Dottorato in MEDICINA MOLECOLARE

Titolo della tesi: The development of a circulating biomarker classifier for small cell lung cancer diagnosis

Small cell lung cancer (SCLC) accounts for almost 15% of lung cancer cases and is characterized by early development of metastasis, poor prognosis, and challenging diagnosis. The aberrant expression of circulating microRNAs (miRNAs) and extracellular vesicles (EV) included miRNAs have been reported in many tumors including SCLC, and can help with cancer diagnosis. Our work aimed to build a miRNA-based biomarker classifier for helping SCLC detection. We used NGS and NanoString to profile circulating free and EV-included miRNA, which showed an upregulation in SCLC compared to control and non-small cell lung cancer (NSCLC) samples. We selected and tested, by qRT-PCR, plasma miR-375 and EV-included miR-1285-5p in a Training cohort of samples and found a significant upregulation of miR-375, but not EV-included miR-1285. Using logistic regression, we built a classifier based on miR-375 expression in the Training cohort dataset and applied it to a Validation cohort dataset. The qRT-PCR data showed a significant upregulation of miR-375 in SCLC of the Validation cohort, but the model could not accurately classify the samples of the Validation cohort into histological groups. Further analyses are ongoing to improve the accuracy of the classifier. We have also performed preliminary functional studies of miR-375 in lung cancer cell lines. Through Affymetrix, IPA and western blot analysis, we found that the overexpression of miR-375 induced the activation of the mTOR pathway in both SCLC and NSCLC cell lines. Moreover, we found that the overexpression of miR-375 induced an upregulation of NeuroD1, which is a transcriptional factor that is implicated in SCLC pathology. Our results show that miR-375 is a validated biomarker for SCLC detection, but that this miRNA alone cannot offer an accurate classification. Ongoing analyses aim to study other deregulated miRNAs to improve our classifier for SCLC detection. Moreover, miR-375 overexpression induces the activation of the mTOR pathway and an upregulation of the NeuroD1 protein, which are both involved in SCLC pathology.