Imaging Agents Research Articles

Expanding the Substrate Selectivity of the Fimsbactin Biosynthetic Adenylation Domain, FbsH.

Nonribosomal peptide synthetases (NRPSs) produce diverse natural products including siderophores, chelating agents that many pathogenic bacteria produce to survive in low iron conditions. Engineering NRPSs to produce diverse siderophore analogs could lead to the generation of novel antibiotics and imaging agents that take advantage of this unique iron uptake system in bacteria. The highly pathogenic and antibiotic-resistant bacteria Acinetobacter baumannii produces fimsbactin, an unusual branched siderophore with iron-binding catechol groups bound to a serine or threonine side chain. To explore the substrate promiscuity of the assembly line enzymes, we report a structure-guided investigation of the stand-alone aryl adenylation enzyme FbsH. We report structures bound to its native substrate 2,3-dihydroxybenzoic acid (DHB) as well as an inhibitor that mimics the adenylate intermediate. We produced enzyme variants with an expanded binding pocket that are more tolerant for analogs containing a DHB C4 modification. Wild-type and mutant enzymes were then used in an in vitro reconstitution analysis to assess the production of analogs of the final product as well as several early stage intermediates. This analysis shows that some altered substrates progress down the fimsbactin assembly line to the downstream domains. However, analogs from alternate building blocks are produced at lower levels, indicating that selectivity exists in the downstream catalytic domains. These findings expand the substrate scope of producing condensation products between serine and aryl acids and identify the bottlenecks for chemoenzymatic production of fimsbactin analogs.

Distinct Chemical Determinants are Essential for Achieving Ligands for Superior Optical Detection of Specific Amyloid-β Deposits in Alzheimer's Disease.

Aggregated forms of different proteins are common hallmarks for several neurodegenerative diseases, including Alzheimer's disease, and ligands that selectively detect specific protein aggregates are vital. Herein, we investigate the molecular requirements of thiophene-vinyl-benzothiazole based ligands to detect a specific type of Aβ deposits found in individuals with dominantly inherited Alzheimer's disease caused by the Arctic APP E693G mutation. The staining of these Aβ deposits was alternated when switching the terminal heterocyclic moiety attached to the thiophene-vinyl-benzothiazole scaffold. The most prevalent staining was observed for ligands having a terminal 3-methyl-1H-indazole moiety or a terminal 1,2-dimethoxybenzene moiety, verifying that specific molecular interactions between these ligands and the aggregates were necessary. The synthesis of additional thiophene-vinyl-benzothiazole ligands aided in pinpointing additional crucial chemical determinants, such as positioning of nitrogen atoms and methyl substituents, for achieving optimal staining of Aβ aggregates. When combining the optimized thiophene-vinyl-benzothiazole based ligands with a conventional ligand, CN-PiB, distinct staining patterns were observed for sporadic Alzheimer's disease versus dominantly inherited Alzheimer's disease caused by the Arctic APP E693G mutation. Our findings provide chemical insights for developing novel ligands that allow for a more precise assignment of Aβ deposits, and might also aid in creating novel agents for clinical imaging of distinct Aβ aggregates in AD.

Advancements in Tumor Diagnostics through Carbon Dot‐Assisted Photoacoustic Imaging

AbstractSerendipitously discovered, carbon dots (CDs) have attracted significant attention as a potential contrast agent for photoacoustic imaging (PAI) in the biomedical sector. CDs play an essential role in PAI, contributing significantly to the early detection of diseases and monitoring treatment progress, particularly in tumor imaging. This review emphasizes the role of CDs in the domain of PAI, highlighting their characteristics like biocompatibility, enhanced spatial resolution, optical absorption in the NIR region, and facile surface functionalization for tumor‐ targeted imaging. The study explores the use of CDs for enhancing spatial resolution in PAI for improved detection and visualization of tumors in organs such as the breast, cervical, liver, gastrointestinal, skin, cardiovascular system, nervous system, and others. Challenges associated with PAI, such as optimizing the signal‐to‐noise ratio and ensuring stability under physiological conditions, have also been discussed.

