Nuclear Imaging of Glucose Metabolism: Beyond 18F-FDG

Glucose homeostasis plays a key role in numerous fundamental aspects of life, and its dysregulation is associated with many important diseases such as cancer. The atypical glucose metabolic phenomenon, known as the Warburg effect, has been recognized as a hallmark of cancer and serves as a promising target for tumor specific imaging. At present, 2-deoxy-2-[18F]fluoro-glucose (18F-FDG)-based positron emission tomography/computed tomography (PET/CT) represented the state-of-the-art radionuclide imaging technique for this purpose. The powerful impact of 18F-FDG has prompted intensive research efforts into other glucose-based radiopharmaceuticals for positron emission tomography (PET) and single-photon emission computed tomography (SPECT) imaging. Currently, glucose and its analogues have been labeled with various radionuclides such as 99mTc, 111In, 18F, 68Ga, and 64Cu and have been successfully investigated for tumor metabolic imaging in many preclinical studies. Moreover, 99mTc-ECDG has advanced into its early clinical trials and brings a new era of tumor imaging beyond 18F-FDG. In this review, preclinical and early clinical development of glucose-based radiopharmaceuticals for tumor metabolic imaging will be summarized.


Introduction
Glucose, a common monosaccharide in nature, is the primary source of energy for most living organisms. Glucose homeostasis plays a key role in numerous fundamental aspects of life, and its dysregulation is associated with many important human diseases such as cancer [1][2][3]. Cancer is a class of diseases characterized by their uncontrollable proliferation, invasion, and metastasis. In the course of cancer progression, there is a shift of glucose metabolism from mitochondrial oxidative phosphorylation to a glucosedependent glycolytic pathway, even in the availability of oxygen [4,5]. To maintain the demand of energy for rapid proliferation, cancer cells increase glucose uptake as well as their glycolytic rate, which can be up to 200 times greater than that of normal cells.
is atypical metabolic phenomenon is known as the Warburg effect, which has been recognized as a hallmark of cancer and serves as a promising target for diagnosis and therapy of cancer ( Figure 1) [6][7][8].
At present, 2-deoxy-2-[ 18 F]fluoro-glucose ( 18 F-FDG)based positron emission tomography/computed tomography (PET/CT) represents the state-of-the-art radionuclide imaging technique for this purpose. e 18 F-FDG was synthesized by Pacák and his colleagues in 1969 [9], and it was investigated as a PET tracer in the 1970s and early 1980s [10,11]. Since then, it has been broadly used in the clinic.
Currently, 18 F-FDG is the most popular glucose-based radiopharmaceutical and is honored as the "molecule of the century" in the field of molecular imaging. e 18 F-FDG combined with PET/CT has shown great value in the diagnosis, staging, monitoring therapeutic response, and assessment of prognosis [12,13]. e powerful impact of 18 F-FDG in the clinic has prompted intensive research efforts into other glucose-based radiopharmaceuticals for positron emission tomography (PET) and single-photon emission computed tomography (SPECT) imaging. In the last decades, glucose and its analogues labeled with various radionuclides such as 99m Tc,111 In, 18 F, 68 Ga, and 64 Cu have been successfully investigated for tumor metabolic imaging in many preclinical studies [14][15][16][17]. Moreover, 99m Tc-ECDG has advanced into its early clinical trials and brings a new era of tumor imaging beyond 18 F-FDG. In this review, preclinical and early clinical development of glucose-based radiopharmaceuticals for tumor metabolic imaging will be summarized.

