We have developed a Oligopyridylamide (OP) based 2-Dimensional Fragment-Assisted Structure-based Technique (2D-FAST) to identify potent antagonists of α-Synuclein (αS) aggregation, a process central to Parkinson’s disease (PD). The 2D-FAST utilizes a fragment-based screening of large chemical space in OPs, which led to the identification of NS132 as an antagonist of the multiple facets of αS aggregation. We also identified a better cell permeability analog (NS163) without sacrificing activity. OPs rescue αS aggregation mediated PD phenotypes in muscle cells and dopaminergic (DA) neurons in C. elegans models. OPs prevent the progression of PD phenotypes in a novel post-disease onset PD model.This is one of the first examples of a synthetic mimetic-based 2D-FAST to identify antagonists of toxic αS self-assembly. We envision that 2D-FAST will have tremendous potential as it is expandable for other oligoamide scaffolds and for a much larger chemical space to identify lead therapeutics for various diseases.