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

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.

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.

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.

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.