David Van Mater

<jats:title>Abstract</jats:title> <jats:p>We developed a primary mouse model of embryonal rhabdomyosarcoma and discovered that tissue injury dramatically accelerates sarcoma formation at the site of injury. We were intrigued by this finding as many sarcoma patients anecdotally report a history of trauma prior to sarcoma development. We therefore utilized this mouse model system to explore the mechanism of injury-mediated sarcoma formation. Our preliminary findings suggest that the majority of muscle progenitor cells remain quiescent following mutation of genes with high transformation potential. However, tissue injury “flips a switch”, thereby pushing resting muscle progenitor cells into a proliferative phase. In the setting of a normal mouse, proliferation of muscle cells results in resolution of the injury in a matter of weeks. Conversely, when cells in the vicinity of injury harbor transformative gene mutations, the “switch” does not return to an off-state, and a tumor rapidly develops at the injury site.</jats:p> <jats:p>Our novel mouse model of embryonal rhabdomyosarcoma provides temporospatial control over the deletion of p53 and activation of Kras in muscle progenitor cells. Specifically, Pax7-CreERT2 (P7) mice express a tamoxifen-inducible Cre downstream of the endogenous promoter for the satellite cell transcription factor Pax7. P7 mice were crossed to genetically engineered mice containing a lox-STOP-lox cassette upstream of oncogenic K-rasG12D (K) in addition to 2 floxed p53 alleles (P) to generate P7KP mice. P7KP mice were injected with systemic, intraperitoneal (IP) tamoxifen to mediate Cre-dependent recombination in Pax7-expressing cells. When treated in this manner, sarcomas arise throughout the animal with 100% penetrance with a median onset of 45 days. In contrast, tumor onset is dramatically accelerated when P7KP mice are injured with cardiotoxin, a component of cobra venom that causes myonecrosis. P7KP mice treated with IP tamoxifen along with concurrent intramuscular (IM) cardiotoxin develop sarcomas at the site of injury with a median onset of 15 days. Cardiotoxin was next injected into the gastrocnemius muscle at time points either before, concurrent, or after IP tamoxifen to better define the effect of injury on sarcoma formation. Cardiotoxin promoted highly efficient transformation when administered up to 3 days before and up to 21 days after IP tamoxifen administration. Lineage tracing studies showed no appreciable proliferation of recombined Pax7+ cells in the absence of cardiotoxin, suggesting that sarcoma formation requires factors in addition to loss of p53 and activation of K-ras to cause sarcoma.</jats:p> <jats:p>In order to test the hypothesis that proliferating satellite cells are more prone to sarcoma formation, we treated P7KP mice in the gastrocnemius muscle with hepatocyte growth factor (HGF) along with systemic, IP tamoxifen. HGF has been shown to promote proliferation of resting satellite cells. Interestingly, 12/13 mice treated with IM HGF developed sarcomas at the injection site in a median of 37 days versus 3/12 of mice treated with vehicle control. Current experiments are aimed at dissecting the role of the immune system in the cardiotoxin response.</jats:p> <jats:p>Our experiments support the role of tissue injury acting as a classic promoter in the initiator/promoter model of tumorigenesis. Thus perturbation of the microenvironment has a dramatic effect on sarcoma formation. While the majority of cancer therapeutics are directed against mutations within the tumor, our findings reveal alternative candidates to prevent sarcoma initiation and perhaps maintenance. We anticipate that an understanding of signaling events at the earliest stages of sarcoma formation will provide new drug targets to treat existing tumors and potentially prevent the outgrowth of micrometastases.</jats:p> <jats:p>Citation Format: David Van Mater, Leonor Ano, Jordan Blum, David G. Kirsch. A role for injury in sarcomagenesis. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr A64.</jats:p>

Mutations in the gene CBL were first identified in adults with various myeloid malignancies. Some patients with juvenile myelomonocytic leukemia (JMML) were also noted to harbor mutations in CBL, but were found to have generally less aggressive disease courses compared to other forms of Ras pathway-mutant JMML. Importantly, and in contrast to most reports in adults, the majority of CBL mutations in JMML patients are germline with acquired uniparental disomy occurring in affected marrow cells. Here, we systematically studied a large cohort of 33 JMML patients with CBL mutations and found this disease to be highly diverse in presentation and overall outcome. Moreover, we discovered somatically-acquired CBL mutations in 15% of pediatric patients who presented with more aggressive disease. Neither clinical features nor methylation profiling were able to distinguish somatic CBL patients from germline CBL patients, highlighting the need for germline testing. Overall, we demonstrate that disease courses are quite heterogeneous even among germline CBL patients. Prospective clinical trials are warranted to find ideal treatment strategies for this diverse cohort of patients.

Adjuvant chemotherapy for osteosarcoma and Ewing sarcoma consists of conventional cytotoxic regimens that have changed little over the past decades. There is an urgent need for agents that are more effective and have less long-term toxicity. Receptor tyrosine kinases regulate cell growth and proliferation of these tumors, and small-molecule inhibitors for many of these kinases are now available. In this article, we review published phase II trials for patients with recurrent disease and highlight the pathways targeted by available agents, as well as the toxicity and efficacy results seen to date. We also discuss the difficulties in identifying biomarkers to facilitate rational patient selection, as well as published and proposed strategies for how these inhibitors can be combined with conventional chemotherapy or other targeted agents. It is hoped future trials can capitalize on this growing experience to optimize the use of this exciting class of agents.

BACKGROUND: Disruption of cell-cycle regulators is a potential therapeutic target for brain tumors in children and adolescents. The aim of this study was to determine the maximum tolerated dose (MTD) and describe toxicities related to palbociclib, a selective cyclin-dependent kinase 4/6 (CDK4/6) inhibitor in pediatric patients with progressive/refractory brain tumors with intact retinoblastoma protein. METHODS: Palbociclib was administered orally starting at 50 mg/m2 daily for the first 21 days of a 28-day course. Dose escalation was according to the Rolling-6 statistical design in less heavily (stratum I) and heavily pretreated (stratum II) patients, and MTD was determined separately for each group. Pharmacokinetic studies were performed during the first course, and pharmacodynamic studies were conducted to evaluate relationships between drug levels and toxicities. RESULTS: A total of 21 patients were enrolled on stratum I and 14 patients on stratum II. The MTD for both strata was 75 mg/m2 . Palbociclib absorption (mean Tmax between 4.9 and 6.6 h) and elimination (mean half-life between 11.3 and 19.5 h) were assessed. The most common toxicity was myelosuppression. Higher palbociclib exposure was associated with grade 3/4 neutropenia and leukopenia. Dose limiting toxicities included grade 4 neutropenia and grade 3 thrombocytopenia and dehydration. No patients had an objective response to palbociclib therapy. CONCLUSIONS: Palbociclib was safely administered to children and adolescents at a dosage of 75 mg/m2 for 21 consecutive days followed by seven days of rest in both strata. Future studies will establish its optimal utilization in pediatric patients with brain tumors.