Enabling Visible Light Sensitization of YbIII, NdIII and ErIII in Dimeric LnIII/GaIII Metallacrowns through Functionalization with RuII Complexes for NIR‐II Multiplex Imaging

AbstractMultiplex imaging in the second near‐infrared window (NIR‐II, 1000–1700 nm) provides exciting opportunities for more precise understanding of biological processes and more accurate diagnosis of diseases by enabling real‐time acquisition of images with improved contrast and spatial resolution in deeper tissues. Today, the number of imaging agents suitable for this modality remains very scarce. In this work, we have synthesized and fully characterized, including theoretical calculations, a series of dimeric LnIII/GaIII metallacrowns bearing RuII polypyridyl complexes, LnRu‐3 (Ln=YIII, YbIII, NdIII, ErIII). Relaxed structures of YRu‐3 in the ground and the excited electronic states have been calculated using dispersion‐corrected density functional theory methods. Detailed photophysical studies of LnRu‐3 have demonstrated that characteristic emission signals of YbIII, NdIII and ErIII in the NIR‐II range can be sensitized upon excitation in the visible range through RuII‐centered metal‐to‐ligand charge transfer (MLCT) states. We have also showed that these NIR‐II signals are unambiguously detected in an imaging experiment using capillaries and biological tissue‐mimicking phantoms. This work opens unprecedented perspectives for NIR‐II multiplex imaging using LnIII‐based molecular compounds.

Phage-Templated Synthesis of Targeted Photoactive 1D-Thiophene Nanoparticles.

Thiophene-based nanoparticles (TNPs) are promising therapeutic and imaging agents. Here, using an innovative phage-templated synthesis, a strategy able to bypass the current limitations of TNPs in nanomedicine applications is proposed. The phage capsid is decorated with oligothiophene derivatives, transforming the virus in a 1D-thiophene nanoparticle (1D-TNP). A precise control of the shape/size of the nanoparticles is obtained exploiting the well-defined morphology of a refactored filamentous M13 phage, engineered by phage display to selectively recognize the Epidermal Growth Factor Receptor (EGFR). The tropism of the phage is maintained also after the bioconjugation of the thiophene molecules on its capsid. Moreover, the 1D-TNP proved highly fluorescent and photoactive, generating reactive oxygen species through both type I and type II mechanisms. The phototheranostic properties of this platform are investigated on biosystems presenting increasing complexity levels, from in vitro cancer cells in 2D and 3D architectures, to the in vivo tissue-like model organism Hydra vulgaris. The phage-templated 1D-TNP showed photocytotoxicity at picomolar concentrations, and the ability to deeply penetrate 3D spheroids and Hydra tissues. Collectively the results indicate that phage-templated synthesis of organic nanoparticles represents a general strategy, exploitable in many diagnostic and therapeutic fields based on targeted imaging and light mediated cell ablation.

Development and validation of a controlled heating apparatus for long‐term MRI of 3D microfluidic tumor models

AbstractConventional testing of novel contrast agents for magnetic resonance imaging (MRI) involves cell and animal studies. However, 2D cultures lack dynamic flow and in vivo MRI is limited by regulatory approval of long‐term anesthesia use. Microfluidic tumor models (MTMs) offer a cost‐effective, reproducible, and high throughput platform for bridging cell and animal models. Yet, MRI of microfluidic devices is challenging, due to small fluid volumes generating low sensitivity. For the first time, an MRI of MTMs was performed at low field strength (1 T) using conventional imaging equipment without microcoils. To enable longitudinal MRI, we developed (1) CHAMP‐3 (controlled heating apparatus for microfluidics and portability) which heats MTMs during MRI scans and (2) an MRI‐compatible temperature monitoring system. CHAMP‐3 maintained chip surface temperature at ~37°C and the media inside at ~35.5°C. Enhanced T1‐weighted MRI contrast was achieved in 3D MTMs with free manganese (Mn2+) solutions and Mn2+ labeled tumor cells.

