The Anti-Melanoma Activity of Dinaciclib, a Cyclin-Dependent Kinase Inhibitor, Is Dependent on p53 Signaling

Brijal M Desai,Adina Vultur,Keiran S M Smalley,Keith T Flaherty,Jun Kong,Thierry-Thien K Nguyen,Clemens Krepler,Jessie Villanueva,Min Xiao,Katherine L Nathanson,Mercedes Lioni,Meenhard Herlyn

doi:10.1371/journal.pone.0059588

Abstract

Although cyclin dependent kinase (CDK)-2 is known to be dispensable for the growth of most tumors, it is thought to be important for the proliferation of melanoma cells, where its expression is controlled by the melanocyte-lineage specific transcription factor MITF. Treatment of a panel of melanoma cells with the CDK inhibitor dinaciclib led to a concentration-dependent inhibition of growth under both 2D adherent and 3D organotypic cell culture conditions. Dinaciclib targeted melanoma cell lines regardless of cdk2 or MITF levels. Inhibition of growth was associated with a rapid induction of G2/M cell arrest and apoptosis. Treatment of human melanoma mouse xenografts with dinaciclib led to tumor regression associated with reduced retinoblastoma protein phosphorylation and Bcl-2 expression. Further mechanistic studies revealed that dinaciclib induces p53 expression whilst simultaneously downregulating the expression of the anti-apoptotic factors Mcl-1 and XIAP. To clarify the role of p53 activation in the dinaciclib-induced cell death, we generated melanoma cell lines in which p53 expression was knocked down using a shRNA lentiviral vector. Knockdown of p53 completely abolished the induction of apoptosis seen following dinaciclib treatment as shown by a lack of annexin-V staining and caspase-3 cleavage. Altogether, these data show that dinaciclib induces apoptosis in a large panel of melanoma cell lines through a mechanism requiring p53 expression.

Highlights

The landscape of melanoma therapy has changed dramatically within the last few years with the discovery that the majority of melanomas harbor activating mutations in BRAF [1,2] and that virtually all melanomas exhibit constitutive activity in the MAP kinase pathway [3]
In an initial series of studies, we examined the relationship between CDK2/MITF mRNA and protein expression levels and response to treatment by dinaciclib
As BRAF/NRAS mutational status is thought to predict targeted therapy response in melanoma, we further sub-stratified the groups according to CDK2 melanoma cell lines that harbored mutations in either BRAF V600E or NRAS

Summary

Introduction

The landscape of melanoma therapy has changed dramatically within the last few years with the discovery that the majority of melanomas harbor activating mutations in BRAF [1,2] and that virtually all melanomas exhibit constitutive activity in the MAP kinase pathway [3]. Two novel drugs were approved for the treatment of metastatic melanoma and several more are in late stage clinical development [4]. In the class of small molecule kinase inhibitors, the BRAF inhibitor vemurafenib has shown unprecedented clinical activity; but patients eventually develop resistance [5]. The field is increasingly accepting the notion that novel targets outside of the MAPK pathway will be necessary to avoid emergence of resistant cell subpopulations and that combinations of inhibitors are the most likely to provide long lasting results [10]. Increased efforts are focused on finding agents that are able to induce melanoma cell death and that may be advantageous in combination therapies with inhibitors of the MAPK pathway. There are 50% of melanoma patients whose tumors do not harbor BRAF mutations and for whom no effective therapies are currently available

Methods

Results

Conclusion