The phosphatidylinositol-3-kinase inhibitor PX-866 overcomes resistance to the epidermal growth factor receptor inhibitor gefitinib in A-549 human non-small cell lung cancer xenografts

Abstract

Epidermal growth factor receptor (EGFR) inhibitors such as gefitinib show antitumor activity in a subset of non-small cell lung cancer (NSCLC) patients having mutated EGFR. Recent work shows that phosphatidylinositol-3-kinase (PI3-K) is coupled to the EGFR only in NSCLC cell lines expressing ErbB-3 and that EGFR inhibitors do not inhibit PI3-K signaling in these cells. The central role PI3-K plays in cell survival suggests that a PI3-K inhibitor offers a strategy to increase the antitumor activity of EGFR inhibitors in resistant NSCL tumors that do not express ErbB-3. We show that PX-866, a PI3-K inhibitor with selectivity for p110alpha, potentiates the antitumor activity of gefitinib against even large A-549 NSCL xenografts giving complete tumor growth control in the early stages of treatment. A-549 xenograft phospho-Akt was inhibited by PX-866 but not by gefitinib. A major toxicity of PX-866 administration was hyperglycemia with decreased glucose tolerance, which was reversed upon cessation of treatment. The decreased glucose tolerance caused by PX-866 was insensitive to the AMP-activated protein kinase inhibitor metformin but reversed by insulin and by the peroxisome proliferator-activated receptor-gamma activator pioglitazone. Prolonged PX-866 administration also caused increased neutrophil counts. Thus, PX-866, by inhibiting PI3-K signaling, may have clinical use in increasing the response to EGFR inhibitors such as gefitinib in patients with NSCLC and possibly in other cancers who do not respond to EGFR inhibition.

Figure 1. Potentiation of the antitumor activity of gefitnib by PX-866

Female scid mice were implanted subcutaneously with 10⁷ A-549 human non small cell lung cancer cells. A, Tumors were 100 mm³ on day 22 (shown by dashed line) when dosing was begun every other day for 15 doses shown by the arrows with () vehicle alone; (_) gefitinib 75 mg/kg po; (Δ) PX-866 9 mg/kg iv; (_) PX-866 2.5 mg/kg po; (_) PX-866 9 mg/kg iv 4 hr before gefitinib 75 mg/kg po; (_) PX-866 2.5 mg/kg po 4 hr before gefitinib 75 mg/kg po. Values are the mean of 8 mice per group and bars are SE. B. Tumors were 600 mm³ on day 39 (shown by dashed line) when dosing was begun every other day for 14 doses shown by the arrows with (_ ) vehicle alone; (_) gefitinib 75 mg/kg po; (Δ) PX-866 12 mg/kg iv; (_) PX-866 4 mg/kg po; (_) PX-866 12 mg/kg iv 4 hr before gefitinib 75 mg/kg po; (_) PX-866 4 mg/kg po 4 hr before gefitinib 75 mg/kg po. Values are the mean of 8 mice per group and bars are SE.

Figure 2

Inhibition of EGFR and phospho-Akt in A-549 non small cell lung cancer xenografts by gefitinib and PX-866.. Female scid mice with 300 mm³ A-549 non small cell lung cancer xenografts were administered gefitinib 75 mg/kg po, PX-866 12 mg/kg iv or PX-866 3 mg/kg po, alone or with the PX-866 4 hr before the gefitinib, daily every other day for 5 days. 24 hr after the last dose tumors were removed for measurement of phospho-Akt/total Akt shown by the open bars or phospho-EGFR shown by the shaded bars. Values are the mean of 3 mice and bars are SE. * p < 0.05 compared to non treated control.

Figure 3. Effect of PX-866 on plasma insulin and glucose tolerance

A, Plasma insulin in female C57Bl/6 mice fasted for 16 hr. (○) vehicle control and (•) mice administered PX-866 10 mg/kg po. Values are the mean of 3 mice and bars are SE. B, Plasma glucose in female C57Bl/6 mice fasted for 16 hr and administered a dose of glucose of 1 g/kg po. (○) vehicle control; (•) administered PX-866 10 mg/kg 4 hr previously; (_) treated daily for 7 days with pioglitazone 10 mg/kg ip; and (Δ) treated daily for 7 days with pioglitazone 10 mg/kg ip and administered PX-866 10 mg/kg 4 hr previously. Values are the mean of 4 mice and bars are SE.