Search

Nanobodies, or single-domain antibody fragments, are promising candidates for molecular imaging due to their small size, rapid tissue penetration, and high target specificity. However, a significant challenge in their use is high renal uptake and retention, which can limit the therapeutic efficacy and complicate image interpretation. This study compares five different fluorine-18-labeled prosthetic groups for nanobodies, aiming to optimize pharmacokinetics and minimize kidney retention while maintaining tumor targeting. Using an epidermal growth factor receptor (EGFR) targeting nanobody as a model, two labeling approaches were evaluated; direct labeling of RESCA (with and without polyethylene glycol (PEG))-conjugated nanobody using Al[ 18 F]F and indirect labeling using ([ 18 F]F-fluoropyridine ([ 18 F]F-FPy)-based prosthetic groups (site-specific and nonsite-specific). Labeled nanobodies were characterized in vitro for binding affinity and cell uptake with in vivo behavior assessed in EGFR + A431 tumor-bearing mice using PET imaging and biodistribution studies. Labeling with Al[ 18 F]F showed high renal retention, which was partially mitigated by PEGylation. However, PEGylation also led to a decreased tumor uptake, particularly with longer PEG chains. Labeling using [ 18 F]F-FPy prosthetic groups exhibited the most favorable pharmacokinetics, with rapid renal clearance and minimal kidney retention while maintaining high tumor uptake. These constructs showed excellent tumor-to-background contrast as early as 1 h postinjection. The study confirms that the selection of the prosthetic group significantly impacts the in vivo behavior of nanobodies, particularly regarding kidney accumulation. [ 18 F]F-FPy-based prosthetic groups show the most promising results, with high tumor and minimal kidney uptake. Robust production of [ 18 F]F-FPy on Sep-Pak is adaptable to clinical translation. Moreover, the potential substitution of 18 F with therapeutic radioisotopes such as 131 I or 211 At could expand the application of these nanobodies from diagnostics to targeted radionuclide therapy while maintaining a low kidney exposure. These findings have important implications for optimizing nanobody-based radiopharmaceuticals for molecular imaging and targeted radionuclide therapy.

Nanotheranostics based on tumor-selective small molecular prodrugs could be more advantageous in clinical translation for cancer treatment, given its defined chemical structure, high drug loading efficiency, controlled drug release, and reduced side effects. To this end, we have designed and synthesized a reactive oxygen species (ROS)-activatable heterodimeric prodrug, namely, HRC, and nanoformulated it for tumor-selective imaging and synergistic chemo- and photodynamic therapy. The prodrug consists of the chemodrug camptothecin (CPT), the photosensitizer 2-(1-hexyloxyethyl)-2-devinyl pyropheophorbide-a (HPPH), and a thioketal linker. Compared to CPT- or HPPH-loaded polymeric nanoparticles (NPs), HRC-loaded NPs possess higher drug loading capacity, better colloidal stability, and less premature drug leakage. Interestingly, HRC NPs were almost nonfluorescent due to the strong π-π stacking and could be effectively activated by endogenous ROS once entering cells. Thanks to the higher ROS levels in cancer cells than normal cells, HRC NPs could selectively light up the cancer cells and exhibit much more potent cytotoxicity to cancer cells. Moreover, HRC NPs demonstrated highly effective tumor accumulation and synergistic tumor inhibition with reduced side effects on mice.

Biocatalytic reactions in living cells involve complex transformations in the spatially confined microenvironments. Inspired by biological transformation processes, we demonstrate effective biocatalytic cascade driven photodynamic therapy in tumor-bearing mice by the integration of an artificial enzyme (ultrasmall Au nanoparticles) with upconversion nanoparticles (NaYF 4 @NaYb 0.92 F 4 :Er 0.08 @NaYF 4 )zirconium/iron porphyrin metal-organic framework core-shell nanoparticles (UMOF NPs) which act as biocatalysts and nanoreactors. The construction of core-shell UMOF NPs are realized by using a unique "solvent-assisted self-assembly" method. The integration of ultrasmall AuNPs on the UMOFs matrix leads to glucose depletion, providing Au-mediated cancer therapy via glucose oxidase like catalytic activity. Meanwhile, the UMOF matrix acts as a near-infrared (NIR) light photon-activated singlet oxygen generator through a continuous supply of oxygen via hydrogen peroxide decomposition upon irradiation. Such kinds of biocatalysts offer exciting opportunities for biomedical, catalytical ,and energy applications.

In this study, we applied a new strategy to identify sentinel lymph node (SLN) metastasis by combining 68 Ga-NOTA-Evans Blue ( 68 Ga-NEB) for SLN mapping and 68 Ga-NOTA-RM26 for LN metastasis detection in breast cancer patients. A total of 24 female patients with breast cancer diagnosed by core biopsy or suspected by mammography or ultrasonography were recruited and provided informed consent. All patients underwent 68 Ga-NEB and 68 Ga-NOTA-RM26 PET/CT imaging. Visual analysis of 68 Ga-NEB PET/CT images was used to determine SLNs, and then compared with the 68 Ga-NOTA-RM26 results and histopathological findings. SLNs were visualized in 24 of 24 patients (100.0%) within 4.0-10.0 (5.6 ± 1.4) min. All patients were pathologically diagnosed with breast cancer, and 12 patients had ipsilateral lymph node metastasis. By combining 68 Ga-NEB and 68 Ga-NOTA-RM26 images, 7/12 (58.3%) patients showed mild to intense uptake of 68 Ga-NOTA-RM26 in SLNs, 1/12 patient (8.3%) had moderate uptake of 68 Ga-NOTA-RM26 in the non-SLNs rather than SLN, indicating possible bypass lymphatic drainage, partially accounting for the false negatives in SLN biopsy during surgery. No false positives were found. The SUV max of 68 Ga-NOTA-RM26 activity in metastatic SLNs was significantly higher than that in non-metastatic SLNs (2.2 ± 2.3 vs 0.7 ± 0.1, P = 0.047). This study manifests the value of combination of 68 Ga-NEB and 68 Ga-NOTA-RM26 dual tracer PET/CT in preoperative evaluation of SLN metastasis in breast cancer patients, especially in those patients with lymphatic obstruction and bypass drainage. In general, positive 68 Ga-NOTA-RM26 uptake in either SLN or other lymph nodes can apply lymph node dissection rather than intraoperative SLN biopsy.