Current Context of Designing Phototheranostic Cyclometalated Iridium (III) Complexes to Open a New Avenue in Cancer Therapy.

Photo-induced chemotherapy offers the best option for the selective treatment of cancer among all the prevailing modalities. Iridium (III) complexes, flourished with excellent photophysical and photochemical properties, have been considered to be superior for undergoing photo-responsive cancer therapy. Large Stokes shift, long-lived triplet excited state, photostability, and tuneable emission have rendered its excellence as a phototheranostic agent. In particular, the cyclometalated Ir (III) complexes and their respective nanoparticles have made a strong niche in the arena of cancer therapy. In recent years, Ir (III) based complexes have shown promising utilities as both imaging and therapeutic agents as well. Therefore, this review summarises the recent advances in the strategic designing of cyclometalated Ir(III) complexes to augment their phototheranostic applications in precision medicine.

Functional outcome after infectious encephalitis: a longitudinal multicenter prospective cohort study.

ObjectivesWe aimed to describe longitudinal functional outcome among survivors after an infectious encephalitis (IE) and to analyze risk factors for poor functional outcome. MethodsPatients included in the prospective ENCEIF cohort (France, 2016-2019) were followed-up at 6 months and one year after hospital discharge for assessment of i) functional outcome using modified Rankin scale (mRS); ii) cognitive function and abilities to perform activities of daily living. Risk factors for poor outcome on the full distribution of mRS were estimated using multivariable mixed ordinal regression analysis with time between hospital discharge and follow-up as a covariate. ResultsOur follow-up study included 322 patients with 896 mRS evaluations. Median age was 66 [50-74] years, 197/322 were male (61%) and 35/322 were immunocompromised (11%). Causative agents were herpes simplex virus 1 (HSV-1) in 95/322 cases (30%), varicella zoster virus (VZV) in 46/322 cases (14%), others documented IE in 90/322 cases (28%) and unknown in 91/322 cases (28%). Intensive care unit (ICU) admission was necessary for 117/322 patients (36%). Brain imaging was abnormal in 180/311 (58%) of patients.At 6 months, 95/287 (33%) had fully recovered and 181/287 (63%) had persisting symptoms. At 12 months, 124/253 (49%) had fully recovered and 108/253 (43%) had persisting symptoms. The proportion of patients who improved was 41% (117/287) during the first 6 months, and 24% (52/218) between 6 and 12 months. Factors significantly associated with poor functional outcome were age, immunosuppression, ICU admission, abnormal brain imaging and causative agents, notably HSV-1. Follow-up visit during the first 120 days did not detect significant change in functional outcome, but was strongly associated with better outcome at the subsequent evaluation. ConclusionsAfter IE, improvement may take several months. Functional outcome is associated with baseline health status (age, immunosuppression), abnormal brain imaging, ICU admission, and causative agent.

Intraoperative Guidance of Pancreatic Cancer Resection Using a Toll-like Receptor 2-Targeted Fluorescence Molecular Imaging Agent.

Human TLR2 is broadly expressed among pancreatic adenocarcinomas, and the highly specific TLR2L-800 fluorescence molecular imaging agent has potential for use in fluorescence-guided surgery to increase R0 margins and improve patient survival.

Molecular Imaging of Melanoma VEGF-expressing Tumors through [99mTc]Tc-HYNIC-Fab(Bevacizumab).

