Molecular imaging modalities hold great potential as less invasive techniques for diagnosis and management of various diseases. Molecular imaging combines imaging agents with targeting moieties to specifically image diseased sites in the body. Monoclonal antibodies (mAbs) have become increasingly popular as novel therapeutics against a variety of diseases due to their specificity, affinity and serum stability. Because of the same properties, mAbs are also exploited in molecular imaging to target imaging agents such as radionuclides to the cell of interest in vivo. Many studies investigated the use of mAb-targeted imaging for a variety of purposes, for instance to monitor disease progression and to predict response to a specific therapeutic agent. Herein, we highlighted the application of mAb-targeted imaging in three different types of pathologies: autoimmune diseases, oncology and cardiovascular diseases. We also described the potential of molecular imaging strategies in theranostics and precision medicine. Due to the nearly infinite repertoire of mAbs, molecular imaging can change the future of modern medicine by revolutionizing diagnostics and response prediction in practically any disease.

T cell modulation in the clinical background of autoimmune diseases or allogeneic cell and organ transplantations with concurrent preservation of their natural immunological functions (e.g., pathogen defense) is the major obstacle in immunology. An anti-human CD4 antibody (MAX.16H5) was applied intravenously in clinical trials for the treatment of autoimmune diseases (e.g., rheumatoid arthritis) and acute late-onset rejection after transplantation of a renal allograft. The response rates were remarkable and no critical allergic problems or side effects were obtained. During the treatment of autoimmune diseases with the murine MAX.16H5 IgG1 antibody its effector mechanisms with effects on lymphocytes, cytokines, laboratory and clinical parameters, adverse effects as well as pharmacodynamics and kinetics were studied in detail. However, as the possibility of developing immune reactions against the murine IgG1 Fc-part remains, the murine antibody was chimerized, inheriting CD4-directed variable domains of the MAX.16H5 IgG1 connected to a human IgG4 backbone. Both antibodies were studied in vitro and in specific humanized mouse transplantation models in vivo with a new scope. By ex vivo incubation of an allogeneic immune cell transplant with MAX.16H5 a new therapy strategy has emerged for the first time enabling both the preservation of the graft-vs.-leukemia (GVL) effect and the permanent suppression of the acute graft-vs.-host disease (aGVHD) without conventional immunosuppression. In this review, we especially focus on experimental data and clinical trials obtained from the treatment of autoimmune diseases with the murine MAX.16H5 IgG1 antibody. Insights gained from these trials have paved the way to better understand the effects with the chimerized MAX.16H5 IgG4 as novel therapeutic approach in the context of GVHD prevention.

Chronic inflammatory diseases are often progressive, resulting not only in physical damage to patients but also social and economic burdens, making early diagnosis of them critical. Nuclear medicine techniques can enhance the detection of inflammation by providing functional as well as anatomical information when combined with other modalities such as magnetic resonance imaging, computed tomography or ultrasonography. Although small molecules and peptides were mainly used for the treatment and imaging of chronic inflammatory diseases in the past, antibodies and their fragments have also been emerging for chronic inflammatory diseases as they show high specificity to their targets and can have various biological half-lives depending on how they are engineered. In addition, imaging with antibodies or their fragments can visualize the in vivo biodistribution of the probes or help monitor therapeutic responses, thereby providing physicians with a greater understanding of drug behavior in vivo and another means of monitoring their patients. In this review, we introduce various targets and radiolabeled antibody-based probes for the molecular imaging of chronic inflammatory diseases in preclinical and clinical studies. Targets can be classified into three different categories: 1) cell-adhesion molecules, 2) surface markers on immune cells, and 3) cytokines or enzymes. The limitations and future directions of using radiolabeled antibodies for imaging inflammatory diseases are also discussed.

Molecular imaging of CD4⁺ T cells throughout the body has implications for monitoring autoimmune disease and immunotherapy of cancer. Given the key role of these cells in regulating immunity, it is important to develop a biologically inert probe. GK1.5 cys-diabody (cDb), a previously developed anti-mouse CD4 antibody fragment, was tested at different doses to assess its effects on positron emission tomography (PET) imaging and CD4⁺ T cell viability, proliferation, CD4 expression, and function.

The effect of protein dose on image contrast (lymphoid tissue-to-muscle ratio) was assessed by administering different amounts of ⁸⁹Zr-labeled GK1.5 cDb to mice followed by PET imaging and ex vivo biodistribution analysis. To assess impact of GK1.5 cDb on T cell biology, GK1.5 cDb was incubated with T cells in vitro or administered intravenously to C57BL/6 mice at multiple protein doses. CD4 expression and T cell proliferation were analyzed with flow cytometry and cytokines were assayed.

