Comprehensive characterization of RNA cargo of extracellular vesicles in breast cancer patients undergoing neoadjuvant chemotherapy

Lilite Sadovska; Pawel Zayakin; Kristaps Eglītis; Edgars Endzeliņš; Ilze Radoviča-Spalviņa; Elīza Avotiņa; Jānis Auders; Laura Keiša; Inta Liepniece-Karele; Mārcis Leja; Jānis Eglītis; Aija Linē

doi:10.3389/fonc.2022.1005812

Comprehensive characterization of RNA cargo of extracellular vesicles in breast cancer patients undergoing neoadjuvant chemotherapy

Front Oncol. 2022 Oct 26:12:1005812. doi: 10.3389/fonc.2022.1005812. eCollection 2022.

Authors

Affiliations

¹ Cancer Biomarker group, Latvian Biomedical Research and Study Centre, Riga, Latvia.
² Latvian Oncology Center, Riga Eastern Clinical University Hospital, Riga, Latvia.
³ Genera Ltd., Riga, Latvia.
⁴ Institute of Clinical and Preventive Medicine, University of Latvia, Riga, Latvia.
⁵ Department of Pathology, Riga Stradins University, Riga, Latvia.
⁶ University of Latvia, Faculty of Medicine, University of Latvia, Riga, Latvia.

Abstract

Extracellular vesicles (EVs) are g7aining increased attention as carriers of cancer-derived molecules for liquid biopsies. Here, we studied the dynamics of EV levels in the plasma of breast cancer (BC) patients undergoing neoadjuvant chemotherapy (NAC) and explored the relevance of their RNA cargo for the prediction of patients' response to the therapy. EVs were isolated from serial blood samples collected at the time of diagnosis, at the end of NAC, and 7 days, 6, and 12 months after the surgery from 32 patients with locally advanced BC, and 30 cancer-free healthy controls (HCs) and quantified by nanoparticle tracking analysis. The pre-treatment levels of EVs in BC patients were higher than in HCs, significantly increased during the NAC and surgery, and decreased to the levels found in HCs 6 months after surgery, thus showing that a substantial fraction of plasma EVs in BC patients are produced due to the disease processes and treatment. RNA sequencing analysis revealed that the changes in the EV levels were associated with the alterations in the proportions of various RNA biotypes in EVs. To search for RNA biomarkers that predict response to the NAC, patients were dichotomized as responders and non-responders based on Miller-Payne grades and differential expression analyses were carried out between responders and non-responders, and HCs. This resulted in the identification of 6 miRNAs, 4 lncRNAs, and 1 snoRNA that had significantly higher levels in EVs from non-responders than responders at the time of diagnosis and throughout the NAC, and significantly lower levels in HCs, thus representing biomarkers for the prediction of response to NAC at the time of diagnosis. In addition, we found 14 RNAs representing piRNA, miRNA, lncRNA, snoRNA, and snRNA biotypes that were induced by NAC in non-responders and 2 snoRNAs and 1 piRNA that were induced by NAC in patients with early disease progression, thus warranting further functional studies on their role in chemoresistance and metastasis.

Keywords: RNA biotypes; RNA sequencing; breast cancer; extracellular vesicles; liquid biopsy; neoadjuvant chemotherapy; prognostic biomarker; snoRNA.