Amplificazione genomica nel cancro: circolarizzazione degli ampliconi tramite CRISPR-C, conseguenze trascrizionali e implicazioni terapeutiche

In the complex landscape of cancer, gene amplification emerges as a pivotal mechanism driving tumor development and progression. This thesis discusses the multifaceted role of gene amplification across various cancer types. The investigation focuses initially on metastatic gastric cancer, where analysis of patient tumor samples reveals that overexpression of the Vascular Endothelial Growth Factor A (VEGFA) gene, in some cases due to the genomic amplification, may serve as a potential biomarker to predict the response to anti-angiogenic therapies, like that with Ramucirumab. Furthermore, interesting evidence regarding the overexpression of circular RNAs originating from the MDM2 amplified proto-oncogene allows to identify a possible difference in the transcriptional signature between malignant and non-malignant lipomatous tumors in both patient RNA and cell lines, implying a role in aggressive tumor development. The thesis further describes the setup of in vitro cell models to investigate the functional role of the PVT1 long non-coding RNA gene, coamplified and overexpressed with MYC in a subset of small cell lung cancer tumors. Additionally, the genesis of circular extrachromosomal DNA (ecDNA) harboring oncogenes such as CDK4, MDM2, and ABCB1 is investigated. Through the adaptation of CRISPR-C technology, successful modeling of ecDNA generation and amplification is achieved. Finally, CRISPR technology was also attempted to trigger chromothripsis, a catastrophic chromosomal rearrangement often leading to amplification. In summary, the research work described in the present thesis contributes interesting information on the mechanisms behind the genesis of genomic amplification in cancer and its transcriptional consequences, aiming to understand tumor initiation and evolution better.