Angiogenesis, a well-known process by which new blood vessels sprout from the pre-existing capillaries, includes vascular endothelial cell activation, the degradation of vascular basement membrane during the activation, proliferation and migration of endothelial cells, and the construction of new blood vessels and vascular networks. It is associated with a long list of physiological and pathological events (1, 2). Under normal physiological conditions, such as embryonic development, wound healing, and the menstrual cycle, angiogenesis serves primarily to ensure adequate nutrient and oxygen supply and waste removal. Of course, normal angiogenesis also occurs in pathological conditions, such as tissue damage after reperfusion of ischemic tissue or cardiac failure (3, 4). Unlike the normal angiogenesis described above, both solid and hematologic tumors feature a high dependence on persistent abnormal angiogenesis (5–7). Solid tumors initially form as avascular masses that rely on the vascular system in the host microenvironment, but upon reaching a diameter of 1–2 mm, these tumors exhibit excessive dysfunctional angiogenesis to obtain essential oxygen and nutrients (8–11). The initial recognition of angiogenesis as an interesting process in oncology began in the early 1970s, when Drs. Folkman and Denekamp put forward the idea that highly immature neovascularized tumors are vulnerable via their blood supply. The immature vessels are characterized by an undifferentiated endothelium and a lack of smooth muscles, and act as the vehicles for peripheral sprouting of new capillaries, invasion and distant metastasis (12–15). These mechanisms are facilitated by the synergistic effects of transmission signals from the extracellular matrix (ECM) to endothelial cells, among which integrin binding to endothelial cell recognition sites is the first process to occur. Importantly, the tripeptide sequence of Arg-Gly-Asp (RGD) was identified as a common recognition site in the ECM and blood, and can bind to a variety of adhesion proteins and integrins (16).

The contribution of the RGD tripeptide sequence to cell attachment was first recognized in fibronectin by Pierschbacher and Ruoslahti in 1984 and later confirmed in more ligands, namely, vitronectin, fibrinogen, decorsin, von Willebrand factor, thrombospondin, disintegrins, laminin, entactin, tenascin, prothrombin, osteopontin, bone sialoprotein, adenovirus penton base protein, and collagens (16–18). Combinations of different alpha (α) and beta (β) subunits, such as αvβ3, αvβ5, αvβ8, α5β1 and α8β1, act as receptors for many of the above-mentioned proteins, mainly acting on angiogenesis, inflammation, thrombosis, lung development, cell differentiation, and other pathophysiological processes (19–22). Among these, αvβ3 is highly expressed on angiogenic endothelial cells and several tumor cells, but is expressed only at low levels by resting endothelial cells of normal tissues (23–26). The αvβ3 expression level has also been reported to be a promising factor in tumor diagnosis, staging and therapy (27–29). Due to the high specific affinity between RGD and αvβ3, radiolabeled RGD peptides have been intensively studied for their ability to represent αvβ3 expression density and the tumor angiogenesis state noninvasively (30, 31). Combinations of RGD-containing peptides with radionuclides such as ¹⁸F, ⁶⁴Cu, and ⁶⁸Ga, have been introduced successfully in positron emission tomography/computed tomography (PET/CT) imaging, among which ¹⁸F has near ideal decay characteristics for PET (t_1/2 = 110 min, β⁺ = 0.64 MeV). A linear RGD peptide was first labeled with ¹⁸F via solid-phase synthesis in 2001, but this radiotracer showed nonspecific accumulation within tumor tissue and early breakdown (32). Then, the cyclic pentapeptide cyclo (Arg-Gly-Asp-D-Phe-Val) (denoted as cyclic RGD peptide) was prepared and found to be a highly potent, stable and selective inhibitor of integrin signaling (33, 34). In addition, based on the polyvalency effect, different types of dimeric and multimeric cyclic RGD peptides were prepared to improve αvβ3-binding affinities for intense tumor accumulation and increased tumor/background ratios compared with the monomeric compounds (35–37). During the optimization of radiotracers, strategies such as cyclization, glycosylation, pegylation, multimerization, and insertion of spacers/linkers have been adopted to optimize the in vitro and in vivo behaviors of radiolabeled RGD peptides (38–41). However, few of these RDG peptide labeling methods have been successfully translated from the bench to clinical use.

