Targeted Imaging Agents Research Articles

In vitro and ex vivo anti-myeloma effects of nanocomposite As4S4/ZnS/Fe3O4

Nanoparticles in medicine can integrate actively targeted imaging agents and drug delivery vehicles, and combining multiple types of therapeutics in a single particle has numerous advantages, especially in multiple myeloma. MM is an incurable hematological disorder characterized by clonal proliferation of plasma cells in the bone marrow. In this study, we evaluated the anti-myeloma activity of 3 nanocomposites (3NPs): As4S4/ZnS/Fe3O4 (1:4:1), As4S4/ZnS/Fe3O4 with folic acid (FA), and As4S4/ZnS/Fe3O4 with FA and albumin with reduced survival MM cell lines and primary MM samples by each of 3NP. Cytotoxic effects of 3NPs were associated with caspase- and mitochondria-dependent apoptosis induction and reduced c-Myc expression. Modulation of cell cycle regulators, such as p-ATM/ATM and p-ATR/ATR, and increases in p-Chk2, cyclin B1, and histones were accompanied by G2/M arrest triggered by 3NPs. In addition, 3NPs activated several myeloma-related signaling, including JNK1/2/3, ERK1/2 and mTOR. To overcome BM microenvironment-mediated drug resistance, nanocomposites retained its anti-MM activity in the presence of stroma. 3NPs significantly decreased the stem cell-like side population in MM cells, even in the context of stroma. We observed strong synergistic effects of 3NPs combined with lenalidomide, pomalidomide, or melphalan, suggesting the potential of these combinations for future clinical studies.

The Performance of FDA-Approved PET Imaging Agents in the Detection of Prostate Cancer

Positron emission tomography (PET) incorporated with X-ray computed tomography (PET/CT) or magnetic resonance imaging (PET/MRI) is increasingly being used as a diagnostic tool for prostate cancer (PCa). In this review, we describe and evaluate the clinical performance of some Food and Drug Administration (FDA)-approved agents used for visualizing PCa: [18F]FDG, [11C]choline, [18F]FACBC, [68Ga]Ga-PSMA-11, [18F]DCFPyL, and [18F]-NaF. We carried out a comprehensive literature search based on articles published from 1 January 2010 to 1 March 2022. We selected English language articles associated with the discovery, preclinical study, clinical study, and diagnostic performance of the imaging agents for the evaluation. Prostate-specific membrane antigen (PSMA)-targeted imaging agents demonstrated superior diagnostic performance in both primary and recurrent PCa, compared with [11C]choline and [18F]FACBC, both of which target dividing cells and are used especially in patients with low prostate-specific antigen (PSA) values. When compared to [18F]-NaF (which is suitable for the detection of bone metastases), PSMA-targeted agents were also capable of detecting lesions in the lymph nodes, soft tissues, and bone. However, a limitation of PSMA-targeted imaging was the heterogeneity of PSMA expression in PCa, and consequently, a combination of two PET tracers was proposed to overcome this obstacle. The preliminary studies indicated that the use of PSMA-targeted scanning is more cost efficient than conventional imaging modalities for high-risk PCa patients. Furthering the development of imaging agents that target PCa-associated receptors and molecules could improve PET-based diagnosis of PCa.

First-in-Humans Evaluation of Safety and Dosimetry of 64Cu-LLP2A for PET Imaging.

