Combination therapy with the CDK7 inhibitor and the tyrosine kinase inhibitor exerts synergistic anticancer effects against MYCN‐amplified neuroblastoma

Abstract

Patients with neuroblastoma due to MYCN oncogene amplification and consequent N‐Myc oncoprotein overexpression have very poor prognosis. The cyclin‐dependent kinase 7 (CDK7)/super‐enhancer inhibitor THZ1 suppresses MYCN gene transcription, reduces neuroblastoma cell proliferation, but does not cause significant cell death. The protein kinase phosphatase 1 nuclear targeting subunit (PNUTS) has recently been shown to interact with c‐Myc protein and suppresses c‐Myc protein degradation. Here we screened the U.S. Food and Drug Administration‐Approved Oncology Drugs Set V from the National Cancer Institute, and identified tyrosine kinase inhibitors (TKIs), including ponatinib and lapatinib, as the Approved Oncology Drugs exerting the best synergistic anticancer effects with THZ1 in MYCN‐amplified neuroblastoma cells. Combination therapy with THZ1 and ponatinib or lapatinib synergistically induced neuroblastoma cell apoptosis, while having little effects in normal nonmalignant cells. Differential gene expression analysis identified PNUTS as one of the genes most synergistically reduced by the combination therapy. Reverse transcription polymerase chain reaction and immunoblot analyses confirmed that THZ1 and the TKIs synergistically downregulated PNUTS mRNA and protein expression and reduced N‐Myc protein but not N‐Myc mRNA expression. In addition, PNUTS knockdown resulted in decreased N‐Myc protein but not mRNA expression and decreased MYCN‐amplified neuroblastoma cell proliferation and survival. As CDK7 inhibitors are currently under clinical evaluation in patients, our data suggest the addition of the TKI ponatinib or lapatinib in CDK7 inhibitor clinical trials in patients.

Super-enhancer in prostate cancer: transcriptional disorders and therapeutic targets

“…CDK7 activates MED1 via ligand-specific phosphorylation, and the IDRs (intrinsically disordered region) of MED1 form phase-separation properties to be recruited to the ARbound SEs, resulting in the high-density assembly of the SE-related transcription apparatus. CDK7 inhibition has been effectively tested in several aggressive cancer types, including MYCN-amplified neuroblastoma, T-cell acute lymphoblastic leukemia, triplenegative breast cancer, and small-cell lung cancer 50,[57][58][59][60] . The CDK7-related SEs mediate the recruitment of AR and RNA polymerase II to boost the expression of a host of target genes, known as the "Achilles cluster" genes, thus becoming transcriptional addictive and sensitive to CDK7 inhibition 50 .…”

The RNA‐helicase DDX21 upregulates CEP55 expression and promotes neuroblastoma

“…Cell lines were validated and maintained as previously described [19]. Briefly, BE(2)‐C and SHEP‐21N neuroblastoma cells were maintained in Dulbecco's Modified Eagle's medium (DMEM) containing 10% FBS.…”

Neuroblastoma and the epigenome

“…Moreover, genetic ablation of PNUTS resulted in decreased MYCN protein, but not RNA expression. This was associated with reduced cell proliferation and cell survival, indicating another potential drug for use as an inhibitor of MYCN [158].…”