Developing hyperpolarized metabolic contrast agents at high field dDNP for large animal research

• Developed high-polarization dDNP formulations for [1– 13 C]2-ketoisocaproate and [1,4– 13 C 2 ]fumarate useful for large animal studies. • More than 25 % polarization in less than one hour for 90 mM in 30 ml [1,4– 13 C 2 ]fumarate. • More than 30 % polarization in less than one hour for 150 mM in 30 ml [1– 13 C]2-ketoisocaproate. • Validated scalable [1– 13 C]pyruvate and [1– 13 C]2-ketoisocaproate dDNP for large animal and clinical research at high-field conditions. • Non-isotope labelled [1– 13 C]pyruvate polarized to more than 30 % in 90 minutes. Dissolution Dynamic Nuclear Polarization (dDNP) polarizers have achieved high field strengths and large sample volumes. These advancements necessitate new formulations of hyperpolarized metabolic contrast agents (HMCAs) to enable large animal studies. While several metabolic substrates have been investigated at lower field dDNP and tested as HMCAs in rodents, 13 C-labeled pyruvate remains the most studied HMCA and is currently the only one actively used in clinical trials due to its favorable biological and physical properties. Effective human and large animal dDNP formulations require high molar substrate concentrations, low DNP sample viscosity for efficient dissolution, and adequate dilution of the DNP sample to minimize signal decay and maximize HMCA concentration. We present substrate formulations optimized for high-field polarization and large-volume dissolution. Specifically, we validate the upscaling of [1– 13 C]pyruvate under high-field conditions and demonstrate that [1– 13 C]2-keto-isocaproate and [1,4– 13 C 2 ]fumarate, which have been proven successful in rodent studies, can be formulated to yield high polarization at suitable concentrations and volumes for large animal metabolic MR imaging.