There remains an unmet need for molecularly targeted imaging agents for multiple myeloma (MM). The integrin very late antigen 4 (VLA4), is differentially expressed in malignant MM cells and in pathogenic inflammatory microenvironmental cells. [64Cu]Cu-CB-TE1A1P-LLP2A (64Cu-LLP2A) is a VLA4-targeted, high-affinity radiopharmaceutical with promising utility for managing patients diagnosed with MM. Here, we evaluated the safety and human radiation dosimetry of 64Cu-LLP2A for potential use in MM patients. Methods: A single-dose [natCu]Cu-LLP2A (Cu-LLP2A) tolerability and toxicity study was performed on CD-1 (Hsd:ICR) male and female mice. 64Cu-LLP2A was synthesized in accordance with good-manufacturing-practice-compliant procedures. Three MM patients and six healthy participants underwent 64Cu-LLP2A-PET/CT or PET/MRI at up to 3 time points to help determine tracer biodistribution, pharmacokinetics, and radiation dosimetry. Time-activity curves were plotted for each participant. Mean organ-absorbed doses and effective doses were calculated using the OLINDA software. Tracer bioactivity was evaluated via cell-binding assays, and metabolites from human blood samples were analyzed with analytic radio-high-performance liquid chromatography. When feasible, VLA4 expression was evaluated in the biopsy tissues using 14-color flow cytometry. Results: A 150-fold mass excess of the desired imaging dose was tolerated well in male and female CD-1 mice (no observed adverse effect level). Time-activity curves from human imaging data showed rapid tracer clearance from blood via the kidneys and bladder. The effective dose of 64Cu-LLP2A in humans was 0.036 ± 0.006 mSv/MBq, and the spleen had the highest organ uptake, 0.142 ± 0.034 mSv/MBq. Among all tissues, the red marrow demonstrated the highest residence time. Image quality analysis supports an early imaging time (4-5 h after injection of the radiotracer) as optimal. Cell studies showed statistically significant blocking for the tracer produced for all human studies (82.42% ± 13.47%). Blood metabolism studies confirmed a stable product peak (>90%) up to 1 h after injection of the radiopharmaceutical. No clinical or laboratory adverse events related to 64Cu-LLP2A were observed in the human participants. Conclusion: 64Cu-LLP2A exhibited a favorable dosimetry and safety profile for use in humans.

Protein Cages: From Fundamentals to Advanced Applications.

Proteins that self-assemble into polyhedral shell-like structures are useful molecular containers both in nature and in the laboratory. Here we review efforts to repurpose diverse protein cages, including viral capsids, ferritins, bacterial microcompartments, and designed capsules, as vaccines, drug delivery vehicles, targeted imaging agents, nanoreactors, templates for controlled materials synthesis, building blocks for higher-order architectures, and more. A deep understanding of the principles underlying the construction, function, and evolution of natural systems has been key to tailoring selective cargo encapsulation and interactions with both biological systems and synthetic materials through protein engineering and directed evolution. The ability to adapt and design increasingly sophisticated capsid structures and functions stands to benefit the fields of catalysis, materials science, and medicine.

Preparation of the Biodegradable Lymphatic Targeting Imaging Agent Based on the Indocyanine Green Mesoporous Silicon System.

The lymphatic system plays a crucial role in the immune system’s recognition and response to disease. Therefore, the imaging of the lymphatic system, especially lymphatic vessels, has emerged as a valuable tool for the diagnosis of metastasis. FDA-approved small-molecule dyes, namely, indocyanine green (ICG), have been widely applied to lymphatic vessels imaging. However, due to the small physical size, such molecule-based agents show no selectivity, and rapid clearance from lymph nodes. Herein, a biodegradable lymphatic targeting imaging agent based on the ICG-mesoporous silicon system (ICG@HMONs-HA) was obtained, which not only could target lymph vessels but also had a long residence time. The reported work provides a practical way for lymph vessel fluorescence imaging and paves the way for clinical translation of nanomaterial-based tracers.

A novel dual-labeled small peptide as a multimodal imaging agent for targeting wild-type EGFR in tumors.

The epidermal growth factor receptor (EGFR) is over-expressed in various human cancer. The over-expression of EGFR in tumors is an excellent target for the development of cancer imaging agents. In the present study, we developed Tc-99m SYPIPDT-GHEG-ECG-K-tetramethylrhodamine (SYPIPDT-ECG-TAMRA) as a molecular imaging agent targeting wild-type EFGR (wtEGFR)-positive tumor cells, and verified its feasibility as molecular imaging agent. SYPIPDT-ECG-TAMRA was synthesized using Fmoc solid-phase peptide synthesis. The radiolabeling of SYPIPDT-ECG-TAMRA with Tc-99m was accomplished using ligand exchange via tartrate. Cellular uptake and binding affinity studies were performed. In vivo gamma camera imaging, ex vivo imaging and biodistribution studies were performed using NCI-H460 and SW620 tumor-bearing murine models. After radiolabeling procedures with Tc-99m, Tc-99m SYPIPDT-ECG-TAMRA complexes were prepared at high yield (> 95%). The binding affinity value (Kd) of Tc-99m SYPIPDT-ECG-TAMRA for NCI-H460 cells was estimated to be 76.5 ± 15.8 nM. In gamma camera imaging, the tumor to normal muscle uptake ratios of Tc-99m SYPIPDT-ECG-TAMRA increased with time (2.7 ± 0.6, 4.0 ± 0.9, and 6.2 ± 1.0 at 1, 2, and 3 h, respectively). The percentage injected dose per gram of wet tissue for the NCI-H460 tumor was 1.91 ± 0.11 and 1.70 ± 0.22 at 1 and 3 h, respectively. We developed Tc-99m SYPIPDT-ECG-TAMRA, which is dual-labeled with both radioisotope and fluorescence. In vivo and in vitro studies demonstrated specific uptake of Tc-99m SYPIPDT-ECG-TAMRA into wtEGFR-positive NCI-H460 cells and tumors. Thus, the results of the present study suggest that Tc-99m SYPIPDT-ECG-TAMRA is a potential dual-modality imaging agent targeting wtEGFR.