For immunoPET imaging, the lowest protein dose of 2 μg of ⁸⁹Zr-labeled GK1.5 cDb resulted in significantly higher % injected dose/g in inguinal lymph nodes (ILN) and spleen compared to the 12-μg protein dose. In vivo administration of GK1.5 cDb at the high dose of 40 μg caused a transient decrease in CD4 expression in spleen, blood, lymph nodes, and thymus, which recovered within 3 days postinjection; this effect was reduced, although not abrogated, when 2 μg was administered. Proliferation was inhibited in vivo in ILN but not the spleen by injection of 40 μg GK1.5 cDb. Concentrations of GK1.5 cDb in excess of 25 nM significantly inhibited CD4⁺ T cell proliferation and interferon-γ production in vitro. Overall, using low-dose GK1.5 cDb minimized biological effects on CD4⁺ T cells.

Low-dose GK1.5 cDb yields high-contrast immunoPET images with minimal effects on T cell biology in vitro and in vivo and may be a useful tool for investigating CD4⁺ T cells in the context of preclinical disease models. Future approaches to minimizing biological effects may include the creation of monovalent fragments or selecting anti-CD4 antibodies which target alternative epitopes.

Infection is ubiquitous. However, its management is challenging for both the patients and the health-care providers. Scintigraphic imaging of infection dates back nearly half a century. The advances in our understanding of the pathophysiology of disease at cellular and molecular levels have paved the way to the development of a large number of radiopharmaceuticals for scintigraphic imaging of infection. These include radiolabeling of blood elements such as serum proteins, white blood cells (WBCs), and cytokines, to name a few. Infectious foci have also been imaged using a radiolabeled sugar molecule by taking advantage of increased metabolic activity in the infectious lesions. Literature over the years has well documented that none of the radiopharmaceuticals and associated procedures that facilitate imaging infection are flawless and acceptable without a compromise. As a result, only a few compounds such as 99mTc-hexamethylpropyleneamineoxime, 18F-FDG, the oldest but still considered as a gold standard 111In-oxine, and, yes, even 67Ga-citrate in some countries, have remained in routine clinical practice. Nonetheless, the interest of scientists and physicians to improve the approaches to imaging and to the management of infection is noteworthy. These approaches have paved the way for the development of numerous, innovative radiopharmaceuticals to label autologous WBCs ex vivo or even those that could be injected directly to image infection or inflammation without direct involvement of WBCs. In this review, we briefly describe these agents with their pros and cons and place them together for future reference.

Early diagnosis and effective monitoring of rheumatoid arthritis (RA) are important for a positive outcome. Instant treatment often results in faster reduction of inflammation and, as a consequence, less structural damage. Anatomical imaging techniques have been in use for a long time, facilitating diagnosis and monitoring of RA. However, mere imaging of anatomical structures provides little information on the processes preceding changes in synovial tissue, cartilage, and bone. Molecular imaging might facilitate more effective diagnosis and monitoring in addition to providing new information on the disease pathogenesis. A limiting factor in the development of new molecular imaging techniques is the availability of suitable probes. Here, we review which cells and molecules can be targeted in the RA joint and discuss the advances that have been made in imaging of arthritis with a focus on such molecular targets as folate receptor, F4/80, macrophage mannose receptor, E-selectin, intercellular adhesion molecule-1, phosphatidylserine, and matrix metalloproteinases. In addition, we discuss a new tool that is being introduced in the field, namely the use of nanobodies as tracers. Finally, we describe additional molecules displaying specific features in joint inflammation and propose these as potential new molecular imaging targets, more specifically receptor activator of nuclear factor κB and its ligand, chemokine receptors, vascular cell adhesion molecule-1, αVβ₃ integrin, P2X7 receptor, suppression of tumorigenicity 2, dendritic cell-specific transmembrane protein, and osteoclast-stimulatory transmembrane protein.

T-cell-located CD4 antigen represents one of the therapeutic targets in rheumatoid arthritis (RA). However, up to now there is no established imaging tool to visualize this target in vivo. The aim of our study was to assess the safety and tolerability of a technetium-99m labelled murine anti-human CD4 IgG1-Fab fragment ([(99m)Tc]-anti-CD4-Fab, [(99m)Tc]-EP1645) in patients with active synovitis due to RA, and to evaluate its potential as a marker of disease activity.

