Synthesis and biological assessment of BUB1B inhibitors for the treatment of clear cell renal cell carcinoma

Abstract

Clear cell renal cell carcinoma (ccRCC) presents substantial therapeutic challenges due to its molecular heterogeneity, limited response to conventional therapies, and widespread drug resistance. Recent advancements in molecular research have identified novel targets, such as BUB1B, which has been identified through global transcriptomic profiling and gene co-expression network analysis as critical in ccRCC progression. In this study, we synthesized 40 novel derivatives of TG-101209 to modulate BUB1B expression and activity, leading to the induction of apoptosis in Caki-1 cells. The molecular structures of all compounds were confirmed via 1 H and 13 C-NMR and mass spectrometry. Computational docking studies were conducted using Schrödinger Maestro software. The efficacy of the compounds on cell viability was screened using the MTT assay and further validated by the LDH assay. The expression of the target protein BUB1B and apoptosis-related proteins was analyzed via western blotting. BUB1B activity was assessed through an enzymatic assay, and compound binding efficacy was evaluated using a cellular thermal shift assay (CETSA). The results indicated that four compounds ( 7h, 8h, 8i, and 8j ) demonstrate stronger molecular interactions and better conformational fit within the target cavity, leading to improved binding affinity. These compounds also exhibited more potency in reducing the viability of Caki-1 cells compared to TG-101209. In particular, compound 8h was identified as the most effective, exhibiting the strongest inhibitory effect on BUB1B and inducing apoptosis. Compound 8h demonstrated intracellular binding with BUB1B, similar to TG-101209, but through a different binding moiety that destabilizes the BUB1B protein structure, whereas TG-101209 stabilizes it. In conclusion, compound 8h , by destabilizing BUB1B and inducing apoptosis, shows promise as a potent therapeutic candidate for clear cell renal cell carcinoma (ccRCC) treatment. • Standard chemotherapy has limited effectiveness against clear cell renal cell carcinoma (ccRCC) because of its significant molecular diversity and widespread drug resistance • BUB1B, identified through transcriptomic analysis, plays a crucial role in ccRCC progression, making it a novel target for therapy. • 40 novel TG-101209 derivatives were designed and synthesized to inhibit BUB1B expression and activity, to induce apoptosis in the ccRCC cell line. • Compounds 8h and 8i were identified as the most effective, exhibiting the strongest inhibitory effect on BUB1B and inducing apoptosis compared to TG-101209 in ccRCC treatment, yielding promising results.