Abstract
Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in childhood and adolescence. Refractory/relapsed RMS patients present a bad prognosis that combined with the lack of specific biomarkers impairs the development of new therapies. Here, we utilize dynamic BH3 profiling (DBP), a functional predictive biomarker that measures net changes in mitochondrial apoptotic signaling, to identify anti-apoptotic adaptations upon treatment. We employ this information to guide the use of BH3 mimetics to specifically inhibit BCL-2 pro-survival proteins, defeat resistance and avoid relapse. Indeed, we found that BH3 mimetics that selectively target anti-apoptotic BCL-xL and MCL-1, synergistically enhance the effect of clinically used chemotherapeutic agents vincristine and doxorubicin in RMS cells. We validated this strategy in vivo using a RMS patient-derived xenograft model and observed a reduction in tumor growth with a tendency to stabilization with the sequential combination of vincristine and the MCL-1 inhibitor S63845. We identified the molecular mechanism by which RMS cells acquire resistance to vincristine: an enhanced binding of BID and BAK to MCL-1 after drug exposure, which is suppressed by subsequently adding S63845. Our findings validate the use of DBP as a functional assay to predict treatment effectiveness in RMS and provide a rationale for combining BH3 mimetics with chemotherapeutic agents to avoid tumor resistance, improve treatment efficiency, and decrease undesired secondary effects.
Highlights
Rhabdomyosarcoma (RMS) is a highly malignant cancer that, despite being relatively rare, is the most frequent soft-tissue sarcoma in children, accounting for 5% of all pediatric tumors[1]
Using Annexin V and propidium iodide (PI) or DAPI staining, we analyzed by flow cytometry cell death after 96 h of exposure to the same chemotherapeutic agents as a proof of principle to evaluate the correlation between dynamic BH3 profiling (DBP) predictions and later cell death
We observed that the area under the curve (AUC) for our experiments was 0.81 (Fig. 1d), indicating that DBP presents a good predictive capacity for chemotherapy cytotoxicity in the RMS cell lines tested
Summary
Rhabdomyosarcoma (RMS) is a highly malignant cancer that, despite being relatively rare, is the most frequent soft-tissue sarcoma in children, accounting for 5% of all pediatric tumors[1]. There is a slightly higher prevalence of this disease in males than in females, and it is often associated with genetic disorders, such as Li–Fraumeni familiar cancer syndrome and RMS tumors are subdivided into two main groups, embryonal (ERMS) and alveolar (ARMS). ERMS account for 60% of all RMS, affecting children under the age of 10, especially around the head and neck regions[2,3]. The current treatment strategies for RMS include chemotherapy, radiation, and surgery[4]. The derived toxicities from current treatments and the lack of biomarkers[5], highlight the need for new therapies to enhance RMS clinical outcomes
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