Preclinical Development of Glucose-Based Radiopharmaceuticals
2.1. SPECT Imaging with 99m Tc-Labeled Glucose Analogues. e availability of commercial generators and kit chemistry to prepare 99m Tc-labeled radiopharmaceuticals has had a great impact on nuclear medicine. Considerable efforts have been seen on the development of 99m Tc-labeled glucose analogues for tumor imaging. In this section, the summary of these 99m Tc-labeled glucose analogues is provided, as shown in Figure 2.
2.1.1. 99m Tc-EC-DG. Because of the widely used 99m Tcethylenedicysteine conjugates, 99m Tc-ethylenedicysteinedeoxyglucose (EC-DG) was designed, synthesized, and investigated for tumor imaging by Yang and his colleagues in 2003 [18]. e positive result was achieved in hexokinase assay, which suggested that EC-DG is phosphorylated by hexokinase. e uptake of 99m Tc-EC-DG is comparable to that of 18 F-FDG and was decreased in the presence of D-glucose in lung cancer cells. is finding confirmed the uptake of 99m Tc-EC-DG is mediated by glucose transporters. Subsequently, the biodistribution of 99m Tc-EC-DG in lung tumor-bearing mice was studied. e results showed the tumor uptake of 99m Tc-EC-DG is 2-3 times lower than that of 18 F-FDG (0.41 ± 0.16 vs 1.60 ± 0.18%ID/g). In view of tumor-to-brain tissue and tumor-to-muscle tissue ratios, 99m Tc-EC-DG was superior to 18 F-FDG. e feasibility of 99m Tc-EC-DG for SPECT imaging of tumor was evaluated. e smallest tumor with 3 mm in diameter could be better imaged by 99m     corresponding nontumorous region ratio obtained by SPECT was determined to be 1.82 ± 0.07 for the small tumors and 2.88 ± 0.10 for the medium-sized tumors. In addition, the uptake of 99m Tc-EC-DG was decreased in rats pretreated with FDG, while it was increased in the group pretreated with insulin, which is consistent with in vitro results. ese preliminary results demonstrated the potential of 99m Tc-EC-DG for tumor metabolic imaging. Currently, 99m Tc-EC-DG has advanced into Phase II/III clinical trials (NCT00865319 and NCT01394679), and the corresponding results would be discussed in the following section in detail.

99m
Tc-DTPA-DG. In 2006, diethylenetriaminepentaacetic acid-deoxyglucose (DTPA-DG) was synthesized and then labeled with 99m Tc using a kit formula with a radiochemical purity of 99.2% [19]. e significant uptake of 99m Tc-DTPA-DG was observed at 4 h in vitro, which was comparable with that of 18 F-FDG. e biodistribution of 99m Tc-DTPA-DG in breast tumor-bearing rats showed a marked tumor uptake of 99m Tc-DTPA-DG, which is 1-2 times higher than that of 18 F-FDG. Rapid blood clearance of 99m Tc-DTPA-DG was visualized with the renal excretion. Compared with 18 F-FDG, 99m Tc-DTPA-DG has higher tumor-to-muscle and tumor-to-brain ratios but a lower tumor-to-blood ratio (4.30 ± 0.89, 19.88 ± 3.45, and 3.24 ± 0.65, respectively). Compared with 99m Tc-DTPA, the tumor could be better imaged by 99m Tc-EC-DG SPECT imaging with a tumor to nontumor ratio of 2.46 ± 1.02 and 3.54 ± 1.36 at 0.5 and 3 h, respectively. e feasibility of 99m Tc-DTPA-DG for tumor imaging has been demonstrated, and 99m Tc-DTPA-DG enables visualization of the tumors up to 4 h after injection. In addition, 99m Tc-DTPA-DG has been involved in evaluating early chemotherapy response and differentiating the tumor from inflamed tissues [20][21][22][23]. Considering these positive results, 99m Tc-DTPA-DG may be a potential radiopharmaceutical for tumor imaging. However, the mechanism underlying cellular uptake of 99m Tc-DTPA-DG is not clearly understood. Further studies in various animal models and humans are needed.  [24]. 99m TcN-DGDTC was demonstrated to be hydrophilic and neutral. e biodistribution study showed the high accumulation of 99m TcN-DGDTC in tumors with good retention (1.16 ± 0.57%ID/g at 4 h). Because of the faster blood and muscle clearance, the tumor/blood and tumor/muscle ratios increased with time and reached 2.32 and 1.68 at 4 h after injection. Further studies of the biological characteristics of this radiopharmaceutical may lead to identify a promising candidate for tumor imaging. Similarly, the same group described the radiolabeling of DGDTC by ligand-exchange reaction with 99m Tc-glucoheptonate containing the [ 99m TcO] 3+ core [25]. 99m TcO-DGDTC was prepared under neutral condition and at 100°C for 15 min to achieve high radiochemical purity (>90%). 99m TcO-DGDTC was hydrophilic and positively charged. e cell uptake of 99m TcO-DGDTC increased over time and reached the highest at 4 h. Moreover, the uptake was decreased by the presence of D-glucose, which indicated 99m TcO-DGDTC and D-glucose share a similar mechanism of uptake. A significant tumor uptake was observed in biodistribution studies with a long time of retention (2.73 ± 0.72, 2.85 ± 0.63, and 3.53 ± 0.85%ID/g at 0.5 h, 2 h, and 4 h after injection, respectively). Moreover, the tumorto-blood and tumor-to-muscle ratios were increased over time. As compared with 99m TcN-DGDTC, 99m TcO-DGDTC had a higher tumor uptake and tumor to muscle ratio but a lower tumor to blood ratio. In contrast, the tumor uptake and tumor-to blood-ratio of 99m TcO-DGDTC was lower than that of 18 F-FDG, but its tumor to muscle ratio is higher. Additionally, the tumor could be clearly detected by SPECT imaging in tumor-bearing mice. ese good biological features endow 99m TcO-DGDTC as a potential tumor imaging agent.