Therefore, in this review, we focus on the preliminary clinical applications of PET radionuclide-conjugated RGD-containing peptides that target RGD-binding integrins (mainly αvβ3 integrin), discussing their distribution characteristics and potential use in diagnosis, delineation of tumor subvolume, evaluation response of chemoradiotherapy or antiangiogenic therapy, and the comparative results achieved with traditional ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG) imaging agent in the context of functional tumor imaging.

Monomeric and multimeric RGD-based PET tracers that have been investigated in preliminary clinical studies are listed in Supplementary Table 1 and Figure 1 . ¹⁸F-Galacto-RGD, which is a glycosylated RGD-peptide, was initially reported by Haubner et al. in 2005 and was the first monomeric integrin-specific PET tracer used in patients (56). It is well tolerated, as all studies reported neither drug-related adverse events nor important clinical indicators. The dose distribution results were consistent for a variety of tumor types, with similar evidence of predominant renal excretion, fast blood elimination, rapid tumor tracer accumulation, and low background concentration in most organs, namely, the lung, muscle tissue and brain, allowing imaging of tumor lesions with good contrast (56–58). A relatively small amount of ¹⁸F-Galacto-RGD is also excreted via the hepatobiliary pathway, and thus, in some patients, tracer uptake is seen in the gallbladder. Similar biodistribution results were observed for the other monomeric radionuclide tracers such as ¹⁸F-fluciclatide (denoted as ¹⁸F-AH111585) (59–61) and ¹⁸F-RGD-K5 (43). Consistent with the polyvalency effect caused by dimerization and multimerization of cyclic RGD peptides, series of RGD radiotracers were found to show improved radiosynthesis yields, relatively higher tumor integrin-specific accumulation and favorable in vivo pharmacokinetic properties compared with monomeric ones in both preclinical and clinical studies (44, 45, 59, 62–68). However, the multistep, time-consuming and low-yield synthetic procedures make an automated production process, which is mandatory for routine clinical use, extremely difficult, and thus, the commercial clinical viability of this tracer is limited. Chen et al. successfully established a simplified labeling procedure for ¹⁸F-fluoride-aluminum complex-labeled RGD tracer ¹⁸F-AlF-NOTA-PRGD2 (denoted as ¹⁸F-Alfatide) and ¹⁸F-AlF-NOTA-E[PEG4-c(RGDfk)]2 (denoted as ¹⁸F-Alfatide II) in one single step of radiosynthesis (46, 54, 69). Upon successful introduction of a simple, one-step, lyophilized kit for ¹⁸F-alfatide preparation, with which complete radiosynthesis including purification can be accomplished within 30 min with a radiochemical purity of more than 95%, the potential for clinical transformation of this radiotracer was greatly improved. The pharmacokinetics and imaging properties of ¹⁸F-Alfatide were comparable to those of ¹⁸F-FP-PEG3-E[c(RGDyK)]2 (denoted as ¹⁸F-FPPRGD2) (54, 70). In addition, the positive effect of cyclic mono or multimeric peptides was also successfully developed into ⁶⁸Ga-labeled RGD analogs, which have shown promise for imaging integrin αvβ3 expression (48, 71–79).

Unlike the phenomenon of decreasing standard uptake values (SUVs) in normal organs over time, tumor retention of RGD-based PET tracers did not stop increasing until a plateau period was reached, which showed no obvious fluctuation within nearly 40–60 min after tracer injection. It appeared as either a distinct increase in primary tumors and lung, pleura, bone, and lymph node metastases compared with surrounding normal tissue or as moderate or reduced accumulation in the cases of kidneys, bladder, liver, spleen, and intestine. Sixty minutes after injection was recommended as the appropriate imaging time for optimal lesion-to-background contrast, according to the data from RGD-based fluorine-labeled monomeric or dimeric radiotracer studies (44, 45, 57, 59, 60). For biodistribution in normal tissues, RGD tracers can be used to image brain tumors or brain metastases with a clear background, due to their inability to pass through the healthy blood–brain barrier (BBB) but ability to cross the damaged BBB created by the tumor (43, 46, 61). Primary high uptakes were shown in organs such as the kidneys, bladder, liver, spleen and intestine organs, along with diffuse uptake in the bilateral ventricles and thyroid. High uptake in the kidneys and bladder resulted predominantly from urinary excretion. The uptake values of liver and spleen were thought to be physiological, high liver background may result from the densely vascularized nature of the organ and metabolism of RGD-based imaging tracers by the liver (60). Prominent accumulation of RGD peptide ligands in the liver reported in mice and humans also supported this assertion (80, 81). The uptake values in the intestine maybe associated with physiological expression of αvβ3 on intestinal smooth muscle cells, because the uptake of these tracers did not change significantly during an observation period of 90 min (57, 58, 82). While an initial ¹⁸F-FPPRGD2 increase followed by a pattern of decreasing uptake in the intestine was also observed, this may reflect initial dimeric RGD accumulation was cleared from hepatobiliary excretion (45, 83). Overall, this finding indicates the unsuitability of RGD imaging for hepatocellular carcinoma (HCC) tumors or diseases with liver metastases.

