Abstract 464: Methylation of a single CpG site in the CD95-ligand promoter is a biomarker predicting the response to therapy with APG101 in glioblastoma
Authors: Gieffers Christian , Kunz Claudia , Sykora Jaromir , Merz Christian , Thiemann Meinolf , Fricke Harald , Wiestler Benedikt , Wick Wolfgang 
Published: July 2016

CD95 (APO-1/Fas) is a member of the Tumor Necrosis Factor Receptor Super Family. Binding of CD95-ligand (CD95L) to CD95 triggers intracellular signal transduction that is critically involved in the invasive growth of glioblastoma cells. Invasion of malignant glioblastoma cells into the brain parenchyma is responsible for poor therapeutic outcome of currently available treatments. The inhibition of CD95/CD95L mediated invasive growth of glioblastoma cells represents an attractive novel therapeutic concept. Apogenix has developed APG101, a fully human fusion protein consisting of the extracellular domain of CD95 and the Fc-domain of an IgG. APG101 has been confirmed as a potent inhibitor of CD95L induced invasion of glioblastoma cells in vitro.

In a randomized phase 2 study in glioblastoma patients with 1st or 2nd relapse the combined therapy of APG101 plus radiotherapy (RT) was found to be superior to RT alone in a clinically relevant order of magnitude in all efficacy endpoints (i.e. PFS-6, PFS and OS). At the same time APG101 exhibited an excellent safety profile and was well tolerated.

To identify potential biomarkers we used available tissue sections originating from archived primary tumor of the study patients and analyzed them for the expression of CD95L as well as for the DNA methylation status. A genome-wide assessment of DNA methylation identified a single CpG-site (CpG2) upstream of the CD95L-promotor that showed differential methylation between APG101 responders (PFS&gt;5 months) and non-responders (PFS &lt; 2 months). Available patient DNAs were in addition analyzed by MassARRAY and Pyro-sequencing to confirm differential CpG2 methylation. Based on this data we used the median of the CpG2 methylation level as a threshold to analyze for a correlation of CpG2 methylation and response to APG101 therapy. Patients showing a low level of CpG2 methylation responded best to therapy with APG101 whereas patients with a high level of CpG2 methylation did not show a relevant benefit when treated with APG101 compared to the control RT-group. The analysis shows a significant survival benefit achieved in patients with low CpG2 methylation (median OS: 16.1 vs 7.3 months, p = 0.029).

Hence, the level of CpG2 methylation in the CD95L promoter in the patients' glioblastoma tissue is a prognostic biomarker predicting response to therapy with APG101. Based on the observations described, Apogenix currently develops a qPCR-based assay to quantify CpG2 methylation. This assay is intended as companion diagnostic to identify patients that may respond best to APG101 treatment.

Citation Format: Christian Gieffers, Claudia Kunz, Jaromir Sykora, Christian Merz, Meinolf Thiemann, Harald Fricke, Benedikt Wiestler, Wolfgang Wick. Methylation of a single CpG site in the CD95-ligand promoter is a biomarker predicting the response to therapy with APG101 in glioblastoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 464.