Oncogenic KRAS signaling is the main driving force behind pancreatic ductal adenocarcinoma (PDAC); however, targeting this pathway has proven to be difficult. Conversely, the PI3K/Akt pathway represents an exciting new target, because it has been associated with poor prognosis and chemoresistance, and several inhibitors are under development. In particular, perifosine prevents Akt translocation to the cell membrane, while MK-2206 is an Akt allosteric inhibitor and BEZ-235 is a dual PI3K/mTOR inhibitor. Therefore, we investigated the prognostic role of phospho (p)-Akt in PDAC tissues, as well as the molecular mechanisms underlying the interaction of Akt inhibitors with gemcitabine, using PDAC cells, primary cultures and spheroids.
Immunohistochemistry of tissue microarrays with specimens from radically-resected patients (n=100) revealed a correlation between high p-Akt1 expression and worse outcome. Patients with low p-Akt expression (as detected by digital scoring) had a median overall survival (OS) of 16.2 months (95% CI, 14.8-20.1), while patients with high expression had a median OS of 12.0 months (95% CI, 9.0-14.9, P=0.03). Parallel immunocytochemistry studies revealed high expression levels in LPC028 primary cells, while LPC006 were characterized by low p-Akt1. Akt inhibitors reduced cancer cell growth in monolayers and spheroids, and synergistically enhanced the antiproliferative activity of gemcitabine in LPC028 (e.g., combination index CI of 0.2, in the gemcitabine-perifosine combination for 72h, at fixed IC50 ratio), while this combination was antagonistic in LPC006 cells. The synergistic effect was paralleled by a 5-fold reduction in the expression of the main gemcitabine target ribonucleotide reductase. Inhibition of Akt decreased cell migration and invasion, which was additionally reduced by the combination with gemcitabine. However, the combination of Akt inhibitors with gemcitabine increased apoptosis, associated with induction of caspase-3/6/8/9, PARP and BAD, and inhibition of Bcl-2 and NF-kB in LPC028, but not in LPC006 cells. The Akt signaling is involved in the expression/localization of the key glucose transporter Glut1, and increased glucose metabolism was associated to resistance to axitinib (Hudson et al, Cell Death Dis 2014). Remarkably, the resistant LPC006 cells were characterized by overexpression of Glut1, which was not reduced after exposure to Akt inhibitors. However, the novel Glut1 inhibitor PGL13 enhanced perifosine and perifosine/gemcitabine-induced cell death. In conclusion, our findings support the analysis of phospho-Akt1 expression as both a prognostic and a predictive biomarker, for the rational development of new combination therapies targeting the Akt pathway in PDAC. Finally, inhibition of Glut1 might overcome resistance to these therapies and warrants further studies.
Citation Format: Daniela Massihnia, Amir Avan, Niccola Funel, Carlotta Granchi, Filippo Minutolo, Leticia Leon, Godefridus Peters, Elisa Giovannetti. Phospho-akt: a potential resistance marker to chemotherapy and a therapy-target to restore sensitivity in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4731. doi:10.1158/1538-7445.AM2017-4731