Phototheranostics refers to advanced photonics-mediated theranostic methods for cancer and includes imaging-guided photothermal/chemotherapy, photothermal/photodynamic therapy, and photodynamic/chemotherapy, which are expected to provide a paradigm of modern precision medicine. In this regard, various phototheranostic drug delivery systems with excellent photonic performance, controlled drug delivery/release, and precise photoimaging guidance have been developed. In this study, we reported a special "in situ framework growth" method to synthesize novel phototheranostic hollow mesoporous nanoparticles by ingenious hybridization of perylene diimide (PDI) within the framework of small-sized hollow mesoporous organosilica (HMO). The marriage of PDI and HMO endowed the phototheranostic silica nanoparticles (HMPDINs) with largely amplified fluorescence and photoacoustic signals, which can be used for enhanced fluorescence and photoacoustic imaging. The organosilica shell can be chemically chelated with isotope 64 Cu for positron emission tomography imaging. Moreover, in situ polymer growth was introduced in the hollow structure of the HMPDINs to produce thermosensitive polymer (TP) in the cavity of HMPDINs to increase the loading capacity and prevent unexpected leakage of the hydrophobic drug SN38. Furthermore, the framework-hybridized PDI generated heat under near-infrared laser irradiation to trigger the deformation of TP for controlled drug release in the tumor region. The fabricated hybrid nanomedicine with organic-inorganic characteristic not only increases the cancer theranostic efficacy but also offers an attractive solution for designing powerful theranostic platforms.

A major concern about glucose oxidase (GOx)-mediated cancer starvation therapy is its ability to induce serious oxidative damage to normal tissues through the massive production of H 2 O 2 byproducts in the oxygen-involved glucose decomposition reaction, which may be addressed by using a H 2 O 2 scavenger, known as an antioxidation agent. Surprisingly, H 2 O 2 removal accelerates the aerobic glycometabolism of tumors by activating the H 2 O 2 -dependent "redox signaling" pathway of cancer cells. Simultaneous oxygen depletion further aggravates tumor hypoxia to increase the toxicity of a bioreductive prodrug, such as tirapazamine (TPZ), thereby improving the effectiveness of cancer starvation therapy and bioreductive chemotherapy. Herein, a "nitrogen-protected silica template" method is proposed to design a nanoantioxidant called an organosilica-based hollow mesoporous bilirubin nanoparticle (HMBRN), which can act as an excellent nanocarrier to codeliver GOx and TPZ. In addition to efficient removal of H 2 O 2 for self-protection of normal tissues via antioxidation, GOx/TPZ-coloaded HMBRN can also rapidly deplete intratumoral glucose/oxygen to promote a synergistic starvation-enhanced bioreductive chemotherapeutic effect for the substantial suppression of solid tumor growth. Distinct from the simple combination of two treatments, this study introduces antioxidation-activated self-protection nanotechnology for the significant improvement of tumor-specific deoxygenation-driven synergistic treatment efficacy without additional external energy input, thus realizing the renaissance of precise endogenous cancer therapy with negligible side effects.

Glutathione (GSH), one of the most significant reducing species in vivo, plays important roles in a variety of diseases and cellular functions. Precise quantification of GSH via advanced noninvasive photoacoustic imaging (PAI) is of vital significance for the early diagnosis and prompt treatment of GSH-related deep-seated diseases, which stresses the need for custom-design of GSH-sensitive PAI probes with changeable near-infrared spectroscopy (NIR) absorption. In this work, a novel intelligent tumor microenvironment-activated ratiometric PAI nanoprobe is first developed with the intention of specific ultrasensitive detection of intratumoral GSH by overcoming the limitations of previously reported fluorescent or PA imaging sensors. This special ratiometric PAI nanoprobe (CR-POM) is synthesized through the self-assembly of croconaine (CR) dye and molybdenum-based polyoxometalate (POM) clusters with opposite NIR absorbance change in response to GSH. The resulting amplified ratiometric absorbance (Ab 866 /Ab 700 ), the relatively low limit of detection value (0.51 mM), and the unique acidity-activated self-aggregation contribute to the prolonged intratumoral retention and enhanced tumor accumulation of CR-POM for accurate quantification of intratumoral GSH (0.5-10 mM). Featuring the additional merit of 64 Cu radiolabeling for whole-body positron-emission tomography imaging, the smartly designed CR-POM nanoprobe will open new horizons for real-time noninvasive monitoring of biodistribution and simultaneous accurate quantification of GSH levels, especially in tumor and other GSH-related pathophysiological processes.

As highly expressed in insulinomas, the glucagon-like peptide-1 receptor (GLP-1R) is believed to be an attractive target for diagnosis, localization, and treatment with radiolabeled exendin 4. However, the high and persistent radioactivity accumulation of exendin 4 in the kidneys limits accurate diagnosis and safe, as well as effective, radiotherapy in insulinomas. In this study, we intend to reduce the renal accumulation of radiolabeled exendin 4 through degradation mediated by brush border membrane enzymes. A new exendin 4 ligand NOTA-MVK-Cys 40 -Leu 14 -Exendin 4 containing Met-Val-Lys (MVK) linker between the peptide and 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) chelator was synthesized and labeled with 68 Ga. The in vitro mouse serum stability and cell binding affinity of the tracer were evaluated. Initial in vitro cleavage of the linker was determined by incubation of a model compound Boc-MVK-Dde with brush border membrane vesicles (BBMVs) with and without the inhibitor of neutral endopeptidase (NEP). Further cleavage studies were performed with the full structure of NOTA-MVK-Cys 40 -Leu 14 -Exendin 4. Kidney and urine samples were collected in the in vivo metabolism study after intravenous injection of 68 Ga-NOTA-MVK-Cys 40 -Leu 14 -Exendin 4. The microPET images were acquired in INS-1 tumor model at different time points; the radioactivity uptake of 68 Ga-NOTA-MVK-Cys 40 -Leu 14 -Exendin 4 in tumor and kidneys were determined and compared with the control radiotracer without MVK linker. 68 Ga-NOTA-MVK-Cys 40 -Leu 14 -Exendin 4 was stable in mouse serum. The MVK modification did not affect the affinity of NOTA-MVK-Cys 40 -Leu 14 -Exendin 4 toward GLP-1R. The in vitro cleavage study and in vivo metabolism study confirmed that the MVK sequence can be recognized by BBM enzymes and cleaved at the amide bond between Met and Val, thus releasing the small fragment containing Met. MicroPET images showed that the tumor uptake of 68 Ga-NOTA-MVK-Cys 40 -Leu 14 -Exendin 4 was comparable to that of the control, while the kidney uptake was significantly reduced. As a result, more favorable tumor to kidney ratios were achieved. In this study, a novel exendin 4 analogue, NOTA-MVK-Cys 40 -Leu 14 -Exendin 4, was successfully synthesized and labeled with 68 Ga. With the cleavable MVK sequence, this ligand could be cleaved by the enzymes on kidneys, and releasing the fragment of 68 Ga-NOTA-Met-OH, which will rapidly excrete from urine. As the high and consistent renal radioactivity accumulation could be significantly reduced, NOTA-MVK-Cys 40 -Leu 14 -Exendin 4 shows great potential in the diagnosis and radiotherapy for insulinoma.