In the present phase I proof of principle study five patients with RA were examined. Planar scans of the whole body, hands, and feet were taken 30 min up to 24h after application of 550 ± 150 MBq [(99m)Tc]-anti-CD4-Fab, followed by visual analyses, comparison with clinical data in 68 joints per patient and semiquantitative analysis of hand and wrist joints.

Neither infusion related adverse events nor adverse events during follow up were observed. No increase in human anti-murine antibody titres was seen. All patients had positive scans in almost 70% of clinically affected joints. Positive scans were also found in 8% of joints without evidence of swelling or tenderness.

Scintigraphy with [(99m)Tc]-anti-CD4-Fab is a promising technique for evaluation of inflammatory activity in patients with RA, pre-therapeutical evaluation of CD4 status and therapy control. Tracer uptake in clinically inconspicuous joints strongly indicates diagnostic potential of [(99m)Tc]-anti-CD4-Fab. Whether this technique is eligible as a prognostic factor in RA needs to be analysed in further studies as well as the pathophysiological background of clinically affected joints lacking tracer uptake.

The closing of the last century opened a wide variety of approaches for inflammation imaging and treatment of patients with rheumatoid arthritis (RA). The introduction of biological therapies for the management of RA started a revolution in the therapeutic armamentarium with the development of several novel monoclonal antibodies (mAbs), which can be murine, chimeric, humanised and fully human antibodies. Monoclonal antibodies specifically bind to their target, which could be adhesion molecules, activation markers, antigens or receptors, to interfere with specific inflammation pathways at the molecular level, leading to immune-modulation of the underlying pathogenic process. These new generation of mAbs can also be radiolabelled by using direct or indirect method, with a variety of nuclides, depending upon the specific diagnostic application. For studying rheumatoid arthritis patients, several monoclonal antibodies and their fragments, including anti-TNF-alpha, anti-CD20, anti-CD3, anti-CD4 and anti-E-selectin antibody, have been radiolabelled mainly with (99m)Tc or (111)In. Scintigraphy with these radiolabelled antibodies may offer an exciting possibility for the study of RA patients and holds two types of information: (1) it allows better staging of the disease and diagnosis of the state of activity by early detection of inflamed joints that might be difficult to assess; (2) it might provide a possibility to perform 'evidence-based biological therapy' of arthritis with a view to assessing whether an antibody will localise in an inflamed joint before using the same unlabelled antibody therapeutically. This might prove particularly important for the selection of patients to be treated since biological therapies can be associated with severe side-effects and are considerably expensive. This article reviews the use of radiolabelled mAbs in the study of RA with particular emphasis on the use of different radiolabelled monoclonal antibodies for therapy decision-making and follow-up.

Chronic musculoskeletal diseases such as arthritis, malignancy, and chronic injury and/or inflammation, all of which may produce chronic musculoskeletal pain, often pose challenges for current clinical imaging methods. The ability to distinguish an acute flare from chronic changes in rheumatoid arthritis, to survey early articular cartilage breakdown, to distinguish sarcomatous recurrence from posttherapeutic inflammation, and to directly identify generators of chronic pain are a few examples of current diagnostic limitations. There is hope that a growing field known as molecular imaging will provide solutions to these diagnostic puzzles. These techniques aim to depict, noninvasively, specific abnormal cellular, molecular, and physiologic events associated with these and other diseases. For example, the presence and mobilization of specific cell populations can be monitored with molecular imaging. Cellular metabolism, stress, and apoptosis can also be followed. Furthermore, disease-specific molecules can be targeted, and particular gene-related events can be assayed in living subjects. Relatively recent molecular and cellular imaging protocols confirm important advances in imaging technology, engineering, chemistry, molecular biology, and genetics that have coalesced into a multidisciplinary and multimodality effort. Molecular probes are currently being developed not only for radionuclide-based techniques but also for magnetic resonance (MR) imaging, MR spectroscopy, ultrasonography, and the emerging field of optical imaging. Furthermore, molecular imaging is facilitating the development of molecular therapies and gene therapy, because molecular imaging makes it possible to noninvasively track and monitor targeted molecular therapies. Implementation of molecular imaging procedures will be essential to a clinical imaging practice. With this in mind, the goal of the following discussion is to promote a better understanding of how such procedures may help address specific musculoskeletal issues, both now and in the years ahead.