A Method ofTumor In Vivo Imaging with a New Peptide-Based Fluorescent Probe.

Precise surgical resection directly influences the prognosis and survival of patients with solid tumors. However, it is often difficult to distinguish tumor from normal tissue during resection without any intraoperative imaging guidance. Image-guided surgery particularly when coupled with a near-infrared (NIR) fluorescent agent may improve positive-margin rate thereby improving the overall prognosis. We have developed a unique tumor-targeting fluorescence imaging agent that can aid in the accurate localization of human cancer cells in preclinical settings. The NIR imaging agent, ICG-p28, a water-soluble, nontoxic, and pan-tumor targeting probe consisting of a cell-penetrating peptide (p28) conjugated to indocyanine green (ICG), can accurately localize tumors in vivo. Development of the noninvasive, targeted imaging agent can potentially improve in the resections of tumors by enabling the localization of lesions that are currently difficult or impossible to detect by visual observation or palpation. Here, we describe the methodsof preclinical animal imaging models by using NIR fluorescence imager coupled with a new tumor-targeting agent.

Design, synthesis, and evaluation of a novel PET imaging agent targeting lipofuscin in senescent cells.

Promoting a senescent phenotype to suppress tumour progression may present an alternative strategy for treating cancer and encourages the development of positron emission tomography (PET) imaging biomarkers for assessing response to treatment. The accumulation of lipofuscin deposits in senescent cells is visualised using the pathology stain Sudan Black B (SBB) which is an emerging biomarker of senescence. We describe the design, synthesis and evaluation of [18F]fluoroethyltriazole-SBB ([18F]FET-SBB), a fluorine-18 radiolabelled derivative of SBB. The in vitro uptake of [18F]FET-SBB in a senescent cell line corelated with lipofuscin deposits; in vivo PET imaging and metabolite analysis confirm a favourable pharmacokinetic and metabolic profile for further studies of in vivo models of senescence.

A novel fluorescent c-met targeted imaging agent for intra-operative colonic tumour mapping: Translation from the laboratory into a clinical trial

BackgroundThe c-Met protein is overexpressed in many gastrointestinal cancers. We explored EMI-137, a novel c-Met targeting fluorescent probe, for application in fluorescence-guided colon surgery, in HT-29 colorectal cancer (CRC) cell line and an in vivo murine model. MethodsHT-29 SiRNA transfection confirmed specificity of EMI-137 for c-Met. A HT-29 CRC xenograft model was developed in BALB/c mice, EMI-137 was injected and biodistribution analysed through in vivo fluorescent imaging. Nine patients, received a single intravenous EMI-137 bolus (0.13 mg/kg), 1–3 h before laparoscopic-assisted colon cancer surgery (NCT03360461). Tumour and LN fluorescence was assessed intraoperatively and correlated with c-Met expression in eight samples by immunohistochemistry. Findingsc-Met expression HT-29 cells was silenced and imaged with EMI-137. Strong EMI-137 uptake in tumour xenografts was observed up to 6 h post-administration. At clinical trial, no serious adverse events related to EMI-137 were reported. Marked background fluorescence was observed in all participants, 4/9 showed increased tumour fluorescence over background; 5/9 had histological LN metastases; no fluorescent LN were detected intraoperatively. All primary tumours (8/8) and malignant LN (15/15) exhibited high c-Met protein expression. InterpretationEMI-137, binds specifically to the human c-Met protein, is safe, and with further refinement, shows potential for application in fluorescence-guided surgery.

