Identification Of Effective New Drugs Combinations Exploiting The Ability Of trabectedin To Modulate Transcription

Ewing’s sarcoma (ES) is the second most common malignant bone tumor of childhood, characterized by the chimeric protein EWS-FLI1 (hallmark of this pathology), that alters the transcription of different genes. The five-year overall survival of patients with localized disease is approximately 70%; unfortunately the majority of patients with a metastatic disease have a poor prognosis with a five-year overall survival around the 30%. Among the recently registered drugs for the therapy of sarcomas, trabectedin could be of potential great interest as it seems very active in some “translocated sarcomas”. Trabectedin exerts its antitumor activity with different mechanisms of action. One of the most important is related to its ability to interfere with DNA repair mechanisms (NER and HR), that cause cell cycle perturbations. Furthermore trabectedin is able to displace the oncogenic EWS-FLI1 chimera from its target promoters, modulating the transcription of these genes, in ES cells. Although trabectedin has shown some activity against ES, the overall clinical results indicated only a marginally activity of trabectedin given as single agent in ES. The thesis is aimed at using the available knowledge on trabectedin mechanism of action to identify some effective combinations. Since trabectedin induces cell cycle perturbations I speculated that its activity could be increased by checkpoint inhibitors. The studies performed on the combination of trabectedin and the WEE1 inhibitor, AZD-1775, have shown that the inhibition of WEE1 enhances the trabectedin activity, thus the combination is synergic. Since trabectedin affects the transcription of several genes, I have developed a new approach based on the use of silencing RNA (siRNA) libraries to identify whether the downregulation of some genes was synthetically lethal when ES cells were pretreated with trabectedin. An important part of the thesis was the development, validation and initial application of this approach