Radiopharmaceuticals used for in vivo imaging of inflammatory conditions can be conveniently classified into six categories according to the different phases in which the inflammatory process develops. The trigger of an inflammatory process is a pathogenic insult (phase I) that causes activation of endothelial cells (phase II); there is then an increase of vascular permeability followed by tissue oedema (phase III). Phase IV is characterised by infiltration of polymorphonuclear cells, and a self-limiting regulatory process called apoptosis is observed (phase V). If the inflammatory process persists, late chronic inflammation takes place (phase VI). In some pathological conditions, such as organ-specific autoimmune diseases, chronic inflammation is present early in the disease. The aim of nuclear medicine in the field of inflammation/infection is to develop noninvasive tools for the in vivo detection of specific cells and tissues. This would allow early diagnosis of initial pathophysiological changes that are undetectable by clinical examination or by other diagnostic tools, and could also be used to evaluate the state of activity of the disease during therapy. These potential applications are of great interest in clinical practice. In this review, we describe the various approaches that have been developed in the last 25 years of experience. Recent advances in the diagnosis of inflammatory processes have led to the development of specific radiopharmaceuticals that are intended to allow specific stage-related diagnosis.

The aim of this study was to test the applicability of 99mTc-hexamethylpropylene amine oxime (99mTc-HMPAO) labelled leukocyte joint scintigraphy in the assessment of disease activity in 21 patients with rheumatoid arthritis, and to compare leukocyte scintigraphy with the Disease Activity Score (DAS), a validated activity index developed by the European League Against Rheumatism (EULAR). Twenty-one patients with rheumatoid arthritis were investigated by using 99mTc-HMPAO labelled leukocyte joint scintigraphy. The clinical and laboratory data were recorded, and the DAS was calculated and compared with the scintigraphic results in each case. A relatively high DAS score (4.71+/-1.07) was found in the majority of patients. The degree of accumulation of 99mTc-HMPAO leukocytes showed no correlation with a patient's age, gender, duration of disease, use of disease modifying anti-rheumatic drugs (DMARDs), visual analogue scale (VAS), Richie index, DAS, or any laboratory parameters. In contrast, a significant correlation was found between the global regional accumulation of the labelled leukocytes of the hands and feet, and the swollen-joint count. It is concluded that radiolabelled leukocyte scintigraphy could become one of the promising methods in the assessment of disease activity in patients with rheumatoid arthritis.

Radiopharmaceuticals have been used as investigative tools for the detection and treatment of arthritis activity in rheumatoid arthritis (RA) since the 1950s. Against the background of the pathophysiology of RA, the current status of joint scintigraphy and possible future developments are reviewed. Both non-specific (radiolabelled leucocytes and technetium-99m labelled human immunoglobulin) and specific targeting radiopharmaceuticals (including radiolabelled antibodies) are considered. The use of radiopharmaceuticals in the detection of arthritis activity has the advantages of allowing direct imaging of joints by means of whole-body scintigraphy and of joints that are difficult to assess clinically or radiographically. Promising results have been obtained with radiolabelled anti-CD4 and anti-E-selectin antibodies and with somatostatin receptor imaging, but more data are available regarding 99mTc-IgG scintigraphy, which differentiates between the various degrees of arthritis activity and thus facilitates the choice of antirheumatic drug. Newer promising approaches to the imaging of RA include the use of radiolabelled J001 and cytokines, though studies on these are limited at present.

To study the effect of chimeric anti-CD4 monoclonal antibody (MAb) therapy on synovial inflammation, in order to interpret the clinical experience with anti-CD4 treatment.

The immunohistologic features of synovial biopsy specimens before and 4 weeks after anti-CD4 MAb (cM-T412) therapy were studied in patients with rheumatoid arthritis. The patients received intravenous doses of either placebo (n = 1) or 10 mg (n = 4), 25 mg (n = 2), or 50 mg (n = 1) of cM-T412 daily for 5 consecutive days.

Although the patients did not experience clinical improvement, significant decreases in the number of circulating CD4+ cells, the degree of synovial inflammatory infiltration, and the mean scores for expression of adhesion molecules were found in the 7 patients 4 weeks after receiving cM-T412. The scores for infiltration with CD4+ and other inflammatory cells were particularly reduced following treatment with either 25 mg or 50 mg cM-T412. Cytokines, such as interleukin-1 beta and tumor necrosis factor alpha, could still be detected in the synovial tissue after treatment.

The decline in the numbers of inflammatory cells and adhesion molecules in synovial tissue after CD4+ cell depletion supports the view that CD4+ T cells orchestrate local cellular infiltration. The lack of clinical effect of anti-CD4 therapy might be explained by an insufficient decrease in the number of synovial CD4+ cells and by the persistence of cytokines. Determination of whether more adequate dosing would lead to a clinical improvement must await further study.