In addition to integrin αvβ3 expression, glucose metabolism is also believed to correlate with tumor aggressiveness and progression (84, 85). In tumor lesions, RGD-based tracer uptake is significantly lower than that of FDG, as commonly observed in studies comparing FDG and RGD tracers (52, 63, 65, 66, 86–92), but this difference was altered in some cases, such as with the higher maximum SUV (SUV_max) and tumor-to-blood ratio (TBR) of ⁶⁸Ga-NOTA-3P-TATE-RGD compared with ¹⁸F-FDG in neuroendocrine tumor (NET) cases and also in both human epidermal growth factor receptor 2 negative [HER2 (–)] and estrogen receptor positive [ER (+)] breast cancer (52, 88). One possible reason responsible for this difference may be related to the different binding mechanisms of the tracers. ¹⁸F-FDG mainly depends on the hypermetabolism of glucose in malignant tumor cells, whereas RGD PET imaging predominantly binds to avβ3 highly expressed in tumor neovascular endothelial cells. Also, within each lesion, the number of endothelial cells expressing integrin αvβ3 is smaller than the abundant number of tumor cells containing glucose transporters (93). In addition, both tracers showed strong uptake in inflammatory lesions similar to malignancies. RGD was thought to be more beneficial than FDG for differentiating inflammation, as it demonstrated significantly lower retention than FDG after comparable high uptake levels in inflammation (56, 94–96). With regard to the correlation between RGD and FDG data, inconsistent results were observed when tumor lesions overall were considered. Moderate or significant correlations between RGD and FDG parameters were reported in locally advanced rectal cancer (LARC) and squamous cell carcinoma of head and neck (HNSCC) (87, 88, 90, 97), and for subgroups of tumors, namely, non-small cell lung cancer (NSCLC), triple-negative breast cancer (TNBC), and FDG-avid tumors (86, 89). However, other studies failed to demonstrate a correlation between RGD and FDG uptake (65, 91, 98–100).

These results overall may indicate the coexistence of interrelated pathophysiological phenomena within the tumor, such as cell proliferation and neoangiogenesis, but the lack of a significant correlation between RGD and FDG uptake suggests that these imaging modalities provide complementary rather than similar information (101). RGD imaging offers indispensable value in terms of evaluating tumor angiogenesis and planning αvβ3-based therapy. As the next logical step, many research groups focused on exploring the clinical characteristics of RGD-based imaging agents.

In a pan-cancer study with 9 patients (5 with malignant melanomas, 2 with sarcomas, 1 with osseous metastasis from renal cell carcinoma [RCC], and 1 with villonodular synovitis), ¹⁸F-Galacto-RGD SUVs showed inter- and intra-individual variances ranging from 1.2 to 10.0 (56). Consistently, Beer et al. reported the heterogeneous accumulation of ¹⁸F-Galacto-RGD from 1.2 to 9.0 in 19 patients. Distribution volume values for tumor tissue (1.5 ± 0.8) were 4 times higher than those for muscle tissue (0.4 ± 0.1), suggesting intense specific receptor binding in tumor tissues and only minimal free and bound (specific or nonspecific) tracer in muscle tissue (57). Similarly, studies of other imaging agents such as ¹⁸F-AH111585 and ⁶⁸Ga-NODAGA-E[(cRGDyK)₂] (denoted as ⁶⁸Ga-NODAGA-RGD2) also reported heterogeneous tracer uptake in tumor and metastatic sites and superior tumor-to-background ratios in patients with malignant disease (58, 61, 76, 86, 102). However, the sensitivity of ¹⁸F-Galacto-RGD (76%) for lesion identification was lower than that of conventional staging with contrast-enhanced CT and ¹⁸F-FDG PET in a comparative study (86). This is not surprising because, unlike the glycolytic effect of entire tumor cell, αvβ3 is expressed at elevated levels on angiogenic blood vessels and on malignant tumors (103, 104). Interestingly, the related research demonstrated high inter- and intraindividual variances in tracer accumulation in different types of lesions, indicating great diversity in receptor expression.

