Introduction: Tumors are a major public health problem. The resistence to pharmaceuticals increases day after day and that imposes the need for the synthesis of the new ones. The newly synthesized N-9 sulfonyurea derivatives have a strong anti-inflammatory potential. The aims of the study: To investigate the effects of the newly synthesized N-9 sulfonylurea (R-191 and D7-23) derivatives on HeLa and K562 cells in in vitro conditions. Materials and methods: The newly synthesized N-9-sulfonylurea derivatives have been synthesized at the Department of Organic Chemistry and Biochemistry in Zagreb, at the Ruđer Bošković Institute. The R-191 and D7-23 compounds, used in the study, have manifested themselves as the most specific. The commercial HeLa (2D and 3D) and K562 cells have been cultivated in the bottles for the cultivation in a CO2 incubator at 37° C. After having been cultivated, the cells have been separated by the trypsin and the RNA has been isolated by RNeasy Mini Kit (50). For the purposes of this research, we have used PrimeScript First Strand cDNA Synthesis Kit for the RNA transcription in the cDNA and QuantStudio 5 RT-PCR for the gene expression analysis. Results: The R-191 derivative at a concentration of 5x10-6 M promotes gene expression of cytochrome c in the HeLa cells, grown in the monolayer and in 3D cell cultures. The R-191 derivative has no effect on the gene expression at the K562 cell line. The D7-23 derivative shows opposite effects in 2D and 3D culture of the HeLa cells and at the same time stimulates expression of cytochrome c in the K562 cells. Conclusion: It has been proved that the studied derivatives change the level of the cytochrome c gene expression under the influence of the N-9 sulfonylurea derivatives