Supplements
Appendix A1: Cell proliferation in primary cells
A) Number of cells after 36h, 48h, 72h and 96h after treatment with 2.5μg/ml, 5 μg/ml and
10μg/ml Cetuximab. Application of the monoclonal antibody to EGFR Cetuximab did not lead to
a reduction in cell number independent of the applied concentration or incubation time in any of
the 5 probed primary cultures.
B) Gefitinib: Number of cells after 36h, 48h, 72h and 96h after adding Gefitinb. The number of
treated cells was negatively correlated to the applied inhibitor concentration for 0.1 μM and 1
μM compared to the control group. A highly significant reduction of cells was detected after 72h
and 96h of 5μM Gefitinib treatment compared to control in any of the 5 probed primary cultures.
PTPIP51 levels in glioblastoma cells depend on inhibition of the EGF-receptor, Journal of
Neuro-Oncology, M.K. Petri, A. Brobeil, J. Planz, A. Bräuninger, S. Gattenlöhner, U. Nestler, A.
Stenzinger, A. Paradowska, M. Wimmer. meike.petri@gmx.de
Appendix A2: Concentration gradient of Gefitinib and reference gel
A) Gefitinib concentration gradient: Quantitative PCR of PTPIP51 and 14-3-3 in Gefitinib
(0.1µM, 1µM, 2µM, 3µM, 4µM and 5µM) treated U87 cells after 24h. No significant differences
in mRNA of either protein were seen. B) Reference gel: bands exclusively detecting mRNA with
the expected amplification size.
PTPIP51 levels in glioblastoma cells depend on inhibition of the EGF-receptor, Journal of
Neuro-Oncology, M.K. Petri, A. Brobeil, J. Planz, A. Bräuninger, S. Gattenlöhner, U. Nestler, A.
Stenzinger, A. Paradowska, M. Wimmer. meike.petri@gmx.de
Appendix A3: Erlotinib
Immunostaining of Erlotinib treated U87 cells. The application of 1 µM Erlotinib for 24 h
resulted in dispersed granular cytoplasmic distribution of PTPIP51. After 96 h treatment with 1
µM Erlotinib a more attenuated granular pattern of the reduced PTPIP51 protein (A3B) was
seen. The application of 20 µM Erlotinib resulted in a distinct subcellular localization of
PTPIP51 with highest concentrations in the perinuclear region (A3 C and F). With increasing
incubation time (96 h) 20 µM Erlotinib treatment resulted in a further redistribution of PTPIP51
to a dispersed granular pattern (A3 D).Bar=20µm
PTPIP51 levels in glioblastoma cells depend on inhibition of the EGF-receptor, Journal of
Neuro-Oncology, M.K. Petri, A. Brobeil, J. Planz, A. Bräuninger, S. Gattenlöhner, U. Nestler, A.
Stenzinger, A. Paradowska, M. Wimmer. meike.petri@gmx.de
Appendix A4: Primary cell cultures quantitative real time PCR of PTPIP51 and 14-3-3
mRNA
A) Quantitative PCR of PTPIP51 in Gefitinib treated primary cells: No significant differences in
PTPIP51 were seen after 24h of treatment with 0.1µM, 1µM and 5µM Gefitinib compared to
controls. Likewise no significant differences in PTPIP51 mRNA were detected after incubation
periods of 48h, 72h and 96h.
B) Quantitative PCR of 14-3-3 in Gefitinib treated primary cells: No significant differences in
14-3-3 mRNA were seen after 24h of treatment with 0.1µM, 1µM and 5µM Gefitinib compared
to controls. Likewise no significant differences in 14-3-3 mRNA were detected after incubation
periods of 48h, 72h and 96h. Values were analyzed by using the Dunnett’s multiple comparison
test.
PTPIP51 levels in glioblastoma cells depend on inhibition of the EGF-receptor, Journal of
Neuro-Oncology, M.K. Petri, A. Brobeil, J. Planz, A. Bräuninger, S. Gattenlöhner, U. Nestler, A.
Stenzinger, A. Paradowska, M. Wimmer. meike.petri@gmx.de