De novo discovery and preclinical evaluation of cyclic radiopeptide [203/212Pb]Pb-PSV-359 targeting human fibroblast activation protein for alpha-particle radiotherapy in cancers (2024)

Abstract

241158

Introduction: Fibroblast activation protein alpha (i.e.,FAP), a type II transmembrane serine protease, has emerged as a promising target for theranostic radiopharmaceuticals due to its overexpression in various cancers and relative absence in normal tissues. ²¹²Pb (alpha-particle therapy; half-life 11 h) and ²⁰³Pb (SPECT imaging; half-life 52 h) are increasingly recognized as an ideal (elementally identical) radioisotope pair for targeted-image-guided alpha-particle radioligand therapies. In this study, a novel cyclic peptide targeting human FAP was discovered via phage display methods with site-specific cyclization. The peptide was further optimized into final candidate PSV-359 by conjugation to a lead (Pb)-specific chelator for ²¹²Pb via a molecular linker. The purpose of this study was to evaluate the in vitro and in vivo performance of [^203/212Pb]Pb-PSV-359 in preclinical xenograft models.

Methods: Radiolabeling of ²¹²Pb and ²⁰³Pb in PSV-359 was achieved by in-house protocols. The binding affinity and specificity of PSV-359 was determined by surface plasmon resonance (SPR) and enzyme inhibition assays. Serum stability was determined after incubating [²⁰³Pb]Pb-PSV-359 in fresh human serum at 37˚C. In vivo biodistribution of [²⁰³Pb]Pb-PSV-359 was determined in female athymic nude mice bearing HT1080-hFAP fibrosarcoma or U87MG glioma xenografts (n=3). [²¹²Pb]Pb-PSV-359 was imaged in athymic nude mice bearing HT080-FAP tumors. Alpha-particle radiotherapy of [²¹²Pb]Pb-PSV-359 was performed in athymic nude mice bearing HT1080-hFAP or U87MG xenografts with 100 mm³ initial tumor volume. In the HT1080-hFAP model, 4.9 MBq [²¹²Pb]Pb-PSV-359 was administered via IV injection over 3 fractions (n=10) at two-week intervals. An additional dose of 1.9 MBq was given when tumor recurrence was observed. In U87MG xenograft model, 4.8 MBq [²¹²Pb]Pb-PSV-359 was administered with over 3 fractions (n=9) at two-week intervals. Body weight and tumor volume was determined twice per week. Upon conclusion of the studies, blood samples were collected from survivors to perform CBC and serum chemistry analysis against age-matched naïve mice.

Results: Superior binding affinity and specificity of PSV-359 against hFAP was observed (K_d=1.8 nM, K_i=0.4 nM), with no observed binding to prolyl endopeptidase or dipeptidyl peptidase IV. [²⁰³Pb]Pb-PSV-359 remained stable in serum for 96 hours. In the HT1080-hFAP xenograft model, high uptake and long retention was observed in tumors (1.5 h: 20% ID/g, 24 h: 14% ID/g), along with fast blood clearance (0.2%ID/g at 3 h) via the renal system (4.7%ID/g at 24 h). In the U87MG tumors, despite low endogenous hFAP expression, high uptake of [²⁰³Pb]Pb-PSV-359 was observed (2 h: 11% ID/g, 6 h: 15% ID/g), indicating that expression of murine FAP in the U87MG tumor microenvironment might be found in stromal tissue. Strong tumor retention was observed with planar imaging of [^203/212Pb]Pb-PSV-359 in athymic nude mice bearing HT1080-FAP tumors. In vivo therapy studies revealed strong anti-tumor efficacy in both xenograft models. In the HT1080-hFAP model, all control mice reached tumor volume endpoint within 3 weeks, while 100% survival was observed in treated mice upon conclusion of the study (90 days) including 80% complete tumor remission. In the U87MG model, 89% survival rate was found upon conclusion of the study (60 days) in the treated group with half of the remaining mice having minimal to no residual tumor, compared with 22% survival rate in control cohorts. No significant change was found in CBC and serum chemistry analysis compared with naïve mice.