Imaging the invaders: TREM1 as a novel PET imaging biomarker of peripheral infiltrating myeloid cells and potential therapeutic target in multiple sclerosis. (2019)

1506b Objectives: Myeloid cells (e.g. macrophages, neutrophils, microglia) of the innate immune system are vital to the initiation and progression of multiple sclerosis (MS)1. Existing imaging strategies for detecting myeloid cells lack cell specificity and cannot distinguish between beneficial and toxic immune responses. Thus, new imaging biomarkers that specifically identify myeloid cells and their activation states are critical for disease staging and therapy monitoring. We recently identified triggering receptor expressed on myeloid cells 1 (TREM1) as a highly specific marker of pro-inflammatory myeloid cells and subsequently created the first positron emission tomography (PET) tracer for this target. Here, we validate TREM1 as a biomarker of maladaptive myeloid-driven immune responses in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. We then demonstrate the ability of TREM1 PET to monitor active disease with increased sensitivity compared to the gold-standard imaging approach for detecting neuroinflammation - translocator protein 18 kDa (TSPO) PET. Finally, we illustrate the therapeutic potential of manipulating TREM1 signaling.
 Methods: Wild-type (WT) and TREM1 knockout (KO) C57BL/6 mice were induced with EAE using MOG35-552 and grouped by disease severity: pre-symptomatic (pre; score 0, >1g weight loss in 48 h), low (score 0.5-2), and high EAE (score 2.5-4.5). Flow cytometry: CNS tissues were stained for TREM1, CD45, CD11b, CD11c, CD3, and Ly-6G. In vivo: Anti-TREM1 monoclonal antibody (mAb) was DOTA-conjugated and radiolabeled with 64Cu. PET imaging was performed 20 h post-injection of [64Cu]TREM1-mAb (95-120 μCi, >99% RCP) and 50-60 min after [18F]GE-180 (231-269 μCi, >99% RCP). Ex vivo: Radioactivity in perfused tissues was measured via a gamma-counter and central nervous system (CNS) tissues were further analyzed with high-resolution autoradiography. Treatment studies: LP17, a decoy receptor peptide previously shown to attenuate TREM1 signaling3, was administered daily to pre EAE mice (10, 15 mg/kg, or saline i.p) for 10 days.
 Results: Flow cytometry revealed TREM1 to be selectively expressed on peripheral myeloid cells but not microglia. TREM1-positive myeloid cell infiltration was dramatically increased in CNS tissues of EAE versus TREM1 KO and naive mice (Suppl.). Interestingly, increased infiltration was evident in pre EAE prior to clinical onset. In vivo TREM1 PET images reflected flow cytometry data (Fig. 1a, b). PET quantification revealed increased tracer uptake in the brain, spinal cord, spleen and femur of WT (pre, low, high) EAE compared to TREM1 KO EAE and naive mice (p<0.001-p<0.0001, n=5-11). Specificity and sensitivity of [64Cu]TREM1-mAb was further confirmed by ex vivo spinal cord autoradiography showing a lack of tracer signal in clinically scoring TREM1 KO EAE mice. Notably, TREM1 PET was significantly more sensitive than TSPO PET (up to 18-fold higher) for detecting activated myeloid cells in EAE. Ex vivo gamma counting of CNS tissues revealed increased EAE-to-naive tracer binding ratios for [64Cu]TREM1-mAb compared to [18F]GE-180 (Fig. 1c, brain; p=0.015, cervical/thoracic; p=0.004 and lumbar spinal cord; p=0.003, n=5-7). Although 100% of WT mice developed EAE, only 50% of TREM1 KO mice developed EAE and with a delayed onset of symptoms (Fig. 1d, p=0.01-p<0.001, KO n=25, WT n=58). Additionally, LP17 treatment significantly ameliorated disease severity compared to saline-treated EAE mice (Fig. 1e, p=0.01-p=0.05).
 Conclusions: To our knowledge, this is the first report of a highly specific PET imaging strategy for detecting peripheral CNS-infiltrating myeloid cells. TREM1 PET has high potential for clinical impact on early-stage diagnosis and therapy selection/monitoring. Moreover, TREM1 appears to play a critical role in driving EAE/MS, making it a functionally relevant biomarker and viable therapeutic target.