99m Tc-MAG 3 -Glucose Analogues.
As a well-known bifunctional chelator, MAG 3 has been involved in the development of 99m Tc-labeled glucose analogues. In 2006, MAG 3 -DG was designed, synthesized, and radiolabeled via ligand-exchange reaction with 99m Tc-glucoheptonate to produce 99m Tc-MAG 3 -DG [26]. e biodistribution of 99m Tc-MAG 3 -DG was performed in breast tumor-bearing mice. A moderate tumor uptake was observed and estimated to be 0.82 ± 0.06%ID/g. e tumor-to-muscle ratio and tumor-to-blood ratio was determined to be 4.35 ± 1.41 and 0.94 ± 0.13, respectively. In addition, 99m Tc-S-DG and 99m Tc-MAMA-BA-DG were also synthesized and evaluated in this work.
ere were significant similarities in the biodistribution of these three radiopharmaceuticals. e only difference is hepatobiliary excretion for 99m Tc-MAMA-BA-DG. Among them, 99m Tc-MAG 3 -DG showed the most favorable characteristics and could be further studied as potential tumor imaging agents. In 2009, de Barros et al. reported the synthesis of glucose analogue MAG 3 -G and radiolabeled it with 99m Tc-tartarate via ligand-exchange reaction [27]. e radiochemical purity was higher than 90%.
e biodistribution of 99m Tc-MAG 3 -G in Ehrlich tumor-bearing mice showed that the highest tumor uptake (1.64 ± 0.19%ID/g) is obtained at 0.5 h after injection. However, the tumor-to-muscle ratio and tumor-to-blood ratio increased with time and reached 5.03 ± 0.98 and 2.42 ± 0.50 at 8 h after injection. In addition, 99m Tc-MAG 3 -G was excreted rapidly through the liver and kidneys. e feasibility of 99m Tc-MAG 3 -G for tumor imaging needs to be further evaluated. Similarly, the same group synthesized three compounds MAG 3 -Gl, MAG 3 -Ga, and MAG 3 -NG and successfully radiolabeled them with 99m Tc in high radiochemical purities [28]. ese three complexes were rapidly excreted through kidneys and had a similar biodistribution in normal mice. Subsequently, the biodistribution of 99m Tc-MAG 3 -Gl in tumor mice was carried out [29]. e tumor uptake was high (2.25%ID/g) at 5 min after injection and decreased over time. However, the target-to-nontarget ratio was always greater than 2.0. SPECT imaging showed a marked uptake of 99m Tc-MAG 3 -Gl in tumor with a targetto-nontarget ratio of about 2.0, which was in agreement with biodistribution studies. ese preliminary results suggested 99m Tc-MAG 3 -Gl would possess the potential for tumor imaging.

