In B-cell acute lymphoblastic leukemia (B-ALL), activation of Notch signaling leads to cell-cycle arrest and apoptosis. We aimed to harness knowledge acquired by understanding a mechanism of Notch-induced cell death to elucidate a therapeutically viable target in BALL. To this end, we identified that Notch activation suppresses Polo-like kinase 1 (PLK1) in a BALL specific manner. We identified that PLK1 is expressed in all subsets of BALL and is highest in Philadelphia-like (Ph-like) ALL, a high-risk subtype of disease. We biochemically delineated a mechanism of Notch-induced PLK1 downregulation that elucidated stark regulation of p53 in this setting. Our findings identified a novel posttranslational cascade initiated by Notch in which CHFR was activated via PARP1-mediated PARylation, resulting in ubiquitination and degradation of PLK1. This led to hypophosphorylation of MDM2 Ser260 , culminating in p53 stabilization and upregulation of BAX. shRNA knockdown or pharmacologic inhibition of PLK1 using BI2536 or BI6727 (volasertib) in BALL cell lines and patient samples led to p53 stabilization and cell death. These effects were seen in primary human BALL samples in vitro and in patient-derived xenograft models in vivo. These results highlight PLK1 as a viable therapeutic target in BALL. Efficacy of clinically relevant PLK1 inhibitors in BALL patient-derived xenograft mouse models suggests that use of these agents may be tailored as an additional therapeutic strategy in future clinical studies.

<p>Primary Ph-like B-ALL (Pt.B1) was treated with PLK1 inhibitor volasertib (100 nm) for 30 min. Cells were harvested and subjected to single cell time of flight mass cytometry (CyTOF) and analyzed using the Spanning Tree Progression of Density Normalized Events (SPADE) algorithm as previously described (ref). SPADE analysis shows an increased median level expression of the p53 protein and the pro-apoptotic Bax protein in cells subjected to PLK1 inhibition.</p>

<div>Abstract<p>In B-cell acute lymphoblastic leukemia (B-ALL), activation of Notch signaling leads to cell-cycle arrest and apoptosis. We aimed to harness knowledge acquired by understanding a mechanism of Notch-induced cell death to elucidate a therapeutically viable target in B-ALL. To this end, we identified that Notch activation suppresses Polo-like kinase 1 (PLK1) in a B-ALL–specific manner. We identified that PLK1 is expressed in all subsets of B-ALL and is highest in Philadelphia-like (Ph-like) ALL, a high-risk subtype of disease. We biochemically delineated a mechanism of Notch-induced PLK1 downregulation that elucidated stark regulation of p53 in this setting. Our findings identified a novel posttranslational cascade initiated by Notch in which CHFR was activated via PARP1-mediated PARylation, resulting in ubiquitination and degradation of PLK1. This led to hypophosphorylation of MDM2^Ser260, culminating in p53 stabilization and upregulation of BAX. shRNA knockdown or pharmacologic inhibition of PLK1 using BI2536 or BI6727 (volasertib) in B-ALL cell lines and patient samples led to p53 stabilization and cell death. These effects were seen in primary human B-ALL samples <i>in vitro</i> and in patient-derived xenograft models <i>in vivo</i>. These results highlight PLK1 as a viable therapeutic target in B-ALL. Efficacy of clinically relevant PLK1 inhibitors in B-ALL patient-derived xenograft mouse models suggests that use of these agents may be tailored as an additional therapeutic strategy in future clinical studies.</p></div>

<p>Primary Ph-like B-ALL (Pt.B1) was treated with PLK1 inhibitor volasertib (100 nm) for 30 min. Cells were harvested and subjected to single cell time of flight mass cytometry (CyTOF) and analyzed using the Spanning Tree Progression of Density Normalized Events (SPADE) algorithm as previously described (ref). SPADE analysis shows an increased median level expression of the p53 protein and the pro-apoptotic Bax protein in cells subjected to PLK1 inhibition.</p>

<p>U133A microarray (Affymetrix) gene expression data for 172 B-ALL patients, 34 T-ALL patients.</p>