Osteosarcoma patient-derived xenografts derived from naive and pretreated metastatic patients with high-risk CDK4/6 hyperactivation signatures are sensitive to dual inhibition of CDK4/6 and PI3K/mTOR

Abstract Precision genomics studies have demonstrated hyperactivation of cyclin-dependent kinases 4 and 6 (CDK4/6) as a top actionable marker in children, as well as adolescents and young adults (AYA) with osteosarcoma (OS). CDK4/6 binds to cyclin D resulting in a complex that mediates RB phosphorylation leading to cell cycle progression. Preclinical modeling approaches are critical for identification of tumor adaptive responses to CDK4/6 inhibitors (CDK4/6i) as well as validation of alternative or combination therapies. Although CDK4/6i are clinically well-validated, cytostatic effects make combination treatments essential. Moreover, concomitant dysregulation of CDK4/6 and PI3K/mTOR pathways are observed in aggressive OS. Multiple positive feedback loops between these pathways exacerbate the hyperactivation of CDK4/6 and PI3K/mTOR signaling. Thus, we hypothesize that dual inhibition of CDK4/6 and PI3K/mTOR will be efficacious in RB+ OS PDXs. In this study, OS PDX models TT2-77 (pretreated patient) and HT96 (treatment-naïve patient) with molecular signatures indicative of therapeutic sensitivity to palbociclib (RB+, CDKN2A null, CCND3 amplified) were treated long-term with CDK4/6i (palbociclib) (50 mg/kg), PI3K/mTOR inhibitor (PI3K/mTORi; voxtalisib) (50 mg/kg) or combination palbociclib+voxtalisib. In both PDXs, growth was significantly reduced in single-agent and combination groups compared to vehicle (p<0.05, two-way ANOVA). Importantly, combination palbociclib + voxtalisib was more efficacious than single-agents following prolonged treatment and well tolerated based on histological analyses. Kinome profiling analysis of long-term treated HT96 PDX demonstrated that compared to single agents, dual inhibition of CDK4/6+PI3K/mTOR significantly decreased PI3K pathway activity, including downregulation of Pik3ca, mTOR, and the G2 to M transition regulator CDK1 (-log10[p] ≥1.3). OS metastatic lesion 143B model indicated increased survival based on body scoring criteria in combo versus single agent. In RB+ OS cell lines and TT2-77 xenoline, palbociclib+voxtalisib caused additive-to-synergistic cell growth inhibition, G1 arrest, and minimal apoptosis at clinically relevant doses. Increased activity of senescence biomarker beta-galactosidase indicated that inhibition of CDK4/6 but not PI3K/mTOR induced significant levels of senescence in OS cells. Mechanistic siRNA RB studies indicated CDK4/6i effect was partially dependent on RB status. These data provide evidence that combination palbociclib+voxtalisib therapy is safe, efficacious, and increases CDK4/6i efficiency in both pretreated and naive PDX models of OS. These studies provide rationale for earlier therapeutic intervention in pediatric and AYA OS patients with CDK4/6 hyperactivation signatures. Citation Format: Farinaz Barghi, M. Reza Saadatzadeh, Erika Dobrota, Rada Malko, Barbara J Bailey, Courtney Young, Harlan E. Shannon, Ryli Justice, Niknam Riyahi, Khadijeh Bijangi-Vishehsaraei, Melissa Trowbridge, Kathy Coy, Felicia M Kennedy, Anthony L Sinn, Amber Mosley, Steve Angus, Michael J. Ferguson, Pankita H. Pandya, Karen E. Pollok. Osteosarcoma patient-derived xenografts derived from naive and pretreated metastatic patients with high-risk CDK4/6 hyperactivation signatures are sensitive to dual inhibition of CDK4/6 and PI3K/mTOR. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6728.