Human Anti-FGFR2 Recombinant Antibody (clone M048-D01)

As shown antibodies M048-D01-hIgGl and M047-D08-hIgGl increase the ELISA signal of P-FGFR2 by a factor greater 4 fold compared with untreated control cells. In contrast neither the control IgG antibody nor anti FGFR2 antibodies showed any significant effect on P-FGFR2 levels after short-term incubation. These results reveal an agonistic effect of anti FGFR2 antibodies on FGFR2 after short-term incubation.

As shown the antibodies M048-D0I-hIgGl and M047-D08-hIgGl reduce the level of P-FGFR2 which can be achieved after FGF7 stimulation very pronounced. In cells treated without antibody treatment as well as in cells treated with isotype control IgG stimulation with FGF7 lead to an about 4 fold increase of P-FGFR2 levels. In contrast, in samples pretreated with anti FGFR2 antibodies for 24h, FGF7 only induced P-FGFR2 levels by 1.37-1.4 fold. Taken together these results show that prolonged incubation of cells with anti FGFR2 antibodies leads to desensitization towards stimulation with FGF7,

As shown antibodies M048-D01-hIgGl and M047-D08-hIgGl are the only antibodies that reduce FGFR2 surface expression with FGFR2 overexpressing cell lines (MFM223, SNU16) and cells lines having FGFR2 mutations (AN3-CA, MFE-296). Antibodies like MAB684 and MAB6843 only reduce FGFR2 surface expression with cell lines which do not overexpress FGFR2. Antibodies like GAL-FR21 do not reduce FGFR2 surface expression with cell lines having FGFR2 mutations.

As shown antibodies M048-D01-hIgGl (white) and M047-D08-hIgGl (striped) decrease the total FGFR2 levels significantly after 96h in a dose dependent manner. A non-binding control antibody (black) does not show any effects. These results indicate that anti FGFR2 antibodies M048-D01-hIgGl and M047-D08-hIgGl do not only lead to a short term decrease in surface FGFR2 levels but also a long term reduction of total FGFR2 levels.

Y is "granule counts per cell". X is "time [min]". Internalization of antibodies was investigated on breast cancer cell line SUM 52PE. The granule counts per cell were measured in a kinetic fashion. As shown antibodies M048-D01-hIgGl (black squares and solid line) and M047-D08-hIgGl (black triangles and dashed line) show a rapid internalization as indicated by increasing granule count per cell. An isotype control antibody (stars and dashed line) does not show any internalization.

Mean + standard deviation are plotted. X is "time after tumor inoculation [days]". Y is "tumor area [mm2]". Treatment with M048-D0 I-hlgG I resulted in a very significant tumor growth inhibition.

At this time point mice recived treatment with PBS alone (A), 5 mg/kg of M048-D01-hIgGl twice weekly i.v. (B), 100 mg/kg Lapatinib p.o. (C) or with 5 mg/kg of M048-D01-hIgGl twice weekly i.v. and 100 mg/kg Lapatinib p.o. (D). Y is tumor area [mm²] at day 13, dotted lines indicate the mean values, solid lines indicate the medians. Treatment with M048-D01-higGl alone resulted in a significant reduction of tumor area, while Lapatinib alone did not significantly affect tumor area. Combination of M048-D01-hIgGl with Lapatinib resulted in a significantly additive anti-tumor activity.

Mean ± standard error of the means are plotted. X is "time under treatment [days]". Y is "tumor volume [mm³]". Treatment with all three doses of M048-D01-hIgGl resulted in a significant tumor growth inhibition.

As shown after treatment with M048-D01-hIgGl and M047-D08-hIgGl total FGFR2 [total FGFR2] and phosphorylated FGFR2 [P-FGFR2] were reduced significantly in comparison with treatment with control IgGl. Actin served as loading control.