Preclinical Evaluation of the WEE1 Inhibitor MK-1775 as Single-Agent Anticancer Therapy

Abstract

Inhibition of the DNA damage checkpoint kinase WEE1 potentiates genotoxic chemotherapies by abrogating cell-cycle arrest and proper DNA repair. However, WEE1 is also essential for unperturbed cell division in the absence of extrinsic insult. Here, we investigate the anticancer potential of a WEE1 inhibitor, independent of chemotherapy, and explore a possible cellular context underlying sensitivity to WEE1 inhibition. We show that MK-1775, a potent and selective ATP-competitive inhibitor of WEE1, is cytotoxic across a broad panel of tumor cell lines and induces DNA double-strand breaks. MK-1775-induced DNA damage occurs without added chemotherapy or radiation in S-phase cells and relies on active DNA replication. At tolerated doses, MK-1775 treatment leads to xenograft tumor growth inhibition or regression. To begin addressing potential response markers for MK-1775 monotherapy, we focused on PKMYT1, a kinase functionally related to WEE1. Knockdown of PKMYT1 lowers the EC 50 of MK-1775 by five-fold but has no effect on the cell-based response to other cytotoxic drugs. In addition, knockdown of PKMYT1 increases markers of DNA damage, gH2AX and pCHK1 S345 , induced by MK-1775. In a post hoc analysis of 305 cell lines treated with MK-1775, we found that expression of PKMYT1 was below average in 73% of the 33 most sensitive cell lines. Our findings provide rationale for WEE1 inhibition as a potent anticancer therapy independent of a genotoxic partner and suggest that low PKMYT1 expression could serve as an enrichment biomarker for MK-1775 sensitivity.

Cytokinetic effects of Wee1 disruption in pancreatic cancer

“…The effects of MK-1775 on OCIP23 cells were much more dramatic than Wee1 knockdown, with the accumulation of cells in early S-phase showing H3S10 phosphorylation, associated with high levels of Cyclin A2 and Cyclin B1 that are normally highly expressed in late S-phase and G2. These effects have been previously observed, and attributed to premature entry of S-phase cells into mitosis, 19,20 which is consistent with the confocal microscopy features seen in Figure 2B. However, we did not observe them in the OCIP23 xenografts, even at the high dose of 80mg/kg (for comparison, the human maximum tolerated dose of 225mg twice daily 13 is roughly 6mg/kg), and it remains unclear if forced mitotic entry is a significant drug effect in humans.…”

“…18 Subsequently it was reported that treatment with the clinical Wee1 inhibitor MK-1775 (AZD-1775) likewise had striking cytokinetic effects, including features suggestive of premature entry into mitosis during early S-phase. 19,20 Of the Wee1 inhibitors, MK-1775 is currently the most advanced in clinical development, and it is reported to be highly selective toward Wee1 (21). As a single agent it appears to be well tolerated, but has little anti-cancer activity.…”

“…13 Several clinical trials are underway combining MK-1775 with conventional chemotherapy agents such as gemcitabine, where the effects relieving Weel 0 s restraint of Cdk2 during S-phase might be particularly relevant. 7,15,20,22 There is also a longstanding interest combining checkpoint inhibitors with radiotherapy, since this might differentially improve the radiation response of p53-deficient cancers, where the G1 checkpoint that normally facilitates repair is compromised. Here the anticipated effect of Wee1 inhibition would be to abrogate the G2 checkpoint, resulting in cells entering mitosis with unresolved radiation-induced chromosomal damage, and thereby undergoing death through mitotic catastrophe.…”

A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity

“…Inhibition of Wee1 potentiates DNA-damaging chemotherapeutics and radiotherapy and has cytotoxic effects as a single agent (19)(20)(21)(22). The consensus view is that Wee1 inhibition facilitates tumor cell killing through G 2 /M checkpoint inactivation, which would catalyze mitotic catastrophe (23)(24)(25).…”

Manipulating DNA damage-response signaling for the treatment of immune-mediated diseases

“…For this reason, it has been speculated that p53-deficient malignant cells are highly dependent on the S and G 2 /M checkpoints for maintaining their genomic integrity. Accordingly, a variety of kinase inhibitors have been developed as cancer therapeutics that inhibit CHK1, CHK2, or WEE1, the known enforcers of these later checkpoints (21,22). We speculated that even though normal T cells have intact p53, their extraordinarily rapid rate of division would also make them exquisitely dependent on the S and G 2 /M checkpoints for survival.…”