Mouse Anti-VEGFA Recombinant Antibody (clone mAb G6-31) (CAT#: HPAB-0330CQ)

Figure 1 Assessment of G6–31 activity against lung lesions when administrated intraperitoneally.

(A) Experimental procedure; Four month-old K-rasLA1 mice received G6–31 or an isotype control, administrated by i.p injection or aerosol, once a week for 4 wk. At the time of death visible lesions were counted on the whole lungs. (B) Quantification of visible nodules per mouse (n = 15 mice per group; 2.5 mg/kg and 10 mg/kg; *P< 0.05 Mann-Whitney test). Results are expressed as the mean ± SEM of nodules. (C) Quantification of lung lesions on H&E stained sections from control and G6–31 treated group (n = 15 mice per group; 2.5 mg/kg and 10 mg/kg; P< 0.05 Mann-Whitney test). Results are expressed as the mean ± SEM of the number of lesions.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 2 Assessment of aerosolized G6–31 activity against lung lesions.

(A) Quantification of visible nodules per mouse (n = 15 mice per group; 2.5 mg/kg and 10 mg/kg; *P < 0.05 Mann-Whitney test). Results are expressed as the mean ± SEM of nodules. (B) Quantification of lung lesions on H&E stained sections from control and G6–31 treated group (n =15 mice per group; 2.5 mg/kg and 10 mg/kg; P < 0.05 Mann-Whitney test). Results are expressed as the mean ± SEM of the number of lesions.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 3 Effect of G6–31 at 10 mg/kg administered by i.p. injection or aerosol on K-ras LA1 lung tumors according to each lesion type and effect on microvascular density.

(A) Quantification of AAH (atypical alveolar hyperplasia), Ad (adenoma) and ADC (adenocarcinoma), on H&E stained lung sections from control and G6–31-treated (10 mg/kg dose) mice (n = 30 mice per group; *P < 0.05 Mann-Whitney test). Results are expressed as the mean ± SEM of the number of each lesion type. (B) Representative image of vWFimmunostaining in the lung of K-ras LA1 mice. Bold arrow shows large vessel while standard arrow shows small vessel. (C) Quantification of the microvascular density, from vWFimmunostaining in control and G6–31-treated (10 mg/kg dose) mice (n = 30 mice per group; *P < 0.05 Mann-Whitney test). Results are expressed as the mean ± SEM.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 4 Detection of PCNA by western blotting in whole lung protein extracts from control mice (mice 1–4) and G6–31-treated (10 mg/kg) mice (mice 5–9).

Pulmonary delivery is on the right and i.p. delivery is on the left.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 5 CC-3 immunostaining.

Representative images of the control (left) and G6–31 (10 mg/kg dose) delivered either by i.p. (middle) or by pulmonary route (right). Brown cells are positive for CC-3 (arrows show some representative cells). The slices were counterstained with hematoxylin (× 400 magnification).

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 6 Quantification of apoptosis in tumor lesions of the right lung of K-ras LA1 mice (n = 15 mice per group; *P < 0.05 Mann-Whitney test).

Results are expressed as the mean staining scores of CC-3 positive cells relative to the control group ± SEM.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 7 Blockade of VEGFR2 signaling pathway.

Phospho-VEGFR2 Y1175, total VEGFR2, Phospho-PI3K, b-actin, Phospho-AKT, total AKT, Phospho-ERK and total ERK were analyzed by western blotting in whole lung protein extracts from control mice (mice 1–4) and from mice treated with G6–31 at 10 mg/kg (mice 5–9). Pulmonary delivery is on the right and i.p. delivery is on the left.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 8 Pharmacokinetics of G6–31.

Serum concentration of G6–31 mAb was measured by ELISA in the serum collected at different time points from mice that received a single dose of the mAb (10 mg/kg) either by the pulmonary route (left, n = 13) or intravenously (right, n = 15). Gray area and solid line show the 5th-95th and 50th percentiles of the model-predicted time-concentration profiles, respectively.

Hervé, V., Rabbe, N., Guilleminault, L., Paul, F., Schlick, L., Azzopardi, N., ... & Paintaud, G. (2014, November). VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations. In MAbs (Vol. 6, No. 6, pp. 1638-1648). Taylor & Francis.

Figure 9 Mab G6-31 treated Drd2-/- female mice.

A) Pituitary weight in vehicle (N=5) and Mab G6-31 treated (N=7) Drd2-/- female mice, * p <0.05 vs vehicle Drd2-/- treated mice. B) Serum prolactin levels in female Drd2-/- vehicle (N=5) or Drd2-/- Mab G6-31 treated mice (N=7). C) Pituitary prolactin concentration (ng/gland) in control Drd2-/-, Mab G6-31 treated Drd2-/- and wild-type (WT) female mice (N=5, 5 and 5, respectively). D) Percent of prolactin positive area by confocal immunohistochemistry in control Drd2-/- and Mab G6-31 treated Drd2-/- and wild-type (WT) female mice (N=6, 7 and 5 respectively), * p<0.05 vs. Drd2-/- vehicle treated mice.

Luque, G. M., Perez-Millán, M. I., Ornstein, A. M., Cristina, C., & Becu-Villalobos, D. (2011). Inhibitory effects of antivascular endothelial growth factor strategies in experimental dopamine-resistant prolactinomas. Journal of Pharmacology and Experimental Therapeutics, 337(3), 766-774.

Figure 10 Percent of PCNA stained cells (positive PCNA nuclei X 100/ total nuclei) evaluated by immunohistochemistry in control Drd2-/-, Mab G6-31 treated Drd2-/-, and wild-type (WT) female mice (N=6, 6 and 4 respectively), * p <0.05 vs. Drd2-/- vehicle treated mice.

Luque, G. M., Perez-Millán, M. I., Ornstein, A. M., Cristina, C., & Becu-Villalobos, D. (2011). Inhibitory effects of antivascular endothelial growth factor strategies in experimental dopamine-resistant prolactinomas. Journal of Pharmacology and Experimental Therapeutics, 337(3), 766-774.