Università degli studi di Padova
Identification of New Biomarkers of Disease Resistance in ALCL of childhood: the role of exosomes.
Università degli studi di Padova
Supervisor: Lara MUSSOLIN
Liquid biopsy analysis has grown exponentially in the last few years, impacting various areas of research including cancer diagnostics and prognostication. In particular, small extracellular vesicles (EVs) formed inside endosomal compartments (i.e., exosomes) are actively released (carrying tumour RNA, DNA and protein) and can function as inter-cellular messengers.
Exosomes and other EVs are particularly interesting as cancer biomarkers since they are stable carriers of genetic material and proteins from their cell of origin. They are also thought to be part of the disease process, for example, tumour exosomes have been shown to stimulate tumour cell growth, suppress the immune response and induce angiogenesis and even be part of the metastatic process. Exosome-associated cancer biomarkers have been extensively described in a great number of adult solid cancers. With respect to paediatric ALCL, genomic and proteomic analyses have been performed of pre-treatment patient samples from primary tumour cells, but very few data are available of plasma EV samples.
Recently we profiled by RNAseq the small RNA (sRNA) content of circulating exosomes of a small cohort of paediatric patients with ALCL and a series of healthy controls. Our analysis disclosed that non-miRNA derived sRNAs constitute the prominent fraction of sRNA loaded in exosomes and identified 180 sRNAs significantly more abundant in exosomes of ALCL patients compared to controls. YRNA fragments, accounted for most of the exosomal content and were significantly increased in ALCL patients, and hence were prioritized for further investigation by qRT-PCR.
Quantification of RNY4 fragments and full-length sequences disclosed that the latter are massively loaded into exosomes of ALCL patients with more advanced and aggressive disease. Recent findings underlined the role of RNY4 in particular in the modulation of the tumour microenvironment.
The main goals we aim to achieve are:
- to isolate exosomes from an existing bioresource of plasma samples of ALCL patients at diagnosis and before the last cycle of chemotherapy;
- to characterize exosomes at a transcriptional level;
- to analyse the association of significant exosomal sRNAs with clinical and biological characteristics;
to investigate exosomal cargo modification in response to different therapeutic conditions (chemotherapy alone /chemotherapy + tyrosine kinase inhibitor). The identification of exosomal biomarkers from a simple blood test and without invasive procedures might permit rapid characterization of tumour biology and guide treatment, giving a better insight into molecular targeted therapies.
Based on the experimental design proposed, a wide and complete investigation of the potential role of EVs for the management of ALCL will be obtained. ALK represents a rational therapeutic target for paediatric patients with ALCL, however due to the resistance to ALK inhibitors fostered by multiple different molecular mechanisms, new diagnostic strategies are needed to assess resistance mechanisms and provide patients with the most effective therapy. The identification of exosome-associated biomarkers is expected to provide important information about the molecular characteristics of aggressive ALCL, which might be used to improve diagnosis, treatment and prognosis of affected children.
Year 1: Galseq: Sequencing of exosomal cargo using equipment and technologies available in the host lab in which use, training will be provided (4 months)
Year 2: Universitätsklinikum Hamburg-Eppendorf: Access to a second independent cohort of samples for data validation and training in analysis of the immune cell profile (4 months)