Preclinical Evaluation of RYM1, a Matrix Metalloproteinase-Targeted Tracer for Imaging Aneurysm

Abstract

Matrix metalloproteinases (MMPs) play a key role in abdominal aortic aneurysm (AAA) development. Accordingly, MMP-targeted imaging provides important information regarding vessel wall biology in the course of aneurysm development. Given the small size of the vessel wall and its proximity with blood, molecular imaging of aneurysm optimally requires highly sensitive tracers with rapid blood clearance. To this end, we developed a novel hydrosoluble zwitterionic MMP inhibitor, RYM, on the basis of which a pan-MMP tracer, RYM1, was designed. Here, we describe the development and preclinical evaluation of RYM1 in comparison with RP805, a commonly used pan-MMP tracer in murine models of aneurysm. Methods: The macrocyclic hydroxamate-based pan-MMP inhibitor coupled with 6-hydrazinonicotinamide, RYM1, was synthesized and labeled with ^99mTc. Radiochemical stability of ^99mTc-RYM1 was evaluated by radio-high-performance liquid chromatography analysis. Tracer blood kinetics and biodistribution were compared with ^99mTc-RP805 in C57BL/6J mice (n = 10). ^99mTc-RYM1 binding to aneurysm and specificity were evaluated by quantitative autoradiography in apolipoprotein E-deficient (apoE^-/-) mice with CaCl₂-induced carotid aneurysm (n = 11). Angiotensin II-infused apoE^-/- (n = 16) mice were used for small-animal SPECT/CT imaging. Aortic tissue MMP activity and macrophage marker CD68 expression were assessed by zymography and reverse-transcription polymerase chain reaction. Results: RYM1 showed nanomolar range inhibition constants for several MMPs. ^99mTc-RYM1 was radiochemically stable in mouse blood for 5 h and demonstrated rapid renal clearance and lower blood levels in vivo compared with ^99mTc-RP805. ^99mTc-RYM1 binding to aneurysm and its specificity were shown by autoradiography in carotid aneurysm. Angiotensin II infusion in apoE^-/- mice for 4 wk resulted in AAA formation in 36% (4/11) of surviving animals. In vivo ^99mTc-RYM1 small-animal SPECT/CT images showed higher uptake of the tracer in AAA than nondilated aortae. Finally, aortic uptake of ^99mTc-RYM1 in vivo correlated with aortic MMP activity and CD68 expression. Conclusion: The newly developed pan-MMP inhibitor-based tracer ^99mTc-RYM1 displays favorable pharmacokinetics for early vascular imaging and enables specific detection of inflammation and MMP activity in aneurysm.

Keywords: SPECT/CT; abdominal aortic aneurysm; inflammation; matrix metalloproteinase inhibitor; molecular imaging.