99m
Tc-1-TG and 99m Tc-DGTA. As another analogue of β-D-glucose, 1-thio-β-D-glucose (1-TG) was labeled with 99m Tc in high labeling efficiency (>97%) [30]. e in vitro assay showed the uptake of 99m Tc-1-TG highly depends on 1-TG concentration and was not significantly changed with different glucose concentrations. is finding indicated the tumor uptake mechanism for 99m Tc-1-TG was different from that for 18 F-FDG. Further studies were carried out for early detection of melanoma tumor [31]. e tumor uptake of 99m Tc-1-TG was clearly visualized by scintigraphic imaging, showing potential as a new type of radiopharmaceutical for melanoma imaging. In addition, 99m Tc-1-TG has been successfully used for inflammation imaging [32].
Lee et al. reported the preparation of diglucosediethylenetriamine (DGTA) from diethylenetriamine and natural D-glucose using a single step chemical synthesis and radiolabeled it with 99m Tc in a high radiochemical yield of >95% [33]. e in vitro cell uptake of 99m Tc-DGTA was 1.5-8 times higher than that of 18 F-FDG. However, the uptake of 99m Tc-DGTA was not highly dependent on glucose concentration, which indicated that its uptake mechanism differs from that of 18 F-FDG. Although promising, further investigations in animal models are necessary.
Additionally, Fernández et al. reported the derivatization of glucose at C-2 using the so-called "click chemistry" to form a histidine-like, 1,4-disubstituted triazole molecule for radiolabeling with [ 99m Tc(CO) 3 ] + ligand [39]. A relatively low protein binding of 99m Tc(CO) 3 -glucose-histidine was obtained, correlating with its high in vitro stability and hydrophilicity. Biodistribution was characterized by low blood and liver uptake. Because of its hydrophilicity, renal excretion was observed as expected. e tumor uptake of 99m Tc(CO) 3 -glucose-histidine was moderate and retained for a long time. e tumor-to-muscle ratio was high and was determined to be 2.75 ± 0.06 at 2 h after injection. By comparison, 99m Tc(CO) 3 -glucose-histidine and 18 F-FDG have a similar biodistribution in C57BL/6 mice bearing induced Lewis murine lung carcinoma. However, the tumor uptake and tumor-to-muscle ratio of 99m Tc(CO) 3 -glucosehistidine are much lower than those of 18 F-FDG. Modifications of the structure are needed to improve biological properties. . Recently, a D-glucosamine derivative with an isonitrile group (CN5DG) was synthesized and labeled with 99m Tc to prepare 99m Tc-CN5DG (Figure 3) [40]. 99m Tc-CN5DG was readily obtained with high radiochemical purity (>95%) and specific activity (11.17-335.22 GBq/ mmol).

99m
is hydrophilic radiopharmaceutical exhibited great in vitro stability and metabolic stability in urine. e tumor cell uptake of 99m Tc-CN5DG was significantly blocked in the presence of D-glucose and increased by insulin, which demonstrated that 99m Tc-CN5DG is transported via glucose transporters. Biodistribution studies in mice bearing A549 xenografts showed that 99m Tc-CN5DG had a rapid, high tumor uptake and cleared quickly from normal organs, resulting in a satisfactory tumor-tobackground ratio. e tumor uptake of 99m Tc-CN5DG is comparable to that of 18 F-FDG. However, the tumor-toblood, tumor-to-muscle, and tumor-to-lung ratios of 99m Tc-CN5DG are much higher than those of 18  Tc-CN5DG coupled clearly visualizes the tumor sites for a long time.
e smallest tumor that can be detected by 99m Tc-CN5DG SPECT imaging was about 3 mm. ese excellent biological characteristics confirmed that 99m Tc-CN5DG may be a potential "working horse" and be another breakthrough in glucose-based radiopharmaceuticals for tumor imaging.

