Ago-RIP Sequencing Identifies New MicroRNA-449a-5p Target Genes Increasing Sorafenib Efficacy in Hepatocellular Carcinoma

Reinkens, Thea; Stalke, Amelie; Huge, Nicole; Vajen, Beate; Eilers, Marlies; Schäffer, Vera; Dittrich-Breiholz, Oliver; Schlegelberger, Brigitte; Illig, Thomas; Skawran, Britta

doi:10.7150/jca.66016

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J Cancer 2022; 13(1):62-75. doi:10.7150/jca.66016 This issue Cite

Research Paper

Ago-RIP Sequencing Identifies New MicroRNA-449a-5p Target Genes Increasing Sorafenib Efficacy in Hepatocellular Carcinoma

Thea Reinkens¹, Amelie Stalke¹, Nicole Huge¹, Beate Vajen¹, Marlies Eilers¹, Vera Schäffer¹, Oliver Dittrich-Breiholz², Brigitte Schlegelberger¹, Thomas Illig^1,3, Britta Skawran^{1 ✉}

1. Department of Human Genetics, Hannover Medical School, Hannover, Germany.
2. Research Core Unit Genomics, Hannover Medical School, Hannover, Germany.
3. Hannover Unified Biobank (HUB), Hannover Medical School, Hannover, Germany.

Citation:

Reinkens T, Stalke A, Huge N, Vajen B, Eilers M, Schäffer V, Dittrich-Breiholz O, Schlegelberger B, Illig T, Skawran B. Ago-RIP Sequencing Identifies New MicroRNA-449a-5p Target Genes Increasing Sorafenib Efficacy in Hepatocellular Carcinoma. J Cancer 2022; 13(1):62-75. doi:10.7150/jca.66016. https://www.jcancer.org/v13p0062.htm

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Abstract

BACKGROUND: Patients with hepatocellular carcinoma (HCC) have very limited treatment options. For the last fourteen years, the multi-tyrosine kinase inhibitor sorafenib has been used as standard-of-care therapeutic agent in advanced HCC. Unfortunately, drug resistance develops in many cases. Therefore, we aimed to find a way to mitigate drug resistance and to improve the sorafenib efficacy in HCC cells. MicroRNAs play a significant role in targeting genes involved in tumor control suggesting microRNA/sorafenib combination therapy as a promising treatment option in advanced HCC.

METHODS: MiR-449a-5p target genes were identified by Ago-RIP sequencing and validated by luciferase reporter assays and expression analyses. Target gene expression and survival data were analyzed in public HCC datasets. Tumor-relevant functional effects of miR-449a-5p and its target genes as well as their impact on the effects of sorafenib were analyzed using in vitro assays. An indirect transwell co-culture system was used to survey anti-angiogenic effects of miR-449a-5p.

RESULTS: PEA15, PPP1CA and TUFT1 were identified as direct target genes of miR-449a-5p. Overexpression of these genes correlated with a poor outcome of HCC patients. Transfection with miR-449a-5p and repression of miR-449a-5p target genes inhibited cell proliferation and angiogenesis, induced apoptosis and reduced AKT and ERK signaling in HLE and Huh7 cells. Importantly, miR-449a-5p potentiated the efficacy of sorafenib in HCC cells via downregulation of PEA15, PPP1CA and TUFT1.

CONCLUSIONS: This study provides detailed insights into the targetome and regulatory network of miR-449a-5p. Our results demonstrate for the first time that targeting PEA15, PPP1CA and TUFT1 via miR-449a overexpression could have significant implications in counteracting sorafenib resistance suggesting miR-449a-5p as a promising candidate for a microRNA/sorafenib combination therapy.

Keywords: Liver cancer, microRNA combination therapy, drug resistance, microRNA target genes, Ago-RIP sequencing, multi-tyrosine kinase inhibitor

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APA

Reinkens, T., Stalke, A., Huge, N., Vajen, B., Eilers, M., Schäffer, V., Dittrich-Breiholz, O., Schlegelberger, B., Illig, T., Skawran, B. (2022). Ago-RIP Sequencing Identifies New MicroRNA-449a-5p Target Genes Increasing Sorafenib Efficacy in Hepatocellular Carcinoma. Journal of Cancer, 13(1), 62-75. https://doi.org/10.7150/jca.66016.

ACS

Reinkens, T.; Stalke, A.; Huge, N.; Vajen, B.; Eilers, M.; Schäffer, V.; Dittrich-Breiholz, O.; Schlegelberger, B.; Illig, T.; Skawran, B. Ago-RIP Sequencing Identifies New MicroRNA-449a-5p Target Genes Increasing Sorafenib Efficacy in Hepatocellular Carcinoma. J. Cancer 2022, 13 (1), 62-75. DOI: 10.7150/jca.66016.

NLM

CSE

Reinkens T, Stalke A, Huge N, Vajen B, Eilers M, Schäffer V, Dittrich-Breiholz O, Schlegelberger B, Illig T, Skawran B. 2022. Ago-RIP Sequencing Identifies New MicroRNA-449a-5p Target Genes Increasing Sorafenib Efficacy in Hepatocellular Carcinoma. J Cancer. 13(1):62-75.

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