Abstract 4439: Targeting FANCD2 as a radiosensitizer in pediatric rhabdomyosarcoma.

Abstract

Purpose. Alveolar rhabdomyosarcoma that harbors the PAX3/FOXO1 translocation (t-ARMS) is the most common and lethal subtype of this childhood malignancy. In this report we identify FANCD2 as a potential mediator of radiation resistance and biomarker of increased virulence in t-ARMS. Methods. After the observation that knockdown of mTOR resulted in a loss of FANCD2 expression, we postulated that pharmacologic inhibition of TORC1/TORC2 would result in a similar suppression of this DNA-repair protein and increase cell-kill after radiotherapy. Therefore, an initial set of mice bearing t-ARMS and embryonal rhabdomyosarcoma (ERMS) xenografts were subjected to a dose escalation study, followed by a standard dosing study which tested the concurrent administration of AZD8055 (an mTOR kinase inhibitor) and radiotherapy versus either treatment alone. Molecular studies, focusing on drug action and resultant FANCD2 expression and function were also performed on xenografts treated in parallel. We also performed clonogenic survival assays in RMS cell lines with and without FANCD2 knockdown to determine the impact of this protein on radiation sensitivity in these lines. Lastly, we tested the prognostic significance of FANCD2 in the tumors from two independent patient sets. Results. In a dose escalation study of 30 mice, we first observed the selective radiosensitization within the t-ARMS xenograft line versus the ERMS line. In a second set of 80 mice, subjected to standardized treatment arms, validated this selective radiosensitization. To verify the desired drug action, we analyzed xenografts from mice treated in parallel (collected at various points in treatment) and found that FANCD2 expression and activity is significantly suppressed by the addition of the drug in the sensitized, t-ARMS, but much less so in the ERMS xenograft line. We further observed that direct FANCD2 suppression (via siRNA knockdown) confers sensitivity to radiation therapy in both of these lines in vitro. In the initial patient set (n=108), we show that, on a protein level, FANCD2 expression correlates with PAX3/FOXO1 translocation and that the presence of this marker is additive to the poor prognosis of a translocated status. The co-existence of these two biomarkers is independently prognostic from other known clinical variables and the PAX3/FOXO1 translocation itself. In our second patient set (n=101), we show that based on the mRNA expression of FANCD2, that quartile-FANCD2 levels are prognostic in a univariate analysis, significantly associated with metastatic (Stage 4) disease and that, again, the co-presence of the translocation and high expression of FANCD2 is independently prognostic. Conclusion. Our data demonstrate that FANCD2 may have a significant role in the radiation resistance and virulence of t-ARMS, and indirectly targeting this DNA repair protein, through mTOR inhibition, may represent a novel and selective treatment strategy. Citation Format: Mamata Singh, Justin Leasure, Christopher Chronowski, Ning Li, Kathryn Bondra, Wenrui Duan, Miguel Villalona, Anthony Vergis, Lauren Hensley, Rita Kaplon, Brian Geier, Raushan Kurmasheva, Gary Woods, Sue Hammond, Peter Houghton, Christopher E. Pelloski. Targeting FANCD2 as a radiosensitizer in pediatric rhabdomyosarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4439. doi:10.1158/1538-7445.AM2013-4439

Case report: Highly response to low-dose brachytherapy in recurrent retroperitoneal leiomyosarcoma with FANCD2 frameshift mutation: a unique case study

“…Additionally, aberrations were also found in members of the Fanconi anemia complementation groups, specifically FANCA (27%) and FANCD2 (10%).The most promising approaches for advanced leiomyosarcoma include those targeting the DDR pathway (9). Research demonstrates FANCD2 plays a role in its resistance to treatment in alveolar rhabdomyosarcoma and a specific inhibition of FANCD2 leading to increased sensitivity to radiation in the PAX3-FOXO1 fusion gene cell line (10). However, there is still a lack of research on the effects of FANCD2 deletion and genetic mutations on tumor sensitivity to radiotherapy.…”