The HDAC6/8/10 inhibitor TH34 induces DNA damage-mediated cell death in human high-grade neuroblastoma cell lines

Fiona R Kolbinger,Nikolas Gunkel,Olaf Witt,Ina Oehme,Wolfgang Sippl,Emily Koeneke,Aubry K Miller,Tino Heimburg,Frank Westermann,Michael Müller,Theresa Bayer,Johannes Ridinger,Manfred Jung

doi:10.1007/s00204-018-2234-8

Abstract

High histone deacetylase (HDAC) 8 and HDAC10 expression levels have been identified as predictors of exceptionally poor outcomes in neuroblastoma, the most common extracranial solid tumor in childhood. HDAC8 inhibition synergizes with retinoic acid treatment to induce neuroblast maturation in vitro and to inhibit neuroblastoma xenograft growth in vivo. HDAC10 inhibition increases intracellular accumulation of chemotherapeutics through interference with lysosomal homeostasis, ultimately leading to cell death in cultured neuroblastoma cells. So far, no HDAC inhibitor covering HDAC8 and HDAC10 at micromolar concentrations without inhibiting HDACs 1, 2 and 3 has been described. Here, we introduce TH34 (3-(N-benzylamino)-4-methylbenzhydroxamic acid), a novel HDAC6/8/10 inhibitor for neuroblastoma therapy. TH34 is well-tolerated by non-transformed human skin fibroblasts at concentrations up to 25 µM and modestly impairs colony growth in medulloblastoma cell lines, but specifically induces caspase-dependent programmed cell death in a concentration-dependent manner in several human neuroblastoma cell lines. In addition to the induction of DNA double-strand breaks, HDAC6/8/10 inhibition also leads to mitotic aberrations and cell-cycle arrest. Neuroblastoma cells display elevated levels of neuronal differentiation markers, mirrored by formation of neurite-like outgrowths under maintained TH34 treatment. Eventually, after long-term treatment, all neuroblastoma cells undergo cell death. The combination of TH34 with plasma-achievable concentrations of retinoic acid, a drug applied in neuroblastoma therapy, synergistically inhibits colony growth (combination index (CI) < 0.1 for 10 µM of each). In summary, our study supports using selective HDAC inhibitors as targeted antineoplastic agents and underlines the therapeutic potential of selective HDAC6/8/10 inhibition in high-grade neuroblastoma.

Highlights

Neuroblastoma is the most common extracranial solid tumor in childhood and the most frequently occurring cancer in infancy, accounting for 15% of pediatric cancer mortality (Brodeur 2003; Ward et al 2014)
The histone deacetylase (HDAC) inhibitor TH34 selectively inhibits HDACs 6, 8 and 10. As no such compound has been developed to date, we sought to develop a novel HDAC inhibitor with high selectivity for HDAC8 and HDAC10 over HDACs 1, 2 and 3, and tolerated HDAC6 inhibition during the screening process (Fig. 1a, b)
To further characterize the effects mediated by TH34, we examined the expression of the cell cycle inhibitor CDKN1A (p21WAF1/CIP1)

Summary

Introduction

Neuroblastoma is the most common extracranial solid tumor in childhood and the most frequently occurring cancer in infancy, accounting for 15% of pediatric cancer mortality (Brodeur 2003; Ward et al 2014). Its clinical presentation is diverse, including highly differentiated local tumors with an excellent prognosis, spontaneously regressing metastatic disease and chemotherapy-resistant, invasive masses, which are likely to relapse. Treatment regimens for high-risk tumors involve dose-intensive chemotherapy, surgical resection and a combination of immunotherapy, antibodies and 13-cis retinoic acid Despite high-intensity chemotherapy, overall survival in high-risk neuroblastoma remains poor and chemotherapy-related toxicities are commonly observed. Research has recently focused on the identification of novel, druggable targets and developing respective antineoplastic agents to abolish therapy resistance mechanisms and minimize chemotherapy-related adverse events

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