Podocyte dysfunction and loss is an early event and a hallmark of proteinuric kidney diseases. Podocyte's normal function is maintained via its unique cellular architecture that relies on an intracellular network of filaments, including filamentous actin (F-actin) and microtubules, that provides mechanical support. Damage to this filamentous network leads to changes in cellular morphology and results in podocyte injury, dysfunction, and death. Conversely, stabilization of this network protects podocytes and ameliorates proteinuria. This suggests that stabilization of podocyte architecture via its filamentous network could be a key therapeutic strategy for proteinuric kidney diseases. However, development of podocyte-directed therapeutics, especially those that target the cell's filamentous network, is still lacking, partly due to unavailability of appropriate cellular assays for use in a drug-discovery environment. Here, we describe a new high content screening based methodology that utilizes a multi-parametric approach to analyze phenotypic data. Implementation of this methodology on a podocyte-based screening assay with a library of 2121 compounds identified paullone-derivatives as a novel group of podocyte protective compounds. We find that three compounds - kenpaullone, 1-azakenpaullone, and alsterpaullone - dose-dependently protect podocytes from puromycin aminonucleoside (PAN) mediated injury in vitro, by reducing PAN-induced changes in both the filamentous actin and microtubules, with alsterpaullone providing maximal protection. Mechanistic studies further show that alsterpaullone suppressed PAN-induced activation of signaling downstream of GSK3β and p38 mitogen-activated protein kinase. In vivo, it reduced ADR-induced glomerular injury in a zebrafish model. Together, these results identify paullone-derivatives as novel podocyte protective agents for future therapeutic development.