Selective Small Molecule Inhibition of P110α and δ Isoforms of PI3 Kinase Is Cytotoxic to Human AML Progenitors

Abstract 1556

P110α, β, γ and δ isoforms are catalytic domains of phosphoinosityl-3-kinase (PI-3K) which is frequently constitutively active in blast cells from acute myeloid leukemia (AML) patients. RNA and protein from all 4 isoforms were ubiquitously expressed in 32 AML samples which also showed expression of p-Akt Ser473, indicating PI-3K activation. Treatment of AML patient blast cells (n=45) with the p110α selective inhibitor PI3-Kinase alpha inhibitor 2 and p110δ selective inhibitor PCN5603 (Piramed Pharma,UK) caused dose dependent kill of AML colony forming cells (CFC) and inhibition of p-Akt Ser473 expression. In contrast, the P110β and p110γ selective inhibitors TGX-221 and AS-604850 showed little AML CFC kill. AML samples were more sensitive to PI3-Kinase alpha inhibitor 2 and PCN5603 killing than normal bone marrow or normal peripheral blood (Median IC₅₀=1.8μM for 45 AML-CFC and 4.3μM for 8 normal CFC for PI3-Kinase alpha inhibitor 2. Median IC₅₀=1.9μM for 45 AML-CFC and 6.2μM for 10 normal CFC for PCN5603). There was no significant difference between the IC₅₀ and IC₉₀ of p110α and δ inhibitors against AML CFC from samples with or without the FLT3 ITD. There was a weak correlation between the IC₅₀ of the PI3-Kinase alpha inhibitor 2 or the IC₅₀ of PCN5603 and the ratio of pAkt/total Akt (R² = 0.23 and 0.24, respectively, p<0.05). RNAi inhibition of p110α and p110δ decreased the corresponding isoform's RNA and protein expression and caused AML CFC kill and inhibition of p-Akt Ser473 expression. There was a strong correlation between the percent AML CFC kill and RNA knockdown (R² = 0.67) and between inhibition of p-Akt and p110 protein expression (R² = 0.90 for α and 0.82 for δ). Thus, p110α and δ inhibition may achieve selective targeting of malignant rather than normal CFCs. Furthermore, treatment of AML cells with PCN5603, decreased survival of AML long-term culture- initiating cells (AML LTC-IC). In contrast, less toxicity toward normal bone marrow LTC-IC was observed (IC₅₀=0.08μM, ≤2μM and 1.33μM for 3 AML samples and IC₅₀=3.09μM, 2.63μM, >10μM and >10μM for 4 normal bone marrows). Selective inhibition of p110α and δ isforms of PI-3K kills AML progenitors including candidate leukemic stem cells (AML LTC-IC) with relative sparing of analogous normal cells suggesting that these molecules could be therapeutic targets in AML.