Angiogenesis is a process that many tumors depend on for growth, development, and metastasis. Vascular endothelial growth factor (VEGF) is one of the major players in tumor angiogenesis in several tumor types, including melanoma. VEGF inhibition is achieved by bevacizumab, a humanized monoclonal antibody that binds with high affinity to VEGF and prevents its function. In order to successfully enable in vivo VEGF expression imaging in a murine melanoma model, we previously labeled bevacizumab with [99mTc]Tc. We observed that this was feasible, but it had prolonged blood circulation and delayed tumor uptake. The aim of this study was to develop a radiolabeled Fab bevacizumab fragment, [99mTc]Tc-HYNICFab( bevacizumab), for non-invasive in vivo VEGF expression molecular imaging. Flow cytometry was used to examine VEGF presence in the murine melanoma cell line (B16-F10). Bevacizumab was digested with papain for six hours at 37°C to produce Fab(bevacizumab), which was then conjugated to NHS-HYNIC-Tfa for radiolabeling with [99mTc]Tc. Stability and binding affinity assays were also evaluated. Biodistribution and single photon emission computed tomography/computed tomography (SPECT/CT) were performed at 1, 3, and 6 h (n = 4) after injection of [99mTc]Tc-HYNIC-Fab(Bevacizumab) in normal and B16-F10 tumor-bearing C57Bl/6J mice. Using flow cytometry, it was shown that the B16-F10 murine melanoma cell line has intracellular VEGF expression. Papain incubation resulted in the complete digestion of bevacizumab with good purity and homogeneity. The radiolabeling yield of [99mTc]Tc-HYNIC-Fab(bevacizumab) was 85.00 ± 6.06%, with a specific activity of 291.87 ± 18.84 MBq/mg (n=3), showing in vitro stability. Binding assays demonstrated significant intracellular in vitro VEGF expression. Fast blood clearance and high kidney and tumor uptake were observed in biodistribution and SPECT/CT studies. We present the development and evaluation of [99mTc]Tc-HYNIC-Fab(bevacizumab), a novel molecular VEGF expression imaging agent that may be used for precision medicine in melanoma and potentially in other VEGF-expressing tumors.

Preliminary Study of Radionuclide-Labeled MerTK-Targeting PET Imaging Agents for the Diagnosis of Melanoma.

MerTK PET imaging holds potential as a promising approach for assessing tumor aggressiveness and monitoring treatment response. In this study, we synthesized a series of 18F- and 68Ga-labeled tracers derived from MerTK inhibitors for detection of MerTK expression. Among the synthesized agents, the dimeric compounds [68Ga]10 and [68Ga]12 demonstrated good in vivo and in vitro stability, high affinities to the MerTK receptor, and good MerTK-targeting specificity. Notably, [68Ga]10 exhibited a tumor uptake of 2.6 ± 0.2%ID/g at 1 h p. i. in B16F10 tumor-bearing mice, nearly tripling the uptake of its monomeric counterpart [68Ga]3. A similar enhancement was observed with [68Ga]12 compared to its monomeric analog [68Ga]6. Additionally, [18F]14 achieved a tumor uptake of 7.6 ± 0.5%ID/g at 2 h p. i., outperforming the previously reported [18F]15. Biodistribution analysis further validated the results, highlighting their potential for clinical investigation.

Nanozymes as Glucose Scavengers and Oxygenerators for Enhancing Tumor Radiotherapy.

Insufficient accumulation of reactive oxygen species (ROS) due to tumor hypoxia significantly contributes to increased radiation resistance and the failure of radiotherapy (RT). Therefore, developing methods to alleviate hypoxia and boost ROS levels represents a promising strategy for enhanced radiosensitivity. This study introduced a self-cascade catalytic Pt@Au nanozymes as a radiosensitizer, using glucose oxidase (GOx)-, catalase (CAT)-, and peroxidase (POD)-like activities to improve hypoxia and increase ROS accumulation, thereby affecting glucose metabolism and enhancing the effects of RT. Pt@Au nanozymes exhibit GOx-like activity, which not only depletes glucose to induce starvation therapy, but also generates hydrogen peroxide (H2O2) for cascade reactions. Moreover, Pt@Au nanozymes demonstrate CAT-like activity, catalyzing the conversion of H2O2 to O2. This conversion effectively alleviates hypoxia, stabilizes ROS, increases DNA damage, significantly enhancing RT efficacy and sustaining the effects of starvation therapy. As high-Z materials, Pt@Au nanozymes can deposit more X-ray energy. Furthermore, the POD-like activity catalyzes the conversion of H2O2 into highly reactive hydroxyl radicals (·OH), which increases ROS levels and enhances RT. Pt@Au nanozymes serve as X-ray computed tomography (CT) imaging agents, allowing for clear differentiation between tumor and normal tissue boundaries and enhancing the precision of RT. In summary, Pt@Au nanozymes serve as effective radiosensitizers by depleting glucose to induce starvation therapy, enhancing cascade reactions, and inhibiting tumor proliferation. Through their self-cascade reactions, these nanozymes dramatically increase oxygen levels within tumors, reduce hypoxia, and enhance ROS levels. This advancement addresses the radioresistance associated with hypoxic tumors, paving the way for innovative strategies in RT.

