Figure 3. The correlation between post-treatment PIP₃ levels and the capacity of Akt inhibition to re-sensitize resistant cells to PI3Ki.

A) Each resistant line was treated with escalating doses of the agent to which it was made resistant for 24 hours (BYL719 for 453R and T47DR, and GDC-0941 for MCF7R). Lysates were prepared and were probed with the indicated antibodies.

B) Phospholipids in parental and resistant cells were isolated after 24 hr treatment with vehicle or the indicated PI3Ki (for parental lines) or continuous treatment with PI3Ki (for resistant lines. PIP₃ and PIP₂ levels were measured and the data shown as mean of three independent experiments. *indicates p<0.05 by student's t test.

C) Viability was assessed in parental and resistant 453 and T47D lines treated with escalating doses of MK2206 for 5 days. Resistant lines were also treated with escalating doses of MK2206 in the presence of 1 μM BYL719, and dose response curves were generated. Viability values for each curve were normalized to the measured inhibition value at zero dose of MK2206 (-/+ BYL719 1 μM for 453R and T47DR). Data represent mean of 3 replicates.

D) MCF7 and MCF7R cells were treated with escalating doses of MK2206. MCF7R was also treated with MK2206 in the presence of 1 μM dose of GDC-0941 or BYL719. Viability values for each curve were normalized to the measured inhibition value at zero dose of MK2206 (-/+ GDC-0941 1 μM for MCF7R). Data represent mean of 3 replicates.

E-F) Parental and resistant 453, T47D (E), and MCF7 (F) cells were treated for 24 hours with the specified agents. Lysates were made after 24 hours, and probed with the indicated antibodies.

All error bars in this figure represent +/- SEM.

See also Figure S3.