The effectiveness of numerous molecular drugs is hampered by their poor pharmacokinetics. Different from previous approaches with limited effectiveness, most recently, emerging high-affinity albumin binding moieties (ABMs) for in vivo hitchhiking of endogenous albumin opens up an avenue to chaperone small molecules for long-acting therapeutics. Although several FDA-approved fatty acids have shown prolonged residence and therapeutic effect, an easily synthesized, water-soluble, and high-efficiency ABM with versatile drug loading ability is urgently needed to improve the therapeutic efficacy of short-lived constructs. We herein identified an ideal bivalent Evans blue derivative, denoted as N(tEB) 2 , as a smart ABM-delivery platform to chaperone short-lived molecules, through both computational modeling screening and efficient synthetic schemes. The optimal N(tEB) 2 could reversibly link two molecules of albumin through its two binding heads with a preferable spacer, resulting in significantly extended circulation half-life of a preloaded cargo and water-soluble. Notably, this in situ dimerization of albumin was able to sandwich peptide therapeutics to protect them from proteolysis. As an application, we conjugated N(tEB) 2 with exendin-4 for long-acting glucose control in a diabetic mouse model, and it was superior to both previously tested NtEB-exendin-4 (Abextide) and the newly FDA-approved semaglutide, which has been arguably the best commercial weekly formula so far. Hence, this novel albumin binder has excellent clinical potential for next-generation biomimetic drug delivery systems.

Radioligand therapy (RLT) relies on the use of pharmacophores to selectively deliver ionization energy to cancers to exert its tumoricidal effects. Cancer cells that are not directly targeted by a radioconjugate remain susceptible to RLT because of the crossfire effect. This is significant given the inter- and intra-heterogeneity of tumors. In recent years, reversible albumin binders have been used as simple "add-ons" for radiopharmaceuticals to modify pharmacokinetics and to enhance therapeutic efficacy. In this Review, we discuss recent advances in albumin binder platforms used in RLT, with an emphasis on 4-( p-iodophenyl)butyric acid and Evans blue derivatives. We focus on four biological systems pertinent to oncology that utilize this class of compounds: folate receptor, integrin αvβ3, somatostatin receptor, and prostate-specific membrane antigen. Finally, we offer our perspectives on albumin binders for RLT, highlighting future areas of research that will help propel the technology further for clinical use.

Featured with a large surface area, uniform interpenetrating mesopores, diverse organic framework hybridization, and well-defined surface properties, the hollow mesoporous organosilica nanoparticle (HMON) represents a promising paradigm in drug delivery systems with excellent biocompatibility. However, effective tumor accumulation and precise cancer theranostics of the HMON still remain a challenge. In this study, an "ammonia-assisted hot water etching" method is applied for the successful construction of sub-50 nm thioether/phenylene dual-hybridized HMON with low hemolytic effect. Particularly, the surface modification with Mo(VI)-based polyoxometalate (POM) clusters drives the self-assembly of HMON in the mild acidic tumor microenvironment (TME) to achieve enhanced tumor retention and accumulation. More importantly, the reducibility-activated Mo(VI)-to-Mo(V) conversion within POM not only endows the POM-anchored HMON with outstanding TME-responsive photoacoustic (PA) imaging contrast and photothermal therapy (PTT) performance but also plays an indispensable role in controllably triggering the decomposition of the Mn 2 (CO) 10 payload for CO release, which gives rise to remarkable synergistic PTT-enhanced CO gas therapy for complete tumor eradication. By harnessing the unique acidic and redox properties of TME, the judiciously designed smart POM-anchored HMON nanoplatform is expected to act as a "magic bomb" to selectively destroy cancer without damaging normal tissues. This nanoplatform holds significant potential in realizing TME-responsive self-assembly for enhanced tumor accumulation and precise tumor-specific synergistic therapy, which is very promising for clinical translation.

Prostate cancer is the most frequently diagnosed malignant tumor in men worldwide. Prostate-specific membrane antigen (PSMA) is a surface molecule specifically expressed by prostate tumors that has been shown to be a valid target for internal radionuclide therapy in both preclinical and clinical settings. The most common radiotherapeutic agent is the small molecule 177 Lu-PSMA-617, which is under clinical evaluation in multiple countries. Nevertheless, its efficacy in causing tumor regression is still suboptimal, even when administered in several cycles per patient, perhaps due to poor pharmacokinetics (PK), which limits uptake by the tumor cells. We postulated that the addition of the Evans blue (EB) moiety to PSMA-617 would improve the PK by extending circulation half-life, which would increase tumor uptake and improve radiotherapeutic efficacy. PSMA-617 was modified by conjugation of a 2-thiol acetate group onto the primary amine and thereafter reacted with a maleimide functional group of an EB derivative, to give EB-PSMA-617. The PK and radiotherapeutic efficacy of 90 Y- or 177 Lu-EB-PSMA-617 was compared to the clinically used radiopharmaceutical 90 Y- or 177 Lu- PSMA-617 in PC3-PIP tumor-bearing mice. EB-PSMA-617 retained binding to serum albumin as well as a high internalization rate by tumor cells. Upon injection, metal-labeled EB-PSMA-617 demonstrated an extended blood half-life compared to PSMA-617 and, thereby, prolonged the time window for binding to PSMA. The improved PK of EB-PSMA-617 resulted in significantly higher accumulation in PSMA + tumors and highly effective radiotherapeutic efficacy. Remarkably, a single dose of 1.85 MBq of 90 Y- or 177 Lu-EB-PSMA-617 was sufficient to eradicate established PMSA + tumors in mice. No significant body weight loss was observed, suggesting little to no gross toxicity. The construct described here, EB-PSMA-617, may improve the radiotherapeutic efficacy for patients with PSMA-positive tumors by reducing both the amount of activity needed for therapy as well as the frequency of administration, as compared to PSMA-617.

