Fig. 6. Role of IGF-IR in the sensitization of MCF-7 and T-47D cells to
PD098059. A, MCF-7 and T-47D cells were plated in
triplicate in 24-well plates at densities of 1 x 104
and 2.5 x 104 cells/cm2, respectively.
Cells subsequently were incubated in complete growth medium
supplemented with control antibody/DMSO, control antibody/PD098059,
anti-IGF-IR antibody/DMSO, or anti-IGF-IR antibody/PD098059. After 4
days, triplicate wells were counted for each treatment (Table 1)
number of cells in antibody/PD098059-treated wells was expressed as a
percentage of the number of cells in the respective
antibody/DMSO-treated wells. Means are shown; bars, SD.
The Students paired t test was used to determine that
PD098059-inhibited growth in the presence of anti-IGF-IR blocking
antibody was significantly different from PD098059-inhibited growth
with control IgG antibody. *, P < 0.05. Data
shown are representative of three similar experiments.
B, MCF-7 and T-47D cells were plated at equivalent
subconfluent densities and were treated as in A or were
left untreated. After 3 h, cell lysates were prepared with RIPA
buffer, and 750 µg of lysate protein were immunoprecipitated
(IP) with polyclonal anti-IGF-IR, resolved by SDS-PAGE,
and immunoblotted with anti-phosphotyrosine (anti-pTyr). From the same
lysates, 20 µg of protein were fractionated by SDS-PAGE and
immunoblotted with anti-phosphoERK1/ERK2. The membranes were stripped
and reprobed with anti-IGF-IR or anti-ERK2 to confirm that equivalent
amounts of protein were present in each lane.