Gamma scintigraphy imaging of murine invasive pulmonary aspergillosis with a 111In-labeled cyclic peptide☆
Introduction
Invasive pulmonary aspergillosis (IPA) is an important cause of morbidity and mortality in immunocompromised patients [1], [2], [3], [4]. As the pathogenesis of IPA involves inhalation of airborne conidia by susceptible hosts, pneumonia is the most common clinical manifestation of IPA [5], [6]. In high-risk patients (e.g., those with acute leukemia or those who have received a bone marrow transplant), IPA is associated with a mortality rate of 42–83%, despite the administration of systemic antifungal therapy [7], [8]. The high mortality rate associated with IPA can be attributed to the immunosuppression of the affected patients, the typically late diagnosis and the suboptimal in vivo efficacy of antifungal agents against Aspergillus species. Thus, early detection and early administration of antifungal agents, i.e., when the tissue fungal burden is relatively low, may help improve the outcomes of patients with IPA. Because of the high mortality rate of Aspergillus infection, there is an urgent need to develop new strategies for the early diagnosis of IPA.
In cases of suspected pulmonary infection, computed tomography (CT) provides high-quality anatomic information, but in the absence of structural morphological changes in the lungs, it is difficult to diagnose a pulmonary infection in its early stages by CT alone. Other diagnostic methods, e.g., fungal culturing and non–culture-based methods such as serodiagnosis or polymerase chain reaction, may also be used [5], [9], [10], [11]. The sensitivity and specificity of these methods, however, remain suboptimal. Nuclear imaging techniques may be a valuable alternative in the diagnosis of IPA. Several nuclear imaging techniques have been evaluated for use in the diagnosis of fungal infections. In one approach, diagnosis is made based on structural and physiological changes in the lung induced by the invading microorganisms. An example of this approach is the injection and scanning of 67Ga-citrate, which binds to circulating transferrin and extravasates at the site of infection because of the increased vascular permeability [12]. Radiolabeled white blood cells (WBC) that migrate toward and infiltrate the inflammatory and infectious lesions have also been used for imaging fungal infections. WBC can be labeled either ex vivo [13], [14] or in vivo [15], [16]. In vivo labeling of WBC can be achieved using 99mTc-labeled antigranulocyte antibody tracers [17], [18]; 99mTc-labeled interleukin-8 (IL-8), which binds to IL-8 receptors [15]; or 111In-labeled leukotriene B4 (LTB4) antagonist, which targets LTB4 receptors [16]. However, because in vivo cell labeling depends on radiolabeled materials' attaching to receptors expressed on neutrophils, this approach might have limited utility in the diagnosis of IPA in severely neutropenic patients. [18F]Fluoro-2-deoxy-d-glucose (18F-FDG) accumulates avidly in metabolically active inflammatory cells [19]. Recent evidence suggests that 18F-FDG positron emission tomography (PET) imaging could be a useful tool in the diagnosis and management of opportunistic infections, including fungal infections in immunocompromised patients [20], [21]. However, diagnosis of fungal infection with 18F-FDG PET is not specific and is prone to errors (i.e., patients are often misdiagnosed as having a malignancy) [22].
An alternative approach for nuclear imaging of fungal infection, which may offer better detection specificity as compared to the methods described above, exploits the differences between fungi and normal host tissues or with bacteria. In this study, we investigated a 111In-labeled cyclic peptide that directly targets A. fumigatus in a clinically relevant model of IPA. The cyclic peptide c(CGGRLGPFC)-NH2 was identified through bacteriophage display technology and was found to bind in vitro to the surface of conidia and hyphae of A. fumigatus [23]. Radiolabeled peptides are promising nuclear imaging agents because of their pharmacokinetic properties, rapid binding and relatively low immunogenicity [24]. Our data suggest that 111In-labeled c(CGGRLGPFC)-NH2 can selectively accumulate in infected lungs; therefore it may facilitate diagnostic imaging of A. fumigatus infection.
Section snippets
Materials
All Nα-Fmoc amino acids, 1-hydroxybenzotriazole (HOBt), diisopropylcarbodiimide (DIC), triisopropylsilane (TIS) and Fmoc-Rink linker, were purchased from Novabiochem (San Diego, CA, USA). N,N-Diisopropylethylamine, trifluoroacetic acid (TFA), ethylenediaminetetraacetic acid (EDTA), cyclophosphamide and cortisone acetate were purchased from Sigma-Aldrich Chemical (St. Louis, MO, USA). Aminobenzyl diethylenetriaminepentaacetic acid (DTPA-Bz-NH2) was obtained from Macrocyclics (Dallas, TX, USA).