Several radioligands targeting prostate-specific membrane antigen (PSMA) have been clinically introduced as a new class of radiotheranostics for the treatment of prostate cancer. Among them, ((( R)-1-carboxy-2-mcercaptoethyl)carbamoyl)-l-glutamic acid (MCG) has been successfully labeled with radioisotopes for prostate cancer imaging. The aim of this study is to conjugate MCG with an albumin binding moiety to further improve the in vivo pharmacokinetics. MCG was conjugated with an Evans blue (EB) derivative for albumin binding and a DOTA chelator. PSMA positive (PC3-PIP) and PSMA negative (PC3) cells were used for both in vitro and in vivo studies. Longitudinal PET imaging was performed at 1, 4, 24, and 48 h post-injection to evaluate the biodistribution and tumor uptake of 86 Y-DOTA-EB-MCG. DOTA-EB-MCG was also labeled with 90 Y for radionuclide therapy. Besides tumor growth measurement, tumor response to escalating therapeutic doses were also evaluated by immunohistochemistry and fluorescence microscopy. Based on quantification from 86 Y-DOTA-EB-MCG PET images, the tracer uptake in PC3-PIP tumors increased from 22.33 ± 2.39%ID/g at 1 h post-injection (p.i.), to the peak of 40.40 ± 4.79%ID/g at 24 h p.i. Administration of 7.4 MBq of 90 Y-DOTA-EB-MCG resulted in significant regression of tumor growth in PSMA positive xenografts. No apparent toxicity or body weight loss was observed in all treated mice. Modification of MCG with an Evans blue derivative resulted in a highly efficient prostate cancer targeting agent (EB-MCG), which showed great potential in prostate cancer treatment after being labeled with therapeutic radioisotopes.

The development of somastatin (SS) peptide analogues for the detection and treatment of neuroendocrine tumors has been successful with the recent FDA approval of 68 Ga-DOTA-TATE and 177 Lu-DOTA-TATE. The structure of these peptide constructs contains the peptide binding motif that binds to the receptor with high affinity, a chelator to complex the radioactive metal, and a linker between the peptide and chelator. However, these constructs suffer from rapid blood clearance, which limits their tumor uptake. In this study, this design has been further improved by incorporating a modification to control the in vivo pharmacokinetics. Adding a truncated Evans Blue (EB) dye molecule into the construct provides a prolonged half-life in blood as a result of its low micromolar affinity to albumin. We compared 177 Lu-DOTA-TATE to the modified 177 Lu Evans Blue compound ( 177 Lu-DMEB-TATE), in vitro and in vivo in mice bearing A427-7 xenografts. The tumor uptake of 177 Lu-DMEB-TATE was significantly greater than the uptake of 177 Lu-DOTA-TATE in the biodistribution and SPECT-imaging studies. The therapeutic effect of the 177 Lu-DMEB-TATE construct was superior to the that of the 177 Lu-DOTA-TATE construct at the doses evaluated.

Nanomedicines have achieved several breakthroughs in cancer treatment over the past decades; however, their potential immunotoxicities are ignored, which results in serious adverse effects and greatly reduces the potential in clinical translation. Herein, we innovatively develop a theranostic supramolecular polymer using β-cyclodextrin as the host and camptothecin (CPT) as the guest linked by a glutathione-cleavable disulfide bond. The supramolecular polymerization remarkably increases the solubility of CPT by a factor of 232 and effectively inhibits its lactone ring opening in physiological environment, which is favorable for intravenous formulation and maintenance of the therapeutic efficacy. Supramolecular nanoparticles can be prepared through orthogonal self-assembly driven by π-π stacking interaction, host-guest complexation, and hydrogen bonds. The sophisticated nanomedicine constructed from the obtained supramolecular polymer can be specifically delivered to tumor sites and rapidly excreted from body after drug release, thus effectively avoiding systemic toxicity, especially long-term immunotoxicity. In vivo investigations demonstrate this supramolecular nanomedicine possesses superior antitumor performance and antimetastasis capability. This pioneering example integrating the advantages of the dynamic nature of supramolecular chemistry and nanotechnology provides a promising platform for cancer theranostics.

Alleviation of tumor hypoxia has been the premise for improving the effectiveness of radiotherapy, which hinges upon the advanced delivery and rapid release of oxygen within the tumor region. Herein, we propose a "bubble-enhanced oxygen diffusion" strategy to achieve whole tumor oxygenation for significant radiation enhancement based on the "bystander effect". Toward this end, sub-50 nm CuS-modified and 64 Cu-labeled hollow mesoporous organosilica nanoparticles were constructed for tumor-specific delivery of O 2 -saturated perfluoropentane (PFP). Through the aid of PFP gasification arising from NIR laser-triggered mild hyperthermia, simultaneous PET/PA/US multimodality imaging and rapid oxygen diffusion across the tumor can be achieved for remarkable hypoxic radiosensitization. Furthermore, the multifunctional oxygen-carrying nanotheranostics also allow for other oxygen-dependent treatments, thus greatly advancing the development of bubble-enhanced synergistic therapy platforms.

