Fuchs Natalie, Kuncewicz Katarzyna, Gaamouch Farida El, Gabr Moustafa T
Molecular Imaging Innovations Institute (MI3), Department of Radiology, Weill Cornell Medicine, 1300 York Avenue, New York, NY, 10065, United States.
Molecular Imaging Innovations Institute (MI3), Department of Radiology, Weill Cornell Medicine, 1300 York Avenue, New York, NY, 10065, United States; Department of Biomedical Chemistry, Faculty of Chemistry, University of Gdansk, Poland.
SLAS Discov. 2025 Sep;35:100255. doi: 10.1016/j.slasd.2025.100255. Epub 2025 Jul 30.
Triggering receptor expressed on myeloid cells 2 (TREM2) is an immunomodulatory receptor implicated in both neurodegenerative diseases and cancer. Depending on the context, TREM2 agonists or inhibitors hold therapeutic potential. To date, the majority of TREM2-targeted strategies have centered on monoclonal antibodies (mAbs), which face limitations such as poor tissue penetration and potential immunogenic side effects. To overcome these challenges and expand the chemical space for TREM2-targeting agents, we developed a high-throughput screening (HTS) platform to identify novel small molecule TREM2 binders. Using temperature-related intensity change (TRIC) technology in a 384-well plate format (NanoTemper Dianthus), we screened two focused compound libraries comprising over 1,200 molecules. From this screen, 18 preliminary hits (1.44 % hit rate) were identified and subsequently validated by dose-response binding studies using microscale thermophoresis (MST), yielding four validated hits (0.32 % hit rate) with binding affinities in the high to medium micromolar range (e.g., T2337, K = 22.4 µM). The binding of the top hit, T2337, was further validated using surface plasmon resonance (SPR). Additionally, we assessed the functional activity of all four validated hits in a cellular assay measuring TREM2-mediated Syk phosphorylation in HEK293 cells co-expressing human TREM2 and its adaptor protein DAP12. These findings establish a robust and scalable platform for the discovery of small molecule TREM2 modulators and serve as a proof-of-concept for broader HTS campaigns targeting TREM2.
髓系细胞触发受体2（TREM2）是一种免疫调节受体，与神经退行性疾病和癌症都有关联。根据具体情况，TREM2激动剂或抑制剂具有治疗潜力。迄今为止，大多数针对TREM2的策略都集中在单克隆抗体（mAb）上，这些单克隆抗体面临着组织穿透性差和潜在免疫原性副作用等局限性。为了克服这些挑战并扩大靶向TREM2药物的化学空间，我们开发了一种高通量筛选（HTS）平台来鉴定新型小分子TREM2结合剂。我们使用384孔板形式（NanoTemper Dianthus）的温度相关强度变化（TRIC）技术，筛选了两个包含1200多个分子的聚焦化合物库。从该筛选中，鉴定出18个初步命中物（命中率为1.44%），随后通过使用微量热泳动（MST）的剂量反应结合研究进行验证，得到4个经过验证的命中物（命中率为0.32%），其结合亲和力在高到中微摩尔范围内（例如，T2337，K = 22.4 µM）。使用表面等离子体共振（SPR）进一步验证了最佳命中物T2337的结合。此外，我们在细胞试验中评估了所有4个经过验证的命中物的功能活性，该试验测量了共表达人TREM2及其衔接蛋白DAP12的HEK293细胞中TREM2介导的Syk磷酸化。这些发现建立了一个强大且可扩展的平台，用于发现小分子TREM2调节剂，并作为针对TREM2的更广泛高通量筛选活动的概念验证。