Advancements in nanotheranostics for glioma therapy.

Gliomas are brain tumors mainly derived from glial cells that are difficult to treat and cause high mortality. Radiation, chemotherapy, and surgical excision are the conventionaltreatments for gliomas. Patients who have surgery or have undergone chemotherapy for glioma treatment have poor prognosis with tumor recurrence. In particular, for glioblastoma,the 5-year averagesurvival rateis 4-7%, and the median survival is 12-18months. Anumber of issues hinder effective treatmentsuch as, poor surgical resection, tumor heterogeneity, insufficient drug penetration across the blood-brain barrier, multidrug resistance, and difficulties with drug specificity. Nanotheranostic-mediated drug delivery is becoming a well-researched consideration, and an efficient non-invasive method for delivering chemotherapeutic drugs to the target area. Theranostic nanomedicines, which incorporate therapeutic drugs and imaging agents forpersonalized therapies, can be used for preventing overdose of non-responders. Through the identification of massive and complicated information from next-generation sequencing, machine learning enables for precise prediction of therapeutic outcomes and post-treatment management for patients with cancer. This article gives a thorough overview of nanocarrier-mediated drug delivery with a brief introduction to drug delivery challenges. In addition, this assessment offers a current summary of preclinical and clinical research on nanomedicines for gliomas. In the future, nanotheranostics will provide personalized treatment for gliomas and other treatable cancers.

Recent Development of Organic Afterglow Probes for Diagnosis and Treatment of Cancer

Afterglow imaging plays a crucial role in the cancer treatment field. In contrast to inorganic afterglow imaging agents, organic afterglow imaging agents possess easily modifiable structures and exhibit excellent biocompatibility, thereby presenting significant prospects for application in tumor diagnosis and management. In this review, we summarize the design principles and applications of afterglow probes in tumor imaging and therapy. Finally, we discuss the future challenges and prospects of organic afterglow probes in cancer diagnosis and therapy.

Rational modifications on N-(4-quinolinoyl)-Gly-2-cyanopyrrolidine to develop fibroblast activation protein-targeted radioligands with improved affinity and tumor uptake

Fibroblast activation protein (FAP) has been an attractive target for cancer imaging and therapy. Radiolabeled FAP-targeting ligands have shown promising results for clinical applications. However, further improvements are ongoing in pursuit of increasing tumor uptake, prolonging tumor residence, and maintenance good tumor–to–background contrast for extensive theranostic application. Achieving a higher affinity of the precursor is one of the ways in research. In this study, we designed a series of FAP inhibitors based on N-(4-quinolinoyl)-Gly-2-cyanopyrrolidine and found compound QI-18 with an IC50 value of 0.50 nM, which is a 6.5-fold increase in potency over that of UAMC-1110 (IC50 of 3.25 nM). QI-18 was then functionalized with a DOTA chelator to obtain the ligand CY03 for further radiolabeling with 68Ga to obtain the radiotracer [68Ga]Ga-CY03. In BALB/c nude mice bearing U87MG tumor models, [68Ga]Ga-CY03 exhibited a high and specific uptake (10.30 ± 0.63 % ID/g at 1 h post-injection and 9.28 ± 1.60 % ID/g at 2 h post-injection), which represented 3.2- and 4.1-fold increases over those for [68Ga]Ga-FAPI-04 (3.24 ± 0.53 % ID/g and 2.25 ± 0.33 % ID/g, respectively). [68Ga]Ga-CY03 also showed higher tumor–to–blood and tumor–to–kidney ratios (7.62 ± 0.44 and 2.59 ± 0.27, respectively) than [68Ga]Ga-FAPI-04 (2.38 ± 0.47 and 0.98 ± 0.19, respectively). The results indicate that [68Ga]Ga-CY03 is a promising imaging agent to target FAP.

