Prolyl Hydroxylase 3 Knockdown Accelerates VHL-Mutant Kidney Cancer Growth In Vivo.

Niki M Zacharias,Li Li,Steven W Millward,Jose A Karam,Neema Navai,Christopher G Wood,Tapati Maity,Lei Wang

doi:10.3390/ijms22062849

Niki M Zacharias, Li Li + Show 6 more

Open Access

https://doi.org/10.3390/ijms22062849

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Abstract

Von Hippel Lindau (VHL) inactivation, which is common in clear cell renal cell carcinoma (ccRCC), leads directly to the disruption of oxygen homoeostasis. VHL works through hypoxia-inducible factors (HIFs). Within this VHL-HIF system, prolyl hydroxylases (PHDs) are the intermediary proteins that initiate the degradation of HIFs. PHD isoform 3′s (PHD3) role in ccRCC growth in vivo is poorly understood. Using viral transduction, we knocked down the expression of PHD3 in the human ccRCC cell line UMRC3. Compared with control cells transduced with scrambled vector (UMRC3-SC cells), PHD3-knockdown cells (UMRC3-PHD3KD cells) showed increased cell invasion, tumor growth, and response to sunitinib. PHD3 knockdown reduced HIF2α expression and increased phosphorylated epidermal growth factor (EGFR) expression in untreated tumor models. However, following sunitinib treatment, expression of HIF2α and phosphorylated EGFR were equivalent in both PHD3 knockdown and control tumors. PHD3 knockdown changed the overall redox state of the cell as seen by the increased concentration of glutathione in PHD3 knockdown tumors relative to control tumors. UMRC3-PHD3KD cells had increased proliferation in cell culture when grown in the presence of hydrogen peroxide compared to UMRC3-SC control cells. Our findings illustrate (1) the variable effect of PHD3 on HIF2α expression, (2) an inverse relationship between PHD3 expression and tumor growth in ccRCC animal models, and (3) the role of PHD3 in maintaining the redox state of UMRC3 cells and their proliferative rate under oxidative stress.

Highlights

About 90% of kidney cancers are renal cell carcinomas (RCCs) [1], the incidence of which has been increasing over the past decade [2]
UMRC3 cells transfected with PHD isoform 3 s (PHD3) shRNA (UMRC3-PHD3KD) showed reduced PHD3 protein expression compared with UMRC3 cells transfected with scrambled shRNA (UMRC3-SC cells) (Figure 2a)
The knockdown of PHD3 in the UMRC3 clear cell renal cell carcinoma (ccRCC) cell line led to increased tumor growth in mouse models which was inhibited by sunitinib treatment. This increased growth rate with PHD3 knockdown is counterintuitive if PHD30 s primary role is the degradation of hypoxia-inducible factors (HIFs) proteins

Summary

Introduction

About 90% of kidney cancers are renal cell carcinomas (RCCs) [1], the incidence of which has been increasing over the past decade [2]. Despite recent advances in therapy, the mean survival duration of patients with metastatic RCC is only 13–27 months [1]. The development of effective therapies to improve these patients’ dismal prognoses requires a better understanding of the mechanisms underlying the carcinogenesis of RCC [3]. Clear cell RCC (ccRCC) accounts for approximately 70% of all RCCs and metabolic reprogramming plays a central role in ccRCC carcinogenesis. 90% of ccRCCs show inactivation of the von Hippel Lindau gene, VHL [1]. VHL inactivation increases the expression levels of hypoxia-inducible factors (HIFs), which drive ccRCC carcinogenesis

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