CRISPR/Cas9 mutagenesis invalidates a putative cancer dependency targeted in on-going clinical trials.

Ann Lin,Nicole M Sayles,Jason M Sheltzer,Christopher J Giuliano

doi:10.7554/elife.24179

Abstract

The Maternal Embryonic Leucine Zipper Kinase (MELK) has been reported to be a genetic dependency in several cancer types. MELK RNAi and small-molecule inhibitors of MELK block the proliferation of various cancer cell lines, and MELK knockdown has been described as particularly effective against the highly-aggressive basal/triple-negative subtype of breast cancer. Based on these preclinical results, the MELK inhibitor OTS167 is currently being tested as a novel chemotherapy agent in several clinical trials. Here, we report that mutagenizing MELK with CRISPR/Cas9 has no effect on the fitness of basal breast cancer cell lines or cell lines from six other cancer types. Cells that harbor null mutations in MELK exhibit wild-type doubling times, cytokinesis, and anchorage-independent growth. Furthermore, MELK-knockout lines remain sensitive to OTS167, suggesting that this drug blocks cell division through an off-target mechanism. In total, our results undermine the rationale for a series of current clinical trials and provide an experimental approach for the use of CRISPR/Cas9 in preclinical target validation that can be broadly applied.

Highlights

Tumors of the breast can be divided into five distinct subtypes based on characteristic gene expression patterns
Over a dozen previous publications have reported that Maternal Embryonic Leucine Zipper Kinase (MELK) is a cancer dependency, as blocking MELK with RNAi or small molecules inhibited the proliferation of cell lines derived from multiple tumor types (Gray et al, 2005; Lin et al, 2007; Kuner et al, 2013; Du et al, 2014; Kig et al, 2013; Speers et al, 2016; Alachkar et al, 2014; Marie et al, 2008; Nakano et al, 2008; Hebbard et al, 2010; Wang et al, 2014; Choi et al, 2011; Xia et al, 2016; Gu et al, 2013)
As a mitotic kinase highly expressed in many cancer types, MELK has been identified as a promising target for therapeutic intervention

Summary

Introduction

Tumors of the breast can be divided into five distinct subtypes based on characteristic gene expression patterns. These breast cancer subtypes are referred to as Luminal A, Luminal B, Her2-enriched, normal-like, and basal (Sørlie et al, 2001). Tumors that lack expression of ER, PR, or HER2 are referred to as ‘triple-negative’ breast cancers, and are irresponsive to hormonal or anti-HER2 therapies that have proven effective against receptor-positive cancers. Due to their resistance to targeted therapies as well as their rapid rate of cell division, basal breast cancers currently have the worst prognosis of any breast cancer subtype. There is an urgent need to develop new therapies that are effective against triple-negative or basal-type tumors

Methods

Results

Conclusion