Radiolabeled bombesin (BBN) analogs have long been used for developing gastrin-releasing peptide receptor (GRPR) targeted imaging probes, and tracers with excellent in vivo performance including high tumor uptake, high contrast, and favorable pharmacokinetics are highly desired. In this study, we compared the 68 Ga-labeled GRPR agonist (Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH 2 , BBN 7-14 ) and antagonist (d-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH 2 , RM26) for the positron emission tomography (PET) imaging of prostate cancer. The in vitro stabilities, receptor binding, cell uptake, internalization, and efflux properties of the probes 68 Ga-1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA)-Aca-BBN 7-14 and 68 Ga-NOTA-poly(ethylene glycol) 3 (PEG 3 )-RM26 were studied in PC-3 cells, and the in vivo GRPR targeting abilities and kinetics were investigated using PC-3 tumor xenografted mice. BBN 7-14 , PEG 3 -RM26, NOTA-Aca-BBN 7-14 , and NOTA-PEG 3 -RM26 showed similar binding affinity to GRPR. In PC-3 tumor-bearing mice, the tumor uptake of 68 Ga-NOTA-PEG 3 -RM26 remained at around 3.00 percentage of injected dose per gram of tissue within 1 h after injection, in contrast with 68 Ga-NOTA-Aca-BBN 7-14 , which demonstrated rapid elimination and high background signal. Additionally, the majority of the 68 Ga-NOTA-PEG 3 -RM26 remained intact in mouse serum at 5 min after injection, while almost all of the 68 Ga-NOTA-Aca-BBN 7-14 was degraded under the same conditions, demonstrating more-favorable in vivo pharmacokinetic properties and metabolic stabilities of the antagonist probe relative to its agonist counterpart. Overall, the antagonistic GRPR targeted probe 68 Ga-NOTA-PEG 3 -RM26 is a more-promising candidate than the agonist 68 Ga-NOTA-Aca-BBN 7-14 for the PET imaging of prostate cancer patients.

This study applies in situ production of hypochlorous acid (HOCl) to improve the therapeutic efficacy of platinum drugs. The phagocytic enzyme myeloperoxidase (MPO) is coated with two functional polyphenol derivatives (platinum prodrug polyphenols and PEG polyphenols) and ferric ion by metal phenolic coordination, which can shield MPO from degradation by other compounds in the blood. Moreover, the platinum prodrug can be reduced to cisplatin in cells and produce hydrogen peroxide (H 2 O 2 ). The MPO catalyzes the conversion of H 2 O 2 to HOCl in the intercellular environment. The as-prepared MPO Pt PEG nanoparticles (MPP NPs) can be employed as a reactive oxygen species cascade bioreaction to enhance platinum drug therapy. The MPP NPs show prolonged blood circulation and high tumor accumulation as evidenced by 89 Zr-based positron emission tomography imaging. The MPP NPs effectively inhibit tumor growth in vivo. As a first-in-class platform to harness the highly toxic HOCl in nanomedicine for cancer therapy, this strategy may open doors for further development of progressive therapeutic systems.

The clearance of nanoparticles (NPs) by mononuclear phagocyte system (MPS) from blood leads to high liver and spleen uptake and negatively impacts their tumor delivery efficiency. Here we systematically evaluated the in vitro and in vivo nanobio interactions of a two-dimensional (2D) model, gold (Au) nanorings, which were compared with Au nanospheres and Au nanoplates of similar size. Among different shapes, Au nanorings achieved the lowest MPS uptake and highest tumor accumulation. Among different sizes, 50 nm Au nanorings showed the highest tumor delivery efficiency. In addition, we demonstrated the potential use of Au naonrings in photoacoustic imaging and photothermal therapy. Thus, engineering the shape, surface area, and size of Au nanostructures is important in controlling NP-MPS interactions and improving the tumor uptake efficiency.

We report a camptothecin (CPT) prodrug that was well formulated in solution and rapidly transformed into long-circulating nanocomplexes in vivo for highly efficient drug delivery and effective cancer therapy. Specifically, using a redox-responsive disulfide linker, CPT was conjugated with an albumin-binding Evans blue (EB) derivative; the resulting amphiphilic CPT-ss-EB prodrug self-assembled into nanostructures in aqueous solution, thus conferring high solubility and stability. By binding CPT-ss-EB to endogenous albumin, the 80 nm CPT-ss-EB nanoparticles rapidly transformed into 7 nm albumin/prodrug nanocomplexes. CPT-ss-EB was efficient at intracellular delivery into cancer cells, released intact CPT in a redox-responsive manner, and exhibited cytotoxicity as potent as CPT. In mice, the albumin/CPT-ss-EB nanocomplex exhibited remarkably long blood circulation (130-fold greater than CPT) and efficient tumor accumulation (30-fold of CPT), which consequently contributed to excellent therapeutic efficacy. Overall, this strategy of transformative nanomedicine is promising for efficient drug delivery.

Reported procedures on the synthesis of gold nanoshells with smooth surfaces have merely demonstrated efficient control of shell thickness and particle size, yet no branch and nanoporous features on the nanoshell have been implemented to date. Herein, we demonstrate the ability to control the roughness and nanoscale porosity of gold nanoshells by using redox-active polymer poly(vinylphenol)-b-(styrene) nanoparticles as reducing agent and template. The porosity and size of the branches on this branched nanoporous gold nanoshell (BAuNSP) material can be facilely adjusted by control of the reaction speed or the reaction time between the redox-active polymer nanoparticles and gold ions (Au 3+ ). Due to the strong reduction ability of the redox-active polymer, the yield of BAuNSP was virtually 100%. By taking advantage of the sharp branches and nanoporous features, BAuNSP exhibited greatly enhanced physico-optical properties, including photothermal effect, surface-enhanced Raman scattering (SERS), and photoacoustic (PA) signals. The photothermal conversion efficiency can reach as high as 75.5%, which is greater than most gold nanocrystals. Furthermore, the nanoporous nature of the shells allows for effective drug loading and controlled drug release. The thermoresponsive polymer coated on the BAuNSP surface serves as a gate keeper, governing the drug release behavior through photothermal heating. Positron emission tomography imaging demonstrated a high passive tumor accumulation of 64 Cu-labeled BAuNSP. The strong SERS signal generated by the SERS-active BAuNSP in vivo, accompanied by enhanced PA signals in the tumor region, provide significant tumor information, including size, morphology, position, and boundaries between tumor and healthy tissues. In vivo tumor therapy experiments demonstrated a highly synergistic chemo-photothermal therapy effect of drug-loaded BAuNSPs, guided by three modes of optical imaging.