To study the ability of technetium-99m hexamethyl propylene amineoxime (HMPAO) labelled lymphocyte scintigraphy to quantify synovial inflammation, and to analyse the kinetics of lymphocyte retention in the joints of patients with rheumatoid arthritis (RA).

After isolation of the lymphocytes, the cells were radiolabelled in vitro with 250 MBq 99mTc-HMPAO. The scans were performed 30 minutes, three hours and 20 hours after injection.

An increase of the scintigram signal obtained at 20 hours was associated with a high joint swelling and joint pain score (F test = 3.07, p < 0.002), but not with the radiological score. A positive joint scintigram was predictive of active synovitis. Although the scintigram variation over time did not reach statistical significance, the kinetics of the scintigram signal tended to differ according to the disease duration: in early RA, active arthritis could be clearly imaged as early as 30 minutes, increased at three hours and the signal intensity persisted at 20 hours. In contrast, in long standing disease, the affected joints were imaged at 30 minutes, persisted unchanged at three hours, and the scintigram score decreased significantly at 20 hours.

The study shows that 99mTc-HMPAO joint scintigraphy may be used to detect and to localise active rheumatoid arthritis.

Over the last three years there has been a dramatic rise in the number of trials using monoclonal antibodies in the treatment of rheumatoid arthritis. So far, the numbers of patients treated in the individual studies have been small, and the study designs not comparable. All these trials have been conducted in a non-blinded, uncontrolled fashion. The patient populations tended to represent the severe end of the disease spectrum, being usually individuals for whom all other conventional and sometimes even unconventional experimental therapeutic approaches have failed. Clearly, therefore, larger controlled double blind studies in patients with less advanced stages of rheumatoid arthritis are needed. In the trials thus far, long-standing diseases afflicting the joints, usually with severe destruction, have frequently made clinical evaluation very difficult. Moreover, apparently with the exception of one or two reagents (16H5 and possibly B-F5) routine laboratory parameters which are helpful in determining disease activity such as CRP or the rheumatoid factor usually remain unaltered with anti-T cell therapy. In addition, in some individuals there was no clinical improvement despite sometimes severe CD4 cell depletion. The notion that the mere depletion of CD4+ cells is not sufficient to permanently suppress disease activity in autoimmune disease is further supported by studies carried out by Conolly and Wofsy in 1990. In a mouse lupus model, these investigators demonstrated that a small subpopulation of CD4+ T cells may be refractory to depletion by anti-CD4 and may be able to promote the full expression of the disease. Similar mechanisms could apply to certain individuals with human autoimmune disorders. Many additional questions remain open. The most important of these is which markers identify clinical responders to therapy. Attempts to correlate clinical response to the level of T cell depletion, modulation of the target antigens or in vitro functional assays so far have not yielded significant results. Other questions relate to the frequency of antibody administration and the amounts needed to permanently suppress disease activity. The initial hope based on animal experiments of inducing a permanent tolerance to certain antigens by anti-CD4 treatment has been clearly shown not to apply to rheumatoid arthritis. Even though there are individual variations, the efficacy of anti-T cell treatment tends to wear off after 3 or even 1 month, necessitating retreatment. Protocols will have to be designed for either longer treatment periods, repeated courses or more frequent single administrations.(ABSTRACT TRUNCATED AT 400 WORDS)

Monoclonal murine antibodies are increasingly used for immunotherapy and in vivo diagnostic procedures such as immunoscintigraphy. The therapeutic or diagnostic reagent however, is a foreign antigen, which may induce host reactivity. This may interfere with the therapeutic or diagnostic reagent in vivo, resulting in a loss of efficacy or the necessity to increase dosages. In addition, there is an important interference to in vitro immunoassays detecting specific antigens utilizing murine monoclonal antibodies. In the present study, sera of patients who had undergone a therapeutic trial using 140 mg of an anti-CD4 antibody, were investigated. Human anti-murine-immunoglobulin-antibodies (HAMA) were detected 2-3 weeks after treatment was started and reached maximal amounts of 0.8 micrograms/ml after a single and 2 micrograms/ml after a repeated treatment course. Parallely raised values of TSH were found in sera containing HAMAs of more than 0.3 micrograms/ml. Elevations of TSH levels up to 13 microU/ml were most pronounced after a repeated trial of the murine antibody and were detectable up to 20 weeks.