A circular RNA derived from PLXNB2 as a valuable predictor of the prognosis of patients with acute myeloid leukaemia

Leilei Lin,Yuhuang Wang,Min Wu,Zhibo Guo,Linlin Zhao,Sicheng Bian,Desheng Kong,Yuying Wang,Yu Wang,Lili Sun,Yinghua Li,Dan Guo,Qi Li

doi:10.1186/s12967-021-02793-7

Abstract

BackgroundAs a common haematological malignancy, acute myeloid leukaemia (AML), particularly with extramedullary infiltration (EMI), often results in a high mortality rate and poor prognosis. Circular RNAs (circRNAs) regulate biological and pathogenic processes, suggesting a potential role in AML. We have previously described the overall alterations in circRNAs and their regulatory networks between patients with AML presenting with and without EMI. This study aims to find new prognostic and therapeutic targets potentially associated with AML.MethodsqRT-PCR was performed on samples from 40 patients with AML and 15 healthy controls. The possibility of using circPLXNB2 (circRNA derived from PLXNB2) as a diagnostic and prognostic biomarker for AML was analysed with multiple statistical methods. In vitro, the function of circPLXNB2 was studied by lentivirus transfection, CCK-8 assays, flow cytometry, and Transwell experiments. Western blotting and qRT-PCR were performed to detect the expression of related proteins and genes. The distribution of circPLXNB2 in cells was observed using RNA fluorescence in situ hybridization (RNA-FISH). We also investigated the role of circPLXNB2 by establishing AML xenograft models in NOD/SCID mice.ResultsBy analysing the results of qRT-PCR detection of clinical samples, the expression of the circPLXNB2 and PLXNB2 mRNAs were significantly increased in patients with AML, more specifically in patients with AML presenting with EMI. High circPLXNB2 expression was associated with an obviously shorter overall survival and leukaemia-free survival of patients with AML. The circPLXNB2 expression was positively correlated with PLXNB2 mRNA expression, as evidenced by Pearson’s correlation analysis. RNA-FISH revealed that circPLXNB2 is mainly located in the nucleus. In vitro and in vivo, circPLXNB2 promoted cell proliferation and migration and inhibited apoptosis. Notably, circPLXNB2 also increased the expression of PLXNB2, BCL2 and cyclin D1, and reduced the expression of BAX.ConclusionIn summary, we validated the high expression of circPLXNB2 and PLXNB2 in patients with AML. Elevated circPLXNB2 levels were associated with poor clinical outcomes in patients with AML. Importantly, circPLXNB2 accelerated tumour growth and progression, possibly by regulating PLXNB2 expression. Our study highlights the potential of circPLXNB2 as a new prognostic predictor and therapeutic target for AML in the future.

Highlights

As a common haematological malignancy, acute myeloid leukaemia (AML), with extramedullary infiltration (EMI), often results in a high mortality rate and poor prognosis
PLXNB2 and circPLXNB2 expression were significantly increased in patients with AML Whole-genome and circRNA microarrays were conducted from 8 samples of AML patients with or without EMI and 4 samples of healthy controls, which indicating PLXNB2, hsa_circ_0004520 and hsa_circ_0001257 were likely to be involved in the regulation of intercellular crosstalk associated with EMI [21]
Immunohistochemical (IHC) staining was performed on bone marrow biopsy specimens obtained from patients above showed that PLXNB2 protein expression were significantly higher in patients with AML, in patients with AML presenting with EMI (Fig. 1a, b). Real‐time quantitative polymerase chain reaction (qRT-PCR) analysis of Bone marrow mononuclear cells (BMMCs) from patients with AML presenting with or without EMI (EMI = 24, non-EMI = 16) and healthy controls (n = 15) revealed that PLXNB2 mRNA was highly expressed in patients with AML (Fig. 1c)

Summary

Introduction

As a common haematological malignancy, acute myeloid leukaemia (AML), with extramedullary infiltration (EMI), often results in a high mortality rate and poor prognosis. CircRNAs function through the following mechanisms: regulating gene expression in cis at the transcriptional level [12], acting as a microRNA “sponge” and inhibiting its function [13], interacting with RNA binding proteins that regulate transcription [14], encoding peptides [15], and other functions. Due to their special covalently closed circular structure, circRNAs are strongly resistant to RNase R, are more stable than mRNAs and are often regarded as potential biomarkers for cancer [16, 17]. Our previous studies confirmed that circRNAs may play an important role in regulating intercellular crosstalk in EMI, and we found that PLXNB2 predicted a poor prognosis of patients with AML and might be involved in EMI regulation [21]

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