Therapeutic Targeting of the Cell Cycle in Pediatric, Adolescent and Young Adult Osteosarcoma

Abstract

Treating pediatric, adolescent, and young adult (AYA) osteosarcoma (OS) is challenging due to its aggressiveness, genetic complexity, lack of standard therapies, and chemotherapy long term side effects. Genetically guided therapies could enhance outcomes. This study explores palbociclib, an inhibitor of Cyclin-Dependent Kinases 4/6 (CDK4/6), targeting the cell cycle based on genomic insights. CDK4/6 forms complexes with cyclin D, facilitating retinoblastoma1 (RB1) phosphorylation, leading to RB1 dissociation from E2F transcription factor and promoting cell cycle progression. CDK4/6 inhibitor (CDK4/6i) monotherapy shows limited efficacy due to cytostatic responses and resistance through compensatory pathways like PI3K/mTOR pathway, common in OS. Hence, dual inhibition of CDK4/6 and PI3K/mTOR could be effective for OS treatment. In this study, OS patient-derived xenografts (PDX) models RHT-96 (from a treatment-naïve patient) and TT2-77 (from a pretreated patient with metastatic disease) were used. Molecular signatures (CDKN2A deletion, CCND3 amplification, RB1 proficiency) indicating sensitivity to CDK4/6i , were verified in both PDX models. Short-term palbociclib treatment in pretreated TT2-77 PDX upregulated PI3K/mTOR pathway via upstream growth factor receptors, highlighting the necessity for combination therapy with CDK4/6i. Both PDXs were treated long-term with CDK4/6i (50 mg/kg palbociclib), PI3K/mTOR inhibitor (50 mg/kg voxtalisib) or combination treatment. In both PDXs, combination treatment was more efficacious than single-agent following prolonged treatment and well-tolerated based on body weight and histological analyses. The increased efficacy of the combination treatment in the naïve RHT-96 PDX was associated with decreased pathway activity of PI3K/mTOR, and autophagy induction. In RB1 proficient OS cells, the combination treatment led to additive-to-synergistic growth inhibition, G1 arrest, and induced senescence and autophagy, as shown by senescence biomarker (beta-galactosidase) and autophagy markers. In the human OS lung colonization 143B model, combination treatment improved survival and reduced metastatic burden compared to the vehicle group, as observed in body scoring, quantification of human tumor cells, and histological analyses. Our data provide evidence that combining palbociclib and voxtalisib is safe, efficacious, and enhances palbociclib efficiency in both naïve and pretreated PDXs, as well as humanized lung colonization models of pediatric and AYA OS. This provides the rationale for earlier therapeutic intervention in patients with CDK4/6 hyperactivation signatures.