Abstract B14: Activation of the MACC1/PIM/cMyc axis confers resistance to PI3K/mTOR inhibition in PIK3CA mutant NSCLC
Authors: Moore Gillian , Elbai Samira , Heavey Susan , Finn Stephen , O'Neill Michael , Gately Kathy 
Published: December 2020

Abstract
Introduction: The PI3K pathway is often dysregulated in lung cancer, making it an ideal therapeutic target. Pan- and isoform-specific inhibitors of the PI3K pathway are currently being evaluated in clinical trials. However, all patients treated thus far develop progressive disease due to the emergence of bypass signaling mechanisms. We developed a panel of lung cancer preclinical models to elucidate potential mechanisms of resistance to PI3K-mTOR inhibitor GDC-0980 (apitolisib). In-depth characterization of the PIK3CA mutant positive H1975 sensitive cells (H1975P) versus apitolisib-resistant cells (H1975GR) identified activation of MACC1 as an early marker of drug resistance and the subsequent activation of all three PIM kinases. Inhibition of MACC1 and PIM kinase by siRNAs and the effect of IBL-301 (a novel PIM/PI3K/mTOR inhibitor) were examined for effects on cell viability/proliferation and downstream signaling pathways in vitro.
Methods: Profiling of H1975P versus H1975GR cells was undertaken using a mRNA/lncRNA array (Arraystar Inc) to quantify the expression of 20,730 mRNAs and 40,173 lncRNAs. Concomitant changes were assessed in the IL6/STAT3 signaling pathway by a QPCR array, immunofluorescence, Western blot and miRNA arrays. The effect of MACC1 and PIM kinase inhibition by siRNAs and targeted agents were examined by Western blot and cell viability assays.
Results: MACC1 mRNA was upregulated 77 fold in H1975GR cells after 1-month exposure to apitolisib, and its regulated proteins HGF and MET were upregulated (10.06 and 1.65 fold, respectively). At 4 months MACC1 mRNA was increased by 44 fold, HGF 2 fold and MET 6.6 fold while several members of the IL-6/STAT3 pathway including c-Myc and PIM1, PIM2 and PIM3 kinases were also significantly activated (25.5, 19.8, 3.3 and 31.9 fold respectively). Results were also validated by Western blot. A miRNA signature of PI3K/mTOR inhibitor resistance was validated that can be used to monitor patients on treatment. Inhibition of PIM kinases by siRNA and targeted therapy restored sensitivity to PI3K-mTOR inhibitors. Novel triple-targeted therapy IBL-301 decreased PIM kinase, c-Myc and MACC1 expression. IC50 values of IBL-301 were significantly less for H1975GR cells (0.02μM) compared to H1975P cells (0.75μM), highlighting activated PIM kinase as a mechanism of resistance to PI3K-mTOR inhibition.
Conclusion: Activated MACC1, PIM kinases and their interacting partners play a role in resistance to PI3K-mTOR inhibitors in PIK3CA mutant NSCLC. Our data provide a rationale for combined or sequential targeting of PI3K-mTOR and PIM kinases to significantly delay resistance and provide durable treatment responses in patients with PIK3CA mutant lung tumors.
Citation Format: Gillian Moore, Samira Elbai, Susan Heavey, Stephen Finn, Michael O'Neill, Kathy Gately. Activation of the MACC1/PIM/cMyc axis confers resistance to PI3K/mTOR inhibition in PIK3CA mutant NSCLC [abstract]. In: Proceedings of the AACR Special Conference on Targeting PI3K/mTOR Signaling; 2018 Nov 30-Dec 8; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(10_Suppl):Abstract nr B14.