Radiolabeling
DTPA-c(CGGRLGPFC)-NH2 was labeled with 111InCl3 in 0.1 M NaAC buffer at pH 5 (Fig. 1). Radiolabeling efficiency was 98.8±0.55%. Maximum specific activity was 8.5×1010 MBq/mol (74 MBq/μg). Fig. 2 shows the radiochromatograph and the UV/Vis chromatograph of 111In-DTPA-c(CGGRLGPFC)-NH2.
Stability of 111In-DTPA-c(CGGRLGPFC)-NH2
Negligible radioactivity was lost from 111In-DTPA-c(CGGRLGPFC)-NH2 after 4-h incubation in PBS containing either EDTA or human serum. Even after 24 h of incubation, greater than 97% of radioactivity was associated
Discussion
Our findings suggest that imaging of A. fumigatus in the lungs is feasible using 111In-DTPA-c(CGGRLGPFC)-NH2. c(CGGRLGPFC)-NH2 could be labeled with 111In through the DTPA chelator with high radiolabeling efficiency and high stability. Using an established murine model of IPA, we found that 111In-DTPA-c(CGGRLGPFC)-NH2 was selectively localized to lungs infected with A. fumigatus. The uptake of the radiotracer in the lungs of the infected mice was more than twice that in the lungs of the healthy
Conclusions
This study is the first to demonstrate the feasibility of specific nuclear imaging of fungal infection in a murine model of IPA using a radiolabeled peptide, c(CGGRLGPFC)-NH2, selected from a bacteriophage display library. The ability of this small cyclic peptide to accumulate in and delineate A. fumigatus colonies in the lungs emphasizes the specificity of its binding. Future work is needed to optimize its imaging properties through peptide mimetics approach, to determine the immunogenicity of
Acknowledgments
The authors thank Dr. Juri Gelovani for helpful discussion, and Lionel Santibañez for editing this manuscript.
References (35)
- et al.
Invasive pulmonary aspergillosis in solid organ and bone marrow transplant recipients
Transplant Proc
(2005) - et al.
Usefulness of sequential aspergillus galactomannan antigen detection combined with early radiologic evaluation for diagnosis of invasive pulmonary aspergillosis in patients undergoing allogeneic stem cell transplantation
Transplant Proc
(2006) - et al.
99mTc-labeled interleukin-8 for scintigraphic detection of pulmonary infections
Chest
(2004) - et al.
Characterization of uptake of 2-deoxy-2-[18F]-fluoro-d-glucose by fungal-associated inflammation: the standardized uptake value is greater for lesions of blastomycosis than for lymphoma in dogs with naturally occurring disease
Mol Imaging Biol
(2002) - et al.
The use of phage display for the development of tumour targeting agents
Adv Drug Deliv Rev
(2000) - Marr KA, Patterson T, Denning D. Aspergillosis. Pathogenesis, clinical manifestations, and therapy. Infect Dis Clin...
- et al.
Nosocomial pneumonia in patients having bone marrow transplant. Attributable mortality and risk factors
Cancer
(1992) - et al.
Multidimensional volumetric imaging of pulmonary infiltrates for measuring therapeutic response to antifungal therapy in experimental invasive pulmonary aspergillosis
Antimicrob Agents Chemother
(2006) - et al.
Epidemiology of aspergillus infections in a large cohort of patients undergoing bone marrow transplantation
J Infect Dis
(1997) - et al.
Invasive aspergillosis in 2002: an update
Eur J Clin Microbiol Infect Dis
(2002)
Aspergillus fumigatus and aspergillosis
Clin Microbiol Rev
Invasive pulmonary aspergillosis following bone marrow transplantation: risk factors and diagnostic aspect
Haematologia (Budap)
Invasive fungal infections; evolving challenges for diagnosis and therapeutics
Mol Immunol
Development and optimization of quantitative pcr for the diagnosis of invasive aspergillosis with bronchoalveolar lavage fluid
BMC Infect Dis
Mechanism of gallium-67 accumulation in inflammatory lesions
J Nucl Med
Combined labeled leukocyte and technetium-99 m sulfur colloid bone marrow imaging for diagnosing musculoskeletal infection
Radiographics
Inflammation and infection: imaging properties of 18F-FDG-labeled white blood cells versus 18f-fdg
J Nucl Med
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This research was supported in part by John S. Dunn Foundation.
- 1
On leave from the Department of Nuclear Medicine, Peking University School of Oncology, Beijing Cancer Hospital, Beijing 100142, China, and from the Isotope Department, China Institute of Atomic Energy, Beijing 102413, China.
- 2
These authors contributed equally to this work.