BACKGROUND. Poly(ADP-ribose) polymerase (PARP) inhibitors are effective in a broad population of patients with ovarian cancer; however, resistance caused by low enzyme expression of the drug target PARP-1 remains to be clinically evaluated in this context. We hypothesize that PARP-1 expression is variable in ovarian cancer and can be quantified in primary and metastatic disease using a novel PET imaging agent. METHODS. We used a translational approach to describe the significance of PET imaging of PARP-1 in ovarian cancer. First, we produced PARP1-KO ovarian cancer cell lines using CRISPR/Cas9 gene editing to test the loss of PARP-1 as a resistance mechanism to all clinically used PARP inhibitors. Next, we performed preclinical microPET imaging studies using ovarian cancer patient–derived xenografts in mouse models. Finally, in a phase I PET imaging clinical trial we explored PET imaging as a regional marker of PARP-1 expression in primary and metastatic disease through correlative tissue histology. RESULTS. We found that deletion of PARP1 causes resistance to all PARP inhibitors in vitro, and microPET imaging provides proof of concept as an approach to quantify PARP-1 in vivo. Clinically, we observed a spectrum of standard uptake values (SUVs) ranging from 2–12 for PARP-1 in tumors. In addition, we found a positive correlation between PET SUVs and fluorescent immunohistochemistry for PARP-1 (r ² = 0.60). CONCLUSION. This work confirms the translational potential of a PARP-1 PET imaging agent and supports future clinical trials to test PARP-1 expression as a method to stratify patients for PARP inhibitor therapy. TRIAL REGISTRATION. Clinicaltrials.gov NCT02637934. FUNDING. Research reported in this publication was supported by the Department of Defense OC160269, a Basser Center team science grant, NIH National Cancer Institute R01CA174904, a Department of Energy training grant DE-SC0012476, Abramson Cancer Center Radiation Oncology pilot grants, the Marsha Rivkin Foundation, Kaleidoscope of Hope Foundation, and Paul Calabresi K12 Career Development Award 5K12CA076931.

Alpha-emitters can be pharmacologically delivered for irradiation of single cancer cells, but cellular lethality could be further enhanced by targeting alpha-emitters directly to the nucleus. PARP-1 is a druggable protein in the nucleus that is overexpressed in neuroblastoma compared with normal tissues and is associated with decreased survival in high-risk patients. To exploit this, we have functionalized a PARP inhibitor (PARPi) with an alpha-emitter astatine-211. This approach offers enhanced cytotoxicity from conventional PARPis by not requiring enzymatic inhibition of PARP-1 to elicit DNA damage; instead, the alpha-particle directly induces multiple double-strand DNA breaks across the particle track. Here, we explored the efficacy of [²¹¹At]MM4 in multiple cancers and found neuroblastoma to be highly sensitive in vitro and in vivo. Furthermore, alpha-particles delivered to neuroblastoma show antitumor effects and durable responses in a neuroblastoma xenograft model, especially when administered in a fractionated regimen. This work provides the preclinical proof of concept for an alpha-emitting drug conjugate that directly targets cancer chromatin as a therapeutic approach for neuroblastoma and perhaps other cancers.

Purpose: [131I]meta-iodobenzylguanidine ([131I]MIBG) is a targeted radiotherapeutic administered systemically to deliver beta particle radiation in neuroblastoma. However, relapses in the bone marrow are common. [211At]meta-astatobenzylguanidine ([211At] MABG) is an alpha particle emitter with higher biological effectiveness and short path length which effectively sterilizes microscopic residual disease. Here we investigated the safety and antitumor activity [211At]MABG in preclinical models of neuroblastoma. Experimental Design: We defined the maximum tolerated dose (MTD), biodistribution, and toxicity of [211At]MABG in immunodeficient mice in comparison with [131I]MIBG. We compared the antitumor efficacy of [211At]MABG with [131I]MIBG in three murine xenograft models. Finally, we explored the efficacy of [211At]MABG after tail vein xenografting designed to model disseminated neuroblastoma. Results: The MTD of [211At]MABG was 66.7 MBq/kg (1.8 mCi/kg) in CB17SC scid−/− mice and 51.8 MBq/kg (1.4 mCi/kg) in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice. Biodistribution of [211At]MABG was similar to [131I]MIBG. Long-term toxicity studies on mice administered with doses up to 41.5 MBq/kg (1.12 mCi/kg) showed the radiotherapeutic to be well tolerated. Both 66.7 MBq/kg (1.8 mCi/kg) single dose and fractionated dosing 16.6 MBq/kg/fraction (0.45 mCi/kg) × 4 over 11 days induced marked tumor regression in two of the three models studied. Survival was significantly prolonged for mice treated with 12.9 MBq/kg/fraction (0.35 mCi/kg) × 4 doses over 11 days [211At]MABG in the disseminated disease (IMR-05NET/GFP/LUC) model (P = 0.003) suggesting eradication of microscopic disease. Conclusions: [211At]MABG has significant survival advantage in disseminated models of neuroblastoma. An alpha particle emitting radiopharmaceutical may be effective against microscopic disseminated disease, warranting clinical development.

The currently available therapeutic radiopharmaceutical for high-risk neuroblastoma, ¹³¹I-metaiodobenzylguanidine, is ineffective at targeting micrometastases because of the low–linear-energy-transfer (LET) properties of high-energy β-particles. In contrast, Auger radiation has high-LET properties with nanometer ranges in tissue, efficiently causing DNA damage when emitted near DNA. The aim of this study was to evaluate the cytotoxicity of targeted Auger therapy in preclinical models of high-risk neuroblastoma. Methods: We used a radiolabled poly(adenosine diphosphate ribose) polymerase (PARP) inhibitor called ¹²⁵I-KX1 to deliver Auger radiation to PARP-1, a chromatin-binding enzyme overexpressed in neuroblastoma. The in vitro cytotoxicity of ¹²⁵I-KX1 was assessed in 19 neuroblastoma cell lines, followed by in-depth pharmacologic analysis in a sensitive and resistant pair of cell lines. Immunofluorescence microscopy was used to characterize ¹²⁵I-KX1–induced DNA damage. Finally, in vitro and in vivo microdosimetry was modeled from experimentally derived pharmacologic variables. Results:¹²⁵I-KX1 was highly cytotoxic in vitro across a panel of neuroblastoma cell lines, directly causing double-strand DNA breaks. On the basis of subcellular dosimetry, ¹²⁵I-KX1 was approximately twice as effective as ¹³¹I-KX1, whereas cytoplasmic ¹²⁵I-metaiodobenzylguanidine demonstrated low biological effectiveness. Despite the ability to deliver a focused radiation dose to the cell nuclei, ¹²⁵I-KX1 remained less effective than its α-emitting analog ²¹¹At-MM4 and required significantly higher activity for equivalent in vivo efficacy based on tumor microdosimetry. Conclusion: Chromatin-targeted Auger therapy is lethal to high-risk neuroblastoma cells and has the potential to be used in micrometastatic disease. This study provides the first evidence for cellular lethality from a PARP-1–targeted Auger emitter, calling for further investigation into targeted Auger therapy.