A magic bullet for targeting cancers with supernumerary centrosomes

Abstract

Centrosome amplification is frequently observed in aggressive human cancers, and it has been proposed as a promising target in cancer therapy. Work by Mariappan et al (2019) in this issue details the identification of a small‐molecule inhibitor that perturbs the CPAP–tubulin interaction, prevents efficient centrosome clustering during mitosis, and exhibits in vivo anti‐tumor activity. This study demonstrates the potential of developing tumor‐specific treatment by specifically targeting centrosome amplification.

As the primary microtubule‐organizing center of cells, centrosomes play primordial roles in a number of cellular processes that include cell signaling, motility, and the regulation of cell shape and polarity. Centrosomes are composed of an orthogonally arranged pair of ninefold symmetric centrioles, surrounded by pericentriolar material (PCM). PCM harbors cell cycle regulators and proteins involved in microtubule nucleation and mitotic spindle assembly, contributing to a commitment of accurate cell division (Nigg & Holland, 2018). To maintain those functions, centrosome must be duplicated once and only once per cell cycle, and alterations in this process contribute to many human disorders including cancer and microcephaly (Nigg & Holland, 2018).

A wide range of human cancers display prominent numerical and structural centrosomal aberrations (Prosser & Pelletier, 2017). Centrosome loss, through a failure of centriole duplication, in normal cells leads to robust cell cycle arrest, whereas cancer cells can survive and circumvent this surveillance pathway by inhibiting p53 function (Lambrus & Holland, 2017). In contrast, centrosome amplification is frequently found in solid tumors and it is well established that centrosome amplification can lead to the formation of multipolar spindles, chromosome segregation errors, and an increase in aneuploidy. Those abnormalities in healthy cells lead to mitotic spindle assembly checkpoint (SAC) activation, which in turn leads to the activation of a so‐called mitotic catastrophe pathway and induced cell death. Cancer cells can bypass the SAC by clustering extra centrosomes into two spindle poles thereby achieving seemingly normal bipolar division. However, centrosome clustering mechanisms are error‐prone, causing merotelic chromosome attachments (e.g., chromosomes captured from microtubules emanating from the same spindle pole) which result in low rates of aneuploidy and genome instability which subsequently can contribute to tumor formation (Ganem et al, 2009). Surprisingly, it still remains unclear whether centrosome amplification can alone drive tumorigenesis or if simply a passenger phenotype. Toward this, a recent study nicely shows that centrosome amplification by itself is sufficient to trigger aneuploidy and promote the initiation of tumors in in vivo mouse model, when p53 pathway is partially inactivated (Levine et al, 2017). Intriguingly, centrosome amplification has been linked to another cellular‐invasion mechanism through increasing centrosomal microtubule nucleation (Godinho et al, 2014). Those findings support the notion that centrosome amplification plays a causative role in tumor initiation and progression, which has been found as a potential target to treat cancer.

In this context, inhibition of centrosome clustering is proposed as an attractive therapeutic strategy against cancer cells with amplified centrosomes. In the past decades, several studies report that a set of microtubule‐based motors and microtubule‐bundling proteins that maintain spindle poles are responsible for centrosome clustering, such as HSET (microtubule plus‐end motor), NUMA (nuclear mitotic apparatus), dynein (microtubule minus‐end motor), KIF2C (centromere‐associated kinesin), and TACC3–chTOG complex (spindle microtubule stabilizers) (Leber et al, 2010). Genetic and chemical perturbation of those factors has been described to inhibit centrosome clustering and trigger apoptosis in cancer cells with supernumerary centrosomes (Leber et al, 2010). However, it still remains elusive about how this protective mechanism actually operates in those tumor cells. It also remains unknown whether an intrinsic centrosomal mechanism involves in centrosome clustering that can be manipulated in cancer cells exhibiting extra centrosomes.

Mitotic centrosomal PCM undergoes a dramatic expansion to achieve a concomitant increase in microtubule nucleation capacity when compared to interphase PCM that remains inactive (Piehl et al, 2004; Lawo et al, 2012). In this issue, Mariappan et al (2019) further explored the interplay between the microtubule nucleating activity of centrosomes and their clustering during mitosis. The authors hypothesized that enhancing interphase microtubule nucleation may inhibit the following mitotic centrosome clustering. To test this, the authors utilized their previously characterized CPAP F375A mutant that is defective for tubulin binding, and its overexpression has been shown to increase the level of interphase PCM and thus enhance microtubule nucleation (Gopalakrishnan et al, 2012; Zheng et al, 2016). In an elegant time‐lapse experiment, they found that expressing CPAP F375A mutant in extra centrosome containing MCF10A cells significantly increased microtubule nucleation in G2 phase and prevented centrosome clustering during mitosis, ultimately leading to mitotic delay and cell death. Similar defects have been observed in breast cancer MDA‐MB‐231 and NSCLC cells with amplified centrosomes. Importantly, this manipulation has no adverse effect on normal MCF10A cells with two centrosomes, suggesting that CPAP–tubulin interaction could serve as a selective therapeutic target for supercentrosomal cancer cells.

Based on those findings, the authors performed a high‐throughput small‐molecule screen, to hunt for compounds that would specifically disrupt CPAP–tubulin interaction, and the most promising candidate molecules were further tested for their impact on centrosome clustering during mitosis. As a result, a compound of increased solubility (CCB02) was synthesized where the alkylamino residue was replaced with a methoxy group (Fig 1). Mariappan et al (2019) then showed that this compound has the ability to significantly enhance interphase centrosome‐based microtubule nucleation and to inhibit mitotic centrosome clustering. NMR and ITC assays revealed that CCB02 is a novel tubulin binder that effectively competes with CPAP for the same binding site on tubulin, although it remains a possibility that CCB02 might also compete for binding with additional microtubule binding proteins to impact centrosome clustering. The authors assessed the impact of CCB02 on several extra centrosomes containing cancer lines when cultured in 2D and 3D. In a short‐term treatment, CCB02 is sufficient to perturb centrosome clustering, leading to multipolar mitosis with apparent cell cycle delay. Consistently, a long‐term CCB02 treatment would only allow the survival of cancer cells with two centrosomes. Those findings suggest that CCB02 is a new drug that selectively eliminates cancer cells with amplified centrosomes. A final mouse xenograft experiment further supports this notion that CCB02 exhibits a significant in vivo anti‐tumor activity. However, it still remains unclear about the underlying mechanism to account for CCB02's manipulation on cancer. It will also be of importance, for example, to consider its alternative mechanism on cancer stem cells, investigate CCB02 in vivo off‐target effects, and optimize the drug delivery system in order to enhance its impact on tumor growth.

Figure 1. Impact of the CCB02 inhibitor on cancer cells with extra centrosomes.

CCB02 treatment disrupts the CPAP–tubulin interaction thereby increasing interphase microtubule nucleation through increased PCM recruitment around centrosomes. This activation further inhibits centrosome clustering during mitosis, leading to mitotic delay and cell death.

In summary, this study by Mariappan et al (2019) describes a novel small molecule with the potential to selectively target cancer cells with amplified centrosomes without harming normal cells. Importantly, this compound can also be used as a research tool to further decipher the molecular basis of centrosome clustering and its role in tumorigenesis. This work also suggests the tantalizing possibility that CCB02 treatment along with other anti‐tumor agents might help improve treatment of malignant and resistant cancers.

The EMBO Journal (2019) 38: e101134