While most pediatric sarcomas respond to front-line therapy, some bone sarcomas do not show radiographic response like soft-tissue sarcomas (rhabdomyosarccomas) but do show 90% necrosis. Though, new therapies are urgently needed to improve survival and quality of life in pediatric patients with sarcomas. Complex chromosomal aberrations such as amplifications and deletions of DNA sequences are frequently observed in pediatric sarcomas. Evaluation of copy number variations (CNVs) associated with pediatric sarcoma patients at the time of diagnosis or following therapy offers an opportunity to assess dysregulated molecular targets and signaling pathways that may drive sarcoma development, progression, or relapse. The objective of this study was to utilize publicly available data sets to identify potential predictive biomarkers of chemotherapeutic response in pediatric Osteosarcoma (OS), Rhabdomyosarcoma (RMS) and Ewing’s Sarcoma Family of Tumors (ESFTs) based on CNVs following chemotherapy (OS n = 117, RMS n = 64, ESFTs n = 25 tumor biopsies). There were 206 CNV profiles derived from pediatric sarcoma biopsies collected from the public databases TARGET and NCBI-Gene Expression Omnibus (GEO). Through our comparative genomic analyses of OS, RMS, and ESFTs and 22,255 healthy individuals called from the Database of Genomic Variants (DGV), we identified CNVs (amplifications and deletions) pattern of genomic instability in these pediatric sarcomas. By integrating CNVs of Cancer Cell Line Encyclopedia (CCLE) identified in the pool of genes with drug-response data from sarcoma cell lines (n = 27) from Cancer Therapeutics Response Portal (CTRP) Version 2, potential predictive biomarkers of therapeutic response were identified. Genes associated with survival and/recurrence of these sarcomas with statistical significance were found on long arm of chromosome 8 and smaller aberrations were also identified at chromosomes 1q, 12q and x in OS, RMS, and ESFTs. A pool of 63 genes that harbored amplifications and/or deletions were frequently associated with recurrence across OS, RMS, and ESFTs. Correlation analysis of CNVs from CCLE with drug-response data of CTRP in 27 sarcoma cell lines, 33 CNVs out of 63 genes correlated with either sensitivity or resistance to 17 chemotherapies from which actionable CNV signatures such as IGF1R, MYC, MAPK1, ATF1, and MDM2 were identified. These CNV signatures could potentially be used to delineate patient populations that will respond versus those that will not respond to a particular chemotherapy. The large-scale analyses of CNV-drug screening provides a platform to evaluate genetic alterations across aggressive pediatric sarcomas. Additionally, this study provides novel insights into the potential utilization of CNVs as not only prognostic but also as predictive biomarkers of therapeutic response. Information obtained in this study may help guide and prioritize patient-specific therapeutic options in pediatric bone and soft-tissue sarcomas.

In cancer, the mouse double minute 2 (MDM2) is an oncoprotein that contributes to the promotion of cell growth, survival, invasion, and therapeutic resistance. The impact of MDM2 on cell survival versus cell death is complex and dependent on levels of MDM2 isoforms, p53 status, and cellular context. Extensive investigations have demonstrated that MDM2 protein–protein interactions with p53 and other p53 family members (p63 and p73) block their ability to function as transcription factors that regulate cell growth and survival. Upon genotoxic insults, a dynamic and intricately regulated DNA damage response circuitry is activated leading to release of p53 from MDM2 and activation of cell cycle arrest. What ensues following DNA damage, depends on the extent of DNA damage and if the cell has sufficient DNA repair capacity. The well-known auto-regulatory loop between p53-MDM2 provides an additional layer of control as the cell either repairs DNA damage and survives (i.e., MDM2 re-engages with p53), or undergoes cell death (i.e., MDM2 does not re-engage p53). Furthermore, the decision to live or die is also influenced by chromatin-localized MDM2 which directly interacts with the Mre11-Rad50-Nbs1 complex and inhibits DNA damage-sensing giving rise to the potential for increased genome instability and cellular transformation.

