NOX2 oxidase expressed in endosomes promotes cell proliferation and prostate tumour development.

Ian P Harrison,Ian R.D Johnson,Christopher G Sobey,Tony Tiganis,Raymond Luong,Doug A Brooks,Antony Vinh,Stavros Selemidis,Elizabeth D Williams,Grant R Drummond,John J O’Leary

doi:10.18632/oncotarget.26237

Abstract

Reactive oxygen species (ROS) promote growth factor signalling including for VEGF-A and have potent angiogenic and tumourigenic properties. However, the precise enzymatic source of ROS generation, the subcellular localization of ROS production and cellular targets in vivo that influence tumour-promoting processes, are largely undefined. Here, using mRNA microarrays, we show increased gene expression for NOX2, the catalytic subunit of the ROS-generating NADPH oxidase enzyme, in human primary prostate cancer compared to non-malignant tissue. In addition, NOX4 gene expression was markedly elevated in human metastatic prostate cancers, but not in primary prostate tumours. Using a syngeneic, orthotopic mouse model of prostate cancer the genetic deletion of NOX2 (i.e. NOX2-/y mouse) resulted in reduced angiogenesis and an almost complete failure in tumour development. Furthermore, pharmacological inhibition of NOX2 oxidase suppressed established prostate tumours in mice. In isolated endothelial cells, and in human normal and prostate cancer cells, NOX2 co-located to varying degrees with early endosome markers including EEA1, Appl1 and Rab5A and the late endosome marker Rab7A, and this correlated with significant VEGF-A-dependent ROS production within acidified endosomal compartments and endothelial cell proliferation that was NOX2 oxidase- and hydrogen peroxide dependent. We concluded that NOX2 oxidase expression and endosomal ROS production were important for prostate cancer growth and that this was required to positively regulate the VEGF pathway. The research provides a paradigm for limiting tumour growth through a better understanding of NOX2 oxidase's effect on VEGF signalling and how controlling the development of tumour vasculature can limit prostate tumour development and metastasis.

Highlights

A new paradigm in cancer cell research is that vascular endothelial growth factor VEGF signalling is dependent on receptor internalisation into early endosomes
NOX2 oxidase was expressed at higher levels in human primary prostate cancers when compared to normal tissue, suggesting that this Reactive oxygen species (ROS) generating enzyme plays a role in tumour development
Using a mouse model of prostate cancer, we show evidence that NOX2 oxidase is crucial for angiogenesis and tumour development

Summary

Introduction

A new paradigm in cancer cell research is that vascular endothelial growth factor VEGF signalling is dependent on receptor internalisation into early endosomes. Delays in the trafficking of VEGFR2-containing endosomes leads to VEGFR2 dephosphorylation, resulting in receptor de-activation and decreased arterial morphogenesis [18]. In both vascular and non-vascular cells, VEGFR2 undergoes constitutive recycling between peripheral endosomes and the cell surface [16]. VEGF in turn stimulates the production of reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (H2O2) that promote cell proliferation and angiogenesis [32, 33]. H2O2 stimulates VEGF production in vascular smooth muscle cells [26], and upregulates VEGF mRNA to induce endothelial cell proliferation and migration [34]. The exact enzymatic source of ROS generation and the subcellular location of ROS production remain largely unknown

Methods

Results

Conclusion