Validating PLCG2 as a target for the treatment of Alzheimer's Disease through cell biology and pharmacological tools

PROJECT SUMMARY/ABSTRACT
Genetic studies have identified novel Alzheimer’s Disease (AD) protective and risk associated genes and
genetic variants. PLCG2 is one such gene with protective (P522R) and risk (M28L) variants making it a useful
protein to target for the treatment of AD. However, the particular phenotype to be targeted through small molecule
modulation of PLCG2 is unknown. Without this knowledge, the validation of PLCG2 as a target for the treatment
of AD will be greatly impeded. Therefore, we hypothesize that activation of PLCG2 will mimic the protective effect
of PLCG2P522R by inducing molecular signaling changes to PLCG2WT in a manner similar to the protective variant.
We will address this hypothesis through the following Specific Aims: Aim 1: Determine how PLCG2 variants
associated with AD risk and protection affect PLCG2 related signaling, in vitro. We hypothesize that these
variants cause their respective effects (protection and risk) by modulating the preference of PLCG2 to be
recruited to the cell membrane, its phosphorylation state, its preferred interaction complexes, and its turnover
rate. We will test this hypothesis by stimulating human induced pluripotent stem cell (iPSC) derived microglia
like cells (MGLs) expressing either wildtype (WT), protective, or risk variants of PLCG2, and then use both mass
spectrometry and traditional biochemical methods to measure these molecular effects. Aim 2: Determine how
modulation of PLCG2 activity affects PLCG2WT microglia, in vitro, compared to protective and risk PLCG2
variants. We hypothesize that activation of PLCG2 will produce an activation profile that overlaps with
PLCG2P522R microglia. We will test this hypothesis by comparing pAKT levels and Aβ uptake between WT MGLs
treated with already developed, novel, small molecule modulators of PLCG2, and vehicle treated MGLs bearing
the protective and risk variants. With respect to outcomes as a consequence of the work proposed, we expect
to identify changes in the phosphorylation state, subcellular localization, interactome, and turnover rate of PLCG2
caused by the protective and risk variants. We also expect to identify the effect of small molecule modulation of
PLCG2 activity on pAKT levels and Aβ uptake with reference to the protective and risk variants. These results
will advance our understanding of how PLCG2P522R protects from AD and provide insight into the pharmacological
requirements of a commercial PLCG2 modulator as a potential AD therapeutic. Additionally, we expect that this
study will guide the design of therapeutic interventions that mimic the protective functions of the PLCG2P522R
variant. Finally, this study will contribute to the continued understanding of the role of microglia in response to
AD pathology. Overall, this study will have a positive impact because it will provide a biological basis for
understanding the pharmacology of novel therapeutics targeting PLCG2 for the treatment of AD.

Grant Number: 5F31AG089990-02
NIH Institute/Center: NIH
Principal Investigator: Logan Bedford