Osteosarcoma (OS) patients exhibit poor overall survival, partly due to copy number variations (CNVs) resulting in dysregulated gene expression and therapeutic resistance. To identify actionable prognostic signatures of poor overall survival, we employed a systems biology approach using public databases to integrate CNVs, gene expression, and survival outcomes in pediatric, adolescent, and young adult OS patients. Chromosome 8 was a hotspot for poor prognostic signatures. The MYC-RAD21 copy number gain (8q24) correlated with increased gene expression and poor overall survival in 90% of the patients (n = 85). MYC and RAD21 play a role in replication-stress, which is a therapeutically actionable network. We prioritized replication-stress regulators, bromodomain and extra-terminal proteins (BETs), and CHK1, in order to test the hypothesis that the inhibition of BET + CHK1 in MYC-RAD21+ pediatric OS models would be efficacious and safe. We demonstrate that MYC-RAD21+ pediatric OS cell lines were sensitive to the inhibition of BET (BETi) and CHK1 (CHK1i) at clinically achievable concentrations. While the potentiation of CHK1i-mediated effects by BETi was BET-BRD4-dependent, MYC expression was BET-BRD4-independent. In MYC-RAD21+ pediatric OS xenografts, BETi + CHK1i significantly decreased tumor growth, increased survival, and was well tolerated. Therefore, targeting replication stress is a promising strategy to pursue as a therapeutic option for this devastating disease.

Establishment of clinically annotated, molecularly characterized, patient-derived xenografts (PDXs) from treatment-naïve and pretreated patients provides a platform to test precision genomics-guided therapies. An integrated multi-OMICS pipeline was developed to identify cancer-associated pathways and evaluate stability of molecular signatures in a panel of pediatric and AYA PDXs following serial passaging in mice. Original solid tumor samples and their corresponding PDXs were evaluated by whole-genome sequencing, RNA-seq, immunoblotting, pathway enrichment analyses, and the drug–gene interaction database to identify as well as cross-validate actionable targets in patients with sarcomas or Wilms tumors. While some divergence between original tumor and the respective PDX was evident, majority of alterations were not functionally impactful, and oncogenic pathway activation was maintained following serial passaging. CDK4/6 and BETs were prioritized as biomarkers of therapeutic response in osteosarcoma PDXs with pertinent molecular signatures. Inhibition of CDK4/6 or BETs decreased osteosarcoma PDX growth (two-way ANOVA, p < 0.05) confirming mechanistic involvement in growth. Linking patient treatment history with molecular and efficacy data in PDX will provide a strong rationale for targeted therapy and improve our understanding of which therapy is most beneficial in patients at diagnosis and in those already exposed to therapy.

Despite improved therapeutic and clinical outcomes for patients with localized diseases, outcomes for pediatric and AYA sarcoma patients with high-grade or aggressive disease are still relatively poor. With advancements in next generation sequencing (NGS), precision medicine now provides a strategy to improve outcomes in patients with aggressive disease by identifying biomarkers of therapeutic sensitivity or resistance. The integration of NGS into clinical decision making not only increases the accuracy of diagnosis and prognosis, but also has the potential to identify effective and less toxic therapies for pediatric and AYA sarcomas. Genome and transcriptome profiling have detected dysregulation of the CDK4/6 cell cycle regulatory pathway in subpopulations of pediatric and AYA OS, RMS, and EWS. In these patients, the inhibition of CDK4/6 represents a promising precision medicine-guided therapy. There is a critical need, however, to identify novel and promising combination therapies to fight the development of resistance to CDK4/6 inhibition. In this review, we offer rationale and perspective on the promise and challenges of this therapeutic approach.

Cancer cells tend to develop resistance to chemotherapy and enhance aggressiveness. A counterintuitive approach is to tame aggressiveness by an agent that acts opposite to chemotherapeutic agents. Based on this strategy, induced tumor-suppressing cells (iTSCs) have been generated from tumor cells and mesenchymal stem cells. Here, we examined the possibility of generating iTSCs from lymphocytes by activating PKA signaling for suppressing the progression of osteosarcoma (OS). While lymphocyte-derived CM did not present anti-tumor capabilities, the activation of PKA converted them into iTSCs. Inhibiting PKA conversely generated tumor-promotive secretomes. In a mouse model, PKA-activated CM suppressed tumor-induced bone destruction. Proteomics analysis revealed that moesin (MSN) and calreticulin (Calr), which are highly expressed intracellular proteins in many cancers, were enriched in PKA-activated CM, and they acted as extracellular tumor suppressors through CD44, CD47, and CD91. The study presented a unique option for cancer treatment by generating iTSCs that secret tumor-suppressive proteins such as MSN and Calr. We envision that identifying these tumor suppressors and predicting their binding partners such as CD44, which is an FDA-approved oncogenic target to be inhibited, may contribute to developing targeted protein therapy.