title: Detection of Aspergillus fumigatus pulmonary fungal infections in mice with (99m)Tc-labeled MORF oligomers targeting ribosomal RNA.
abstract: OBJECTIVE
  Invasive aspergillosis is a major cause of infectious morbidity and mortality in immunocompromised patients. The fungus Aspergillus fumigatus (A. fumigatus) is the primary causative agent of invasive aspergillosis. However, A. fumigatus infections remain difficult to diagnose particularly in the early stages due to the lack of a rapid, sensitive and specific diagnostic approach. In this study, we investigated (99m)Tc labeled MORF oligomers targeting fungal ribosomal RNA (rRNA) for the imaging detection of fungal infections.
  
  
  METHODS
  Three phosphorodiamidate morpholino (MORF) oligomer (a DNA analogue) probes were designed: AGEN, complementary to a sequence of the fungal 28S ribosomal RNA (rRNA) of Aspergillus, as a genus-specific probe; AFUM, complementary to the 28S rRNA sequence of A. fumigatus, as a fungus species-specific probe; and cMORF, irrelevant to all fungal species, as a control probe. The probes were conjugated with Alexa Fluor 633 carboxylic acid succinimidyl ester (AF633) for fluorescence imaging or with NHS-mercaptoacetyl triglycine (NHS-MAG3) for nuclear imaging with (99m)Tc and then evaluated in vitro and in vivo.
  
  
  RESULTS
  The specific binding of AGEN and AFUM to fungal total RNA was confirmed by dot blot hybridization while specific binding of AGEN and AFUM in fixed and live A. fumigatus was demonstrated by both fluorescent in situ hybridization (FISH) analysis and accumulation in live cells. SPECT imaging of BALB/c mice with pulmonary A. fumigatus infections and administered (99m)Tc labeled AGEN and AFUM showed immediate and obvious accumulation in the infected lungs, while no significant accumulation of the control (99m)Tc-cMORF in the infected lung was observed. Compared to non-infected mice, with sacrifice at 1h, the accumulation of (99m)Tc-AGEN and (99m)Tc-AFUM in the lungs of mice infected with A. fumigatus was 2 and 2.7 fold higher respectively.
  
  
  CONCLUSIONS
  In vivo targeting fungal ribosomal RNA with (99m)Tc labeled MORF probes AGEN and AFUM may be useful for A. fumigatus infection imaging and may provide a new strategy for the noninvasive diagnosis of invasive aspergillosis and other fungal infections.
authors.affiliation: Division of Nuclear Medicine, Department of Radiology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA. Yuzhen.Wang@umassmed.edu
authors.forename: Yuzhen
authors.initials: Y
authors.lastname: Wang
authors.forename: Ling
authors.initials: L
authors.lastname: Chen
authors.forename: Xinrong
authors.initials: X
authors.lastname: Liu
authors.forename: Dengfeng
authors.initials: D
authors.lastname: Cheng
authors.forename: Guozheng
authors.initials: G
authors.lastname: Liu
authors.forename: Yuxia
authors.initials: Y
authors.lastname: Liu
authors.forename: Shuping
authors.initials: S
authors.lastname: Dou
authors.forename: Donald J
authors.initials: DJ
authors.lastname: Hnatowich
authors.forename: Mary
authors.initials: M
authors.lastname: Rusckowski
mesh_terms: D000818:Animals; D001232:Aspergillus fumigatus / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D008168:Lung / Q000000981:diagnostic imaging / Q000382:microbiology; D008172:Lung Diseases, Fungal / Q000000981:diagnostic imaging*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D060172:Morpholinos / Q000235:genetics / Q000378:metabolism* / Q000493:pharmacokinetics; D064847:Multimodal Imaging; D015345:Oligonucleotide Probes / Q000235:genetics / Q000378:metabolism / Q000493:pharmacokinetics; D015609:Organotechnetium Compounds; D049268:Positron-Emission Tomography; D012331:RNA, Fungal / Q000378:metabolism*; D012339:RNA, Ribosomal, 28S / Q000378:metabolism*; D013045:Species Specificity; D013172:Spores, Fungal / Q000235:genetics / Q000502:physiology; D014057:Tomography, X-Ray Computed
chemical_list: D060172:Morpholinos; D015345:Oligonucleotide Probes; D015609:Organotechnetium Compounds; D012331:RNA, Fungal; D012339:RNA, Ribosomal, 28S