SPECT Imaging with 111 In-Labeled Glucose Analogues.
e c-emitting radioisotope indium-111 ( 111 In) (t 1/2 � 2.83 d, 171 KeV (90%), 245 KeV (94%)) is of great practical interest for clinical SPECT [41][42][43]. Because of its large size, the macrocyclic chelators such as diethylenetriaminepentaacetic acid (DTPA), 1,4,7,10-tetraazacyclodode cane-1,4,7,10tetraacetic acid (DOTA), and 1,4,7-triazacyclononane-1,4,7triacetic acid (NOTA) have usually been used to form stable metal complexes for further conjugation of biomolecules [44]. In 2012, Yang and his coworkers reported the development of radiopharmaceutical 111 In-DOTA-DG from precursor compound DOTA-DG and 111 In with the labeling efficiency of >95% and radiochemical purity of >96% (Figure 3) [45]. Nude mice bearing MDA-MB-468 mammary tumors were employed to evaluate the pharmacokinetics and targeting ability of 111 In-DOTA-DG. e prominent accumulation of radioactivity in the liver, kidneys, and urinary bladder was observed, indicating that this radiopharmaceutical is mainly through the renal excretory pathway. As seen from the SPECT images, the tumors were visualized at 120 min after injection. e results suggest that 111 In-DOTA-DG may be a promising glucose-based radiopharmaceutical for SPECT imaging. Further detailed evaluation is required to elucidate its metabolic mechanism.

SPECT Imaging with Radioiodine-Labeled Glucose
Analogues. Because of the excellent physical properties, easy accessibility, and low manufacturing cost, radioiodination of glucose analogues for SPECT imaging is of great clinical interest. So far, several radioiodine-labeled glucose analogues have been proposed [46,47]. Iodine-123 ( 123 I) is the halogen isotope of choice due to its excellent physical properties that make it ideal for imaging. Of them, 2-deoxy-2-[ 123 I] iodo-glucose (IDG) is the most logical form of iodinated glucose analogues. However, this small molecule is not stable under physiological conditions, making it unsuitable for imaging applications [46]. Subsequently, 2deoxy-2-fluoro-2-iodo-D-mannose (FIM) and 2-deoxy-2fluoro-2-iodo-D-glucose (2-FIG) were synthesized and shown to be stable for several days in saline, which demonstrated that the presence of fluorine on position 2 enables the stability of iodine atom in glucose analogues [48]. erefore, the FIM was labeled with 123 I to produce 2-fluoro-2-[ 123 I]iodo-D-mannose ( 123 I-FIM), which was stable in vitro for 24 hours (Figure 4). Unfortunately, in vivo studies showed 123 I-FIM has a rapid blood clearance and high stomach and thyroid uptake, indicating its rapid deiodination after injection [49]. In addition, iodination of glucose isomers in positions 3, 4, and 6 were investigated, and none of them exhibited the similar biological features of 2-deoxy-D-glucose [50]. e iodinated glucose analogues have not been found to be metabolic markers for in vivo studies.

PET Imaging with 18 F-Labeled Glucose Analogues.
e 18 F-FDG is the only Food and Drug Administration-(FDA-) approved glucose-based radiopharmaceutical and has been used worldwide. e success of 18 F-FDG leads to the development of other 18 F-labeled glucose analogues for tumor imaging ( Figure 5).

18 F-6FDG.
With the goal of developing the radiopharmaceuticals similar to 18  . e preliminary in vitro and in vivo studies demonstrated 18 F-6FDG may be a more representative candidate for the glucose transporter than 18 F-FDG. Interestingly, because of the substitution of fluorine at C-6 position, 18 F-6FDG is just transported through glucose transporters and cannot be phosphorylated for subsequent metabolism [52]. Meanwhile, 1-[ 18 F]fluorodeoxyfructose, 1-[ 18 F]-fluoroalkyldeoxyglucose, and other glucose analogues have been evaluated as novel candidates for PET imaging [53].

