Objective: To assess feasibility of developing a CNS LRRK2 PET tracer.

Background: LRRK2 enzyme belongs to the leucine-rich repeat kinase family. Pathogenic mutations in the LRRK2 gene alter enzyme level, activity, and susceptibility to Parkinson’s disease (PD). So, inhibition of LRRK2 may be beneficial to treatment of PD. Positron Emission Tomography (PET) is a non-invasive imaging technique, wildly used for assessing target occupancy/efficacy of drug in vivo. A CNS LRRK2 PET tracer supports development of LRRK2 inhibitor. But, is it feasible to develop a CNS LRRK2 PET tracer?

Methods: Hot saturation binding assay was done with human LRRK2 enzyme, and tissue homogenates prepared from various tissues, including brain and Kidney. In vitro autoradiography (ARG) was done with frozen tissue slices. [3H]A* and [3H]B* were synthesized in house. Non-displaceable binding (NDB) was defined using self-block.

Results: [3H]A and [3H]B showed displaceable and saturable binding to one site with very high affinities (Kd = 56pM and 57pM respectively), and large displaceable binding window to human WT full-length LRRK2 enzyme. However, in tissue homogenates, [3H]A showed minimal or no displaceable binding (rat, rhesus monkey and human CPu), while only very modest specific binding of [3H]B was observed. [3H]B bound to more than one site in rhesus CPu and cortex. The Kd values of high affinity binding site of [3H]B were 90pM (CPu) and 50pM (cortex) with the Bmax values of 0.3nM (wet tissue weight) for both tissues. [3H]B also showed modest binding in WT rat kidney homogenates, but not in LRRK2 KO rat kidney. In ARG study, [3H]A showed no displaceable binding in brain slices of rat, rhesus monkey, and human.

Conclusions: The data show [3H]A and [3H]B are radioligands with very high binding affinity to human WT full-length LRRK2. But in brain CPu homogenates of rat, rhesus monkey and human, [3H]A shows no specific binding, consistent with [3H]A ARG, which shows minimal specific binding in brain slices of rat, rhesus monkey and human. Only very modest specific binding of [3H]B is presented in CPu and cortex of rhesus monkey brain. The data demonstrate low binding site densities of [3H]A and [3H]B in brain tissues examined, indicating low feasibility to develop a CNS PET tracer for LRRK2.

References: *: the structures of compounds will be disclosed at presentation.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/study-on-feasibility-to-develop-a-central-nervous-system-cns-leucine-rich-repeat-kinase-2-lrrk2-pet-tracer/