212Pb-DOTAM-peptide targeting LDLR-receptors in glioblastoma and pancreatic ductal adenocarcinoma

Introduction: RMX-VH is a DOTAM-modified peptide with high affinity to the low-density lipoprotein receptors (LDLR). The overexpression of LDLR has been reported in multiple solid tumors and correlated with the prognosis of the disease. RMX-VH conjugate has been selected to selectively deliver the alpha-emitters to LDLR(+) cancer cells in GBM and PDAC. DOTAM chelator allows for stable coordination of both ²⁰³Pb and ²¹²Pb/daughter isotopes. Herein, we report radiosynthesis of ²¹²Pb-RMX-VH, the protocol of SPECT ²¹²Pb imaging, and results of in vitro/in vivo pre-clinical studies in relevant glioblastoma (GBM) and pancreatic ductal adenocarcinoma (PDAC) models.

Methods: ²¹²Pb-labeling of RMX-VH (25ug) was done using a manual approach in 0.4 sodium acetate buffer pH=6.0 at room temp in the presence of 40mg/ml. ascorbate used as a scavenger. The radiochemical and chemical purity was analyzed using iTLC chromatography, radio/UV HPLC, and Perkin Elmer gamma counter normalized for ²¹²Pb and ²¹²Bi energy windows. The uptake studies were completed in multiple GBM cancer cell lines ( U87MG, A172, U373) and PDAC (HPAF-II, BxPC-3, and MiaPaCa2 ) showing various levels of expression of LDLR. The SPECT images were acquired using a gamma-eye camera (Bioemtech, Greece) at 2h, 4h, and 24h post-injection with follow-up biodistribution studies. The organs and tumor were collected, weighed, and the tissue radioactivity was measured with Wizard2 Gamma Counter (Perkin-Elmer, Waltham, MA). The percentage of injected dose per gram of tissue (%ID/g) was calculated and decay-corrected.

Results: The radiochemical purity of ²¹²Pb-RMX-VH was higher than 98%. There was no need for additional post-purification of the final product. The selection of cell lines for studies was based on the FACS results showing the expression of LDLR. The cellular uptake studies confirmed time-dependent accumulation agent reaching the highest uptake at 24h timepoint for HPAFII (13.7± 1.6 %ID/g), BXPC3 10.9 ±0.9%ID/g and lower for MiapAca (3.6 ±0.37%ID/g) and Colo357 (2.99± 0.79%ID/g). Multi-point SPECT imaging studies showed selective accumulation of the agent in the PDAC models generated in athymic nude mice as early as 2h post-injection. The uptake correlated with bioluminescent data and PET/CT imaging studies using diagnostics analog, ⁶⁸Ga-RMX-VH. The bioD studies have shown (7.05 ±1.31%ID/g) retention of the agent in the orthotopic PDAC at 2h post-injection, slightly reduced at (4.56± 0.66%ID/g) and (5.44± 0.18%ID/g) and 4h and 24h time pint with renal excretion from the bloodstream as the main route elimination.

Conclusions: ²¹²Pb-RMX-VH is a novel radiopharmaceutical for TAT of LDLR overexpressing solid tumors. The TAT agent has shown favorable LDLR-tumor targeting properties in vitro and in vivo. We optimized radiolabeling process and established a protocol for imaging of ²¹²Pb-agent using SPECT system. The companion diagnostic ⁶⁸Ga-RMX-VH is currently evaluated in eIND clinical studies in the US ( Houston).

• Download figure
• Open in new tab
• Download powerpoint