Pazopanib and Trametinib as a Synergistic Strategy against Osteosarcoma: Preclinical Activity and Molecular Insights
Giulia Chiabotto, Giovanni Grignani, Maja Todorovic, Valentina Martín, Maria Laura Centomo, Elisa Prola, G Giordano, Alessandra Merlini, Umberto Miglio, Enrico Berrino, Lucia Napione, Claudio Isella, Federica Capozzi, Marco Basiricò, Cristina Marsero, Ilaria Gerardi, Tiziana Venesio, Dario Sangiolo, Massimo Aglietta, Lorenzo D’Ambrosio, Ymera Pignochino
Abstract
Receptor tyrosine kinases (RTKs) inhibitors' activity in advanced osteosarcoma is significant but short-lived. To prevent or at least delay drug resistance, we explored a vertical inhibition by combining drugs acting at different levels of the RTK pathways (pazopanib + trametinib). We studied pazopanib + trametinib antitumor activity both in vitro and in vivo (MNNG-HOS and KHOS xenografts in NOD/SCID mice) investigating the molecular mechanisms and potential escapes. The involvement of MAPK-PI3K pathways was validated by Nanostring technology, western blot and by silencing/overexpression experiments. Pazopanib targets were expressed on seven osteosarcoma cell lines and their pathways were activated. Pazopanib + trametinib exhibited synergistic antitumor activity by inducing apoptosis and inhibiting ERK1/2 and Akt. In vivo antitumor activity was shown in osteosarcoma-bearing mice. The drug combination significantly down-modulated RTK Ephrin Type-A Receptor 2 (EphA2) and Interleukin-7 Receptor (IL-7R), whereas induced mitogen-activated protein-kinase kinase (MAPKK) MEK6. EphA2 silencing significantly reduced osteosarcoma cell proliferation and migration, while impeding MEK6 up-regulation in the treated cells significantly increased the antitumor effect of the studied drugs. Moreover, the up-regulation of MEK6 reduced combination activity. Pazopanib + trametinib demonstrated synergistic antitumor effects in osteosarcoma models through ERK and Akt inhibition and EphA2 and IL-7R down-modulation. MEK6 up-regulation might evoke escaping mechanism.