Overcoming the Challenges of Hyperpolarizing Substrates with Parahydrogen‐Induced Polarization in an MRI System

Hyperpolarization of 13C nuclei in biomolecules and their administration as imaging agents enables in‐vivo monitoring of metabolism. This approach has demonstrated potential for deriving imaging biomarkers for cancer detection, differentiation, and therapy efficacy assessment. The in situ generation of polarized substrates using a permanent addition of parahydrogen to an unsaturated precursor inside the bore of an MRI system used for subsequent imaging circumvents the need for a dedicated external polarizer. This approach reduces polarization loss associated with sample transfer, minimizes hardware requirements and cost, and results in reduced spatial requirements. However, performing INEPT‐like pulsed sequences for heteronuclear spin‐order transfer in the bore of an MRI system is challenged by poor uniformity of static and excitation magnetic field and molecular convection during the polarization transfer. Therefore, here we characterize these effects, implement a robust modification to the pulse sequence, and measure experimentally the polarization improvement upon modification of the sequence. After rigorous optimization of the parameters, we obtained a 13C polarization of 44.5% for 50 mM of the 1‐13C site of ethyl acetate‐d6. Our parahydrogen‐induced polarization approach enhances the accessibility to hyperpolarized MRI, circumventing the need for an external polarizer.

Evaluating the impact of gadolinium contamination on the marine bivalve Donax trunculus: Implications for environmental health

Gadolinium (Gd), commonly used in contrast agents for medical imaging, has been detected in hospital wastewater and aquatic environments, raising environmental concerns. This study examined the accumulation and cellular impacts of Gd in the clam species Donax trunculus, commonly used as bioindicator of contamination. Gadolinium accumulation in clams increased with exposure and over time. Biological responses varied with Gd levels: low concentrations (10 and 50 µg/L) led to low metabolic activity and glycogen content, but high antioxidant activities and lipid peroxidation levels (LPO); high concentrations (250 and 500 µg/L) resulted in increased metabolic activity, while antioxidant enzyme activity was inhibited and LPO levels were the lowest. Metabolic activity decreased after two weeks, suggesting limited long-term metabolic resilience. The study underscores D. trunculus as an effective early warning species for Gd pollution and highlights the ecological risks of rising Gd levels, emphasizing the need for environmental monitoring and regulation.

Role of physicochemical properties in silica nanoparticle-mediated immunostimulation

Immunostimulation caused by nanoparticles may be beneficial or adverse depending on their intended application. Activation of immune cells is beneficial for indications targeting the immune system for therapeutic purposes, such as tumor microenvironment reprogramming, immunotherapy, and vaccines. When it is unwanted, however, immunostimulation may lead to excessive inflammation, cytokine storm, and hypersensitivity reactions. The increasing use of silica nanoparticles (SiNPs) for the delivery of drugs, imaging agents, and antigens warrants preclinical studies aimed at understanding carrier-mediated effects on the number, activation status, and function of immune cell subsets. Herein, we present an in vitro study utilizing primary human peripheral blood mononuclear cells (PBMC) to investigate the proinflammatory properties of four types of SiNPs varying in size and porosity. Cytokine analysis was performed in resting and LPS-primed PBMC cultures to understand the ability of silica nanoparticles to induce de novo and exaggerate preexisting inflammation, respectively. Changes in the number and activation status of lymphoid and myeloid cells were studied by flow cytometry to gain further insight into SiNP-mediated immunostimulation. Nonporous SiNPs were found to be more proinflammatory than mesoporous SiNPs, and larger-sized particles induced greater cytokine response. LPS-primed PBMC resulted in increased susceptibility to SiNPs. Immunophenotyping analysis of SiNP-treated PBMC resulted in T and B lymphocyte, natural killer cell, and dendritic cell activation. Additionally, a loss of regulatory T cells and an increase in γδ TCR T cell population were observed with all particles. These findings have implications for the utility of SiNPs for the delivery of drugs and imaging agents.

