Combined Targeting of Pathogenetic Mechanisms in Pancreatic Neuroendocrine Tumors Elicits Synergistic Antitumor Effects

Abstract

Pancreatic neuroendocrine neoplasms (PanNENs) are the second most common malignancy of the pancreas. Surgery remains the only curative treatment for localized disease. For patients with inoperable advanced or metastatic disease, few targeted therapies are available, but their efficacy is unpredictable and variable. Exploiting prior knowledge on pathogenetic processes involved in PanNEN tumorigenesis, we tested buparlisib (PI3K inhibitor) and ribociclib (CDK4/6 inhibitor), as single agents or in combination, in different preclinical models. First, we used cell lines representative of well-differentiated (INS-1E, NT-3) and poorly differentiated (BON-1) PanNENs. The combination of buparlisib with ribociclib reduced the proliferation of 2D and 3D spheroid cultures more potently than the individual drugs. Buparlisib, but not ribociclib, induced apoptosis. The anti-proliferative activity of the drugs correlated with downstream target inhibition at mRNA and protein levels. We then tested the drugs on primary islet microtissues from a genetic PanNET animal model (Men1-defective mice) and from wild-type mice: the drug combination was effective against the former without altering islet cell physiology. Finally, we treated PanNET patient-derived islet-like 3D tumoroids: the combination of buparlisib with ribociclib was effective in three out of four samples. Combined targeting of PI3K and CDK4/6 is a promising strategy for PanNENs spanning various molecular and histo-pathological features.

Two-Dimensional and Spheroid-Based Three-Dimensional Cell Culture Systems: Implications for Drug Discovery in Cancer

“…The treatment with 5-fluorouracil induced less cell death in spheroids of the HepG2 (hepatocarcinoma) cell line than in monolayers due to increased intercellular adhesion through E-cadherin and CD44v6, resistance due to differentiated phenotypes, and greater tolerance to stressful stimuli [22][23][24][25][26]. The inhibitory concentration of 50% (IC 50 ) for MCF-7 (breast cancer) spheroids showed an approximate 2.24-fold increase compared to the monolayer culture upon arsenic administration [23].…”

Three Dimensional Models of Endocrine Organs and Target Tissues Regulated by the Endocrine System

“…Cell lines such as BON-1, QGP-1 and NT-3 are widely used in pNEN research. Compared to 2D culture, spheroids from BON-1 and NT-3 display resistance or require higher concentrations of certain drugs to elicit effects on cellular viability and apoptosis [ 145 ]. These attributes of spheroids versus monolayer cultures are thought to arise from diffusion kinetics and hypoxia within the 3D aggregates, as mentioned above.…”

“…To date, the only pNEN organoid line was reported by Kawasaki et al and was one out of twenty-five NEN organoid lines established by the group [ 151 ]. Short-lived 3D tumoroids from human primary pNETs have been reported and used to investigate response to current treatments (such as sunitinib, everolimus and temezolomide) and identify novel therapies targeted against PI3K and CDK4/6 signaling pathways [ 145 , 152 ].…”