18 F-Labeled
Glucosamine. N-[ 18 F]Fluoroacetyl-D-glucosamine ( 18 F-FAG) was the first D-glucosamine analogue to be radiolabeled with 18 F by Fujiwara and his colleagues in 1990 [54]. C3H/HeMsNRS mice with spontaneous hepatomas were used for PET imaging. e high uptakes of 18 F-FAG were observed in the tumor, liver, and kidney at 60 min after injection, whose mean value was estimated to be 5.16%ID/g, 3.71%ID/g, and 3.27%ID/g, respectively. Among all tissues, the tumor has the highest radioactivity with a long retention (5.51%ID/g at 5 min after injection and 5.16%ID/g at 60 min after injection). Furthermore, the tumor was clearly visualized by PET imaging in the rabbit VX-2 tumor model. erefore, 18 F-FAG is a promising PET radiotracer for tumor imaging. Another glucosamine-based radiopharmaceutical, of note, is N-(2-[ 18 F]fluoro-4-nitrobenzoyl)glucosamine ( 18 F-FNBG) [55]. e biodistribution study in mice models showed 18 F-FNBG mainly accumulates in the tumor, liver, and kidney. e tumor uptake of 18 F-FNBG became the highest at 5 min after injection with a value of 1.68 ± 0.05%ID/g and was decreased with time. At 120 min, the tumor still has an uptake of 0.21 ± 0.02%ID/g, which is comparable to that of the liver and kidney at the same time. Besides, the tumor uptake of 18  was 4.32 ± 0.79%ID/g for 18 F-FAG. However, the tumor/ blood and tumor/muscle ratios of 18 F-FNBG were similar with those of 18 F-FAG. In addition, Carroll et al. reported the synthesis of three novel 18 F-labeled glucosamine analogues ( 18 F-5, 18 F-8 and 18 F-13) and the evaluation of their tumor uptake in vivo by PET imaging [56]. Among them, 18 F-13 showed a discernible tumor uptake of 2.80 ± 0.51%ID/g at 60 min after injection, which is 5.12 ± 1.59%ID/g for 18 F-FDG. In view of these primary results, 18 F-labeled glucosamine analogues might be promising candidates for tumor PET imaging. More studies are needed to further investigate their imaging property.

