Data from <i>CDK4</i> Amplification Reduces Sensitivity to CDK4/6 Inhibition in Fusion-Positive Rhabdomyosarcoma

Abstract

<div>Abstract<p><b>Purpose:</b> Rhabdomyosarcoma (RMS) is the most common pediatric soft tissue sarcoma and includes a <i>PAX3–</i> or <i>PAX7–FOXO1</i> fusion-positive subtype. Amplification of chromosomal region 12q13–q14, which contains the <i>CDK4</i> proto-oncogene, was identified in an aggressive subset of fusion-positive RMS. CDK4/6 inhibitors have antiproliferative activity in <i>CDK4</i>-amplified liposarcoma and neuroblastoma, suggesting CDK4/6 inhibition as a potential therapeutic strategy in fusion-positive RMS.</p><p><b>Experimental Design:</b> We examined the biologic consequences of CDK4 knockdown, CDK4 overexpression, and pharmacologic CDK4/6 inhibition by LEE011 in fusion-positive RMS cell lines and xenografts.</p><p><b>Results:</b> Knockdown of CDK4 abrogated proliferation and transformation of 12q13–14-amplified and nonamplified fusion-positive RMS cells via G<sub>1</sub>-phase cell-cycle arrest. This arrest was mediated by reduced RB phosphorylation and E2F-responsive gene expression. Significant differences in E2F target expression, cell-cycle distribution, proliferation, or transformation were not observed in RMS cells overexpressing CDK4. Treatment with LEE011 phenocopied CDK4 knockdown, decreasing viability, RB phosphorylation, and E2F-responsive gene expression and inducing G<sub>1</sub>-phase cell-cycle arrest. Although all fusion-positive cell lines showed sensitivity to CDK4/6 inhibition, there was diminished sensitivity associated with CDK4 amplification and overexpression. This variable responsiveness to LEE011 was recapitulated in xenograft models of <i>CDK4</i>-amplified and nonamplified fusion-positive RMS.</p><p><b>Conclusions:</b> Our data demonstrate that CDK4 is necessary but overexpression is not sufficient for RB–E2F–mediated G<sub>1</sub>-phase cell-cycle progression, proliferation, and transformation in fusion-positive RMS. Our studies indicate that LEE011 is active in the setting of fusion-positive RMS and suggest that low CDK4-expressing fusion-positive tumors may be particularly susceptible to CDK4/6 inhibition. <i>Clin Cancer Res; 21(21); 4947–59. ©2015 AACR</i>.</p><p><i>See related commentary by Gatz and Shipley, p. 4750</i></p></div>