Semiconducting molecules of perylene diimide (PDI) with strong light absorption properties in the near-infrared region and good biocompatibility have received increasing attention in the field of theranostics, especially as photoacoustic (PA) imaging agents. Herein, we report a series of [ 64 Cu]-labeled PDI nanoparticles (NPs) of different sizes (30, 60, 100, and 200 nm) as dual positron emission tomography (PET) and PA imaging probes and photothermal therapy agents. The precise size control of the PDI NPs can be achieved by adjusting the initial concentration of PDI molecules in the self-assembly process, and the photophysical property of different sized PDI NPs was studied in detail. Furthermore, we systematically investigated the size-dependent accumulation of the PDI NPs in the lymphatic system after local administration and in tumors after intravenous injection by PA and PET imaging. The results revealed that 100 nm is the best size for differentiating popliteal and sciatic LNs since the interval is around 60 min for the NPs to migrate from popliteal LNs to sciatic LNs, which is an ideal time window to facilitate surgical sentinel LN biopsy and pathological examination. Furthermore, different migration times of the different-sized PDI NPs will provide more choices for surgeons to map the specific tumor relevant LNs. PDI NP theranostics can also be applied to imaging-guided cancer therapy. The NPs with a size of 60 nm appear to be the best for tumor imaging and photothermal cancer therapy due to the maximum tumor accumulation efficiency. Thus, our study not only presents organic PDI NP theranostics but also introduces different-sized NPs for multiple bioapplications.

HER2, a cell membrane protein overexpressed in many types of cancers, is correlated with poor diagnosis, suboptimal treatment outcome, and low survival rate. Multiple HER2-targeted drugs have been developed for the treatment of HER2-overexpressing tumor, which can in turn down-regulate HER2 expression. It is thus significant to profile HER2 expression for cancer prognosis, patient stratification, and monitoring therapy response. Aptamers, a class of single-stranded DNA/RNA (ssDNA/ssRNA) ligands, are promising for molecular biomarker imaging. Aptamers typically have strong binding affinity, high selectivity, batch-to-batch reproducibility, and low toxicity, and systemically injected aptamers often have high tumor-to-background ratio within a short time. However, current aptamers have been mostly screened in vitro, and these aptamers may lose binding ability in vivo due to conformational change under physiological environments. Here, a DNA library was combinatorially screened in vitro and in vivo, to select HER2-targeting DNA aptamers, termed Heraptamers, and labeled with 18 F for positron emission tomography (PET) imaging of HER2 in ovarian cancer. Specifically, using systematic evolution of ligands by exponential enrichment (SELEX), Heraptamer candidates were first selected and validated in vitro using HER2 extracellular domain (ECD) and HER2-positive SKOV3 cancer cells; then, aptamer candidates were modified with alkyne, radiolabeled with 18 F using azide-functionalized precursors by click chemistry, and screened in SKOV3-tumor-bearing mice using PET. Two aptamers, Heraptamer1 and Heraptamer2, reached high tumor uptake ratios within as short as 1 h. At 1.5 h post injection, the tumor uptake ratio of these two aptamers was up to 0.5%ID/g (injection dose/gram tissue), with tumor-to-muscle ratio of 4.55 ± 1.63 in SKOV3 tumor. In contrast, these aptamers have low uptake ratios in control MDA-MB-231 tumors. These preclinical studies showed that Heraptamers are promising for specific HER2 imaging.

One of the major design considerations for a drug is its pharmacokinetics: a drug with short blood half-life is less available at a target organ which in turn dictates treatment with either high or more frequent doses, and increases the likelihood of undesirable side effects. One method to improve drug pharmacokinetics is adding functional chemical groups to the drug molecule that can increase the half-life in the blood, hopefully, without significantly affecting its desired biological activity. Evans Blue (EB) dye reversibly binds to serum albumin with moderate affinity and has a long blood half-life. The binding of EB to albumin has been exploited to quantify protein leakage as an indicator of increased vascular permeability. Design of new chemical entities based on EB structure and coupling them to drugs, enables the usage of albumin as a reversible carrier in the blood and improves drug's half-life. This Topical Review summarizes the recent developments of various EB derivatives for molecular imaging and therapy applications.

It is essential to control the size and morphology of nanoparticles strictly in nanomedicine. Protein cages offer significant potential for templated synthesis of inorganic nanoparticles. In this study, we successfully synthesized ultrasmall copper sulfide (CuS) nanoparticles inside the cavity of ferritin (Fn) nanocages by a biomimetic synthesis method. The uniform CuS-Fn nanocages (CuS-Fn NCs) showed strong near-infrared absorbance and high photothermal conversion efficiency. In quantitative ratiometric photoacoustic imaging (PAI), the CuS-Fn NCs exhibited superior photoacoustic tomography improvements for real-time in vivo PAI of entire tumors. With the incorporation of radionuclide (64)Cu, (64)CuS-Fn NCs also served as an excellent PET imaging agent with higher tumor accumulation compared to free copper. Following the guidance of PAI and PET, CuS-Fn NCs were applied in photothermal therapy to achieve superior cancer therapeutic efficiency with good biocompatibility both in vitro and in vivo. The results demonstrate that the bioinspired multifunctional CuS-Fn NCs have potential as clinically translatable cancer theranostics and could provide a noninvasive, highly sensitive, and quantitative in vivo guiding method for cancer photothermal therapies in experimental and clinical settings.

In the treatment of type 2 diabetes mellitus, it is very important to develop therapeutics with prolonged circulation half-life. Exendin-4 is a glucagon like peptide-1 receptor (GLP-1R) agonist that has been modified in different ways for imaging insulinoma and for treating type-2 diabetes. In this work, we synthesized a maleimide derivative of truncated Evans blue dye (MEB-C3-Mal) to conjugate with (Cys(40))exendin-4 to obtain a highly stable MEB-C3-(Cys(40))exendin-4 (denoted as Abextide II). Through in situ binding with endogenous albumin, Abextide II lowers blood glucose level and prolongs the hypoglycemic effect in a type 2 diabetes mouse model more than the FDA approved Albiglutide., Competing Interests: The authors declare no competing financial interest.

Hexafluorobenzene (HFB) reacts with free thiols to produce a unique and selective perfluoroaromatic linkage between two sulfurs. We modified this chemical reaction to produce dimeric (18)F-RGD-tetrafluorobenzene (TFB)-RGD, an integrin αvβ3 receptor ligand. (18)F-HFB was prepared by a fluorine exchange reaction using K(18)F/K2.2.2 at room temperature. The automated radiofluorination was optimized to minimize the amount of HFB precursor and, thus, maximize the specific activity. (18)F-HFB was isolated by distillation and subsequently reacted with thiolated c(RGDfk) peptide under basic and reducing conditions. The resulting (18)F-RGD-TFB-RGD demonstrated integrin receptor specific binding, cellular uptake, and in vivo tumor accumulation.(18)F-HFB can be efficiently incorporated into thiol-containing peptides at room temperature to provide novel imaging agents., Competing Interests: The authors declare no competing financial interest.

