Romidepsin (FK228) fails in counteracting the transformed phenotype of rhabdomyosarcoma cells but efficiently radiosensitizes, in vitro and in vivo, the alveolar phenotype subtype

Abstract

Purpose

Herein we describe the in vitro and in vivo activity of FK228 (Romidepsin), an inhibitor of class I HDACs, in counteracting and radiosensitizing embryonal (ERMS, fusion-negative) and alveolar (ARMS, fusion-positive) rhabdomyosarcoma (RMS).

Methods

RH30 (ARMS, fusion-positive) and RD (ERMS, fusion-negative) cell lines and human multipotent mesenchymal stromal cells (HMSC) were used. Flow cytometry analysis, RT-qPCR, western blotting and enzymatic assays were performed. Irradiation was delivered by using an x-6 MV photon linear accelerator. FK228 (1.2 mg/kg) in vivo activity, combined or not with radiation therapy (2 Gy), was assessed in murine xenografts.

Results

Compared to HMSC, RMS expressed low levels of class I HDACs. In vitro, FK228, as single agents, reversibly downregulated class I HDACs expression and activity and induced oxidative stress, DNA damage and a concomitant growth arrest associated with PARP-1-mediated transient non-apoptotic cell death. Surviving cells upregulated the expression of cyclin A, B, D1, p27, Myc and activated PI3K/Akt/mTOR and MAPK signaling, known to be differently involved in cancer chemoresistance. Interestingly, while no radiosensitizing effects were detected, in vitro or in vivo, on RD cells, FK228 markedly radiosensitized RH30 cells by impairing antioxidant and DSBs repair pathways in vitro. Further, FK228 when combined with RT in vivo significantly reduced tumor mass in mouse RH30 xenografts.

Conclusion

FK228 did not show antitumor activity as a single agent whilst its combination with RT resulted in radiosensitization of fusion-positive RMS cells, thus representing a possible strategy for the treatment of the most aggressive RMS subtype.

Keywords:

• Rhabdomyosarcoma
• radiotherapy
• FK228
• romidepsin
• radioresistance

Acknowledgments

SC thanks AIRC for support through a fellowship for Italy “Raffaele Anastasio e Lucia Rizzetta in Anastasio” (Rif. 24017) and SP thanks Fondazione Veronesi for providing support.

Ethical approval

This article does not contain any studies with human participants performed by any of the authors. Studies on animal models were performed according to the guidelines established by the University of L’Aquila, Medical School and Science and Technology School Board Regulations, in compliance with the Italian government regulation n.116 27 January 1992 for the use of laboratory animals.

Informed consent

This article does not contain any studies with human participants performed by any of the authors.

Author contributions

AR, FP, LM, FV and GM conceptualized, designed the study, performed the experiments and acquired data. SC, SP, MC, IF and CA performed the experiments and acquired result data. FC, LM, FV and GM performed irradiation. LDL, RM, CF, FDF and MV analyzed and interpre-ted the data. AF, RR, AP, VT, GLG and FM drafted the manuscript and critically revised the manuscript. All authors read and approved the final version of the manuscript.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

The research leading to these results has received funding from AIRC under IG 2020 project [24696] – P.I. Marampon Francesco. This work was also supported by the University of Roma, Sapienza (Fondi Ateneo 2019).

Notes on contributors

Alessandra Rossetti

Alessandra Rossetti graduated as a Biotechnologist from the University of L’Aquila, and has a PhD from the University of L’Aquila, L’Aquila, Italy.

Francesco Petragnano

Francesco Petragnano graduated as a Biotechnologist from the University of L’Aquila, and has a PhD from the University of L’Aquila, L’Aquila, Italy.

Luisa Milazzo

Luisa Milazzo graduated as a Biologist from the University of Rome, Sapienza, works at Istituto Superiore della Sanità Italiana, Rome, Italy.

Francesca Vulcano

Francesca Vulcano graduated as a Biologist from the University of Rome, Sapienza, works at Istituto Superiore della Sanità Italiana, Rome, Italy.

Giampiero Macioce

Giampiero Macioce is a Technician at the Istituto Superiore della Sanità Italiana, Rome, Italy.

Silvia Codenotti

Silvia Codenotti graduated as a Biotechnologist from the University of Brescia, and has a PhD from the University of Brescia, Brescia, Italy.

Matteo Cassandri

Matteo Cassandri graduated as a Biologist from the University of Rome, Sapienza, and is a Post-Doc at the Department of Oncohematology, Bambino Gesù Children’s Hospital, Rome, Italy.

Silvia Pomella

Silvia Pomella graduated as a Biologist from the University Alma Mater Studiorum of Bologna, and is a Post-Doc at Department of Oncohematology, Bambino Gesù Children’s Hospital, Rome, Italy.

Francesca Cicchetti

Francesca Cicchetti is a Radiologic Technologist at Policlinico Umberto I, Rome, Italy.

Irene Fasciani

Irene Fasciani graduated as a Biotechnologist from the University of L’Aquila, and is a Post-Doc at University of L’Aquila, L’Aquila, Italy.

Cristina Antinozzi

Cristina Antinozzi graduated as a Biologist from the University of Rome, Sapienza, and is a Post-Doc at University of Rome “Foro Italico”, Rome, Italy.

Luigi Di Luigi

Luigi Di Luigi graduated as Medical Doctor specialized in Endocrinology and in Sports Medicine from the University of Rome, Sapienza, and is a Full Professor of Endocrinology at University of Rome “Foro Italico”, Rome, Italy.

Claudio Festuccia

Claudio Festuccia graduated as a Biologist from the University of L’Aquila, and is a Technician at University of L’Aquila, L’Aquila, Italy.

Francesca De Felice

Francesca De Felice graduated as Medical Doctor specialized in Radiotherapy from the University of Rome, Sapienza, is a Researcher of Radiotherapy at University of Rome, Sapienza, Rome, Italy.

Massimo Vergine

Massimo Vergine graduated as Medical Doctor specialized in Abdominal Surgery from the University of Rome, Sapienza, is a Researcher of Surgery at University of Rome, Sapienza, Rome, Italy.

Alessandro Fanzani

Alessandro Fanzani graduated as a Biologist from the University of Milano, and is an Associate Professor at University of Brescia, Brescia, Italy.

Rossella Rota

Rossella Rota graduated as a Biologist from the University of L’Aquila, and is a Group Leader at Bambino Gesù Children’s Hospital, Rome, Italy.

Roberto Maggio

Roberto Maggio graduated as a Medical Doctor specialized in Pharmacology from the University of Cagliari, and is a Full Professor of Pharmacology at University of L’Aquila, L’Aquila, Italy.

Antonella Polimeni

Antonella Polimeni graduated as a Medical Doctor specialized in Odontostomatology from the University of Rome, Sapienza, and is a Full Professor of Odontostomatology at University of Rome, Sapienza, Rome, Italy.

Vincenzo Tombolini

Vincenzo Tombolini graduated as a Medical Doctor specialized in Radiotherapy from the University of Rome, Sapienza, and is a Full Professor of Radiotherapy at University of Rome, Sapienza, Rome, Italy.

Giovanni Luca Gravina

Giovanni Luca Gravina graduated as a Medical Doctor specialized in Radiotherapy from the University of L’Aquila, and is an Associate Professor of Radiotherapy at University of L’Aquila, L’Aquila, Italy.

Francesco Marampon

Francesco Marampon graduated as a Medical Doctor specialized in Radiotherapy from the University of L’Aquila, and is an Associate Professor of Radiotherapy at University of Rome, Sapienza, Rome, Italy.