18 F-FDG-2-NIm and 18 F-GAZ.
In 2002, Patt and his coworkers synthesized a new glucose-coupled 2-nitroimidazole derivative, 18 F-FDG-2-NIm, from the peracetylated 2-[ 18 F]FDG in good radiochemical yields [57]. In comparison of 18 F-FDG, in vitro and in vivo studies demonstrated much lower uptake of 18 F-FDG-2-NIm, which suggested that the accumulation into tumor cells via glucose transporters is unlikely to occur. Another similar radiopharmaceutical is an azomycin-glucose conjugate 18 F-GAZ [58]. PET imaging showed the accumulation of 18 F-GAZ was observed at 5-6 min after injection (0.66 ± 0.05%ID/g) and decreased in a time-dependent manner. At 60 min after injection, the tumor uptake was measured to be 0.24 ± 0.04% ID/g with a tumor/muscle ratio of 1.87 ± 0.18. However, competitive experiment showed F-GAZ is a weaker competitive inhibitor of 18 F-FDG compared with D-glucose and unlabeled 2-FDG. 18 F-GAZ seems not to be uptaken by glucose transporters, and further studies should be carried out.  [59]. In comparison with conventional routes, the click labeling method spent shorter time to obtain the radiopharmaceutical with higher decay-corrected radiochemical yield and specific activity. Unfortunately, this radiopharmaceutical was demonstrated to be incompatible for hexokinase phosphorylation and independent of glucose transporter.
However, the radiolabeling process is the multistep chemical manipulation and time-consuming, which is incompatible with the short half-life of 11 C (t 1/2 � 20.3 min). erefore, direct 11 C labeling strategies are still needed. In 2003, Bormans et al. developed a nonmetabolizable 11 C-labeled α/β-methyl-D-glucoside ( 11 C-αMDG and 11 C-βMDG) that is selectively transported by sodium dependent glucose transporters (SGLTs) [68]. ese radiotracers were prepared by straightforward methylation of glucose with 11 C-methyl triflate in a total synthesis time of 20 min and a yield of 30% (decay corrected). In vivo PET imaging showed 11 C-labeled α/β-methyl-D-glucoside accumulated in the kidney, which depends on the functionality of SGLTs in the luminal membrane of renal proximal tubules. Consequently, 11 Cmethyl-D-glucoside is a promising PET tracer for the in vivo visualization of SGLTs in kidney malfunction. Future studies are needed to elucidate whether 11 C-methyl-D-glucoside may be used to detect various human tumors with high level of SGLT transporters. 68 Ga-Labeled Glucose Analogues. e growth and worldwide spread of positron emitting radionuclide gallium-68 ( 68 Ga) in preclinical and clinical research has proven its potential for PET imaging during last two decades. e advantages of 68 Ga such as favorable physical and chemical properties, commercially available generators, robust labeling chemistry diversity have been presented in detail in many literatures and strongly motivate researchers to develop new 68 Ga-based radiopharmaceuticals [69,70].

PET Imaging with
In 2012, Yang et al. radiolabelled 1,4,7,10tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-2deoxy-D-glucosamine (DOTA-DG) with 68 Ga to produce radiopharmaceutical 68 Ga-DOTA-DG with the labeling efficiency of ∼85% and radiochemical purity of 98% in ten minutes assisting with microwave ( Figure 6) [71]. e percentage uptake of 68 Ga-DOTA-DG in A431 cells at 60 min is comparable to that of 18 F-FDG (15.7% and 16.2%, respectively). In a human tumor xenograft model, the tumor uptake of 68 Ga-DOTA-DG was determined to be 0.39%ID/g, which is much lower than that of 18 F-FDG (4.26%ID/g) after 60 min injection. However, the tumor-to-heart, tumor-tobrain, and tumor-to-muscle ratios are higher in comparison with 18 F-FDG. Additionally, the PET images showed renal excretion and accumulation in the bladder. Significant researches are needed to elucidate the potential of 68 Ga-DOTA-DG as a candidate for clinical tumor imaging.
e precursor Zn-ATSE/A-G was synthesized and radiolabeled by 64 Cu via copper-zinc exchange with a radiochemical yield of 71.7% (Figure 7) [76,77]. In the PET images, 64 CuATSE/A-G displayed moderate tumor uptake and a divergent pattern of biodistribution compared with 18 F-FDG. Renal excretion and accumulation in the bladder were observed. In particular, the distinctive brain and heart uptake of 18 F-FDG was not obtained in the images of 64 Cu-ATSE/A-G, which demonstrate that it does not participate in glucose-specific transport and is not a surrogate for glucose metabolism imaging. In addition, the uptake of 64 Cu-ATSE/ A-G in proportion to the O 2 concentration in the HeLa cells demonstrated its hypoxia selectivity and feasibility for hypoxia imaging.

