Circular RNA LARP4 correlates with decreased Enneking stage, better histological response, and prolonged survival profiles, and it elevates chemosensitivity to cisplatin and doxorubicin via sponging microRNA-424 in osteosarcoma

Abstract

Background: This study aimed to evaluate the association of circular RNA La-related RNA-binding protein 4 (circ-LARP4) with clinical features and prognosis in osteosarcoma patients, and further explore its effect on chemosensitivity in osteosarcoma cells.

Methods: Seventy-two osteosarcoma patients with Enneking stage IIA-IIB who underwent resection were consecutively enrolled, and then, tumor tissues and non-tumor tissues were obtained. Circ-LARP4 in tumor tissue/non-tumor tissue was detected by quantitative polymerase chain reaction. After circ-LARP4 overexpression and negative control overexpression plasmid transfection, relative cell viability (%) was evaluated by Cell Counting Kit-8 in MG63 cells treated by different concentrations of cisplatin, methotrexate, and doxorubicin, and IC₅₀ was calculated.

Results: Circ-LARP4 was downregulated in tumor tissue compared with non-tumor tissue and had a good value in distinguishing tumor tissue from non-tumor tissue with an area under curve of 0.829 (95% CI: 0.762-0.859). Meanwhile, tumor circ-LARP4 was negatively correlated with the Enneking stage. After resection, circ-LARP4 high expression patients showed an increased tumor cell necrosis rate to adjuvant chemotherapy compared to circ-LARP4 low expression patients, and circ-LARP4 high expression correlated with prolonged disease-free survival and overall survival. In vitro experiments revealed that circ-LARP4 overexpression elevated the chemosensitivity of MG63 cells to cisplatin and doxorubicin but not methotrexate, with decreased cisplatin IC₅₀ and doxorubicin IC₅₀ concentrations than negative control. Besides, miR-424 overexpression attenuated the chemosensitivity in circ-LARP4 overexpression-treated MG63 cells.

Conclusion: Circ-LARP4 high expression correlates with decreased Enneking stage and prolonged survival profiles, and it elevates chemosensitivity to cisplatin and doxorubicin via sponging miR-424 in osteosarcoma.

Keywords: Enneking stage; chemosensitivity; circ-LARP4; osteosarcoma; survival profile.

Figure 1

Circ‐LARP4 expression in tumor and non‐tumor tissue from osteosarcoma patients. The expression of circ‐LARP4 in tumor and non‐tumor tissue (A) and the ROC curve analysis of circ‐LARP4 expression for differentiating tumor tissue from non‐tumor tissue (B). Comparison was determined by Wilcoxon signed‐rank test. ROC curve was used for assessing the value of circ‐LARP4 expression for differentiating tumor tissue from non‐tumor tissue. P value <.05 was considered statistically significant. Circ‐LARP4, circular RNA La‐related RNA‐binding protein 4; ROC, receiver operative characteristics; AUC, area under curve

Figure 2

Correlation of circ‐LARP4 with good response rate. The good response rate in patients with circ‐LARP4 high expression and in patients with circ‐LARP4 low expression. Comparison was determined by chi‐square test. P value <.05 was considered statistically significant. Circ‐LARP4, circular RNA La‐related RNA‐binding protein 4

Figure 3

Correlation of circ‐LARP4 expression with DFS and OS. The correlation of circ‐LARP4 expression with DFS and OS. Survival curves were estimated using the Kaplan‐Meier method, and the difference of survival was determined by the log‐rank test. P value <0.05 was considered statistically significant. Circ‐LARP4, circular RNA La‐related RNA‐binding protein 4; DFS, disease‐free survival; OS, overall survival

Figure 4

Effect of circ‐LARP4 on chemosensitivity of MG63 cells. Effect of circ‐LARP4 on relative cell viability of MG63 cells under different concentrations of cisplatin treatment (A), the IC₅₀ value of cisplatin in blank control group, NC(+) group and Circ‐LARP4(+) group (B), effect of circ‐LARP4 on relative cell viability under different concentrations of methotrexate treatment (C), the IC₅₀ of methotrexate in blank control, NC(+) group and circ‐LARP4(+) group (D), effect of circ‐LARP4 on relative cell viability under different concentrations of doxorubicin treatment (E), and the IC₅₀ value of doxorubicin in blank control, NC(+) group and circ‐LARP4(+) group (F). Comparison was determined by t test. P value <.05 was considered statistically significant. Circ‐LARP4, circular RNA La‐related RNA‐binding protein 4; NC, negative control

Figure 5

Effect of circ‐LARP4 and miR‐424 on chemosensitivity of MG63 cells. MiR‐424 expression in MG63 cells after circ‐LARP4 overexpression plasmid transfection (A), in further rescue experiments, the miR‐424 expression after transfections (B), the effect of circ‐LARP4 and miR‐424 on relative cell viability in osteosarcoma cells treated by cisplatin (C), methotrexate (D), or doxorubicin (E). Comparison was determined by t test. P value <.05 was considered statistically significant. Circ‐LARP4, circular RNA La‐related RNA‐binding protein 4; miR, microRNA