Lipoprotein-biomimetic nanostructure enables tumor-targeted penetration delivery for enhanced photo-gene therapy towards glioma

Glioma is one of the most malignant primary tumors affecting the brain. The efficacy of therapeutics for glioma is seriously compromised by the restriction of blood-brain barrier (BBB), interstitial tumor pressure of resistance to chemotherapy/radiation, and the inevitable damage to normal brain tissues. Inspired by the natural structure and properties of high-density lipoprotein (HDL), a tumor-penetrating lipoprotein was prepared by the fusion tLyP-1 to apolipoprotein A-I-mimicking peptides (D4F), together with indocyanine green (ICG) incorporation and lipophilic small interfering RNA targeted HIF-1α (siHIF) surface anchor for site-specific photo-gene therapy. tLyP-1 peptide is fused to HDL-surface to facilitate BBB permeability, tumor-homing capacity and -site accumulation of photosensitizer and siRNA. Upon NIR light irradiation, ICG not only served as real-time targeted imaging agent, but also provided toxic reactive oxygen species and local hyperthermia for glioma phototherapy. The HIF‐1α siRNA in this nanoplatform downregulated the hypoxia‐induced HIF‐1α level in tumor microenvironment and enhanced the photodynamic therapy against glioma. These studies demonstrated that the nanoparticles could not only efficiently across BBB and carry the payloads to orthotopic glioma, but also modulate tumor microenvironment, thereby inhibiting tumor growth with biosafety. Overall, this study develops a new multifunctional drug delivery system for glioma theranostic, providing deeper insights into orthotopic brain tumor imaging and treatment.

Identification of a Suitable Untargeted Agent for the Clinical Translation of ABY-029 Paired-Agent Imaging in Fluorescence-Guided Surgery.

Non-specific uptake and retention of molecular targeted agents and heterogeneous tissue optical properties diminish the ability to differentiate between tumor and normal tissues using molecular targeted fluorescent agents. Paired-agent imaging (PAI) can increase the diagnostic ability to detect tumor tissue by mitigating these non-specific effects and providing true molecular contrast by co-administration of an untargeted control imaging agent with a targeted agent. This study evaluates the suitability of available clinically translatable untargeted agents for the translation of PAI in fluorescence-guided surgery using an affibody-based targeted imaging agent (ABY-029). DESIGN: Three untargeted agents that fluoresce near 700nm and exhibit good clinical safety profiles (methylene blue, IRDye 700DX, and IRDye 680LT) were tested in combination with the clinically tested IRDye 800CW-labeled anti-epidermal growth factor receptor (EGFR) affibody molecule, ABY-029 (eIND 122,681). Properties of the untargeted agent important for human use and integrity of PAI were tested: (1) plasma protein binding; (2) fluorescence signal linearity in in vitro whole blood dilution; (3) in vivo pharmacokinetic matching to targeted agent in negative control tissue; and (4) in vivo diagnostic accuracy of PAI vs single agent imaging (SAI) of ABY-029 alone in orthotopic oral head and neck squamous cell carcinomas. IRDye 680LT outperformed IRDye 700DX and methylene blue with the highest signal linearity (R2 = 0.9998 ± 0.0002, 0.9995 ± 0.0004, 0.91 ± 0.02, respectively), the highest fluorescence yield in whole blood at 1μM (104.42 ± 0.05, 103.68 ± 0.09, 101.9 ± 0.2, respectively), and the most closely matched ABY-029 pharmacokinetics in EGFR-negative tissues (binding potential error percentage = 0.31% ± 0.37%, 10.25% ± 1.30%, and 8.10% ± 5.37%, respectively). The diagnostic ability of PAI with ABY-029 and IRDye 680LT outperformed conventional SAI with an area-under-the-receiver-operating-characteristic curve (AUC) value of 0.964 vs. 0.854, and 0.978 vs. 0.925 in the Odyssey scanning system and Pearl wide field imaging system, respectively. PAI is a highly promising methodology for increasing detection of tumors in fluorescence-guided surgery. Although not yet clinically approved, IRDye 680LT demonstrates promise as an untargeted agent when paired with ABY-029. The clinical translation of PAI to maximize tumor excision, while minimizing normal tissue removal, could improve both patient survival and life quality.

