The “LRRK2 Detection in PBMC Consortium” is a pre-competitive collaboration between MJFF and select industry partners with the goal of optimizing the measurement of pLRRK2 in human PBMCs. MJFF launched the consortium in response to the discussions at the LRRK2 Industry Summit in 2016. Each company provided in-kind analysis for pLRRK2 and total LRRK2 using in-house immunoassay platforms while PBMCs collected through the two MJFF PBMC biobanking initiatives served as the matrix for the analysis. The consortium activities were split into three phases and goals for each phase of the study were defined and refined through discussions with consortium members. Phase 1 sought to determine which PBMC collection protocol(s) were best suited for each of the five company’s pLRRK2 assays. A small set of PBMCs were analyzed by consortium members using their preferred phospho and total LRRK2 assays. Based on the Phase 1 data and conversations with consortium members to identify field-enabling and program-enabling questions, MJFF selected the Pfizer pS935 assay for Phase 2 analysis and Merck pS935 assay for Phase 3 analysis.
Phase 2 sought to determine whether pS935 levels differ in iPD, HC, and G2019S manifesting and non-manifesting subjects. The Pfizer assay was re-established at the Quanterix Accelerator Lab, and a selection of PD subject samples from Columbia University was used to assess technical and biological variance to enable power calculations to ensure adequate sample sizes had been collected. Finally, a sample set comprised of 22 G2019S- healthy controls, 16 G2019S+ healthy controls, 46 G2019S- PD, and 33 G2019S+ PD PBMCs was analyzed for total and pS935 LRRK2.
Phase 3 sought to determine the potency of three LRRK2 kinase inhibitor tool compounds across the G2019S carrier and non-carrier groups using Merck’s MSD pS935 assay. Large volume (100ccc) of whole blood was collected from 6 G2019S- healthy controls, 7 G2019S- PD, 4 G2019S+ healthy controls and 8 G2019S+ PD subjects and shipped directly to Merck for same day PBMCs isolation. Immediately after isolation, PBMCs were treated with MLi-2, PFE-360 or GNE-7915 and pSer935 and total LRRK2 levels determined. Ex-vivo pSer935 IC50s for MLi-2, PFE-360 and GNE-7915 were determined in duplicate from each of the donor samples.
The Consortium is currently discussing the potential to expand on these findings in other matrices using the recently developed Rab antibodies to optimize and develop new target engagement and patient enrichment biomarkers.
Authors: Shalini Padmanabhan1, Marco Baptista1, Alyssa Reimer1, Kalpana Merchant1, Roy Alcalay2, Najah Levers2, Chris Liong2, Thomas Lanz3, Donal Gorman4, Julie Lee5, Payal Sheth5 and Matthew Fell5
Affiliations: 1The Michael J. Fox Foundation for Parkinson’s Research, 2Columbia University, 3Biogen Inc., 4Pfizer Inc., 5Merck & Co.