**1. Introduction**

High-density lipoproteins (HDLs) are hydrophobic lipid micelles (endogenous nanoparticles; diameter of 5–12 nm) that carry cholesterol esters, free cholesterol, phospholipids, and triglycerides. HDLs transport and release their lipidic content in the liver and into the cytoplasm of different cells by interacting with specific cell membrane proteins. Due

to the presence of apolipoprotein AI (Apo AI) as the main HDL peripheral component for targeting the scavenger receptor class B type I (SR-BI), synthetic or reconstituted HDL nanoparticles (rHDL), based on phospholipids and Apo AI, have been used as hydrophobic drug transporters, promoting the research of rHDL as a vehicle for the administration of chemotherapeutic agents [1,2].

The SR-BI is overexpressed on the cell surface of many cancer cells (e.g., ovarian, liver, and prostate cancer) and is a selective and specific receptor for HDL (through Apo AI). The SR-BI–HDL interaction triggers the release of cholesterol from HDL to the cell cytoplasm. Cholesterol is necessary for cell growth and is highly required by cancer cells, because it takes part in the synthesis of new cytoplasmic membranes [2,3].

Radiolabeled rHDLs have been employed for molecular imaging of cancer cells by targeting the SR-BI protein. Perez-Medina et al. attached <sup>89</sup>Zr-deferoxamine to the ApoE/phospholipids of rHDL; preclinical PET images showed high <sup>89</sup>Zr-rHDL uptake in tumor-associated macrophages of breast cancer [4]. Recently, preclinical studies of 99mTc-rHDL for SPECT imaging of prostate cancer metastasis have also been reported [5].

Alpha particles are an extraordinary option for targeted radiotherapy due to their capacity to cause damage to cancer cells, because the corresponding deposition of energy at the cellular level is one-hundred times greater than that of β-particles [6]. Targeted alpha-particle therapy (TAT) with <sup>225</sup>Ac has demonstrated considerable potential in the treatment of advanced prostate cancer [6].

Despite dozens of previous articles on <sup>225</sup>Ac labeling of metallic nanoparticles, liposomes, and micelles, the preparation of <sup>225</sup>Ac-HDL nanosystems has not been reported so far. The <sup>225</sup>Ac is produced primarily from a <sup>229</sup>Th/225Ac generator with subsequent <sup>225</sup>Ac purification. The <sup>255</sup>Ac is characterized by decay into multiple alpha-emitting daughter radionuclides (four effective alpha emissions from <sup>225</sup>Ac, <sup>221</sup>Fr, <sup>217</sup>At, and <sup>213</sup>Po). One possible disadvantage of <sup>225</sup>Ac-complexes, however, is the recoil energy (~100 keV) of the nucleus during decay, which could induce the breaking of the bond with the chelator or transporter molecule with the consequent release of the daughter radionuclide to healthy tissues [7].

Therefore, the encapsulation of <sup>225</sup>Ac in rHDL nanoparticles (225Ac-rHDL) would prevent the release of daughter radionuclide from the transporter biomolecule due to the recoil energy effect and, at the same time, the specific molecular recognition of <sup>225</sup>Ac-rHDL by cancer cells that overexpress the SR-BI protein would allow the cytoplasmic internalization of <sup>225</sup>Ac in tumor cells to produce ablative radiation doses.

This research aimed to prepare <sup>225</sup>Ac-rHDL and evaluate its preclinical in vitro and in vivo capability as a potential agent for targeted α-therapy of tumors overexpressing SR-BI receptors.
