RT Journal Article SR Electronic T1 Molecular imaging of collagen remodeling in a murine model of lung fibrosis JF Journal of Nuclear Medicine JO J Nucl Med FD Society of Nuclear Medicine SP 242026 OP 242026 VO 65 IS supplement 2 A1 Ahmad, Azmi A1 Ghim, Mean A1 Kukreja, Gunjan A1 Neishabouri, Afarin A1 Zhang, Zhengxing A1 Li, Jie A1 Salarian, Mani A1 Toczek, Jakub A1 Gona, Kiran A1 Hedayatyanfard, Keshvad A1 Zhang, Jiasheng A1 Huang, Yiyun A1 Yu, Michael A1 Sadeghi, Mehran YR 2024 UL http://jnm.snmjournals.org/content/65/supplement_2/242026.abstract AB 242026 Introduction: Pulmonary fibrosis is the characteristic feature in interstitial lung disease. The current diagnostic methods for pulmonary fibrosis neither provide information on disease activity, nor distinguish between established disease and ongoing matrix remodeling. We hypothesized that targeting denatured collagen as a marker of collagen turnover could provide insight into the remodeling process. The goal of this study was to develop and validate a new collagen-hybridizing SPECT imaging probe to track fibrosis and its resolution in vivo in a murine model of pulmonary fibrosis.Methods: To access novel radiotracers that can target and hybridize to denatured collagen, we designed an imaging agent consisting of a poly-histidine (H6) or a poly-histidine-glutamic acid (HE)3 peptide connected to a N-terminal targeting moiety with 9 glycine-proline-hydroxyproline (GPO) repeats through a flexible 3 glycine linker. The peptide was labeled with 99mTc(CO)3+ and the unbound 99mTc was separated by Sep-Pak C18. The formation of 99mTc-bound peptide was confirmed with thin layer chromatography and high-performance liquid chromatography. Tracer stability in vitro in blood and PBS was evaluated for up to 4 h. Biodistributions of 99mTc-H6-(GPO)9 and 99mTc-(HE)3-(GPO)9 were assessed in C57BL/6J mice. To induce pulmonary fibrosis, lungs of both sexes were exposed to 1.5 U/kg bleomycin sulphate or saline (control). At 3 weeks post induction, ~1 mCi of 99mTc-(HE)3-(GPO)9 was injected to bleomycin-exposed (n=16) and control (n=6) mice. SPECT and contrast-enhanced CT imaging were carried out at 1 h post-injection. A subset of animals was euthanized after 2 h, and different organs were harvested to assess tracer biodistribution by gamma well counting and to evaluate lung histology. Lung tracer uptake and blood activity were quantified as % injected dose (ID)/mL in vivo and as % ID/gram of tissue ex vivo. Specificity of tracer uptake was evaluated using a scrambled homologue. A fluorescent homologue of the tracer, collagen hybridizing peptide (R-CHP), was used to quantify denatured collagen by histology.Results: Peptide synthesis and its purity (99%) were confirmed with liquid chromatography-mass spectrometry. The specific activity of the final radiolabeled products was 1.7±0.3 Ci/µmol, and the tracer remained stable in vitro in PBS and blood during the testing period. Biodistribution of 99mTc-H6-(GPO)9 and99mTc-(HE)3-(GPO)9 showed comparable blood clearances, but 99mTc-(HE)3-(GPO)9 had lower liver uptake (p<0.0001) at 2 h post injection and was selected for the next set of studies. 99mTc-(HE)3-(GPO)9 uptake was higher in the lungs of bleomycin-exposed mice compared to control animals, as determined by in vivo SPECT/CT (p<0.01, Fig. 1A-B) and ex vivo gamma well counting (p<0.05), with significant correlation between the two methods (Pearson R=0.96, p<0.0001). No significant difference was observed in the heart, spleen, kidney, bone, and blood activity between bleomycin and saline groups. A significant difference (p<0.01) between lung uptake of the control and targeted tracers in bleomycin-exposed mice that underwent imaging with both tracers established the specificity of 99mTc-(HE)3-(GPO)9 uptake (p<0.01, Fig. 1C-D). The development of lung fibrosis and single stranded collagen were confirmed by histology (Fig. 2A). Finally, there was a significant correlation between denatured collagen expression in the lungs quantified by R-CHP staining and in vivo tracer uptake (R=0.80, p<0.001, Fig. 2B).Conclusions: A novel tracer with favorable imaging characteristics, 99mTc-(HE)3-(GPO)9, that can hybridize to denatured collagen was developed and shown to detect collagen remodeling activity in bleomycin-induced pulmonary fibrosis. 99mTc-(HE)3-(GPO)9 imaging of collagen remodeling can potentially track the progression of fibrosis and its reversal in response to anti-fibrotic therapies. Ultimately, this can lead to better management of patients with fibrotic disorders.