by the bodily immune responses. Radiolabeled EVs requires trace amounts of
nanomaterials, much less than pharmacologic amounts, and therefore, in vivo
effects of radiolabeled EVs will be negligible and different from chemical and
biological properties of physiological amount of EVs [2, 32].
Radionuclide imaging for monitoring biodistribution and in vivo fate of EVs
could have facilitated more clinical trials for EVs as a drug-carrier and regenerative
therapeutics. Aforementioned concept of using just trace amount of EVs for
identifying kinetics and distribution is closely related to phase 0 clinical trial. The
clinical trial platform requires in vivo evaluation on drug effects and pharmacokinetics for proper target drug selection in drug developments [33]. Radiolabeled EVs
could be applied to this phase 0 model to select appropriate candidates among so
many subtypes of EVs for further trials. The physiology of EVs could be different
according to the cell origin, extraction methods and environmental conditions.
Among various subtypes of EVs, radiolabeled EVs could help select appropriate
subtypes of EVs for further therapeutic usage [34]. In order to reveal in vivo
kinetics and fate of EVs precisely, robust methods for radiolabeling is required.
Strategy for radiolabeling and proper selection of radionuclides in accordance with
purposes is needed.
7.3 Radiolabeling Methods for Extracellular Vesicles
7.3.1 Radiolabeling Using Streptavidin
Most of the EV imaging studies employed optical probes of fluorescence or bioluminescence. Applying previous bioluminescence labeling methods to radionuclide labeling, Morishita et al. reported
125 I-labeled EVs [35]. Previously, they
developed bioluminescence reporter system for imaging EVs using Gaussia luciferase (gLuc), a reporter protein that emits chemiluminescence, and lactadherin
located on the outer surface of EVs [36]. They applied this reporter system to
radiolabel EVs by employing the streptavidin and biotin. As these two molecules
are strongly bound to each other, they designed EVs with a fusion protein of
streptavidin and lactadherin. The fusion protein was made by replacing the epidermal growth factor-like domain of lactadherin with streptavidin. The product
vector was transfected into the B16BL6 murine melanoma cells and from these
cells, EVs were collected that have the fusion protein in their outer membrane. For
radiolabeling, they incubated
125 I-iodobenzoyl norbiotinamide with EVs and then
125 I-labeled biotin was bound with streptavidin of EVs (Fig. 7.1). As radiolabel is
located at the membrane of EVs, it could trace the fate of EVs even after their
vesicle forms were degraded. Multimodal imaging can also be done if biotin is
combined either with luciferases, nanoparticles as well as radionuclides [37]. Other
radioiodine such as
123 I and
124
I could also be used for SPECT and PET imaging,
respectively. However, this method requires protein modification and vector
insertion and per se cannot be used in humans or in clinical applications.
144
H. Choi and D. W. Hwang
nanomaterials, much less than pharmacologic amounts, and therefore, in vivo
effects of radiolabeled EVs will be negligible and different from chemical and
biological properties of physiological amount of EVs [2, 32].
Radionuclide imaging for monitoring biodistribution and in vivo fate of EVs
could have facilitated more clinical trials for EVs as a drug-carrier and regenerative
therapeutics. Aforementioned concept of using just trace amount of EVs for
identifying kinetics and distribution is closely related to phase 0 clinical trial. The
clinical trial platform requires in vivo evaluation on drug effects and pharmacokinetics for proper target drug selection in drug developments [33]. Radiolabeled EVs
could be applied to this phase 0 model to select appropriate candidates among so
many subtypes of EVs for further trials. The physiology of EVs could be different
according to the cell origin, extraction methods and environmental conditions.
Among various subtypes of EVs, radiolabeled EVs could help select appropriate
subtypes of EVs for further therapeutic usage [34]. In order to reveal in vivo
kinetics and fate of EVs precisely, robust methods for radiolabeling is required.
Strategy for radiolabeling and proper selection of radionuclides in accordance with
purposes is needed.
7.3 Radiolabeling Methods for Extracellular Vesicles
7.3.1 Radiolabeling Using Streptavidin
Most of the EV imaging studies employed optical probes of fluorescence or bioluminescence. Applying previous bioluminescence labeling methods to radionuclide labeling, Morishita et al. reported
125 I-labeled EVs [35]. Previously, they
developed bioluminescence reporter system for imaging EVs using Gaussia luciferase (gLuc), a reporter protein that emits chemiluminescence, and lactadherin
located on the outer surface of EVs [36]. They applied this reporter system to
radiolabel EVs by employing the streptavidin and biotin. As these two molecules
are strongly bound to each other, they designed EVs with a fusion protein of
streptavidin and lactadherin. The fusion protein was made by replacing the epidermal growth factor-like domain of lactadherin with streptavidin. The product
vector was transfected into the B16BL6 murine melanoma cells and from these
cells, EVs were collected that have the fusion protein in their outer membrane. For
radiolabeling, they incubated
125 I-iodobenzoyl norbiotinamide with EVs and then
125 I-labeled biotin was bound with streptavidin of EVs (Fig. 7.1). As radiolabel is
located at the membrane of EVs, it could trace the fate of EVs even after their
vesicle forms were degraded. Multimodal imaging can also be done if biotin is
combined either with luciferases, nanoparticles as well as radionuclides [37]. Other
radioiodine such as
123 I and
124
I could also be used for SPECT and PET imaging,
respectively. However, this method requires protein modification and vector
insertion and per se cannot be used in humans or in clinical applications.
144
H. Choi and D. W. Hwang