Inhibition of Polo-like Kinase 4 Induced Both Cell Intrinsic and Non-Cell Intrinsic Anti-Leukemia Effects on TP53-Mutated Acute Myeloid Leukemia

Acute myeloid leukemia (AML) carrying TP53 mutations is a distinct subtype characterized by genomic and chromosomal instability and a complex and monosomy karyotype (CK/MK). TP53-mutated AML portends an extremely grave prognosis and is refractory to conventional chemotherapy and allogeneic hematopoietic stem cells transplantation (HSCT). There is an unmet clinical need to develop novel therapeutic strategy for this disease.

In an in-silico analysis to identify specific gene signatures of TP53-mutated AML, Polo-Like Kinase 4 (PLK4) was found to be highly expressed in this AML subtype. PLK4 is the master regulator of centriole duplication and cytokinesis and has been investigated as a target for therapeutic intervention in cancers. We hypothesized that PLK4 inhibition may perturb the oncogenic pathway of TP53-mutated AML. Twelve-day treatment with PLK4 inhibitor CFI-400945 as well as gene knockout by CRISPR/Cas9 suppressed cellular proliferation of TP53-mutated AML cell lines in vitro. CFI-400945 treatment for two days induced DNA damage as evident by γH2AX staining and cellular senescence by β-galactosidase (SA-β-Gal) staining. There was progressive increase in DNA ploidy and number of microtubule organizing center (MTOC), consistent with defective cytokinesis, which was demonstrable by time-lapsed microscopy. Cytoplasmic chromatin could be readily identified as micronuclei in the polyploid cells.

Transcriptome analyses of TP53 mutated K052 cell line treated with CFI-400945 demonstrated induction of senescence-associated secretory phenotype (SASP), occurring before the onset of polyploidy. Quantitative RT-PCR and ELISA assay confirmed the increase of CCL2, CXCL8, IL10, IL6, TNF-α and IFN-γ upon CFI-400945 treatment. Both M1 macrophages and T-cells were activated upon co-culture with K052 in the presence of CFI-400945, as evident by the respective increases in phagocytic activity and intracellular IFN-γ/TNF-α. Mechanistically, CFI-400945 induced increase in STING dimers and cGAMP levels and cellular senescence could be ameliorated by STING and cGAS inhibitors. Therapeutically, CFI-400945 treatment in vivo significantly reduced bioluminescence of NSG mice transplanted with luciferase tagged K052 cells, resulting in prolonged survival of recipient animals. The in vivo effects could be potentiated by concomitant treatment with anti-CD47 monoclonal antibody (B6H12), which binds to human CD47 and blocks the interaction with macrophages receptor SIRPα, leading to phagocytosis of leukemic cells. Our results demonstrated that PLK4 inhibition exerted both cell intrinsic and non-cell intrinsic therapeutic effects on TP53 mutated AML with the latter being mediated by activation of cGAS/STING pathway hence both innate and adaptive immunity.

No relevant conflicts of interest to declare.