Abstract 2011: Targeting CDK4/6 inhibitor resistance in relapsed RB-proficient osteosarcoma patient-derived xenografts via PI3 Kinase/mTOR inhibition
Authors: Barghi Farinaz , Pandya Pankita H , Saadatzadeh M. Reza , Bijangi-Vishehsaraei Khadijeh , Bailey Barbara J. , Dobrota Erika A. , Young Courtney , Trowbridge Melissa A. , Coy Kathryn L. , Mang Henry , Wohlford Reagan K. , Sinn Anthony L. , Sims Emily C. , Repass Matt J. , Damayanti Nuri , Riyahi Niknam , Shannon Harlan E. , Angus Steve P , Ferguson Michael J , Renbarger Jamie L. , Pollok Karen E. 
Published: June 2022

Abstract
In children, adolescents, and young adults (AYA), osteosarcoma (OS) is the most common type of bone cancer and ~35% patients relapse following frontline cytotoxic therapy. Thus, there is a critical need to identify therapies targeting specific molecular signatures in OS. Hyperactivation of cyclin-dependent kinases 4 and 6 (CDK4/6) has been identified by us and others as a top actionable marker in OS. CDK4/6 binds to cyclin D resulting in a complex that mediates RB phosphorylation leading to cell cycle progression. While CDK4/6 inhibitors (CDK4/6i) have shown promise clinically, one drawback is that CDK4/6i induces cell cycle arrest rather than cell death. Furthermore, prolonged CDK4/6 inhibitor therapy can confer therapeutic resistance in RB1-proficient (RB+) tumors where compensatory pathways such as PI3K/mTOR are activated. To mitigate such CDK4/6i resistance in OS, we hypothesized that dual inhibition of CDK4/6 and PI3K pathways will promote cytotoxicity in hyperactivated CDK4/6 OS models. RB+ OS cell lines and a TT2-77 xenoline were evaluated in vitro. Combination index and Bliss independence analyses indicated that inhibition of OS growth by exposure to CDK4/6i (Palbociclib or Abemaciclib) and PI3K/mTOR inhibitor (PI3K/mTORi-Voxtalisib or LY3023414) was additive-to-synergistic and lead to increased apoptosis at clinically relevant concentrations. Short-term pharmacodynamic study of vehicle- versus Palbociclib-treated TT2-77 patient-derived xenograft (PDX) was analyzed by global/phospho-proteomics and kinome profiling. RB1 and MKI67 phosphopeptides as well as the total protein levels of CDK1 were reduced by Palbociclib, thus, confirming modulation of the cell cycle. Kinome profiling analysis of PDX from Palbociclib-treated mice indicated increased activity of AXL, a receptor tyrosine kinase linked to PI3K pathway activation. Increased activity of the autophagy marker PIK3C3 was also evident. OS PDX models TT2-77 and HT96 (RB+, CDKN2Anull, CCND3 amplified) were treated with Palbociclib (50 mg/kg), Voxtalisib (50 mg/kg) or Palbociclib + Voxtalisib. In TT2-77 PDXs treated for four weeks, tumor growth was significantly reduced in single-agent and combo groups compared to vehicle (p&lt;0.05, Two-way ANOVA; Holm-Sidak). We observed a trend that the combo was more efficacious than single agent, but statistical differences were not evident. Increasing the dosing timeline to six weeks may be beneficial. In HT96 PDXs, tumor growth was significantly decreased in single-agent and combo groups compared to vehicle. Notably, Palbociclib + Voxtalisib was more efficacious than single-agent (p&lt;0.05, Two-way ANOVA; Holm-Sidak). These data highlight the need to optimize CDK4/6i+PI3K/mTORi dosing schedules and provide evidence that Palbociclib + Voxtalisib therapy is safe, efficacious, and can decrease CDK4/6i resistance in aggressive PDX models of OS.
Citation Format: Farinaz Barghi, Pankita H Pandya, M. Reza Saadatzadeh, Khadijeh Bijangi-Vishehsaraei, Barbara J. Bailey, Erika A. Dobrota, Courtney Young, Melissa A. Trowbridge, Kathryn L. Coy, Henry Mang, Reagan K. Wohlford, Anthony L. Sinn, Emily C. Sims, Matt J. Repass, Nuri Damayanti, Niknam Riyahi, Harlan E. Shannon, Steve P Angus, Michael J Ferguson, Jamie L. Renbarger, Karen E. Pollok. Targeting CDK4/6 inhibitor resistance in relapsed RB-proficient osteosarcoma patient-derived xenografts via PI3 Kinase/mTOR inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2011.