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The DEAD-box family of ATPases plays a critical role in nearly all stages of RNA metabolism, from transcription to degradation, and serves as a major regulator of biomolecular condensates. Dysregulation of DEAD-box proteins is well-established in a variety of diseases, including cancer and neurodegenerative disorders, making them attractive therapeutic targets. However, their classification as "undruggable" has historically hindered small-molecule-based modulation. In this study, we focus on DDX21, a member of the DEAD-box family involved in ribosome biogenesis and transcription regulation. As a proof of concept for targeting such RNA-binding proteins, we developed a lysate-based small-molecule microarray platform to identify compounds that directly bind DDX21. This screen led to the discovery of KI-DX-014, a small-molecule compound capable of inhibiting the interaction of DDX21 with RNA. KI-DX-014 modulated the RNA-dependent functions of DDX21, including its ATPase activity and biomolecular condensate formation. Furthermore, KI-DX-014 attenuated the DDX21-dependent release of P-TEFb from the 7SK snRNP complex , suppressed P-TEFb-dependent phosphorylation of the RNA polymerase II CTD, and induced developmental defects in zebrafish embryos. These findings reveal a previously unexploited therapeutic avenue and establish KI-DX-014 as a chemical probe for dissecting the biological functions of DDX21 in both normal physiology and disease states.