Abstract 4606: Combinatorial effects of a sequence of trabectedin (T) and CD13-targeted tissue factor tTF-NGR on human vascular endothelial cells (HUVEC) and in soft tissue sarcoma (STS) models
Authors: Brand Caroline , Pavelka Stefanie , Hessling Kathrin , Hintelmann Heike , Berdel Andrew F. , Bäumer Sebastian , Bäumer Nicole , Lenz Georg , Schliemann Christoph , Berdel Wolfgang E. , Schwöppe Christian 
Published: March 2024

Abstract
Trabectedin represents a standard treatment option for relapsed or refractory soft tissue sarcomas. CD13 is a neutral aminopeptidase expressed on remodeling and invasive endothelial cells such as in the tumor vasculature. CD13-targeted tissue factor (tTF-NGR) is a recombinant pro-coagulatory fusion protein which accumulates in the tumor vasculature leading to tumor vascular occlusion and ultimately to tumor infarction. Giving both compounds in sequence could trap trabectedin inside tumors and thus increase its efficacy. This vice versa can optimize the activity of tTF-NGR. To prepare a multicenter clinical trial of a combination of trabectedin and tTF-NGR in patients with advanced soft tissue sarcomas refractory to 1st line systemic therapy (TRABTRAP), preclinical experiments on the mechanism of action of this combination, its efficacy and pharmacokinetics (PK) were performed. We first tested the pro-apoptotic function of trabectedin on human vascular endothelial cells (HUVEC) and human HT1080 sarcoma cells in vitro. Trabectedin significantly (p&lt;.0001) increased the presence of phosphatidylserine (PS) on the outer leaflet of the phospholipid bilayer of the cell membrane in a dose- and time-dependent manner. The resulting optimized phospholipid milieu significantly potentiated the pro-coagulatory efficacy of tTF-NGR within the factor X:factor VIIa:tTF-NGR:CD13 complex on the outer cell membrane when compared to cultures without trabectedin (p&lt;.05). This effect was specifically dependent on the presence of PS as it could be abolished by masking PS via preincubation with annexin V, whereas corticosteroids had no effect on the pro-coagulatory response of HUVEC in the presence of tTF-NGR. In an in vivo HT1080 human sarcoma xenograft model in athymic mice, systemic combination treatment of trabectedin followed by tTF-NGR (n=11) inhibited tumor growth to a greater extent than either compound alone (trabectedin n=9, tTF-NGR n=9) and compared to a saline control (n=8). The terminal half-life of tTF-NGR infused intravenously over 1 hour, was 9 hours (phase I trial). To further characterize the duration of tTF-NGR presence upon binding to the endothelial cell surface molecule CD13, we performed flow cytometry and fluorescence labeling experiments with tTF-NGR and HUVEC in vitro. After coincubation with HUVEC, the cells rapidly internalized the tTF-NGR:CD13 complex into the cytoplasm of the cells (approx. 50% of the labeled tTF-NGR after 3 h at 37oC), a process which exposed the protein to intracellular degradation. The multicenter TRABTRAP trial is actively recruiting patients (EudraCT 2020-005858-21).
Citation Format: Caroline Brand, Stefanie Pavelka, Kathrin Hessling, Heike Hintelmann, Andrew F. Berdel, Sebastian Bäumer, Nicole Bäumer, Georg Lenz, Christoph Schliemann, Wolfgang E. Berdel, Christian Schwöppe. Combinatorial effects of a sequence of trabectedin (T) and CD13-targeted tissue factor tTF-NGR on human vascular endothelial cells (HUVEC) and in soft tissue sarcoma (STS) models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4606.