MOLECULAR MECHANISMS UNDERLYING CLINICAL BEHAVIOR OF NEUROENDOCRINE TUMORS: STUDY OF NEW POSSIBLE BIOMARKERS PREDICTABLE OF PHARMACOLOGICAL RESISTANCE AND TARGETS

Neuroendocrine tumors (NETs) represent a heterogeneous group of malignances arising from the diffuse neuroendocrine system whose incidence is rising over the years. Due to their complexity and unusual features, first-line therapeutical approach is represented by surgical resection, even if it is not completely resolutive in most of cases, due to recurrence and metastasis accompanied by pharmacological resistance. In this scenario, the lack of effective pharmacological approaches is making hard the treatment of these pathologies. Within the pharmacological context, lot of attention was given to somatostatin and dopamine system, because of prevalent expression of somatostatin receptors (SSTs) and dopamine receptor type 2 (DRD2) in about all NETs that represented, over the years, the main targets for treatment with somatostatin analogs (SSAs) and dopamine agonists (DAs) for these pathologies. Nevertheless, the molecular mechanisms underline pharmacological resistance and the development of novel biomarkers useful to identify patients that may benefit of specific treatments are steel needed, together with the development of novel in vitro pre-clinical model better resembling human tumor behavior. This thesis aimed to investigate possible biomarkers underlining pharmacological resistance and predictable of SSAs/DAs responsiveness in NETs, more specifically focusing on G-proteins and β-arrestins DRD2-mediated signaling pathways in PitNETs and on somatostatin/cortistatin system in novel PanNENs cell models. Briefly, β-arrestin 2 has been highlighted as novel potential biomarker of pharmacological responsiveness to DAs, and that combined therapy with DAs avoids everolimus escape feed-back in PitNETs. The role of -arrestin 2 in regulating both AKT phosphorylation and cell proliferation in tumoral lactotrophs and NF-PitNETs was demonstrated, elucidating a new molecular mechanism that contributes to confer sensitivity or resistance to DAs in PitNETs. In tumoral lactotrophs and NF-PitNETs, the lack of -arrestin 2 prevents the inhibitory effect of DRD2 on AKT pathway activation with a consequent resistance to the antimitotic action of DAs. In accordance, it was showed that cabergoline improved everolimus efficacy by blocking AKT upstream reactivation in NF-PitNETs and tumoral lactotrophs. In fact, the combined treatment with cabergoline strongly reduced the increase of AKT activity, correlating with its antimitotic effect. The role of -arrestin 2 has been highlighted in mediating cabergoline inhibition of both p-AKT activity and cells proliferation after everolimus cotreatment: the lack of β-arrestin 2 reverted the antimitotic effect induced by the combined treatment. Moreover, the cytoskeleton actin binding protein filamin-A (FLNA) emerged as an important player in the regulation of PitNETs drug responsiveness and invasiveness. At this regard our data showed that activation of the cAMP pathway and DRD2 agonist regulated FLNA activity by increasing or decreasing, its phosphorylation, respectively. FLNA phosphorylation has been identified as potential biomarker for tumor responsiveness to DAs, in DRD2 expressing PitNETs, since it participates in mediating DRD2 intracellular signalling, additionally turning out that FLNA phosphorylation prevented DRD2 signaling in PRL- and ACTH-secreting PitNETs. At last, a potential of novel pharmacological compounds of the SS/CST family exerted antiproliferative effects in new human resembling PanNENs models. To sum up, this work identified the role of specific molecular mechanism underlying pharmacological resistance in PitNETs, paving the way for possible combined approaches to overcome drug resistance and the pilot study on novel PanNENs cell lines strongly suggests that testing new pharmacological compounds of the SS/CST family can lead to an innovative discovery to expand the therapeutic arsenal to treat PanNENs and set novel approaches with a personalized perspective.