ECE2023 Rapid Communications Rapid Communications 7: Pituitary and Neuroendocrinology 2 (6 abstracts)
Splicing factor 3B subunit 1 (SF3B1) role in PRL-secreting PitNETs
Federica Mangili 1 , Genesio Di Muro 2,3 , Federico Arlati 2 , Anna Maria Barbieri 2 , Miguel E. G-Garcia 4,5,6 , Emanuela Esposito 7 , Donatella Treppiedi 1 , Rosa Catalano 2 , Giusy Marra 2 , E mma Nozza 7 , Marco Locatelli 8,9 , Andrea Lania 10,11 , Emanuele Ferrante 12 , Elisa Sala 12 , Justo P. Castaño 4,5,6 , Raul M. Luque 4,5,6 , Maura Arosio 12 , Erika Peverelli 2,12 & Giovanna Mantovani 2,12
1Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico dImilano, Endocrinology and Diabetology UO, Milan, Italy; 2University of Milan, Department of Clinical Sciences and community Health, Milan, Italy; 3Sapienza University of Rome, PhD program in Endocrinology Science, Rome, Rome, Italy; 4Maimonides Biomedical Research Institute of Cordoba (IMIBIC), Cordoba, Spain; 5University of Cordoba, Department of Cell Biology, Physiology and Immunology, Cordoba, Spain; 6Reina Sofia University Hospital, Cordoba, Spain; 7University of Milan, PhD program in experimental medicine, Department of Clinical Sciences and community Health, Milan, Italy; 8University of Milan, Department of Pathophysiology and Transplantation, Milan, Italy; 9Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico dImilano, Neurosurgery Unit, Milan, Italy; 10IRCCS Humanitas Research Hospital, Endocrinology, Diabetology and Medical Andrology Unit, Milan, Italy; 11Humanitas University, Department of Biomedical Sciences, Milan, Italy; 1Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico dImilano, Endocrinology and Diabetology UO, Milan, Italy
Dopamine receptor type 2 (DRD2) represents the main target for pharmacological therapy with dopamine agonists (DAs) in PRL-secreting neuroendocrine pituitary tumors (PRL-PitNET), even if about 10% of patients is resistant. A single paper recently described a somatic mutation in the gene encoding splicing factor 3B subunit 1 (SF3B1) in about 20% of patients with PRL-PitNET, that was associated with PRL hypersecretion, increased cell proliferation and invasion and reduced progression free survival. SF3B1 is a key component of pre-mRNA processing, crucial for splicing machinery assembly. Aims of the study were: 1) to characterize the genetic profile of PRL-PitNET, searching for somatic mutations in SF3B1R625H hotspot region; 2) to test in tumoral lactotroph cells the effects of SF3B1 inhibitor pladienolide-B and the role of SF3B1 in affecting DRD2 agonist cabergoline antitumoral effects. SF3B1R625H hot-spot region has been sequenced in PRL-PitNET tissues and rat PRL-secreting pituitary tumoral cells mmQ. SF3B1 was inhibited with pladienolide-B or silenced by siRNA technique, and cells have been analysed to measure cell proliferation, apoptosis, secretion and cell cycle markers activation. No mutation in SF3B1 gene was found in our cohort of PRL-PitNET (n=40), nor in rat lactotroph mmQ cells. Pladienolide-B 50 nM treatment strongly reduced cell proliferation (-92±5%, P<0.001) and increased apoptosis (+10-fold, P<0.001) in mmQ, and similar results were found in primary PRL-PitNET cultured cells, wild-type for SF3B1. Moreover, pladienolide-B reduced PRL secretion in primary cells (-45±6.5%, P<0.001). SF3B1 silencing in mmQ cells abolished cabergoline inhibitory effects on cell proliferation (-22±4.8%, P<0.001), AKT phosphorylation (-31±24.6%, P<0.01), cyclin D3 (-23±7.6%, P<0.05) and increase of p27 levels (+20±8.6%, P<0.05). In addition, SF3B1 knock down was associated with DRD2 reduction (-66.4±8% of DRD2 transcript, P<0.05, and -51±13.2% of DRD2 protein, P<0.001), and the same effects were observed after pladienolide-B incubation. On the other hand, cabergoline reduced SF3B1 protein expression levels in mmQ (-60±40%, P<0.05) and in 2 PRL-PitNET (-43±6.1%, P<0.01). Our data did not confirm the presence of SF3B1R625H mutations in a multicentre cohort of PRL-PitNET. In contrast, we observed that SF3B1 seems to play a relevant role in tumoral lactotroph cells. Indeed, SF3B1 inhibitor pladienolide-B exerted antiproliferative, proapoptotic and antisecretory effects in PRL-PitNET cells wild-type for SF3B1. Moreover, SF3B1 is required for expression and signalling of DRD2.
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Endocrine Abstracts
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