Afuco™ Anti-MET ADCC Recombinant Antibody, ADCC Enhanced (PABZ-155) (CAT#: PABZ-155)

Figure 1 Onartuzumab functionally inhibits HGF/MET signaling.

The relative inhibition of HGF binding (B), MET phosphorylation by KIRA in A549 cells treated with HGF (C), and cell proliferation in Ba/F3-huMET cells (D) is shown for chOA5D5, huOA5D5.v1, and huOA5D5.v2 [onartuzumab (MetMAb)]. (E) Onartuzumab inhibits HGF-dependent MET phosphorylation and downstream activation of GAB1, AKT, ERK1/2, and P70-S6 in a dose-dependent fashion but does not induce any MET signaling in the absence of HGF. (F) Onartuzumab inhibits HGF-dependent migration and wound closure in HGF-treated A549 cells with an IC₅₀ of 20 nM. All plots reflect group mean plus and minus the SEM (n = 3).

Merchant, M., Ma, X., Maun, H. R., Zheng, Z., Peng, J., Romero, M., ... & Santell, L. (2013). Monovalent antibody design and mechanism of action of onartuzumab, a MET antagonist with anti-tumor activity as a therapeutic agent. Proceedings of the National Academy of Sciences, 110(32), E2987-E2996.

Figure 2 The one-armed 5D5 antibody and onartuzumab have prolonged exposure in mice and antitumor activity in HGF-dependent tumor models.

(A) The chOA5D5 and 5D5 Fab proteins were dosed at 5 mg/kg i.v. in nude mice (n = 5), and their serum concentrations were measured over a 2-wk period. The serum half-life for the 5D5 Fab was a few minutes versus 5.8 d for the chOA5D5 antibody. (B) Serum drawn from nude mice dosed in A at day 7 posttreatment was assessed via Western blot using anti-Fab antibodies in nonreduced (Left) or reduced (Right) conditions, demonstrating stability of chOA5D5 in vivo relative to the 5D5 Fab. (C) Nude mice (n = 10 per group) bearing s.c. U-87 MG xenograft tumors were treated with onartuzumab at doses between 1–60 mg/kg, given once via i.p. injection. Data are plotted as group mean tumor volumes, with SEM indicated. Overall tumor response rates were calculated based upon the combined number of partial responses (PRs) and complete regressions (CRs), defined as>50% or 100% drop in tumor volumes, respectively, at any time during the study, and were determined for each dose group to be as follows: 1 mg/kg, 5 PRs and 0 CRs; 3.75 mg/kg, 5 PRs and 0 CRs; 7.5 mg/kg, 90% (7 PRs and 2 CRs); 15 mg/kg, 100% (5 PRs and 5 CRs); 30 mg/kg, 70% (2 PRs and 5 CRs); and 60 mg/kg, 100% (6 PRs and 4 CRs).

Merchant, M., Ma, X., Maun, H. R., Zheng, Z., Peng, J., Romero, M., ... & Santell, L. (2013). Monovalent antibody design and mechanism of action of onartuzumab, a MET antagonist with anti-tumor activity as a therapeutic agent. Proceedings of the National Academy of Sciences, 110(32), E2987-E2996.

Figure 3 The one-armed 5D5 antibody and onartuzumab have prolonged exposure in mice and antitumor activity in HGF-dependent tumor models.

(D) Immunoprecipitation–immunoblot analysis for p-MET (IP-MET, IB-pTyr) and total MET (IP-MET, IB-MET) from U-87 MG xenograft tumors from nude mice following treatment with 30 mg/kg, i.p., once at the indicated time points. (E) MRI evaluation of intracranial U-87 MG xenograft tumors (n = 10 per group) treated with vehicle (black circles), onartuzumab at doses of 7.5 mg/kg (purple triangles), 30 mg/kg (blue circles), 60 mg/kg (green squares), or 120 mg/kg (red diamonds), dosed i.p., once at pretreatment (P), 1 and 2 wk postdose. The mean starting tumor volume for the vehicle group is indicated by the dashed line. Group mean and SEM for each group is shown overlaying the individual animal tumor volumes. (F) Kaplan–Meier analysis for intracranial U-87 MG tumors treated with vehicle (black line) or onartuzumab at 7.5 mg/kg (purple line), 30 mg/kg (blue line), 60 mg/kg (green line), and 120 mg/kg (red line). The median survival for each group was as follows: vehicle, 21 d; 7.5 mg/kg, 37 d; 30 mg/kg, 52 d; 60 mg/kg, 53 d; 120 mg/kg, 51 d. (G) hHGFtg-C3H-SCID mice (n = 10 per group) bearing s.c. NCI-H596 xenograft tumors were treated with either vehicle (white circles) or onartuzumab dosed at 15 mg/kg (light blue diamonds), 30 mg/kg (medium blue squares), or 60 mg/kg mg/kg (dark blue stars). Growth of NCI-H596 in littermate C3H-SCID mice is shown (black circles). Data are shown as group mean tumor volumes, with SEM indicated. (H) Immunoprecipitation–immunoblot analysis for p-MET (IP-MET, IB-pTyr) and total Met (IP-MET, IB-MET) from NCI-H596 xenograft tumors from hHGFtg-C3H-SCID mice following treatment with 30 mg/kg, i.p., once at 0 and 72 h.

Merchant, M., Ma, X., Maun, H. R., Zheng, Z., Peng, J., Romero, M., ... & Santell, L. (2013). Monovalent antibody design and mechanism of action of onartuzumab, a MET antagonist with anti-tumor activity as a therapeutic agent. Proceedings of the National Academy of Sciences, 110(32), E2987-E2996.

Figure 4 Affinity maturation of mouse 5D5 to generate onartuzumab

The on (k_on) and off (k_off) rates and affinity measurements (K_D) are summarized for each clone.

Merchant, M., Ma, X., Maun, H. R., Zheng, Z., Peng, J., Romero, M., ... & Santell, L. (2013). Monovalent antibody design and mechanism of action of onartuzumab, a MET antagonist with anti-tumor activity as a therapeutic agent. Proceedings of the National Academy of Sciences, 110(32), E2987-E2996.