According to the literature, the expression patterns of αvβ3 are specific on malignant carcinomas and benign tumors or inflammatory tissues (95, 105, 106). In benign lesions (e.g., pigmented villonodular synovitis, neurofibroma, and inflammatory tissue), αvβ3 expression is located only on the vasculature. However, in malignant lesions, αvβ3 is expressed not only on the endothelial cells of neovasculature but also on the surface of some tumor cells. In HNSCC, αvβ3 is located predominantly on the neovasculature but shows some low expression on tumor cells (56, 102, 107). Immunohistochemical analyses confirmed αvβ3 expression predominantly on microvessels (5/5) and, to a lesser extent, on tumor cells (3/5) in invasive ductal breast cancer (IDC) (28, 108). In malignant melanoma and clear cell RCC (ccRCC), αvβ3 is highly located on the surface of tumor cells (61, 102, 109, 110). In the other solid tumors, namely, glioma, lung cancer, differentiated thyroid cancer, and breast cancer (56, 102, 108–112), αvβ3 expression is located on the neovasculature and on the tumor cells at varying degrees. For example, a significant correlation between αvβ3 expression and radiotracer uptake was reported by a study employing ¹⁸F-Galacto-RGD PET in melanoma and sarcoma patients (56). Specially, in ccRCC cases, the ¹⁸F-FB-mini-PEG-E[c(RGDyK)]2 (denoted as 18F-FPRGD2) PET signal correlated with integrin αvβ3 expression on tumor cells, and immunohistochemistry results confirmed high expression of αvβ3 on ccRCC cells. In contrast, in papillary RCC (pRCC) cases, the ¹⁸F-FPRGD2 PET signal correlated with the integrin αvβ3 level on tumor vessels (67). These results demonstrate that RGD-based tracer uptake can reflect integrin αvβ3 expression in renal tumors, but represents angiogenesis only when tumor cells do not express integrin αvβ3. In other words, RGD-based PET may be regarded as a surrogate marker of angiogenesis for the visualization and noninvasive quantitation of integrin expression when αvβ3 expression is predominantly confined to the tumor vasculature as in HNSCC (107). Overall, the association between RGD SUVs and TBR and the corresponding αvβ3 expression level demonstrated the value of noninvasive techniques for appropriately diagnosing tumor lesions, although a positive correlation was not always clear (56, 61, 107, 113–116). The heterogeneous distribution pattern among different lesions described above may explain this phenomenon. Next, we provide a review of reported clinical investigations related to the application of RGD-based tracers for diagnosis, definition of tumor subvolume, and therapeutic response prediction according to specific tumor types ( Table 1 ).

For radiation treatment planning, imaging quality is vitally important for biological target volume definition. Beer et al. fused images from ¹⁸F-Galacto-RGD PET and MRI and/or multislice CT (MSCT) scanning in HNSCC patients. The mean gross tumor volume (GTV) was 26.4 ± 20.2 cm³, and when a SUV threshold >3.0 was applied, the mean tumor subvolume was only 3.4 ± 3.4 cm³, accounting for 13.1 ± 11.7% of the GTV (107). These results confirm the imaging quality and heterogeneous subvolume achieved with RGD-containing peptide tracers, which allows for adequate manual fusion with MRI or MSCT using anatomic landmarks. As displayed in a HNSCC patient population by Durante et al., ⁶⁸Ga-NODAGA-RGD and ¹⁸F-FDG PET/CT individually led to inconsistent tumor subvolume delineation. The tracer avid tumor volume (TATV) calculated from RGD imaging tended to be larger than the metabolic tumor volume (MTV) calculated from ¹⁸F-FDG imaging when a 42% SUV_max fixed threshold similarly to MTV delineation was used (p = 0.085) (87). That difference in tumor delineation is assumed to be attributable to the difference in tracer targeting between FDG and RGD, as mostly homogeneous FDG uptake is significantly correlated with glucose metabolism and heterogeneous RGD uptake is associated with αvβ3 on angiogenesis. In addition, the physical properties of radionuclides fluorine-18 and gallium-68 also should be considered. At the same time, the 42% SUV_max of RGD was much lower than that of ¹⁸F-FDG, which may also have contributed to the calculation of a larger volume for the same tumor. In LARC cases examined with ¹⁸F-FPRGD2 imaging, the baseline integrin tumor volume (ITV) 70% (ITV_70%, delineated using a threshold of 70% of the SUV_max) was moderately correlated with the MTV_40% (delineated using a threshold of 40% of the SUV_max) and total lesion glycolysis (calculated as SUV_mean × MTV_40%) (66).

