Retinoic acid and TGF-β signalling cooperate to overcome MYCN-induced retinoid resistance

Abstract

BackgroundRetinoid therapy is widely employed in clinical oncology to differentiate malignant cells into their more benign counterparts. However, certain high-risk cohorts, such as patients with MYCN-amplified neuroblastoma, are innately resistant to retinoid therapy. Therefore, we employed a precision medicine approach to globally profile the retinoid signalling response and to determine how an excess of cellular MYCN antagonises these signalling events to prevent differentiation and confer resistance.MethodsWe applied RNA sequencing (RNA-seq) and interaction proteomics coupled with network-based systems level analysis to identify targetable vulnerabilities of MYCN-mediated retinoid resistance. We altered MYCN expression levels in a MYCN-inducible neuroblastoma cell line to facilitate or block retinoic acid (RA)-mediated neuronal differentiation. The relevance of differentially expressed genes and transcriptional regulators for neuroblastoma outcome were then confirmed using existing patient microarray datasets.ResultsWe determined the signalling networks through which RA mediates neuroblastoma differentiation and the inhibitory perturbations to these networks upon MYCN overexpression. We revealed opposing regulation of RA and MYCN on a number of differentiation-relevant genes, including LMO4, CYP26A1, ASCL1, RET, FZD7 and DKK1. Furthermore, we revealed a broad network of transcriptional regulators involved in regulating retinoid responsiveness, such as Neurotrophin, PI3K, Wnt and MAPK, and epigenetic signalling. Of these regulators, we functionally confirmed that MYCN-driven inhibition of transforming growth factor beta (TGF-β) signalling is a vulnerable node of the MYCN network and that multiple levels of cross-talk exist between MYCN and TGF-β. Co-targeting of the retinoic acid and TGF-β pathways, through RA and kartogenin (KGN; a TGF-β signalling activating small molecule) combination treatment, induced the loss of viability of MYCN-amplified retinoid-resistant neuroblastoma cells.ConclusionsOur approach provides a powerful precision oncology tool for identifying the driving signalling networks for malignancies not primarily driven by somatic mutations, such as paediatric cancers. By applying global omics approaches to the signalling networks regulating neuroblastoma differentiation and stemness, we have determined the pathways involved in the MYCN-mediated retinoid resistance, with TGF-β signalling being a key regulator. These findings revealed a number of combination treatments likely to improve clinical response to retinoid therapy, including co-treatment with retinoids and KGN, which may prove valuable in the treatment of high-risk MYCN-amplified neuroblastoma.Electronic supplementary materialThe online version of this article (doi:10.1186/s13073-017-0407-3) contains supplementary material, which is available to authorized users.

Targeted inhibition of histone H3K27 demethylation is effective in high-risk neuroblastoma

“…We found that a large subset of genes affected by GSK-J4 treatment was similarly affected by RA (Fig. 3A and table S7) (39). We confirmed that this gene overlap was statistically significant both through Fisher’s exact test and gene set enrichment analysis (GSEA) for both the RA up- and down-regulated gene sets (fig.…”

Neuroblastoma: Genetic, Epigenetic Mechanisms and Targeted Therapy Approaches: Review

“…LMO4, CYP26A1, ASCL1, RET, FZD7 ve DKK1 genlerinin MYCN' nin artmış ekspresyonu ile tetiklenmesi ve TGF-β sinyal yolağını indüklemesi güncel bir retinoik asit direnç mekanizmasını tanımlamaktadır. 67 MYCN, mitokondriyal biyogenezi düzenleyen PPA, RGC1A, TFAM genlerinin ve mitokondriyal dinamikleri düzenleyen OPA1, MFN2, DRP1 genlerinin deregülasyonuyla apoptozu inhibe ederek, platin bileşiklerine karşı direnç açısından kritik farklı bir rol oynamaktadır. 68 Farklı yollarla gerçekleşen direnç mekanizmalarını belirleme çalışmaları hâlen devam etmektedir.…”

Molecular Approach to Neuroblastoma

“…A novel retinoic acid resistance mechanism includes LIM domain only 4 (LMO4), cytochrome P450 family 26 subfamily A member 1 (CYP26A1), achaete-scute family BHLH transcription factor 1 (ASCL1), ret protooncogene (RET), frizzled class receptor 7 (FZD7), and dickkopf WNT signaling pathway inhibitor 1 (DKK1) genes are triggered by MYCN overexpression. These causes may lead to targeting of TGF-β signaling pathway, leading to resistance [67]. MYCN plays a different critical role in resistance to platinum compounds by inhibiting apoptosis through deregulation of PPARG coactivator 1 alpha (PPARGC1A), transcription factor A, mitochondrial (TFAM) genes regulating mitochondrial biogenesis and mitochondrial dynamin-like GTPase (OPA1), mitofusin 2 (MFN2), dynamin 1-like (DRP1) genes regulating mitochondrial dynamics [68].…”