Nanoparticles modified with vasculature-homing peptides for targeted cancer therapy and angiogenesis imaging

The two major challenges in cancer treatment include lack of early detection and ineffective therapies with various side effects. Angiogenesis is the key process in the growth, survival, invasiveness, and metastasis of many of cancerous tumors. Imaging of the angiogenesis could lead to diagnosis of tumors in the early stage and evaluation of the therapeutic responses. Angiogenic blood vessels express specific molecular markers different from normal blood vessels (in level or kind). This fact would make the tumor vasculature a suitable site to target therapeutics and imaging agents within the tumor. Surface modified nanoparticles using peptide ligands with high binding affinity to the vasculature markers, provide efficient delivery of therapeutic and imaging agents, while avoiding undesirable side effects. In this review, we discuss discoveries of various tumor targeting peptides useful for tumor angiogenesis imaging and targeted therapy with emphasis on surface modified nanomedicines using vasculature targeting peptides.

Current State of Breast Cancer Diagnosis, Treatment, and Theranostics.

Breast cancer is one of the leading causes of cancer-related morbidity and mortality in women worldwide. Early diagnosis and effective treatment of all types of cancers are crucial for a positive prognosis. Patients with small tumor sizes at the time of their diagnosis have a significantly higher survival rate and a significantly reduced probability of the cancer being fatal. Therefore, many novel technologies are being developed for early detection of primary tumors, as well as distant metastases and recurrent disease, for effective breast cancer management. Theranostics has emerged as a new paradigm for the simultaneous diagnosis, imaging, and treatment of cancers. It has the potential to provide timely and improved patient care via personalized therapy. In nanotheranostics, cell-specific targeting moieties, imaging agents, and therapeutic agents can be embedded within a single formulation for effective treatment. In this review, we will highlight the different diagnosis techniques and treatment strategies for breast cancer management and explore recent advances in breast cancer theranostics. Our main focus will be to summarize recent trends and technologies in breast cancer diagnosis and treatment as reported in recent research papers and patents and discuss future perspectives for effective breast cancer therapy.

A Novel Fluorescent c-Met Targeted Imaging Agent for Intra-Operative Colonic Tumour Mapping: Translation from the Laboratory into a Clinical Trial

Background: Curative resection of colon cancer requires accurate intraoperative identification of tumour margins and lymph node (LN) burden. The c-Met protein is important for cell differentiation and migration and is overexpressed in many gastrointestinal cancers. We explored EMI-137, a novel c-Met targeting fluorescent probe, for application in fluorescence-guided colon surgery, in HT-29 colorectal cancer (CRC) cell line and an in vivo murine model. This informed a first-in-man clinical trial of EMI-137. Methods: HT-29 cells were used as a representative CRC model. SiRNA was used to demonstrate the specificity of EMI-137 for c-Met. A HT-29 CRC xenograft model was developed in female BALB/c mice. Tumours were cultured for 14 days to 10mm diameter. EMI-137 (0.18mg/kg) was injected into tail veins and biodistribution analysed through in vivo fluorescent imaging. Nine patients, aged 67-77 years, received a single intravenous dose of EMI-137 (0.13mg/kg), 1-3 hours before laparoscopic-assisted colon cancer resection surgery (NCT03360461). Tumour and LN fluorescence was assessed with a modified Karl Storz® laparoscope. Immunohistochemistry with a monoclonal antibody analysed c-Met expression in eight resected samples. Findings: c-Met expression in HT-29 cells was effectively silenced and imaged with EMI-137. Strong EMI-137 uptake in tumour xenografts was observed for up to 6 hours post-administration. At clinical trial, no serious adverse events related to EMI-137 were reported. Marked background fluorescence was observed in all trial participants, 4/9 showed an increase in tumour fluorescence over background; 5/9 had histological LN metastases; no fluorescent LN were detected intraoperatively. All primary tumours (8/8) and malignant LN (15/15) exhibited moderate to high c-Met protein expression. Interpretation: EMI-137, binds specifically to the human c-Met transmembrane protein, is safe in patients and with further refinement, shows potential for long-term application in fluorescence-guided colon surgery. Trial Registration: The trial was registered with ClinicalTrials.gov identifier NCT03360461. Funding: Research was supported by internal University of Leeds funding. Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: Ethical approval was granted by the Yorkshire and Humber Leeds West Research and Ethics Committee (Ref 17/YH/0263), on 19th October 2017 and Regulatory approval was granted by the Medicines and Healthcare products Regulatory Agency (MHRA), under The Medicines for Human Use (Clinical Trials) regulations on 29th August 2017.