However, such results have not been reproduced in Lewis lung carcinoma (LLC) tumor-bearing C57BL/6 mice. The GTV determined from ¹⁸F-Alfatide PET was smaller and much closer to the pathological volume than that from ¹⁸F-FDG PET although a similar 40% of the SUV_max threshold was used (146). Several points are worth discussing regarding this difference. First, the expression pattern of αvβ3 varies among cancer types; it is mainly expressed on the neovasculature of HNSCC but on both the neovasculature and tumor cells in lung cancer (107, 111, 147). Secondly, different radionuclides were used in the two studies with ¹⁸F-Alfatide and ⁶⁸Ga-NODAGA-RGD. Additionally, differences in metabolism between animals and humans may also influence the results. Importantly, it is unscientific to define the cutoff value for tumor subvolume delineation in reference to the established threshold for FDG. Instead, threshold optimization for estimating tumor volume based on a spatial comparison of αvβ3 expression on whole-tumor histological slices needs to be performed.

Because RGD is a marker of αvβ3 expression on angiogenesis which is of paramount importance for tumor oxygenation, RGD-containing PET tracers have been considered as a potentially valuable tool for monitoring treatment response and identifying patients less likely to respond to therapy. Several clinical pilot studies have explored their value in assessing treatment efficacy and adverse events, with encouraging results for both antiangiogenic therapy and chemoradiotherapy. Knowledge of the precision of repeated imaging parameters is a prerequisite for serial response measurements and, therefore, response to treatment. In the study by Sharma et al., solid tumor patients underwent two ¹⁸F-AH111585 imaging sessions within an interval of 10 days before commencement of therapy. No significant change in the SUV_peak was observed between the two scans, and the 95% reference range of spontaneous fluctuations was 35–39% (148). Measurements of ¹⁸F-FPPRGD2 uptake in normal organs also showed no significant changes between pre-, 1-week post- and 6-week post-therapy scans (149). Thus, with their protocol, the serial reproducibility of RGD-based tracer uptake supports further investigation of these novel tracers as biomarkers of treatment response ( Table 2 ).

Significant differences in RGD-based tracer uptake before and after antiangiogenic therapy have been reported in several studies (90, 97, 149). In GBM patients receiving bevacizumab-containing therapy, decreases of less than 15% in the SUV_max and angiogenic volumes from ¹⁸F-FPPRGD2 imaging at 1 week predicted recurrence and poor prognosis, while decreases of more than 50% predicted good outcomes to treatment (149). In another study including ovarian and cervical cancer, patients who experienced clinical disease progression, complete response, and partial response showed decreases in the lesional ¹⁸F-FPPRGD2 SUV_mean of 1.6, 25.2, and 7.9%, respectively, with similar decreases in the ¹⁸F-FDG SUV_mean of 9.4, 71.8, and 76.4%, respectively (90). The above studies mainly evaluated the changes in tracer uptake with the application of different antiangiogenic combination therapies, which may confound our understanding of RGD-based imaging for monitoring the response. Li et al. successfully investigated the relationships of ¹⁸F-Alfatide with the response to single antiangiogenic therapy in solid malignancies, higher pretreatment SUV_peak and SUV_mean were seen in responding tumors compared with non-responding tumors (4.98 ± 2.34 vs. 3.59 ± 1.44, p = 0.048; 3.71 ± 1.15 vs. 2.95 ± 0.49, p = 0.036). Cases with an ¹⁸F-Alfatide SUV_mean above 3.82 showed longer progression-free survival (PFS) than those with a SUV_mean less than 3.82. These results provided clinical evidence of the effectiveness of RGD-based PET at baseline for predicting the response to antiangiogenic therapy, suggesting this approach could potentially replace indicators based on change ratios in most previous studies (150). In addition, they also reported the predictive value of RGD-based imaging for antiangiogenic adverse events for the first time. Patients with low maximum ¹⁸F-Alfatide SUVs in the liver, spleen, and cardiac tissue were more likely to experience fatigue, hypertension, and nausea during Apatinib treatment (151). Thus, higher ¹⁸F-Alfatide uptake in specific normal organs predicted less adverse events, whereas higher uptake in tumor lesions predicted a better response to therapy. Higher accumulation in normal organs may indicate more regular, functional vasculature and sufficient blood flow, and higher accumulation within tumors may indicate greater expression of αvβ3 integrins (155, 156).