Early Clinical Development of Glucose-Based Radiopharmaceuticals
Since 1969, when 18 F-FDG was developed for PET imaging in the clinic, intense attempts have been made in the development of other glucose-based radiopharmaceuticals for SPECT and PET imaging, which create a pipeline of exciting tracers for tumor imaging. Among them, 99m Tc-EC-DG is the only glucose-based radiopharmaceutical, which has advanced in Phase II/III clinical trials (NCT00865319, NCT01394679). To assess the biodistribution, radiation dosimetry, and diagnostic efficacy, 99m Tc-EC-DG SPECT imaging and 18 F-FDG PET imaging were performed in seven patients with non-small-cell lung cancer (NSCLC) (Figure 8) [78]. It was found that the uptake of 99m Tc-EC-DG was mainly visualized in the blood pool, kidneys, bladder, and liver over time. Bladder wall was deemed to be the critical organ that receives the highest dose, with an average radiation absorbed dose of 2.47 × 10 −2 mGy/MBq. e mean effective dose equivalent and effective dose were estimated to be 6.20 × 10 −3 mSv/MBq and 5.90 × 10 −3 mSv/MBq for administration of 1,110 MBq activity, which is less than that of 18 F-FDG (3.00 × 10 −2 mSv/MBq). Whole-body images showed that the primary tumor was clearly visualized in six of the seven patients at 4 h after injection with confirmed NSCLC and concordant accumulation of 18 F-FDG. However, tumor-to-background ratios obtained with 99m Tc-EC-DG is lower than that of 18 F-FDG. e patient with negative uptake of 99m Tc-EC-DG and positive uptake of 18 F-FDG was pathologically documented to have a granuloma. In addition, the administration of 99m Tc-EC-DG was well tolerated in this cohort of patients. All these results are encouraging and endow 99m Tc-EC-DG as a diagnostic agent for nuclear medicine imaging. Larger scale clinical studies are now warranted to assess the utility of 99m Tc-EC-DG for tumor imaging.

Conclusions
Diagnosis, staging, monitoring therapeutic response, and assessment of prognosis in the management of patients with cancer pose major challenges to today's medical imaging.
e development of 18   . Uptake is also seen in the blood pool (great vessels; dashed arrows). e three images (left to right) are from the 2, 4, and 6 h SPECT scans, respectively. Adapted from Reference [78] with permission.
represents a major milestone in the field of molecular imaging to overcome these limitations. Along a similar line, glucose and its analogues have been chemically modified by a carefully researched bifunctional chelator with their biochemical properties retained, labeled with various radionuclides, and explored for tumor imaging. Among these glucose-based radiopharmaceuticals for SPECT imaging, 99m Tc-EC-DG, 99m Tc-MAG 3 -DG, and 99m Tc-CN5DG represent a few named diagnostic tracers in this field. Novel glucose-based PET radiopharmaceuticals, such as 18 F-6FDG, 18 F-GAZ, and 68 Ga-DOTA-DG, substantiate their advantages over others, showing great potential for clinical translation. ese radiopharmaceuticals bring a new era of tumor imaging beyond 18 F-FDG. Despite a wealth of preclinical data, only 99m Tc-EC-DG has advanced in Phase II/III clinical trials, and the cases reported so far are few in number. e barriers for developing promising glucose-based radiopharmaceuticals and preventing the translation of them in the clinic are many and not clear. e first challenge lies in the choice of radiochemistry. e introduction of a prosthetic or bulky metal-bearing moiety may have a large impact on the overall biochemical properties of a glucose analogue. erefore, the structureactivity relationship of glucose-based radiopharmaceuticals should be carefully optimized. e simple and fast radiolabeling processes with a high radiochemical purity and specific activity are essential for the development of a promising radiopharmaceutical. In addition, the "theranostics" is new direction of nuclear medicine. Researchers should put efforts to explore the labeling of glucose analogues with 177 Lu, 90 Y, 188 Re, etc. and to investigate them as potential theranostic agents. On the other hand, the design of preclinical studies and clinical trials for clinical translation of glucose-based radiopharmaceuticals are also very important. erefore, the concerted efforts from pharmacologists, radiologists, oncologists, and clinicians are required to validate these well-designed radiopharmaceuticals as clinical diagnostic agents. Furthermore, it is our belief that the increasing radiolabeled glucose analogues will enter clinical trials, progress to authorized approval, and ultimately become widely used imaging agents in the clinic.

Conflicts of Interest
e authors declare no conflicts of interest.