TAR DNA-binding protein 43 (TDP-43) is a key regulator of RNA metabolism, highly abundant in the central nervous system (CNS), and normally localized in the nucleus. Under pathological conditions, TDP-43 is mislocalized to the cytoplasm, where it undergoes phosphorylation, ubiquitination, and subsequently aggregates, forming insoluble inclusions. These aggregates cause loss of normal nuclear function and gain of toxic cytoplasmic function, leading to neuronal cell death. TDP-43 proteinopathy constitutes the principal pathological hallmark of amyotrophic lateral sclerosis (ALS). Despite this central role, no approved ALS therapies directly target TDP-43, leaving a major unmet therapeutic gap. To address this gap directly, we pursued the discovery of novel small-molecule inhibitors that target TDP-43 aggregation. The TDP-43 BiFC cellular assay was developed to visualize its mislocalization and aggregation, enabling high-throughput screening of in-house compound libraries. The initial screening hits were refined through SAR optimization to improve TDP-43 selectivity over other proteinopathies, specifically 4R-tau. This effort yielded five potent and selective inhibitors. Due to early diagnosis for neurodegenerative is critical, we are pursuing to develop PET probe derived from our compound. These findings establish a direct strategy for modulating, and diagnosing TDP-43 pathology and represent a promising foundation for developing disease-modifying therapeutics for ALS.