Synergistic Effect of a Mesothelin-Targeted ²²⁷Th Conjugate in Combination with DNA Damage Response Inhibitors in Ovarian Cancer Xenograft Models

Abstract

Targeted 227Th conjugates (TTCs) represent a new class of therapeutic radiopharmaceuticals for targeted α-therapy. They comprise the α-emitter 227Th complexed to a 3,2-hydroxypyridinone chelator conjugated to a tumor-targeting monoclonal antibody. The high energy and short range of the α-particles induce antitumor activity, driven by the induction of complex DNA double-strand breaks. We hypothesized that blocking the DNA damage response (DDR) pathway should further sensitize cancer cells by inhibiting DNA repair, thereby increasing the response to TTCs. Methods: This article reports the evaluation of the mesothelin (MSLN)-TTC conjugate (BAY 2287411) in combination with several DDR inhibitors, each of them blocking different DDR pathway enzymes. MSLN is a validated cancer target known to be overexpressed in mesothelioma, ovarian, lung, breast, and pancreatic cancer, with low expression in normal tissue. In vitro cytotoxicity experiments were performed on cancer cell lines by combining the MSLN-TTC with inhibitors of ataxia telangiectasia mutated, ataxia telangiectasia and Rad3-related (ATR), DNA-dependent protein kinase, and poly[adenosine diphosphate ribose] polymerase (PARP) 1/2. Further, we evaluated the antitumor efficacy of the MSLN-TTC in combination with DDR inhibitors in human ovarian cancer xenograft models. Results: Synergistic activity was observed in vitro for all tested inhibitors (inhibitors are denoted herein by the suffix “i”) when combined with MSLN-TTC. ATRi and PARPi appeared to induce the strongest increase in potency. Further, in vivo antitumor efficacy of the MSLN-TTC in combination with ATRi or PARPi was investigated in the OVCAR-3 and OVCAR-8 xenograft models in nude mice, demonstrating synergistic antitumor activity for the ATRi combination at doses demonstrated to be nonefficacious when administered as monotherapy. Conclusion: The presented data support the mechanism-based rationale for combining the MSLN-TTC with DDR inhibitors as new treatment strategies in MSLN-positive ovarian cancer.

Preclinical Efficacy of a PSMA-Targeted Thorium-227 Conjugate (PSMA-TTC), a Targeted Alpha Therapy for Prostate Cancer

“…The fully human PSMA-targeting antibody BAY 2315158 specifically recognizing human and cynomolgus PSMA was licensed from Progenics Pharmaceuticals, Inc. (16,17). The nonradiolabeled PSMA antibody-chelator conjugate (BAY 2315493) was manufactured at Bayer AG and Bayer AS by coupling an N-hydroxysuccinimideactivated 3,2-HOPO chelator covalently to the e-amino groups of the lysine residues of the PSMA antibody as described previously (12)(13)(14)(15). The radionuclide thorium-227 was manufactured at IFE and the antibody-chelator conjugate was radiolabeled with thorium-227 in citrate buffer (pH 5.5) at þ22 C for up to 60 minutes resulting in PSMA-TTC (BAY 2315497) as described in Supplementary Methods and previously (14).…”

“…The radionuclide thorium-227 was manufactured at IFE and the antibody-chelator conjugate was radiolabeled with thorium-227 in citrate buffer (pH 5.5) at þ22 C for up to 60 minutes resulting in PSMA-TTC (BAY 2315497) as described in Supplementary Methods and previously (14). A nonbinding radiolabeled isotype control was prepared similarly to PSMA-TTC (12,14,15).…”

“…In vitro cytotoxicity and caspase experiments were performed using CellTiter-Glo or Caspase-3/7 Glo assay (Promega), respectively, in prostate cancer cell lines after a 5-day exposure to PSMA-TTC. Determination of antibodies bound per cell, cell-cycle analysis (staining C4-2 prostate cancer cells with PI RNAse), and DNA DSBs (percentage of phosphorylated histone protein g-H2AX-positive LNCaP cells) were performed by flow cytometry as previously described (12,15). Internalized radioactivity after incubation of C4-2 and PC3 cells with PSMA-TTC was determined as described in Supplementary Methods.…”

“…IHC was performed on paraffin sections of xenograft tumor tissues with anti-human PSMA antibody (clone 3E6, 1:1,000, M3620, Dako Denmark) as described in Supplementary Methods. g-H2AX and cleaved caspase-3 analyses were performed as previously described in ST1273 tumors (12,15).…”

“…PSMA-TTC uses the alpha-particle emitter thorium-227 (half-life of 18.7 days), which is the parent nuclide of radium-223. Thorium-227 can be efficiently complexed by the 3,2-hydroxypyridinone (3,2-HOPO) octadentate chelator (10,11) covalently linked to antibodies (12)(13)(14)(15) at ambient temperature. Strong antitumor efficacy of PSMA-TTC is shown in a panel of prostate cancer models mimicking different clinical stages of prostate cancer including castration-resistant and bone metastatic disease.…”