Abstract LB-213: The antitumor effect of trabectedin (TR) is potentiated by olaparib (OL) in preclinical models of bone and soft tissue sarcomas (STS).
Authors: Pignochino Ymera , Capozzi Federica , Dell'aglio Carmine , Basiricò Marco , D'ambrosio Lorenzo , Galizia Danilo , Palesandro Erica , Benassi Maria Serena , Aglietta Massimo , Grignani Giovanni 
Published: August 2014

Abstract
Background. TR is a DNA-binding alkaloid approved in Europe for the treatment STS as second/third line therapy. TR interferes with gene transcription and nucleotide excision repair, inducing DNA double strand breaks (DSBs). There is a strong clinical interest to increase TR activity by combination with other anti-cancer drugs. PARP-1 inhibitors (PARPi) which disable DNA base-excision repair mechanism causing the accumulation of DSBs, look like a worth to explore TR therapeutic partners. Our preclinical studies were focused on the effects of the combination of TR with the PARPi (OL and veliparib). Methods: We explored DNA damage by comet assay and P-H2AX determination, PARP-1 activity by polyADPribosylation assay in a panel of 18 sarcoma cell lines, evaluating cell viability after 72h treatment with escalating doses of TR, PARPi, and their constant combination. Colony growth, cell cycle, apoptosis and DNA damage were evaluated. Antitumoral activity was checked after 4 weeks of treatment with 3 endovenous injection of TR once a week and OL intraperitoneally 5 days/week, and compared to control and single agents in bone (SJSA-1) and soft tissue sarcoma (DMR) xenografted NOD/SCID mice. TUNEL assay, P-H2AX, PCNA and CD31 immunohistochemistry was done on explanted xenografts. Results: We showed TR-induced DNA damage and PARP-1 activation in sarcoma sensitive cell line. The addition of PARPi completely blocked basal and TR-induced PARP-1 activation. DNA damage response and checkpoints (ATM, ATR, BRCA1, CHK1, CHK2, p53) were activated after single agents and combination treatment inducing cell cycle arrest in S/G2 phase. This block was released after 48h in single agent-treated cells but not in combination-treated cells leading to apoptotic cell death. Colony growth assay and viability tests revealed a strong synergism of the two drugs (Combination Index &lt; 1, based on Chou-Talalay method). Different sarcoma histotypes displayed differences in sensitivity to both drugs and combination (Ewing's sarcoma the most sensitive&gt; myxoid liposarcoma &gt; leiomyosarcoma &gt; osteosarcoma ≥ undifferentiated pleomorphic &gt; fibrosarcoma). The antitumor effect of combined TR and OL was shown after in vivo treatment of SJSA-1 and DMR xenografted mice. A significant tumor volume reduction was observed if compared to both control and single agent and was attributed to increased DNA damage, apoptosis and reduced proliferation, but not to anti-angiogenic potentiation. In fact, a direct effect on tumor vasculature was shown in TR and combination treated group vs control. Conclusions: Our results validate the biological rationale to combine TR and PARPi in sarcoma and suggest exploring this pharmacological approach in the clinical setting.
Citation Format: Ymera Pignochino, Federica Capozzi, Carmine Dell'aglio, Marco Basiricò, Lorenzo D'ambrosio, Danilo Galizia, Erica Palesandro, Maria Serena Benassi, Massimo Aglietta, Giovanni Grignani. The antitumor effect of trabectedin (TR) is potentiated by olaparib (OL) in preclinical models of bone and soft tissue sarcomas (STS). [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 LB-213. doi:10.1158/1538-7445.AM2013-LB-213