P17.16.b investigation of regorafenib tumor sensitivity and underlying treatment induced molecular mechanisms in glioblastoma recurrent tumors

Abstract BACKGROUND Glioblastoma (GB) is the most aggressive brain malignant tumor with a highly unfavorable prognosis. Maximal surgery resection, temozolomide chemotherapy (TMZ), and radiotherapy (RT), currently remain the best treatment option. Recently the phase II REGOMA clinical trial, has shown that regorafenib (REGO), a multikinase inhibitor that targets angiogenic, stromal and oncogenic tyrosine kinase receptors, significantly improves survival in GB patients.The aim of this study was to investigate Regorafenib tumor sensitivity and underlying molecular mechanisms in GB recurrent tumors leveraging on an ex vivo drug response functional precision medicine approach, that we have developed recently consisting of an in vitro vital patient-derived GB 3D organoid model and NAD(P)H Fluorescence Lifetime Imaging (FLIM). MATERIAL AND METHODS In-vitro 3D GB organoids were derived from 17 patients’ resected recurrent GB tumors. For 5 of these we had the matched primary tumor, and for other 3 we collected also the tumor peripheral portion. Totally, we obtained 25 samples. After 3 days from culture, organoids were treated with TMZ and REGO and drug response, using NAD(P)H FLIM, was assessed in the following 72 hours to stratify tumors in Sensitive (S) and Non Sensitive (NS). Whole-exome was performed on 14 tissue samples, comprising 5 primary/recurrence tumor pairs and whole-transcriptome on 11 recurrent samples before and after treatment. RESULTS Stratification of tumor cases in S and NS groups was performed both for TMZ and REGO treatment. We found 32% (n=8) and 76% (n= 19) of TMZ and REGO S tumors, respectively. Interestingly there was never a TMZ S case that was NS to REGO. Heterogeneity was found in the TMZ and REGO responsiveness for the tumor core/peripheral pair portions and in the primary/recurrence tumors pairs. It is noteworthy that all primary tumors were responsive to REGO.The drug-responsive stratification performed using our approach was also well reflected at the molecular level, as shown from transcriptome and exome RESULTS , where differently expressed genes and mutated genes were found in S versus NS tumors. CONCLUSION GB recurrent tumors resulted more responsive to REGO when compared to TMZ treatment, as expected. Thanks to our functional precision medicine approach, we were able to study the molecular alterations occurring due to the treatment, discovering biomarkers of sensitivity or resistance.