8-Anilino-1-naphthalenesulfonate-Conjugated Carbon-Coated Ferrite Nanodots for Fluoromagnetic Imaging, Smart Drug Delivery, and Biomolecular Sensing

Background: Superparamagnetic properties and excitation independence have been incorporated into carbon-decorated manganese ferrite nanodots (MnFe@C) to introduce an economical and safer multimodal agent for use in both T1-T2 MRI and fluorescence-based imaging to replace the conventional highly toxic heavy metal contrast agents. Methods: The surface conjugation of 8-anilino-1-naphthalenesulfonate (ANS) to MnFe@C nanodots (ANS-MnFe@C) enhances both longitudinal and transverse MRI relaxation, improves fluorescence for optical imaging, and increases protein detection sensitivity, showing higher multimodal efficacy in terms of molar relaxivity, radiant efficiencies, and fluorescence sensitivity compared to MnFe@C. Results: The band gap energy was determined using Tauc’s equation to be 3.32 eV, while a 72% quantum yield demonstrated that ANS-MnFe@C was highly fluorescent, with the linear range and association constant calculated using the Stern–Volmer relation. The synthesized ANS-MnFe@C demonstrated excellent selectivity and sensitivity for bovine serum albumin (BSA), with a nanomolar detection limit of 367.09 nM and a broad linear range from 0.015 to 0.225 mM. Conclusions: In conclusion, ANS-MnFe@C holds ease of fabrication, good biocompatibility, as assessed in A375 cells, and an effective pH-sensitive doxorubicin release profile to establish anticancer activity in lung cancer cell line (A549), highlighting its potential as an affordable therapeutic agent for multimodal imaging, drug delivery, and protein sensing.

Optimized method for fluorine-18 radiolabeling of Affibody molecules using RESCA

BackgroundIn recent years, the interest in Al[18F]F as a labeling agent for Positron Emission Tomography (PET) radiotracers has risen, as it allows for fast and efficient fluorine-18 labeling by harnessing chelation chemistry. The introduction of Restrained Complexing Agent (RESCA) as a chelator has also shown that chelator-based radiolabeling reactions can be performed in mild conditions, making the radiolabeling process attractively more facile than most conventional radiofluorination methods. The aim of the study was to establish optimized conditions for Al[18F]F labeling of Affibody molecules using RESCA as a complexing agent, using Z09591 and Z0185, two Affibody proteins targeting PDGFRβ and TNFα, respectively, as model compounds.ResultsThe Al[18F]F labeling of RESCA-conjugated Z09591 was tested at different temperatures (rt to 60 °C) and with varying reaction times (12 to 60 min), and optimal conditions were then implemented on RESCA-Z0185. The optimized synthesis method was: 1.5–2.5 GBq of cyclotron produced fluorine-18 were trapped on a QMA cartridge and eluted with saline solution to react with 12 nmol of AlCl3 and form Al[18F]F. The respective RESCA-conjugated Affibody molecule (14 nmol) in NaOAc solution was added to the Al[18F]F solution and left to react at 60 °C for 12 min. The mixture was purified on a NAP5 size exclusion column and then analyzed by HPLC. The entire process took approximately 35 min, was highly reproducible, indicating the efficiency and reliability of the method. The labeled compounds demonstrated retained biological function for their respective targets after purification.ConclusionsWe present a general and optimized method for Al[18F]F labeling of RESCA-conjugated Affibody molecules, which can be widely applied to this class of peptide-based imaging agents. Moreover, radiochemical yields were improved when the labeling was conducted at 37 °C or above. In vitro and in vivo assessment of the respective tracers was promising, showing retained binding capacity as well as moderate defluorination, which is usually regarded as a potential downside for RESCA-conjugated tracers.Graphical abstract