RNA-seq analysis of plasmatic exosomal miRNAs in pediatric Hodgkin Lymphoma

F Lovisa; E Gaffo; C Elia; A Garbin; I Gallingani; CC Damanti; E Carraro; Re V De; L Caggiari; O Repetto; Zorzi M De; A Sala; S Buffardi; L Vinti; M Pillon; S Bortoluzzi; M Mascarin; L Mussolin

doi:10.1055/s-0040-1701810

Klinische Pädiatrie, Inhaltsverzeichnis

Klin Padiatr 2020; 232(02): 79
DOI: 10.1055/s-0040-1701810

S-I

Session I: Translational Biology

RNA-seq analysis of plasmatic exosomal miRNAs in pediatric Hodgkin Lymphoma

F Lovisa

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

²Unit of Onco-hematology, stem cell transplant and gene therapy, Istituto di Ricerca Pediatrica Città della Speranza, Padova, Italy

,

E Gaffo

³Department of Molecular Medicine, University of Padova, Padova, Italy

,

C Elia

⁴Pediatric Radiotherapy and AYA Unit, Centro di Riferimento Oncologico di Aviano (CRO), IRCCS, National Cancer Institute, Aviano, Italy

,

A Garbin

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

²Unit of Onco-hematology, stem cell transplant and gene therapy, Istituto di Ricerca Pediatrica Città della Speranza, Padova, Italy

,

I Gallingani

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

²Unit of Onco-hematology, stem cell transplant and gene therapy, Istituto di Ricerca Pediatrica Città della Speranza, Padova, Italy

,

CC Damanti

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

²Unit of Onco-hematology, stem cell transplant and gene therapy, Istituto di Ricerca Pediatrica Città della Speranza, Padova, Italy

,

E Carraro

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

,

Re V De

⁵Department of Research and Advanced Tumour Diagnostics, Centro di Riferimento Oncologico di Aviano (CRO), IRCCS, National Cancer Institute, Immunopathology and Tumour Biomarkers Unit/Bio-proteomics Facility, Aviano, Italy

,

L Caggiari

⁵Department of Research and Advanced Tumour Diagnostics, Centro di Riferimento Oncologico di Aviano (CRO), IRCCS, National Cancer Institute, Immunopathology and Tumour Biomarkers Unit/Bio-proteomics Facility, Aviano, Italy

,

O Repetto

⁵Department of Research and Advanced Tumour Diagnostics, Centro di Riferimento Oncologico di Aviano (CRO), IRCCS, National Cancer Institute, Immunopathology and Tumour Biomarkers Unit/Bio-proteomics Facility, Aviano, Italy

,

Zorzi M De

⁵Department of Research and Advanced Tumour Diagnostics, Centro di Riferimento Oncologico di Aviano (CRO), IRCCS, National Cancer Institute, Immunopathology and Tumour Biomarkers Unit/Bio-proteomics Facility, Aviano, Italy

,

A Sala

⁶Department of Paediatrics, Ospedale San Gerardo, University of Milano-Bicocca, Fondazione MBBM, Monza, Italy

,

S Buffardi

⁷Paediatric Haemato-Oncology department, Santobono-Pausilipon Children’s Hospital, Napoli, Italy

,

L Vinti

⁸Department of Pediatric Haemato-Oncology, IRCCS Ospedale Bambino Gesù, Sapienza University of Rome, Roma, Italy

,

M Pillon

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

,

S Bortoluzzi

³Department of Molecular Medicine, University of Padova, Padova, Italy

⁹CRIBI Interdepartmental Research Center for Innovative Biotechnologies (CRIBI), University of Padova, Padova, Italy

,

M Mascarin

⁴Pediatric Radiotherapy and AYA Unit, Centro di Riferimento Oncologico di Aviano (CRO), IRCCS, National Cancer Institute, Aviano, Italy

,

L Mussolin

¹Department of Women’s and Children’s Health, University of Padova, Padova, Italy

²Unit of Onco-hematology, stem cell transplant and gene therapy, Istituto di Ricerca Pediatrica Città della Speranza, Padova, Italy

› Institutsangaben

Abstract

Volltext

Introduction Hodgkin lymphoma (HL) represents around ~10% of all cases of lymphoid neoplasms. Although current risk-adapted and response based treatment approaches can cure >90% of the children, treatment of relapsed/resistant patients remains a challenge [1].

In the last years, a large body of evidence proved the functional involvement of exosomes in cancer progression and spreading, induction of angiogenesis, as well as in chemoresistance and immune response evasion during tumor development [2].

The aim of our study is to characterize miRNA in plasmatic exosomes of pediatric HL patients, for the identification of new non-invasive biomarkers and disease mechanisms, which might be used to improve diagnosis, treatment and prognosis of the affected children.

Methods The study cohort included 36 HL patients treated according to EuroNet-PHL-C2 trial and 7 plasma samples from healthy donors (D) as controls. Exosomes were isolated from 0.5 ml plasma at diagnosis by exoEasy Midi Kit (Qiagen) and validated by Nanoparticle Tracking Analysis (NTA, Malvern Nanosight), Atomic Force Microscopy (ATM) and Western Blotting. Total exosomal RNA was obtained using exoRNeasyMidi Kit (Qiagen) and sequencing libraries were prepared from 10 ng RNA by using the NEBNext® Multiplex Small RNA Library Prep Kit for Illumina® (New England Biolabs). RNA-seq analysis was performed on an Illumina HiSeq 4000 with target depth of 15 M reads per sample. RNA-seq data were analyzed using the software miR&moRe [3]. Differential expression analyses were performed with DESeq2 [4].

Results Overall, >200 expressed miRNAs were identified. Comparing HL and D exosome samples, 11 differentially expressed miRNAs were disclosed. When patients were grouped based on Early Response Assessment (ERA), 15 miRNAs differentially expressed were identified between patients with adequate and inadequate response.

Conclusion Differentially expressed miRNAs will be prioritized based on their functional significance in cancer and/or previous reports in lymphoid malignancies. The most promising candidates will be validated by quantitative real-time PCR in an extended cohort of both HL and D and functionally characterized by in vitro experiments.

These results are expected to provide insights on the role of exosomal miRNAs in HL disease aggressiveness and response to therapy.

Referenzen

References
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