MYB fusions and CD markers as tools for authentication and purification of cancer stem cells from salivary adenoid cystic carcinoma

Alex Panaccione,Yi Zhang,Molly Ryan,Christopher A Moskaluk,Karen S Anderson,Wendell G Yarbrough,Sergey V Ivanov

doi:10.1016/j.scr.2017.05.002

Alex Panaccione, Yi Zhang + Show 5 more

Open Access

https://doi.org/10.1016/j.scr.2017.05.002

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Abstract

Cancer stem cells (CSC) are considered the major cause of aggressive tumor behavior, recurrence, metastases, and resistance to radiation, making them an attractive therapeutic target. However, isolation of CSC from tumor tissue and their characterization are challenging due to uncertainty about their molecular markers and conditions for their propagation. Adenoid cystic carcinoma (ACC), which arises predominantly in the salivary glands, is a slow-growing but relentless tumor that frequently invades nerves and metastasizes. New effective treatment approaches for ACC have not emerged over the last 40years. Previously, based on a highly conserved SOX10 gene signature that we identified in the majority of ACC tumors, we suggested the existence in ACC of SOX10+ cells with neural stem properties and corroborated this hypothesis via isolation from ACC tissue a novel population of CSC, termed ACC-CSC. These cells activated NOTCH1 signaling and co-expressed SOX10 and other ACC-intrinsic neural crest stem cell markers with CD133, a CSC cell surface marker, suggesting that ACC is driven by a previously uncharacterized population of SOX10+/CD133+ cells with neural stem cell properties. Here, we authenticated ACC identity of our primary cultures by demonstrating that most of them harbor MYB-NFIB fusions, which are found in 86% of ACC. We demonstrated using CyTOF, a novel mass cytometry technology, that these cells express high β-catenin and STAT3 levels and are marked by CD24 and CD44. Finally, to streamline development of ACC cell lines, we developed RT-PCR tests for distinguishing mouse and human cells and used immunomagnetic cell sorting to eliminate mouse cells from long-term cell cultures. Overall, this study describes a new population of CSC that activates signaling pathways associated with poor prognosis, validates their ACC identity, and optimizes approaches that can be used for purification of ACC-CSC and generation of cell lines.

Highlights

Adenoid cystic carcinoma (ACC) is a deadly cancer: with a prevalence rate of 1224 cases, 918 patients die from ACC in the U.S every year
In line with a special role of SOX10 in this cancer, we identified in the majority of ACC the expression of a highly conserved SOX10 gene signature that contained a cluster of neural stem cell drivers and markers, such as NOTCH1, MAP2, GPM6B, and FABP7, as well as genes/proteins involved in WNT and NOTCH signaling [13,14]
We asked if ACC cell cultures harbor MYB-NFIB fusions, which are ACC-intrinsic genetic aberrations found in 86% of ACC patients and considered to be primary ACCdriving events [18,19]

Summary

Introduction

Adenoid cystic carcinoma (ACC) is a deadly cancer: with a prevalence rate of 1224 cases, 918 patients die from ACC in the U.S every year (http://www.accoi.org/faq/acc-statistics/). ACC is treated by surgery with or without radiation, but only 40% of patients survive 15 years owing to intrinsic radiation resistance of ACC cells and their propensity to metastasize, relapse, and spread along nerves [1,2]. CSC possess properties of normal stem cells and are widely associated with invasion, recurrence, metastases, and resistance to cytotoxic therapies [4,5,6]. Their identification in ACC will advance understanding of molecular etiology and cell of origin, improving diagnostics, predicting disease outcome, and developing effective therapies

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