Table 1.

Clonogenic survival of pancreatic cancer cells treated with Gem + MK8776 or Gem + AZD1775 concurrent with roscovitine or exogenous nucleosides. Cells treated under equitoxic conditions with gemcitabine followed by either MK8776 or AZD1775 ± nucleosides or 20 μM roscovitine (as illustrated in Figure 1A) were trypsinized and collected at 30 h (MiaPaCa2, BxPC3, AsPC1, Capan1) or 48 h post-Gem (Panc1) (6 or 24 h MK8776 or AZD1775 ± nucleosides or roscovitine, respectively) and assayed for drug-induced loss of clonogenicity. Non-toxic, similarly sensitizing concentrations of each inhibitor were used: 500 nM MK8776 (MiaPaCa2, BxPC3, Panc1, Capan1), 250 nM MK8776 (AsPC1), 200nM AZD1775 (MiaPaCa2, AsPC1, Panc1) or 100nM AZD1775 (BxPC3, Capan1). Data are the mean clonogenic survival ± SEM (n = 3–7; p < 0.05, one-way ANOVA) and are normalized to the plating efficiency for either MK8776 alone, AZD1775 alone or the combinations of MK8776 or AZD1775 with roscovitine or exogenous nucleosides. Two-way analysis of variance was used to test for statistically significant differences in nucleoside or roscovitine protection from chemosensitization by AZD1775 compared to MK8776 (p < 0.05^¥). Under these treatment conditions, roscovitine was not toxic, and did not significantly affect Gem-induced loss of clonogenicity (Suppl. Figure 3).

Cell Line	MiaPaCa2	BxPC3	AsPC1	Panc1 (t = 48h)	Capan1
Gemcitabine	0.65 ± 0.04	0.44 ± 0.06	0.53 ± 0.03	0.49 ± 0.06	0.61 ± 0.02
Gem + MK8776	0.07 ± 0.01*	0.08 ± 0.01*	0.13 ± 0.03*	0.05 ± 0.01*	0.38 ± 0.03*
Gem + MK8776 + roscovitine	0.20 ± 0.01*¥	0.36 ± 0.07ɸ	0.56 ± 0.08ɸ	0.28 ± 0.05*ɸ	0.66 ± 0.06ɸ
Gem + MK8776 + nucleosides	0.19 ± 0.02*¥	0.24 ± 0.04	0.29 ± 0.05*	0.34 ± 0.03ɸ	0.66 ± 0.06ɸ
Gem + AZD1775	0.08 ± 0.01*	0.10 ± 0.01*	0.07 ± 0.01*	0.03 ± 0.01*	0.17 ± 0.03*
Gem + AZD1775 + roscovitine	0.48 ± 0.05*δ¥	0.45 ± 0.08δ	0.56 ± 0.07δ	0.41 ± 0.06δ	0.60 ± 0.05δ
Gem + AZD1775 + nucleosides	0.32 ± 0.03*δ¥	0.33 ± 0.07δ	0.31 ± 0.01*δ	0.10 ± 0.01*	0.55 ± 0.01δ

Versus Control*, Gem+MK8776^ɸ, Gem+AZD1775^δ or Gem+MK8776 vs Gem+AZD1775^¥ (P < 0.05)