Protein-based tumor molecular imaging probes

Abstract

Molecular imaging is an emerging discipline which plays critical roles in diagnosis and therapeutics. It visualizes and quantifies markers that are aberrantly expressed during the disease origin and development. Protein molecules remain to be one major class of imaging probes, and the option has been widely diversified due to the recent advances in protein engineering techniques. Antibodies are part of the immunosystem which interact with target antigens with high specificity and affinity. They have long been investigated as imaging probes and were coupled with imaging motifs such as radioisotopes for that purpose. However, the relatively large size of antibodies leads to a half-life that is too long for common imaging purposes. Besides, it may also cause a poor tissue penetration rate and thus compromise some medical applications. It is under this context that various engineered protein probes, essentially antibody fragments, protein scaffolds, and natural ligands have been developed. Compared to intact antibodies, they possess more compact size, shorter clearance time, and better tumor penetration. One major challenge of using protein probes in molecular imaging is the affected biological activity resulted from random labeling. Site-specific modification, however, allows conjugation happening in a stoichiometric fashion with little perturbation of protein activity. The present review will discuss protein-based probes with focus on their application and related site-specific conjugation strategies in tumor imaging.

Fig. 1

Schematic representation of different antibody formats. Reproduced with permission from Holliger and Hudson (2005)

Fig. 2

Serial microPET images of different xenograft tumor models after intravenous injection of ⁶⁴Cu-DOTA-cetuximab. Decay-corrected whole-body coronal images that contain the tumor were shown and the tumors are indicated by white arrows. Reproduced with permission from Cai et al. (2007a)

Fig. 3

SKBR3-luc cancer cells were inoculated into left flank of athymic nu/nu mice. a Bioluminescence image overlaid on white-light image. White-light image of b mouse taken before fluorescence imaging of (¹¹¹In-DTPA)_n-trastuzumab-(IRDye800)_m (c), 200-fold molar excess of trastuzumab followed by (¹¹¹In-DTPA)_n-trastuzumab-(IRDye800)_m (d), (¹¹¹In-DTPA)_p-IgG-(IRDye800)_q (e), or equivalent dose of IRDye800CW (f) 48 h after administration. Standards placed on side include: (1) (¹¹¹In-DTPA)_n-trastuzumab-(IRDye800)_m, (2) (¹¹¹In-DTPA)_p-IgG-(IRDye800)_q, and (3) IRDye800CW. Reproduced with permission from Sampath et al. (2007)

Fig. 4

a Dynamic small animal PET scans obtained for ¹⁸F-FB-T84.66 diabody with LS 174T tumor-bearing mice and C6 tumor-bearing mice. Coronal whole-body slices that contained tumors are shown; arrows indicate tumors. b Comparison of LS 174T tumor uptake and C6 tumor uptake. Values were determined from ROI analysis of small animal PET imaging data. Differences were significant at all time points examined. *P < 0.05; **P < 0.01. Reproduced with permission from Cai et al. (2007c)

Fig. 5

Gamma camera images of HER2-expressing SKOV-3 xenograft tumors in BALB/c nu/nu mice. The time to image acquisition (a) and controls to prove imaging specificity for HER2 using competition with unlabeled Z_HER2:342 (b), a negative control affibody molecule not targeting HER2 (c), and drug-induced HER2 degradation (d) are shown. All animals were i.v. injected with 3 MBq (3 μg) of ¹¹¹In-DOTA-Z_{HER2:342-pep2} or ¹¹¹In-DOTA-Z_taq4:5. To facilitate interpretation, white contours were superimposed around some animals to indicate the location of the animals on the gamma camera screen. Arrows indicate positions of kidneys (K) or tumors (T). a Imaging of mice at different time points after injection, 1 h (top), 2 h (middle), and 4 h (bottom) after injection of ¹¹¹In-DOTA-Z_{HER2:342-pep2}. b Imaging of mice pre-blocked (bottom) or not blocked (top) with 0.9 mg unlabeled Z_HER2:342 45 min before injection of ¹¹¹In-DOTA-Z_{HER2:342-pep2}. Imaging was done 1 h after injection. c Imaging of mice injected with ¹¹¹In-DOTA-Z_{HER2:342-pep2} (top) or ¹¹¹In-DOTAZ_taq4:5 (bottom) 4 h after injection. d Imaging of mice 4 h after injection, pretreated with 17-AAG (bottom) and controls (top). Reproduced with permission from Orlova et al. (2007)

Fig. 6

MicroPET of ⁶⁴Cu-DOTA-VEGF₁₂₁ in U87MG tumor-bearing mice. a Serial microPET scans of large and small U87MG tumor-bearing mice injected intravenously with 5–10 MBq of ⁶⁴Cu-DOTA-VEGF₁₂₁. Mice injected with ⁶⁴Cu-DOTA-VEGF₁₂₁ 30 min after injection of 100 μg VEGF₁₂₁ are also shown (denoted as “Small tumor + block”). b Two-dimensional whole-body projection of the three mice shown in a at 16 h after injection of ⁶⁴Cu-DOTA-VEGF₁₂₁. Tumors are indicated by arrows. Reproduced with permission from Cai et al. (2006b)

Fig. 7

Schematic illustration of biotin ligase (a), phosphopantetheinyl transferase (PPTase) (b), formylglycine-generating enzyme (FGE) (c), human O⁶-alkylguanine-DNA alkyltransferases (hAGT) (d), and transglutaminase (TGase) (e) catalyzed modification