Abstract 910: Amanitin-based ADCs targeting PSMA as novel therapeutic modality for prostate cancer therapy
Authors: Alexandra Braun, Aniko Palfi, Christoph Mueller, Torsten Hechler, Andreas Pahl, Michael Kulke
Journal: Cancer Research (2021)

Abstract Background: Antibody-targeted amanitin conjugates (ATACs) comprise a new class of antibody-drug conjugates (ADCs) using amanitin as toxic payload. Amanitin binds to the RNA pol II and thereby efficiently inhibits the cellular transcription process. In the current study, in vitro and in vivo data of an ATAC targeting PSMA (prostate specific membrane antigen) are presented. PSMA is a type II integral membrane glycoprotein and used as tumor marker due to its predominant expression on malignant prostate cells in prostate carcinoma. As PSMA expression increases with tumor aggressiveness, metastatic and disease recurrence, it is considered an ideal target for amanitin-based ADCs. Material and methods: Cell lines: PSMA+ cell lines: LNCap, 22RV1; PSMA- cell line: PC3 Antibody: cysteine engineered monoclonal anti-PSMA antibody (Albert Ludwig University Freiburg, medical center; derivatization and production at Heidelberg Pharma) Toxic warhead: Cysteine reactive amanitin-linker constructs were synthesized at Heidelberg Pharma and conjugated site-specifically to engineered cysteine residues of the anti-PSMA antibody yielding ATACs with a DAR of 2. Cell proliferation assay: Quantitative determination of cell viability was performed by BrdU-based chemiluminescent cell proliferation ELISA (Roche). Animal models: Subcutaneous Mouse xenograft tumor models with PSMA-positive cell lines 22RV1 and LNCap were performed in single-dose experiments. Tolerability was assessed in mice and non-human primates (NHP). Results: All anti-PSMA ATACs with optimized amanitin-linker variants for use in solid tumors showed favorable in vitro cytotoxicity with nano- to picomolar activity on PSMA+ cell lines and no cytotoxic activity on PSMA- cells. In mouse xenograft models, the optimized anti-PSMA ATACs caused dose-dependent tumor regression in LNCap s.c. xenografts. Complete remission was achieved after a single i.v. dose of ¼ MTD with 100% overall survival for the full duration of the studies (&amp;gt;160 days). The tolerability of the tested ATACs in mice differed between 15 and &amp;gt; 80 mg/kg Safety profiling in Cynomolgus monkeys revealed a good tolerability and therapeutic index for all selected amanitin-linker variants. Hematology and clinical chemistry parameters were unaffected except liver enzymes and LDH: A moderate and transient increase was observed. Low off target toxicity was confirmed by a non-binding ATAC. Conclusions: Targeted cytotoxic drug delivery to PSMA positive cell lines was achieved by using anti-PSMA ATACs optimized for the use in solid tumors. The mode of action of the payload amanitin led to an efficient anti-tumor potential in vitro and in vivo with good tolerability in non-human primates. Using ATACs in the therapy of PSMA positive prostate cancer is a promising approach, especially by using a cytotoxic agent whose mode of action differs from other commonly used toxins. Citation Format: Alexandra Braun, Aniko Palfi, Christoph Mueller, Torsten Hechler, Andreas Pahl, Michael Kulke. Amanitin-based ADCs targeting PSMA as novel therapeutic modality for prostate cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 910.