Having elucidated the self-immolative release of MB from SIE5, we explored its modulation through CB-adamantane supramolecular interactions. The introduction of 10×CB to a 16 µM solution of SIE5 in pH 6.5 MES resulted in a reduction in ments were conducted at 37 °C in pH 6.5 MES buffer solutions. Irradiation was conducted using a 3000K LED light equipped with a 600 nm long-pass filter (light intensity = 2.2 mW/cm MB release rate by over ninefold (Figure , triangle) compared to free SIE5 (Figure , square). Adding 20×adamantane (320 µM) to the solution of 10×CB/SIE5 restored the rate of MB release (Figure , dot). The attenuation of the release rate from SIE5 by CB was less pronounced in this experiment compared to that observed for SIE2 (Figure ), which can be ascribed to the incomplete CB-SIE5 complexation at 37 o C under the used concentrations, and to structural differences in the leaving groups. While our future research aims to enhance CB-SIE complexation, the reactivity of the SIE5 system was significantly modulated by the CB-adamantane supramolecular chemistry-after 4-hour incubation, 1.88 µM, 0.43 µM, and 2.15 µM of MB were released from SIE5, 10×CB/SIE5, and 10×CB/SIE5/20×Ad, respectively. This level of suppression is sufficient to distinguish the photodynamic cell-killing effects between the gated CB-SIE5 complex and the active forms of SIE5 (the therapeutically effective threshold MB concentration was established to be 2 µM under our experimental conditions; see Figure ). Singlet Oxygen Sensor Green (SOSG) was employed to evaluate the 1 O2-generation ability of SIE5, 10×CB/SIE5, and 10×CB/SIE5/20×Ad. Each sample was incubated at 37 o C for 4 h in pH 6.5 MES, followed by the addition of SOSG. After 5 min of light irradiation, the emission from 1 O2-activated SOSG was measured by fluorescence spectroscopy. The SIE5 solution showed significant fluorescence from the 1 O2-activated SOSG (Figure , red), consistent with its confirmed reactivity toward photosensitizer release. The fluorescence from the 10×CB/SIE5 group was more than 3 times lower compared to SIE5 (Figure , blue) as a result of CB-inhibited MB release. Importantly, the irradiated 10×CB/SIE5/20×Ad sample exhibited comparable fluorescence (Figure , green) as the free SIE5, demonstrating that the adamantane trigger effectively restored MB release and activated its photodynamic effect.