Molecular Profiling Of Blastic Plasmacytoid Dendritic CELL Neoplasm Reveals A Unique Pattern and Suggests Selective Sensitivity To NF-KB Pathway Inhibition
Authors: Piccaluga Pier Paolo , Sapienza Maria Rosaria , Fabio Fuligni , Claudio Agostinelli , Claudio Tripodo , Simona Righi , Laginestra Maria Antonella , Alessandro Pileri , Maura Rossi , Ricci Francesca , Gazzola Anna , Mannu Claudia , Ulbar Francesca , Arpinati Mario , Paulli Marco , Gibellini Davide , Pagano Livio , Pimpinelli Nicola , Cerroni Lorenzo , Croce Carlo M. , Facchetti Fabio , Pileri Stefano A 
Published: October 2019

Abstract
Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare disease of controversial origin recently recognized as a neoplasm deriving from plasmacytoid dendritic cells (pDCs). Nevertheless, it remains an orphan tumor with obscure biology and dismal prognosis.
In this study, we aimed to: 1) molecularly define the cellular counterpart of BPDCN and its relationship with other leukemias; 2) identify genes and cellular programs deregulated in the tumor; and 3) delineate novel potential therapeutic targets.
To address these issues we collected and studied by gene expression profile (GEP) 27 BPDCN cases as well as 8 samples of non neoplastic pDCs. Further, a panel of samples including, myeloid precursors (MPs, N=4), lymphoid precursors (LPs, N=9), acute myeloid leukemias (AMLs, N=132), and acute lymphoblastic leukemias (ALLs, N=155) was analyzed. Validation was performed by immunohistochemistry (IHC) on tissue-microarrays, while functional experiments were carried out by using the CAL-1 cell line (derived from a BPDCN case).
First, we recognized the cellular derivation of BPDCN, which proved to originate from the myeloid lineage and in particular from resting pDCs. Second, by comparing the GEP of BPDCN and resting pDCs, we identified genes and cellular programs deregulated in the tumor. Following, based on an integrated bio-informatic approach, including four different tools, we uncovered the aberrant activation of the NF-kB pathway that was confirmed in independent assays. Interestingly, among other molecules, we identified BCL2 and IRF4, two well known NFkB targets, as aberrantly upregulated in neoplastic samples and confirmed this observation by IHC. We then tested whether NFkB inhibition could represent a potential therapeutic strategy in this setting. We treated BPDCN cells ex vivowith either the proteasome inhibitor bortezomib or the selective IKKB inhibitor BMS-345541 and found them to be effective in inducing cell cycle arrest and apoptosis at relatively low dosage. By contrast, BPDCN cells turned out to be virtually insensitive to cytarabine, one of the most used drug in this condition. GEP and immunocytochemistry were then successfully used to prove that cell death was accompanied by NFkB shut-off.
In conclusion, we identified a molecular signature representative of the transcriptional abnormalities of BPDCN and developed a cellular model proposing the first molecular targeted therapeutic approach in the setting of this currently incurable disease.
Funding
This work was supported by AIRC (IG10519 and 5xMille10007, Prof. Pileri), Centro Interdipartimentale per la Ricerca sul Cancro “G. Prodi”, BolognAIL, RFO (Prof. Pileri, Prof. Piccaluga), FIRB Futura 2011 RBFR12D1CB (Prof. Piccaluga), Fondazione Cassa di Risparmio in Bologna, Fondazione della Banca del Monte e Ravenna, Progetto Strategico di Ateneo 2006 (Prof. Pileri and Dr. Piccaluga) and by MIUR (PRIN 2011, Prof. Facchetti and Prof. Pileri). The authors have no conflicting financial interests to declare.

Acknowledgments
The Authors obtained the CAL-1 cell line from Takahiro Maeda (tmaeda@net.nagasaki-u.ac.jp), Department of Laboratory Medicine, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan.

Disclosures:
No relevant conflicts of interest to declare.