lncRNA CYTOR promotes lung adenocarcinoma gemcitabine resistance and epithelial-mesenchymal transition by sponging miR-125a-5p and upregulating ANLN and RRM2

Abstract

Lung adenocarcinoma (LUAD) is one of the most aggressive types of lung cancer. The prognosis of LUAD patients remains poor, and the overall efficacy of gemcitabine-based chemotherapy is still unsatisfactory. Long noncoding RNAs (lncRNAs) play important roles in several cancer types by interacting with multiple proteins, RNA, and DNA. However, the relationship between lncRNA dysregulation and gemcitabine resistance in LUAD has not been fully elucidated. In this study, lncRNA CYTOR expression and its association with the prognosis of LUAD patients are assessed by quantitative RT-PCR and Kaplan-Meier survival analysis. In vitro and in vivo functional studies are conducted to evaluate the biological functions of CYTOR in LUAD. The underlying mechanism regarding the tumor-promoting effects of CYTOR is explored using RNA immunoprecipitation, biotin-labelled RNA pulldown, luciferase reporter assays, and western blot analysis. We identify that CYTOR is an oncogenic lncRNA and is apparently upregulated in LUAD by analysing TCGA-LUAD data. High CYTOR expression is a poor prognostic factor for LUAD. Functional studies reveal that CYTOR confers LUAD cells with stronger resistance to gemcitabine treatment and upregulates the expression levels of epithelial-mesenchymal transition (EMT)-related proteins. Mechanically, CYTOR acts as a competitive endogenous RNA (ceRNA) to absorb miR-125a-5p, weakens the antitumor function of miR-125a-5p, and ultimately upregulates ANLN and RRM2 expressions. Taken together, this study explains the mechanism of lncRNA in the gemcitabine resistance of LUAD and formulates a theoretical framework for the in depth study of LUAD.

1. Introduction

Lung adenocarcinoma (LUAD) is usually derived from the bronchial epithelium, with some cases originated from the large bronchial mucous glands. LUAD is more likely to occur in women, Asians, and nonsmokers, and the incidence of LUAD has been increasing in recent years [1,2]. Although LUAD typically grows slowly and develops small masses, it easily metastasizes at an early stage and develops resistance to conventional treatment, such as radiotherapy and chemotherapy, which results in poor clinical efficacy and poor prognosis for LUAD patients [3,4]. For LUAD patients with locally advanced or metastatic disease, gemcitabine and gemcitabine-based strategies are the standard chemotherapies [5,6]. Gemcitabine (dFdC) is a pyrimidine antimetabolite that is metabolized into active nucleoside diphosphate (dFdCDP) and nucleoside triphosphate (dFdCTP) by nucleoside kinase. DFdCDP and dFdCTP inhibit DNA synthesis, thus realizing the cytotoxicity of gemcitabine [7,8]. Despite the clinical efficacy of gemcitabine, it is subject to acquired resistance [9]. Therefore, deeply investigating and understanding the mechanisms underlying gemcitabine resistance in LUAD are crucial to improving its treatment effectiveness.

Long noncoding RNA (lncRNA) is a kind of transcript of more than 200 nucleotides (nt) [10]. LncRNA has limited or no protein-coding capacity but exhibits multifaceted regulatory gene expression in the form of RNA [11]. As research has increased, lncRNA has assumed a greater clinical significance. Aside from controlling embryonic development, lncRNA can also play roles in inflammation, immune cell development, and tumor development [12]. LncRNA has been confirmed as a key biological molecule in tumorigenesis and progression in various human tumors, including liver cancer, breast cancer, head and neck cancer, gastric cancer, and cervical cancer [13–18]. In lung cancer, the prognostic value and crucial effects in cancer development of some lncRNAs have been well verified. For example, a novel risk model containing 12 ferroptosis-related lncRNAs has important prognostic value for LUAD and provides a theoretical basis for the clinical development of ferroptosis-related therapeutic targets [19]. LCAT3 is a recently discovered novel lncRNA that recruits FUBP1 (far upstream element-binding protein 1) proteins to the MYC FUSE (far-upstream element) sequence, activates the transcription of the MYC gene, and ultimately promotes LUAD proliferation, migration, and invasion [20]. The lncRNA CYTOR is highly expressed in various cancers, and CYTOR can partially reverse the suppression of glioma proliferation and migration by UPF1 [21]. CYTOR is also highly expressed in colorectal cancer and predicts poor prognosis [22]. We first found that CYTOR is overexpressed in LUAD by bioinformatics analysis. However, the me