We report a hybrid reduced graphene oxide (rGO)-loaded ultrasmall plasmonic gold nanorod vesicle (rGO-AuNRVe) (∼65 nm in size) with remarkably amplified photoacoustic (PA) performance and photothermal effects. The hybrid vesicle also exhibits a high loading capacity of doxorubicin (DOX), as both the cavity of the vesicle and the large surface area of the encapsulated rGO can be used for loading DOX, making it an excellent drug carrier. The loaded DOX is released sequentially: near-infrared photothermal heating induces DOX release from the vesicular cavity, and an intracellular acidic environment induces DOX release from the rGO surface. Positron emission tomography imaging showed high passive U87MG tumor accumulation of (64)Cu-labeled rGO-AuNRVes (∼9.7% ID/g at 24 h postinjection) and strong PA signal in the tumor region. Single intravenous injection of rGO-AuNRVe-DOX followed by low-power-density 808 nm laser irradiation (0.25 W/cm(2)) revealed effective inhibition of tumor growth due to the combination of chemo- and photothermal therapies. The rGO-AuNRVe-DOX capable of sequential DOX release by laser light and acid environment may have the potential for clinical translation to treat cancer patients with tumors accessible by light.

Functionalized quantum dots (QDs) have been widely explored for multimodality bioimaging and proven to be versatile agents. Attaching positron-emitting radioisotopes onto QDs not only endows their positron emission tomography (PET) functionality, but also results in self-illuminating QDs, with no need for an external light source, by Cerenkov resonance energy transfer (CRET). Traditional chelation methods have been used to incorporate the radionuclide, but these methods are compromised by the potential for loss of radionuclide due to cleavage of the linker between particle and chelator, decomplexation of the metal, and possible altered pharmacokinetics of nanomaterials. Herein, we described a straightforward synthesis of intrinsically radioactive [(64)Cu]CuInS/ZnS QDs by directly incorporating (64)Cu into CuInS/ZnS nanostructure with (64)CuCl2 as synthesis precursor. The [(64)Cu]CuInS/ZnS QDs demonstrated excellent radiochemical stability with less than 3% free (64)Cu detected even after exposure to serum containing EDTA (5 mM) for 24 h. PEGylation can be achieved in situ during synthesis, and the PEGylated radioactive QDs showed high tumor uptake (10.8% ID/g) in a U87MG mouse xenograft model. CRET efficiency was studied as a function of concentration and (64)Cu radioactivity concentration. These [(64)Cu]CuInS/ZnS QDs were successfully applied as an efficient PET/self-illuminating luminescence in vivo imaging agents.

Fluorine-18 is the most frequently used radioisotope in positron emission tomography (PET) radiopharmaceuticals in both clinical and preclinical research. Its physical and nuclear characteristics (97% β(+) decay, 109.7 min half-life, 635 keV positron energy), along with high specific activity and ease of large scale production, make it an attractive nuclide for radiochemical labeling and molecular imaging. Versatile chemistry including nucleophilic and electrophilic substitutions allows direct or indirect introduction of (18)F into molecules of interest. The significant increase in (18)F radiotracers for PET imaging accentuates the need for simple and efficient (18)F-labeling procedures. In this review, we will describe the current radiosynthesis routes and strategies for (18)F labeling of small molecules and biomolecules.

N-(2-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)ethyl)-6-fluoronicotinamide ([(18)F]FNEM), a novel prosthetic agent that is thiol-specific, was synthesized using a one-pot two-step strategy: (1) (18)F incorporation by a nucleophilic displacement of trimethylammonium substrate under mild conditions; (2) amidation of the resulting 6-[(18)F]fluoronicotinic acid 2,3,5,6-tetrafluorophenyl ester with N-(2-aminoethyl)maleimide trifluoroacetate salt. The radiosynthesis of the maleimide tracer was completed in 75 min from [(18)F]fluoride with 26 ± 5% decay uncorrected radiochemical yield, and specific activity of 19-88 GBq/μmol (decay uncorrected). The in vitro cell uptake, in vivo biodistribution, and positron emission tomography (PET) imaging properties of its conjugation product with [Cys(40)]-exendin-4 were described. [(18)F]FNEM-Cys(40)-exendin-4 showed specific targeting of glucagon-like peptide 1 receptor (GLP-1R) positive insulinomas and comparable imaging results to our recently reported [(18)F]FPenM-Cys(40)-exendin-4.

Using positron emission tomography (PET) imaging to monitor and quantitatively analyze the delivery and localization of Au nanomaterials (NMs), a widely used photothermal agent, is essential to optimize therapeutic protocols to achieve individualized medicine and avoid side effects. Coupling radiometals to Au NMs via a chelator faces the challenges of possible detachment of the radiometals as well as surface property changes of the NMs. In this study, we reported a simple and general chelator-free (64)Cu radiolabeling method by chemically reducing (64)Cu on the surface of polyethylene glycol (PEG)-stabilized Au NMs regardless of their shape and size. Our (64)Cu-integrated NMs are proved to be radiochemically stable and can provide an accurate and sensitive localization of NMs through noninvasive PET imaging. We further integrated (64)Cu onto arginine-glycine-aspartic acid (RGD) peptide modified Au nanorods (NRs) for tumor theranostic application. These NRs showed high tumor targeting ability in a U87MG glioblastoma xenograft model and were successfully used for PET image-guided photothermal therapy.

