We prepared suspensions with a drug concentration of 100 mg/mL with different particle sizes using wet-milling according to the component and compositions shown in Table , then performed a rat pharmacokinetic (PK) study. Wet-milling is a widely used technique in the pharmaceutical industry for producing injectable nanosuspensions. The two primary technologies for wet-milling are high-pressure homogenization and bead milling. 25) S-892216 is a fine particle with a D50 of 3.3 µm, manufactured via jet-milling. We compared the PK profiles between coarse suspension, where the jet-milled drug substance was simply dispersed, and nanosuspension, where wet-milling was performed. Figure and Table show the results of the rat PK study of subcutaneous (SC) injections at 100 mg/head. The plasma exposure of S-892216 improved as the drug particle size decreased. The area under the plasma concentration-time curve (AUC) from 0 to infinity (AUCinf) of the S-892216 plasma concentration for the coarse suspension was 63,800 ng•hr/mL, whereas suspension with wet-milling showed significantly increased absorption, with AUCs of 163,000 (Z-average: 670 nm) and 316,000 (210 nm). The absorption rate constant (ka, the apparent elimination rate from plasma) showed similar values regardless of drug particle size. Therefore, wet-milling of S-892216 did not affect the sustained-release period, but increased the plasma exposure. This is consistent with the findings of Ho et al., who suggested that smaller particle sizes of drug substances could be widely distributed in the tissue, thereby avoiding the release inhibition effect caused by fibrosis. These results indicate that nanosized drug particles allow higher plasma exposure; therefore, we developed the S-892216 injectable suspension as a wet-milled nanosuspension. modifiers such as surfactants to maintain a consistent particle size. These authors reported seeing no general trends in the physicochemical properties of the drug substance relative to the ease of milling. Hence, we selected an appropriate surfactant to develop the S-892216 injectable suspension. Few surfactants have been confirmed as safe in humans for long-acting injectables. Polysorbate 20 is used in cabotegravir (trade name: Vocabria 600 mg prolonged-release suspension for injection ), and poloxamer P338 is used in rilpivirine (trade name: Rekambys ® 900 mg prolonged-release suspension for injection ). Both polysorbate 20 and poloxamer P338 are representative nonionic surfactants and have been compared in various ways. However, their performance when applied to S-892216 has not been investigated. Therefore, we compared the particle size reduction profile during the wet-milling process using the nanosuspension with a drug concentration of 200 mg/mL (Table ). The concentration of both surfactants was set at 2% w/v. Macrogol 4000 (polyethylene glycol 3350) was added at 2% w/v as a stabilizer. Mannitol was added at 4.5% w/v as a tonicity agent to maintain constant osmotic pressure.