Abstract
Malignant peripheral nerve sheath tumors (MPNSTs) are soft-tissue sarcomas that frequently arise in patients with neurofibromatosis type 1 (NF1). Most of these tumors are unresectable at diagnosis and minimally responsive to conventional treatment, lending urgency to the identification of new pathway dependencies and drugs with potent antitumor activities. We therefore examined a series of candidate agents for their ability to induce apoptosis in MPNST cells arising in nf1/tp53-deficient zebrafish. In this study, we found that DNA topoisomerase I-targeted drugs and mTOR kinase inhibitors were the most effective single agents in eliminating MPNST cells without prohibitive toxicity. In addition, three members of these classes of drugs, either AZD2014 or INK128 in combination with irinotecan, acted synergistically to induce apoptosis both in vitro and in vivo. In mechanistic studies, irinotecan not only induces apoptosis by eliciting a DNA damage response, but also acts synergistically with AZD2014 to potentiate the hypophosphorylation of 4E-BP1, a downstream target of mTORC1. Profound hypophosphorylation of 4E-BP1 induced by this drug combination causes an arrest of protein synthesis, which potently induces tumor cell apoptosis. Our findings provide a compelling rationale for further in vivo evaluation of the combination of DNA topoisomerase I-targeted drugs and mTOR kinase inhibitors against these aggressive nerve sheath tumors.
Highlights
Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive, often metastatic soft-tissue sarcomas associated with neurofibromatosis type 1 (NF1) [1], an autosomal genetic dominant disorder with an incidence of 1 per 3000 individuals [2]
Primary zebrafish MPNST tumor cells are harvested from an nf1a+/−; nf1b−/−; p53m/m; sox10: mCherry zebrafish line, and microinjected into the pericardial cavity of pigmentless Casper recipient zebrafish embryos at 2 days post fertilization (Fig. 1)
This technique provides a uniform mass of tumor cells that can be readily visualized by fluorescence stereomicroscopy at 24 h after injection, in an anatomic region of the fish that is free of autofluorescence and other background signals (Fig. 1a) and is a reported site of both primary and metastatic human MPNSTs [23, 24]
Summary
Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive, often metastatic soft-tissue sarcomas associated with neurofibromatosis type 1 (NF1) [1], an autosomal genetic dominant disorder with an incidence of 1 per 3000 individuals [2]. The histologic features of the MPNSTs in zebrafish are very similar to those of the human tumors [15, 20] To optimize this model for in vivo imaging, we bred the fish to a stable transgenic line expressing mCherry, controlled by the zebrafish sox neural crestspecific promoter, resulting in fluorescent labeling of MPNST tumors in living zebrafish [15]. We show that the mechanisms underlying the synergy between these drugs center around the markedly enhanced inhibition of 4E-BP1 phosphorylation induced by the combination This results in the lack of free eIF4E and an arrest of 5ʹ cap-dependent protein synthesis, which promotes tumor cell apoptosis.
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