Enhanced mitochondrial potential in carcinoma cells is an important characteristic of cancer. It is of great current interest to develop a radiotracer that is sensitive to mitochondrial potential changes at the early stage of tumor growth. In this report, we present the synthesis and evaluation of (64)Cu-labeled Lissamine rhodamine B (LRB), (64)Cu(DOTA-LRB) (DOTA-LRB = 2-(6-(diethylamino)-3-(diethyliminio)-3H-xanthen-9-yl)-5-(N-(2-(2-(4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclo-dodecan-1-yl)acetamido)ethyl)sulfamoyl)benzenesulfonate) as a new radiotracer for imaging tumors in athymic nude mice bearing U87MG human glioma xenografts by positron emission tomography (PET). We also explored its localization mechanism using Cu(DOTA-LRB) as the fluorescent probe in both the U87MG human glioma cell line and the cultured primary U87MG glioma cells. It was found that (64)Cu(DOTA-LRB) had the highest tumor uptake (6.54 ± 1.50, 6.91 ± 1.26, 5.68 ± 1.13, 7.58 ± 1.96, and 5.14 ± 1.50%ID/g at 0.5, 1, 2, 4, and 24 h postinjection, respectively) among many (64)Cu-labeled organic cations evaluated in the same animal model. The cellular staining study indicated that Cu(DOTA-LRB) was able to localize in mitochondria of U87MG glioma cells due to the enhanced negative mitochondrial potential. This statement is completely supported by the results from decoupling experiment with carbonylcyanide-m-chlorophenylhydrazone (CCCP). MicroPET data showed that the U87MG glioma tumors were clearly visualized as early as 30 min postinjection with (64)Cu(DOTA-LRB). (64)Cu(DOTA-LRB) remained stable during renal excretion, but underwent extensive degradation during hepatobiliary excretion. On the basis of the results from this study, it was concluded that (64)Cu(DOTA-LRB) represents a new class of promising PET radiotracers for noninvasive imaging of the MDR-negative tumors.

This report presents the synthesis and evaluation of (64)Cu(DO3A-xy-ACR) (DO3A-xy-ACR = 2,6-bis(dimethylamino)-10-(4-((4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl)methyl)benzyl)acridin-10-ium) as a radiotracer for imaging tumors in athymic nude mice bearing U87MG glioma xenografts by PET (positron emission tomography). The biodistribution data suggested that (64)Cu(DO3A-xy-ACR) was excreted mainly through the renal system with >65% of injected radioactivity being recovered from urine samples at 1 h postinjection (p.i.). The tumor uptake of (64)Cu(DO3A-xy-ACR) was 1.07 ± 0.23, 1.58 ± 0.55, 2.71 ± 0.66, 3.47 ± 1.19, and 3.52 ± 1.72%ID/g at 0.5, 1, 2, 4, and 24 h p.i., respectively. (64)Cu(DO3A-xy-ACR) had very high liver uptake (31.90 ± 3.98, 24.95 ± 5.64, 15.20 ± 4.29, 14.09 ± 6.82, and 8.18 ± 1.27%ID/g at 0.5, 1, 2, 4, and 24 h p.i., respectively) with low tumor/liver ratios. MicroPET studies showed that the tumors were clearly visualized as early as 30 min p.i. in the glioma-bearing mouse administered with (64)Cu(DO3A-xy-ACR). The high liver radioactivity accumulation was also seen. (64)Cu(DO3A-xy-ACR) had a relatively high metabolic stability during excretion via both renal and hepatobiliary routes, but it was completely decomposed in the liver homogenate. We explored the localization mechanism of Cu(DO3A-xy-ACR) using both U87MG human glioma and the cultured primary U87MG glioma cells. The results from the cellular staining assays showed that (64)Cu(DO3A-xy-ACR) is able to localize in the mitochondria of living U87MG glioma cells due to the enhanced negative mitochondrial potential as compared to normal cells. Although (64)Cu(DO3A-xy-ACR) is not an ideal PET radiotracer for tumor imaging due to its high liver uptake, the results from this study strongly suggest that (64)Cu-labeled acridinium cations are indeed able to localize in the energized mitochondria of tumor cells.

Labeling biomolecules with ¹⁸F is usually done through coupling with prosthetic groups, which requires several time-consuming radiosynthesis steps and therefore in low labeling yield. In this study, we designed a simple one-step ¹⁸F-labeling strategy to replace the conventional complex and the long process of multiple-step radiolabeling procedure. Both monomeric and dimeric cyclic RGD peptides were modified to contain 4-NO₂-3-CF₃ arene as precursors for direct ¹⁸F labeling. Binding of the two functionalized peptides to integrin α(v)β₃ was tested in vitro using the MDA-MB-435 human breast cell line. The most promising functionalized peptide, the dimeric cyclic RGD, was further evaluated in vivo in an orthotopic MDA-MB-435 tumor xenograft model. The use of relatively low amount of precursor (~0.5 μmol) gave reasonable yield, ranging from 7 to 23% (decay corrected, calculated from the start of synthesis) after HPLC purification. Overall reaction time was 40 min, and the specific activity of the labeled peptide was high. Modification of RGD peptides did not significantly change the biological binding affinities of the modified peptides. Small animal PET and biodistribution studies revealed integrin specific tumor uptake and favorable biokinetics. We have developed a novel one-step ¹⁸F radiolabeling strategy for peptides that contain a specific arene group, which shortens reaction time and labor significantly, requires low amount of precursor, and results in specific activity of 79 ± 13 GBq/μmol. Successful introduction of 4-fluoro-3-trifluoromethylbenzamide into RGD peptides may be a general strategy applicable to other biologically active peptides and proteins.

Previous studies with the anilinoquinazoline epidermal growth factor receptor (EGFR) irreversible inhibitor [(11)C]-ML03 demonstrated a rapid metabolism of the tracer, which led to its low in vivo accumulation in EGFR overexpressing tumors. To enhance tumor uptake, the chemical structure of the compound was modified, and four new groups of EGFR inhibitors with a wide range of chemical reactivities were synthesized. Chemical reactivity assay of the compounds, performed with reduced glutathione (GSH), revealed that the group C (4-(dimethylamino)-but-2-enoic amide) derivative was the least chemically reactive against the nucleophilic attack of GSH. Nonetheless, it demonstrated a high inhibitory potency and bound irreversibly to the EGFR. Consequently, the blood stability of the group C compound (5a, ML04) labeled with (11)C was studied. In a time frame of 60 min, no radioactive metabolites were detected in blood. The stability of [(11)C]-5a, as indicated both from in vitro blood-stability assays and injection into nude rats, was significantly higher as compared to [(11)C]-ML03. Since group C presented a greater promise for tumor accumulation, it represents, to date, the most suitable candidate for radiolabeling with long-lived positron emission tomography (PET) radioisotopes.