Ala-[1-13C]Gly- <i>d</i> 2-15NMe2: hyperpolarized molecular probe for aminopeptidase N activity with suppressed scalar relaxation

Abstract Aminopeptidase N is a key biomarker associated with various diseases, including cancers and cardiovascular diseases. Nuclear magnetic resonance imaging coupled with dynamic nuclear polarization represents a powerful technique for in vivo detection of enzymatic activity. We previously developed Ala-[1-13C]Gly-d2-NMe2 as a dynamic nuclear polarization-coupled magnetic resonance imaging molecular probe capable of visualizing and mapping aminopeptidase N activity in vivo. However, the broader application of this probe to disease animal models is hindered by significant signal attenuation during sample transfer through low magnetic fields, such as the Earth's magnetic field, primarily due to scalar relaxation via the 13C–14N bond. Here, we present Ala-[1-13C]Gly-d2-15NMe2 as an alternative, incorporating a 15N nucleus in place of 14N. This modification produces markedly enhanced magnetic resonance signal intensity without the need for stringent sample handling to avoid exposure to low magnetic fields. Consequently, this developed probe can potentially offer significant advantages for cellular studies and applications in complex organisms, where elaborate preinjection procedures are often required.