Abstract Transcription factors (TFs) are high value yet challenging therapeutic targets in oncology. Most are considered “undruggable” due to the lack of well-defined small-molecule binding pockets, and the presence of extensive intrinsically disordered regions and shallow protein-protein interaction surfaces. Their active conformations further depend on post-translational modifications, cofactors, and the intracellular milieu - conditions difficult to recapitulate in vitro - therefore, strategies that interrogate TFs in their native cellular environment are preferable. DNA-Encoded Libraries (DELs) provide ultra-high-throughput access to small-molecule binders, but a complicating factor for screening DNA-binding proteins is false positives arising from barcode sequences that resemble native binding motifs. Mutant TF versions engineering to disrupt DNA-binding domains can reduce this issue but risks compromising TF structure and function. Therefore, screening full-length proteins appear highly desirable. Here, clusters of small-molecule hits were identified in an intracellular DEL screen against a full-length, cancer-relevant TF and positively validated in orthogonal assays. Importantly, in the screen, a duplex DNA containing the cognate binding motif was included and successfully suppressed barcode-mediated artifacts. A parallel screen including unrelated duplex DNA confirmed retention of sequence-specific TF-DNA interactions showing that the DNA-binding activity of the TF was intact during the screening. These findings demonstrate that intracellular DEL screening can overcome key mechanistic limitations in targeting transcription factors and enable robust discovery of small-molecule binders to functional, full-length TFs - expanding the druggable space for oncogenic transcriptional regulators previously inaccessible to small-molecule approaches. Citation Format: Leif K. Larsen, Allan B. Christensen, Andrzej Taranta, Charlotte Andersen, Frank A. Sløk, Lars K. Petersen, Ole Kristensen, Rico G. Petersen, Peter Blakskjær, Tobias N. Hansen, Nils J. Hansen. DELs in cells: Small-molecule hits for transcription factor identified by intracellular DNA-encoded library screening abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 471.
Larsen et al. (Fri,) studied this question.