level was significantly higher in ESCC patients with more lymph node metastasis. C. ROC curve of PAI-1. Based on the area under the ROC curve, Youden Index cutoff values that maximized the sum of sensitivity and specificity were determined. D. Kaplan-Meier analysis of the survival rates of patients with ESCC in relation to serum PAI-1 protein expression. E. Representative images of PAI-1 staining according to the expression level in ESCC specimens. ***P<0.001.
Fig 4: Screening of human specific anti-PAI-1mAbs. A. The binding affinity of anti-PAI-1 mAbs to His-PAI-1 proteins was measured by ELISA. PAI-1 protein was coated in the 96-well plate at 50 µl/well overnight. Anti-PAI-1 mAb were added to the wells along gradient of concentration 10-fold increments ranging from e-13M to e-7 M. B. Immunoprecipitation of eukaryotic PAI-1 proteins with anti-PAI-1 mAbs and control mIgG. C. Immunohistochemical staining of ESCC tissue sections with anti-PAI-1 mAbs.
Fig 5: PAI-1 mAb inhibited migration in an LRP1-dependent manner. A. Interaction between PAI-1 and LRP1 detected by co-immunoprecipitation. B. qRT-PCR (left) and Western blot (right) analysis of LRP1 knockdown in KYSE30lm3 cells. C. Knockdown of LRP1 counteracted PAI-1 protein-promoted migration. D. Knockdown of LRP1abolished mAb-1E2 and 2E3-inhibited migration. E. mAb-2E3 inhibited the phosphorylation level of STAT1 and failed to inhibit the level of metastasis-related target proteins which were detected in cells under the treatment of antibody. F. Western blot of LRP1, pSTAT1 and total STAT1 in KYSE30lm3 cells transfected with siRNAs against LRP1. GAPDH was used as a loading control. G. Grayscale analysis of pSTAT1 and total STAT1 levels in KYSE30lm3 cells transfected with siRNAs against LRP1. H. qRT-PCR of STAT1 transcripts in KYSE30lm3 cells transfected with siRNAs against LRP1. *P<0.05, **P<0.01, ***P<0.001, ns =0.05.
Supplier Page from Sino Biological, Inc. for Human SerpinE1 / PAI-1 Protein (His Tag)