Preclinical and clinical development of agents that inhibit cell-cycle progression have brought an understanding of the feasibility of targeting various cell-cycle regulators in patients with cancer. Small molecule inhibitors targeting key proteins that participate in cell-cycle progression including the cyclin-dependent kinases and checkpoint kinases induce cell-cycle arrest and apoptosis in neoplastic cells. Early phase I studies demonstrate targeted inhibitors can be administered safely in adult and pediatric cancer patients, but these agents generally show limited clinical benefits as single agents. In this review, we discuss biological mechanisms that support dual combination strategies of cell-cycle inhibition with chemotherapeutic agents that are anticipated to achieve rationally targeted therapies for cancer patients. The rationale for evaluating these combination strategies is that DNA damage renders tumors highly responsive to irreversible cell-cycle arrest therapy. This approach is predicted to generate less intensive therapies and to maximize the efficacy of individual agents against solid tumors and hematologic malignancies. .

This review describes the pivotal roles of cell cycle and checkpoint regulators and discusses development of specific cell cycle inhibitors for therapeutic use for pediatric cancer. The mechanism of action as well as the safety and tolerability of drugs in pediatric patients, including compounds that target CDK4/CDK6 (palbociclib, ribociclib, abemaciclib), aurora kinases (AT9283 and MLN8237), Wee1 kinase (MK-1775), KSP (ispinesib) and tubulin (taxanes, vinca alkaloids), are presented. The design of mechanism-based combinations that exploit the crosstalk of signals activated by cell cycle arrest, as well as pediatric-focused drug development are critical for the advancement of drugs for rare childhood diseases.

MicroRNA-34a (miR-34a) is a master regulator of signaling networks that maintain normal physiology and disease and is currently in development as a miRNA-based therapy for cancer. Prior studies have reported low miR-34a expression in osteosarcoma (OS); however, the molecular mechanisms underlying miR-34a activity in OS are not well defined. Therefore, this study evaluated the role of miR-34a in regulating signal transduction pathways that influence cell death in OS. Levels of miR-34a were attenuated in human OS cells and xenografts of the Pediatric Preclinical Testing Consortium (PPTC). Bioinformatics predictions identified stathmin 1 (STMN1) as a potential miR-34a target. Biotin pulldown assay and luciferase reporter analysis confirmed miR-34a target interactions within the STMN1 mRNA 3′UTR. Overexpression of miR-34a in OS cells suppressed STMN1 expression and reduced cell growth in vitro. Restoration of miR-34a led to microtubule destabilization and increased βIII-tubulin expression, with corresponding G1/G2 phase cell cycle arrest and apoptosis. Knockdown of the Sp1 transcription factor, by siRNA silencing, also upregulated βIII-tubulin expression in OS cells, suggesting miR-34a indirectly affects Sp1. Validating the coordinating role of miR-34a in microtubule destabilization, when miR-34a was combined with either microtubule inhibitors or chemotherapy, STMN1 phosphorylation was suppressed and there was greater cytotoxicity in OS cells. These results demonstrate that miR-34a directly represses STMN1 gene and protein expression and upregulates βIII-tubulin, leading to disruption of the microtubule network and cell death. Implications The miR-34a/STMN1/βIII-tubulin axis maintains the microtubule cytoskeleton in osteosarcoma, and combining miR-34a with microtubule inhibitors can be investigated as a novel therapeutic strategy.

<p>RT-qPCR measurements of miR-34a in SaOS and 143B cells transfected with non-targeting miR-C (control) oligonucleotides or miR-34a mimics.</p>

<p>HOS and U2OS cells were transfected with miR-34a and Cyclin D1 and p27 protein expression were determined by immunoblot.</p>