Fatty acid-conjugated radiopharmaceuticals for fibroblast activation protein-targeted radiotherapy

Introduction: Radiopharmaceuticals that target cancer-associated fibroblasts (CAFs) have become an increasingly attractive strategy for cancer theranostics. Recently, a series of fibroblast activation protein inhibitor (FAPI)-based radiopharmaceuticals have been successfully applied to the diagnosis of a variety of cancers and exhibited excellent tumor selectivity. Nevertheless, CAF-targeted radionuclide therapy encounters difficulties in cancer treatment, as the tumor uptake and retention of FAPIs are insufficient. To meet this challenge, we tried to conjugate albumin-binding moiety to FAPI molecule for prolonged circulation that may increase the accumulation and retention of radiopharmaceuticals in tumor.

Methods: Two fatty acids, lauric acid (C12) and palmitic acid (C16), were conjugated to FAPI-04 to give two albumin-binding FAPI radiopharmaceuticals, denoted as FAPI-C12 and FAPI-C16, respectively. They had been radiolabeled with gallium-68, yttrium-86, and lutecium-177 for stability study, binding affinity assay, PET and SPECT imaging, biodistribution, and radionuclide therapy study to systematically evaluate their potential for CAF-targeted radionuclide therapy.

Results: FAPI-C12 and FAPI-C16 showed high binding affinity to FAP with the IC₅₀ of 6.80 ± 0.58 nM and 5.06 ± 0.69 nM, respectively. They were stable in both saline and plasma. The tumor uptake of [⁶⁸Ga]Ga-FAPI-04 decreased by 56.9% until 30 h after treated with FAPI-C16 before, and the uptakes of [⁸⁶Y]Y-FAPI-C12 and [⁸⁶Y]Y-FAPI-C16 in HT-1080-FAP tumor were both much higher than that of HT-1080-Vehicle tumor which identified the high FAP specific of these two radiopharmaceuticals. Both FAPI-C12 and FAPI-C16 showed notably longer circulation and significantly enhanced tumor uptake than those of FAPI-04. [¹⁷⁷Lu]Lu-FAPI-C16 had the higher tumor uptake at both 24 h (11.22 ± 1.18%IA/g) and 72 h (6.50 ± 1.19%IA/g) than that of [¹⁷⁷Lu]Lu-FAPI-C12 (24 h, 7.54 ± 0.97%IA/g; 72 h, 2.62 ± 0.65%IA/g); both of them were much higher than [¹⁷⁷Lu]Lu-FAPI-04 with the value of 1.24 ± 0.54%IA/g at 24 h after injection. Significant tumor volume inhibition of [¹⁷⁷Lu]Lu-FAPI-C16 at the high activity of 29.6 MBq was observed, and the median survival was 28 days which was much longer than that of the [¹⁷⁷Lu]Lu-FAPI-04 treated group of which the median survival was only 10 days.

Conclusion: This proof-of-concept study validates the hypothesis that conjugation of albumin binders may shift the pharmacokinetics and enhance the tumor uptake of FAPI-based radiopharmaceuticals. This could be a general strategy to transform the diagnostic FAP-targeted radiopharmaceuticals into their therapeutic pairs.