In vitro and in vivo single-agent efficacy of checkpoint kinase inhibition in acute lymphoblastic leukemia

Ilaria Iacobucci, Andrea Ghelli Luserna Di Rorà, Maria Vittoria Verga Falzacappa, Claudio Agostinelli, Enrico Derenzini, Anna Ferrari, Cristina Papayannidis, Annalisa Lonetti, Simona Righi, Enrica Imbrogno, Silvia Pomella, Claudia Venturi, Viviana Guadagnuolo, Federica Cattina, Emanuela Ottaviani, Maria Chiara Abbenante, Antonella Vitale, Loredana Elia, Domenico Russo, Pier Luigi Zinzani, Stefano Pileri, Pier Giuseppe Pelicci, Giovanni Martinelli

Although progress in children, in adults, ALL still carries a dismal outcome. Here, we explored the in vitro and in vivo activity of PF-00477736 (Pfizer), a potent, selective ATP-competitive small-molecule inhibitor of checkpoint kinase 1 (Chk1) and with lower efficacy of checkpoint kinase 2 (Chk2). The effectiveness of PF-00477736 as single agent in B-/T-ALL was evaluated in vitro and in vivo studies as a single agent. The efficacy of the compound in terms of cytotoxicity, induction of apoptosis, and changes in gene and protein expression was assessed using different B-/T-ALL cell lines. Finally, the action of PF-00477736 was assessed in vivo using leukemic mouse generated by a single administration of the tumorigenic agent N-ethyl-N-nitrosourea. Chk1 and Chk2 are overexpressed concomitant with the presence of genetic damage as suggested by the nuclear labeling for γ-H2A.X (Ser139) in 68 % of ALL patients. In human B- and T-ALL cell lines, inhibition of Chk1/2 as a single treatment strategy efficiently triggered the Chk1-Cdc25-Cdc2 pathway resulting in a dose- and time-dependent cytotoxicity, induction of apoptosis, and increased DNA damage. Moreover, treatment with PF-00477736 showed efficacy ex vivo in primary leukemic blasts separated from 14 adult ALL patients and in vivo in mice transplanted with T-ALL, arguing in favor of its future clinical evaluation in leukemia. In vitro, ex vivo, and in vivo results support the inhibition of Chk1 as a new therapeutic strategy in acute lymphoblastic leukemia, and they provide a strong rationale for its future clinical investigation.