Results suggesting RGD-based tracer uptake at baseline can predict the response to antiangiogenic therapy have also been obtained in patients with platinum-resistant/refractory ovarian cancer (152). Of interest, a negative trend was observed between the baseline SUV_60,mean and PFS. There are several possible explanations for this contradictory observation. First, unlike the study by Li et al., the patients in this study received antiangiogenic combination therapy. It is well established that the addition of chemotherapy also enables a reduction in tumor size, which may account for the lack of association. Secondly, the PET response was assessed after 1 week, while the RECIST 1.1 response was evaluated after 3 months. Finally, preclinical work suggests that normalization of vessels occurs within hours and lasts 7–10 days, it is possible that performing the second PET scan at 7 days may have missed the neovascularization window (157). Overall, further research is needed to explore the potential of RGD-based tracers as biomarkers for response to antiangiogenic and antiangiogenic combination therapies.

The predictive value of RGD-based imaging for the response to chemoradiotherapy was also studied. Twenty-five newly diagnosed GBM patients who were ready for concurrent chemoradiotherapy (CCRT) 3–5 weeks after surgical resection were prospectively enrolled in a study by Zhang et al. The baseline ¹⁸F-Alfatide SUV_max, the 3-week SUV_max, and the TNR were found to be predictive of sensitivity to CCRT. The 3-week SUV_max (AUC = 0.846) was shown to be the best predictive parameter, with a 3-week SUV_max higher than 1.35 predicting worse efficacy (153). Further, Luan et al. explored the similar potential value of ¹⁸F-Alfatide PET/CT in locally advanced NSCLC. From their analysis, the pretreatment SUV_max, SUV_peak, TNR, TBR, and TMR were all significant parameters, with higher SUVs in non-responders than responders, and the TNR was shown to be an independent predictor (nonresponders vs. responders, 8.31 ± 0.61 vs. 6.53 ± 0.78, p <0.001) to CCRT (154). Chen et al. prospectively investigated the potential usefulness of RGD-K5 PET in HNSCC, patients whose primary tumors had lower accumulation of both tracers at 3 months achieved response to CCRT successfully (99). For LARC patients treated with CCRT, baseline ¹⁸F-FPRGD2 SUV_max and SUV_mean were negatively correlated with the tumor regression grade (TRG). The baseline SUV_max of both RGD and FDG imaging were significantly higher in TRG 0 cases than in the other patients. A cut-off value of 5.6 for RGD SUV_max identified TRG 0 status with 100% sensitivity and 66.7% specificity (AUC = 0.84) (66)..

From the research described above, it is interesting to note that RGD-based tracer uptake follows different trends according to differing treatment modalities. For antiangiogenic therapy, the higher uptake values may indicate a high density of effective target receptors and a better response to apatinib therapy. For CCRT, higher uptake values may represent high malignancy and severe hypoxia, which could contribute to increased chemoradiation resistance in tumors (158–160).

A variety of radiolabeled, RGD-based PET imaging agents have been developed and extensively investigated in terms of their value for tumor detection and diagnosis, patient stratification, and monitoring of treatment response. According to promising results, these PET imaging peptides were transferred successfully from preclinical animal studies to clinical human trials. Different RGD-based PET imaging can be used in patients with glioma, lung cancer, esophageal cancer, breast cancer, renal carcinoma and other tumors to differentiate benign and malignant lesions and also pathological types and stages, and also can further distinguish molecular subtypes of breast cancer, delineate tumor volume, and predict the efficacy of radiochemotherapy and antiangiogenic therapy. The results regarding their diagnostic and predictive value are mostly derived from preliminary studies, and continued research is necessary to guarantee their development for clinical application.

