Directed evolution of proteoglycan-modifying enzymes: Functional applications in cervical cancer therapy

The study investigates the therapeutic potential of enzyme variations EP-22, DS-13, and SM-47 in cervical cancer treatment using HeLa and SiHa cell lines, focusing on their effects on cell viability, migration, and molecular targets. The MTT assay findings also show that at a concentration of 50 μg/mL, EP-22 has an IC50 value of 35 % for HeLa cells and 28 % for SiHa cells, a significant dose effect (p < 0.01). EP-22 was not less potent at a lower working concentration of 25 μg/mL and could reduce HeLa cell viability to 78 %. In this case, there were significant changes in the anti-migratory effect, as evidenced by 45 % inhibition of SiHa cell migration and a 12 % wound closure rate compared with 54 % in the untreated cells. The obtained densitometric analysis indicated that in EP-22 treated HeLa cells, syndecan-1 and perlecan protein levels were reduced by approximately 65 % and 57 %, respectively, while the MMP-2 and MMP-9 levels were reduced to about 50 % and 45 %, respectively. Annexin V staining also highlighted a 40 % enhancement in early apoptosis and 25 % in late apoptosis in EP-22 handled cells. These data suggest the potential of EP-22 and its derivatives as therapeutic molecules in cervical cancer treatment, reducing HeLa proliferation by 35 % and SiHa by 28 %, inhibiting SiHa migration by 45 %, and affecting molecular targets involved in adhesion and invasiveness. Future studies must elucidate the effectiveness of in vivo experiments and how these findings were obtained. At 50 μg/mL, EP-22 reduced HeLa and SiHa cell viability by 35 % and 28 %, respectively, with significant dose-dependent effects (p < 0.01). At 25 μg/mL, EP-22 maintained potency, reducing HeLa cell viability to 78 %. EP-22 inhibited SiHa cell migration by 45 % and reduced wound closure rates to 12 % compared to 54 % in untreated cells. This work uses the HeLa and SiHa cell lines to examine the therapeutic potential of enzyme variation EP-22 in cervical cancer. EP-22 showed significant anti-cancer effects at 50 μg/mL doses, reducing cell viability at lower concentrations and achieving an IC50 of 35 % for HeLa cells and 28 % for SiHa cells. It is worth mentioning that EP-22 considerably reduced levels of essential proteins: syndecan-1 (65 %), perlecan (57 %), MMP-2 (50 %), and MMP-9 (45 %), in addition to inhibiting SiHa cell migration by 45 %. Furthermore, annexin V staining showed that treated cells exhibited a 40 % increase in early apoptosis and a 25 % increase in late apoptosis.