Dottorato in MEDICINA MOLECOLARE

Titolo della tesi: OVERCOMING DRUG RESISTANCE IN PROSTATE CANCER: NOVEL BIOMARKERS AND TREATMENT STRATEGIES

Prostate cancer (PCa) is a multifaceted and biologically highly heterogeneous disease. Some patients show an indolent disease with few symptoms while others develop a highly aggressive disease that progresses to metastatic carcinoma refractory to both hormone therapy and chemotherapy (mCRPC). Most prostate cancers are androgen-dependent; thus, androgen deprivation therapy (ADT) is considered the standard treatment. Nevertheless, even in the presence of very low or undetectable androgen levels, androgen receptor (AR) signaling continues to stimulate tumor cell growth and promote disease progression. Thence neoadjuvant and combined therapies have been proposed. Although preliminary results of the use of combination treatments appear to be promising, is still necessary to better understand the biology of prostate cancer with respect to mechanisms of disease progression and therapeutic resistance, including, but not limited to, AR activation via alternative survival signaling pathways or AR splice variants. The first aim of this project was therefore to conduct a retrospective observational study on PCa biopsies to evaluate whether the presence of AR-V7, an androgen receptor splice variant that results in the truncation of the ligand-binding domain, was related to the risk and/or progression to mCRPC. Expression of AR-V7 and AR-full length (FL) was evaluated separately by immunohistochemistry using two monoclonal anti-AR-V7 and anti-AR-FL antibodies on paraffin-embedded tumour tissue sections obtained from 56 patients with histologically proven prostate adenocarcinoma considered for radical prostatectomy (RP). Although the numbers are too low to draw conclusions, we have shown that in a population of untreated, nonmetastatic PCa patients, AR-V7 is detectable by immunohistochemistry in more than 50% of cases. At this early stage, regardless of ADT administration, AR-V7 positivity is associated with risk classification and could therefore predict biochemical and radiological progression after surgery. The second aim of the present thesis was to investigate possible therapeutic strategies with drug combinations in order to overcome drug resistance. For this purpose, we tested the combination of new generation androgen receptor signaling inhibitors (ARSi) with inhibitors of PI3K/AKT/mTOR signalling pathway on prostate cancer cell lines with a different expression of AR and phosphatase and tensin homolog (PTEN). Overall, preliminary results suggest that the action of combinatorial therapy is more effective in an experimental PTEN-loss model of prostate cancer. Translating this observation into clinical practice, proper stratification of patients with PTEN gene deletion could be essential in order to ensure better personalized therapy. The third aim of the current project, carried out in collaboration with University of KU Leuven and based on preliminary data from the ARNEO trial conducted at UZ Leuven, was to provide molecular evidence to support the hypothesis that treatment with Apalutamide (a second-generation antiandrogen) may induce new vulnerabilities in patients with localized prostate cancer. To this end, the combination apalutamide plus ipatasertib (a pan-AKT inhibitor) was studied on three prostate cancer cell lines by evaluating its effects on proliferation, cell cycle and AKT patwhay. The secondary projects conducted in parallel and published in scientific journals during my Ph.D. have been reported at the end of the thesis as appendices. The objectives of the research carried out in collaboration with the University of Tor Vergata (Appendices A-C) were i) to characterize phase-change contrast agents (microbubbles/microdroplets) as promising drug delivery systems, ii) to study the pro-inflammatory effects of medical ultrasound on human keratinocytes, and iii) to analyze the trajectories of adhesive microbubbles approaching endothelial cells. In the latest work (Appendix D), in collaboration with the Department of Pharmaceutical Sciences of Sapienza University of Rome, the antineoplastic properties of a structural analogue of nocodazole were evaluated.