Figure 4.
Activation of SASP and interplay with immune cells upon PLK4 inhibition in AML. (A) Volcano plots showing differentially expressed cytokines (log2FC > 1 or < −1, false discovery rate < 0.05) for each sample pair on day 2 post CFI-400945 treatment (20 nM). (B) GSEA showed positive enrichment of IFN-γ and SASP upon CFI-400945 treatment (20 nM) compared with vehicle control on day 2. (C) Heat map summarizing the ELISA results upon CFI-400945 treatment (10 nM) in K052. The concentration of CXCL8, CCL2, IL-6, IFN-γ, and TNF-α in media cultured with T2N, T4N, and polyploidy of K052 were upregulated, whereas IL-1β was only upregulated in the media from polyploidy cells (supplemental Figure 5D) (n = 3). (D) Addition of IL-1β–neutralizing antibody (0.3 μg/mL) could ameliorate CFI-400945–induced CCL2 and CXCL8 in K052 (n = 3). (E-F) CFI-400945 treatment (10 nM) increased STING dimerization (E) and 2’3’-cGAMP levels (F) in K052 at day 2 (n = 3). (G-H) CFI-400945 (10 nM) increased senescence (G) and expression of CCL2 and CXCL8 (H) on day 2, and the response was ameliorated by cGAS (RU521, 1.6 μM) or STING inhibitors (H-151, 800 nM) (n = 3). (I-K) CFI-400945 treatment (10 nM) increased expression of CCL2 and CXCL8 (I), STING dimerization (J), and cGAMP (K) on day 2, which was rescued by EED226 (PRC2i, 500 nM) (n = 3). (L) CFI-400945 increased the formation of cytoplasmic chromatin in K052 that could be ameliorated by PRC2 inhibitor (supplemental Figure 4I) (n = 3). (M) CFI-400945 treatment (10 nM) increased the secretion of CCL2, CXCL8, and IL-1β in K052 but less in ML2 (n = 3), which was rescued by EED226 (PRC2i, 500 nM). (N) THP-1–derived macrophage showed higher phagocytosis against 2-day CFI-400945–pretreated (10 nM) K052 compared with vehicle-treated K052, which could be ameliorated by RU521 (cGASi, 1.6 μM) and H-151 (STINGi, 800 nM) (n = 3). (O) Activated T cells induced more apoptosis in CFI-400945–pretreated (10 nM) K052 compared with vehicle-treated K052 at day 2 (n = 3). (P) Therapeutic inhibition of RU521 (cGASi, 1.6 μM) and H-151 (STINGi, 800 nM) diminished the CFI-400945 (10 nM) and T cell–induced apoptosis in K052 (n = 3). (Q-R) ML2 isogenic system showed that TP53-knockin line showed relatively higher phagocytosis and (R) T-cell–mediated apoptosis (Q). Phagocytosis and AML cell apoptosis were normalized with respect to those in coculture of ML2 cells with macrophages (Q) and T cells (R) in the vehicle control. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001.