Given the critical role of the PI3K/Akt pathway in cell growth and survival, it is not surprising that constitutive activation of this pathway contributes to the pathogenesis of many types of lymphoid malignancies, including mantle cell lymphoma (MCL), follicular lymphoma (FL), and cutaneous T-cell lymphoma (CTCL). Available drugs aimed to strike the PI3K/Akt pathway in these tumors are unfortunately dampened by relevant toxicities. Therefore, more effective and safer therapeutic options targeting Akt are needed. Herein we demonstrated that the combination of 9-cis-retinoic acid (RA) and Interferon-alpha (IFN-\u3b1) induces marked anti-proliferative and pro-apoptotic effects in MCL cells through the modulation of critical targets. Particularly, IFN-\u3b1 enhances 9-cis-RA-mediated G0/G1 cell accumulation by down-regulating cyclin D1 and increasing p27Kip1 and p21WAF1/Cip1 protein levels. Furthermore, 9-cis-RA/IFN-\u3b1 combination induces MCL apoptosis by triggering both caspase 8 and caspase 9 resulting in Bax and Bak activation, and up-regulating the pro-apoptotic proteins Noxa and PLSCR1. In particular, sequestration of the anti-apoptotic proteins Mcl-1 and A1/Bfl1 by up-regulated Noxa results in the activation of Bid, and the consequent induction of apoptosis is inhibited by Noxa silencing. PLSCR1silencing demonstrated a role for this protein not only in 9-cis-RA/IFN-\u3b1-induced apoptosis but also in 9-cis-RA/IFN-\u3b1-dependent sensitization to anti-tumor agents currently used in the clinical practice for MCL management, such as doxorubicin and bortezomib. These drugs are able to further increase PLSCR1 expression in 9-cis-RA/IFN-\u3b1 pre-treated MCL cells. Moreover, immunohistochemical analysis of MCL tumor biopsies and primary cultures revealed a variable expression of endogenous PLSCR1 in this setting, an heterogeneity that stimulates the search of possible correlations with clinical-pathological parameters, particularly with those related to the response to pro-apoptotic drugs. In future perspective, in fact, analysis of PLSCR1 expression might allow the identification of tumors more prone to undergo apoptosis, and strategies able to up-regulate PLSCR1, like 9-cis-RA/IFN-\u3b1 combination, might successfully complement and improve conventional treatment modalities. Notably, we also found that 9-cis-RA/IFN-\u3b1-induced PLSCR1 up-regulation occurs through STAT1 activation in dependence on Akt pathway. In addition, we demonstrated that 9-cis-RA/IFN-\u3b1 co-treatment is able to trigger apoptotic effects also in FL and CTCL cells, and, more interesting, the 9-cis-RA/IFN-\u3b1-dependent apoptosis is associated with the inhibition of Akt constitutive activation in the different NHL histotypes analyzed. In particular, Noxa up-regulation in MCL, FL, and CTCL lymphoma cells is associated with nuclear translocation of the FOXO3a transcription factor as a consequence of the 9-cis-RA/IFN-\u3b1-induced Akt but not of mTOR inhibition. Indeed, pharmacological suppression of Akt, but not of TORC1, induces apoptosis in FL cell lines, and increases Noxa protein levels in MCL cells, supporting the conclusion that inhibition of the Akt pathway, the resulting FOXO3a activation, and Noxa up-regulation are the critical molecular mechanisms underlying 9-cis-RA/IFN-\u3b1-induced cell death in different type of lymphoid malignancies. These results strengthen the role of Akt as a clinically relevant molecular target and support the potential therapeutic value of RA/IFN-\u3b1 combination to improve the management of Akt-driven non-Hodgking lymphoma