Radiolabeled curcumin as β amyloid imaging and tumor targeting imaging agents

Curcumin, a polyphenolic compound, derived from the rhizomes of Curcuma longa L. Curcumin shows potential pharmacological action against numerous disorders, including cancer, neurodegenerative, and infection diseases. Curcumin-based molecular imaging agents could be useful for early detection of Alzheimer Disease and tumor and monitor the progress of therapy. Radiolabeled curcumin and its derivatives become promising compounds as imaging agents. In this review, radiolabeled curcumin bearing radionuclides including fluorine-18, Technetium-99m, Iodine-125, and Gallium-68 are reviewed as an effort to develop curcumin-based probes not only for β amyloid imaging but also for tumor imaging.

USPIOs as targeted contrast agents in cardiovascular magnetic resonance imaging.

We aim to discuss the diagnostic use of ultra-small superparamagnetic iron oxide (USPIOs) including ferumoxytol in targeted cardiovascular magnetic resonance imaging (MRI). Ferumoxytol is the only USPIO clinically available in the U.S. and is a negatively charged USPIO that has potential use for tracking and characterization of macrophage-infiltrated cardiovascular structures. As an iron supplement that is approved for treatment of iron deficiency anemia, the iron core of ferumoxytol is incorporated into the body once it is phagocytosed by macrophages. In organs or tissues with high inflammatory cellular infiltration, such as atherosclerotic plaques and myocardial infarction, localization of iron-laden macrophages can be visualized on delayed MRI. The iron core of ferumoxytol alters the magnetic susceptibility and results in shortening of T2* and T2 relaxation rates. Areas with high concentration appear hypointense (negative contrast) on T2 and T2* MRI. Recently, in vitro findings support the potential specificity of ferumoxytol interactions with macrophage subtypes, which has implications for therapeutic interventions. With increasing concerns about gadolinium retention in the brain and other tissues, the value of ferumoxytol-enhanced MR for targeted clinical imaging is aided by its positive safety profile in patients with impaired renal function. This paper discusses pharmacokinetic properties of USPIOs with a focus on ferumoxytol, and summarizes relevant in vitro, animal, and human studies investigating the diagnostic use of USPIOs in targeted contrast-enhanced imaging. We also discuss future directions for USPIOs as targeted imaging agents and associated challenges.

Rose bengal conjugated gadolinium complex as a new multimodal imaging agent targeting presynaptic vesicular glutamate transporters

Glutamate is an important excitatory neurotransmitter, and vesicular glutamate transporters (VGLUTs) are critical in regulating the extent of glutamate release. Because initial increase and late stage decrease in VGLUT expression were implicated in progressive Alzheimer’s disease (AD), VGLUTs can be a new imaging target for diagnosing AD. In this study, we have designed and synthesized a new multimodal gadolinium contrast agent conjugated with rose bengal to target VGLUTs (Gd-DO3A-RB). In addition to high kinetic and pH stability, magnetic resonance and fluorescence imaging using Gd-DO3A-RB showed high affinity to VGLUT1. Using the AD animal model (5XFAD), in vivo MRI with a Gd-DO3A-RB showed less MR signal enhancement in 5XFAD mouse brain than that of age-matched normal C57BL/6 mouse, suggesting a late stage decrease in VGLUT expression in AD animal model. Therefore, this new VGLUT targeting gadolinium contrast agent demonstrated a strong potential as a new diagnostic tool for AD with multimodal imaging capability.

18F-C2Am: a targeted imaging agent for detecting tumor cell death in vivo using positron emission tomography