In addition, RGD-based imaging was useful as a noninvasive surrogate parameter of cardiovascular disease. Normal angiogenesis under pathological conditions in cardiovascular diseases such as carotid atherosclerosis, myocardial infarction or reperfusion were also characterized by the overexpression of αvβ3 (3, 4, 161). In atherosclerotic lesions, highly expressed αvβ3 were found in both macrophages and activated endothelial cells (105, 162). The uptakes of ⁶⁸Ga-DOTA-RGD and ¹⁸F-GalactoRGD PET increased in the atherosclerotic plaques in mice (95, 163). Clinically, RGD uptakes were heterogeneously distributed and significantly correlated with avβ3 integrin staining score in human atherosclerotic carotid plaques (164, 165). For ischemia tissue repair after myocardial infarction, αvβ3 is one of the necessary integrins involved in the formation of new capillaries (25, 166). RGD-based imaging of myocardial infarction observed consistent findings from studies of rats, specific RGD tracer accumulation uptake increased in vivo corresponding to infarcted and peri-infarct myocardial regions, and decreased after reperfusion (167–170). Tracer uptakes were expected to be used to monitor the process of myocardial repair (171–173). Makowski et al. confirmed the feasibility of ¹⁸F-Galacto-RGD to monitor vascular remodeling during evolving MI in human studies; tracer uptake was significantly correlated with infarct size and impairment of myocardial blood flow (174). Thus, RGD-based imaging of αvβ3 integrin expression could represent a novel promising imaging approach in determining the angiogenesis of carotid atherosclerosis and myocardial repair, which is potentially affecting the plaque vulnerability and left ventricular remodeling.

The RGD peptides also served as the new target for tumor treatment. RGD peptides could significantly reduce the functional vascular density, inhibit the adhesion and induce apoptosis of tumor cells according to inhibiting the interactions between integrins and their ligands (175–177). The RGD modified gene carriers and target drugs also showed special interest in delivery (178, 179). RGD peptides for imaging or tumor treatment all need to be further investigations for clinical translation.

It is important to note that contradictory reports about the correlation between RGD tracer uptake and αvβ3 expression level exist and that the ultimate meaning of αvβ3 expression in the context of angiogenesis is very complex and requires further characterization. The highly abnormal and dysfunctional vasculature created tumor hypoxic environment and impaired the ability of immune effector cells to penetrate solid tumors. In turn, stimulation of immune cell could also help normalize tumor vessels (180). Other studies have demonstrated the expression of integrin αvβ3 on activated macrophages by different methods (162, 181, 182). Therefore, RGD-based imaging targeting tumor neovascularization may offer a strategy to monitor the tumor microenvironment status and therapeutic response during immune checkpoint blockade or combination treatment. Moreover, the development of the clinically available applications of RGD-based tracers has mainly focus on radionuclide-labeled tracers, approaches to develop molecularly targeted nanoparticles for molecular imaging should be explored. Additionally, the simultaneous acquisition of functional parameters by MRI and PET may help compensate for the shortcomings of these different approaches. A thorough and complete assessment of different aspects of angiogenesis observed via multimodal imaging and biomomics characteristics should be implemented as part of the concept of personalized functional medicine.

SY proposed the conception and edited the manuscript. LL participated and drafted the manuscript. XC and JY participated and edited the manuscript. All authors listed have made a substantial, direct, and intellectual contribution to the work and approved it for publication.

This study was supported in part by the National Natural Science Foundation of China (grant nos. NSFC81872475 and NSFC82073345) and the Jinan Clinical Medicine Science and Technology Innovation Plan (202019060) to SY, the Academic Promotion Program of Shandong First Medical University (grant No. 2019ZL002) to JY, the National University of Singapore Start-up Grant (NUHSRO/2020/133/Startup/08), the NUS School of Medicine Nanomedicine Translational Research Programme (NUHSRO/2021/034/TRP/09/Nanomedicine), and the NUS School of Medicine Kickstart Initiative (NUHSRO/2021/044/Kickstart/09/LOA) to XC.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.