IntroductionTrialing novel cancer therapies in the clinic would benefit from imaging agents that can detect early evidence of treatment response. The timing, extent and distribution of cell death in tumors following treatment can give an indication of outcome. We describe here an 18F-labeled derivative of a phosphatidylserine-binding protein, the C2A domain of Synaptotagmin-I (C2Am), for imaging tumor cell death in vivo using PET.MethodsA one-pot, two-step automated synthesis of N-(5-[18F]fluoropentyl)maleimide (60 min synthesis time, > 98% radiochemical purity) has been developed, which was used to label the single cysteine residue in C2Am within 30 min at room temperature. Binding of 18F-C2Am to apoptotic and necrotic tumor cells was assessed in vitro, and also in vivo, by dynamic PET and biodistribution measurements in mice bearing human tumor xenografts treated with a TRAILR2 agonist or with conventional chemotherapy. C2Am detection of tumor cell death was validated by correlation of probe binding with histological markers of cell death in tumor sections obtained immediately after imaging.Results18F-C2Am showed a favorable biodistribution profile, with predominantly renal clearance and minimal retention in spleen, liver, small intestine, bone and kidney, at 2 h following probe administration. 18F-C2Am generated tumor-to-muscle (T/m) ratios of 6.1 ± 2.1 and 10.7 ± 2.4 within 2 h of probe administration in colorectal and breast tumor models, respectively, following treatment with the TRAILR2 agonist. The levels of cell death (CC3 positivity) following treatment were 12.9–58.8% and 11.3–79.7% in the breast and colorectal xenografts, respectively. Overall, a 20% increase in CC3 positivity generated a one unit increase in the post/pre-treatment tumor contrast. Significant correlations were found between tracer uptake post-treatment, at 2 h post-probe administration, and histological markers of cell death (CC3: Pearson R = 0.733, P = 0.0005; TUNEL: Pearson R = 0.532, P = 0.023).ConclusionThe rapid clearance of 18F-C2Am from the blood pool and low kidney retention allowed the spatial distribution of cell death in a tumor to be imaged during the course of therapy, providing a rapid assessment of tumor treatment response. 18F-C2Am has the potential to be used in the clinic to assess early treatment response in tumors.

A novel molecular magnetic resonance imaging agent targeting activated leukocyte cell adhesion molecule as demonstrated in mouse brain metastasis models

Molecular magnetic resonance imaging (MRI) allows visualization of biological processes at the molecular level. Upregulation of endothelial ALCAM (activated leukocyte cell adhesion molecule) is a key element for leukocyte recruitment in neurological disease. The aim of this study, therefore, was to develop a novel molecular MRI contrast agent, by conjugating anti-ALCAM antibodies to microparticles of iron oxide (MPIO), for detection of endothelial ALCAM expression in vivo. Binding specificity of ALCAM-MPIO was demonstrated in vitro under static and flow conditions. Subsequently, in a proof-of-concept study, mouse models of brain metastasis were induced by intracardial injection of brain-tropic human breast carcinoma, lung adenocarcinoma or melanoma cells to upregulate endothelial ALCAM. At selected time-points, mice were injected intravenously with ALCAM-MPIO, and ALCAM-MPIO induced hypointensities were observed on T2*-weighted images in all three models. Post-gadolinium MRI confirmed an intact blood-brain barrier, indicating endoluminal binding. Correlation between endothelial ALCAM expression and ALCAM-MPIO binding was confirmed histologically. Statistical analysis indicated high sensitivity (80–90%) and specificity (79–83%) for detection of endothelial ALCAM in vivo with ALCAM-MPIO. Given reports of endothelial ALCAM upregulation in numerous neurological diseases, this advance in our ability to image ALCAM in vivo may yield substantial improvements for both diagnosis and targeted therapy.

Visualization of Activated T Cells by OX40-ImmunoPET as a Strategy for Diagnosis of Acute Graft-versus-Host Disease.

Graft-versus-host disease (GvHD) is a major complication of allogeneic hematopoietic cell transplantation (HCT), mediated primarily by donor T cells that become activated and attack host tissues. Noninvasive strategies detecting T-cell activation would allow for early diagnosis and possibly more effective management of HCT recipients. PET imaging is a sensitive and clinically relevant modality ideal for GvHD diagnosis, and there is a strong rationale for the use of PET tracers that can monitor T-cell activation and expansion with high specificity. The TNF receptor superfamily member OX40 (CD134) is a cell surface marker that is highly specific for activated T cells, is upregulated during GvHD, and mediates disease pathogenesis. We recently reported the development of an antibody-based activated T-cell imaging agent targeting OX40. In the present study, we visualize the dynamics of OX40 expression in an MHC-mismatch mouse model of acute GvHD using OX40-immunoPET. This approach enabled visualization of T-cell activation at early stages of disease, prior to overt clinical symptoms with high sensitivity and specificity. This study highlights the potential utility of the OX40 PET imaging as a new strategy for GvHD diagnosis and therapy monitoring. SIGNIFICANCE: OX40-immunoPET imaging is a promising noninvasive strategy for early detection of GvHD, capable of detecting signs of GvHD pathology even prior to the development of overt clinical symptoms.