This study reports the synthesis, characterization, and evaluation of 18 novel carbazole derivatives containing thiosemicarbazide functional groups, which exhibit potent antioxidant, anticancer, and antimicrobial activities. The compounds were synthesized through a series of chemical reactions, including esterification, hydrazination, and coupling with various isothiocyanates. The synthesized compounds were characterized using NMR, mass spectrometry, and other analytical techniques. Antioxidant activity was assessed using the DPPH radical scavenging assay, revealing that compounds 4h and 4o showed the highest antioxidant activity with IC50 values of 0.73 and 0.38 μM, respectively. Anticancer activity was evaluated using MTT assays on MCF-7 cancer cells, with compound 4o showing the highest potency (IC50 = 2.02 μM). Compound 4o also exhibited significant antimicrobial activity against S. aureus, E. coli, and C. albicans, with MIC values of 1.56 μM, 1.56 μM, and 0.39 μM, respectively. The study further investigated the molecular mechanisms underlying the anticancer activity of compound 4o, revealing its ability to target the PI3K/AKT/mTOR signaling pathway, induce apoptosis, and cause cell cycle arrest in MCF-7 cells. Molecular docking simulations supported these findings, showing favorable binding interactions between compound 4o and key enzymes in the PI3K/AKT/mTOR pathway. Overall, the results indicate that compound 4o is a promising candidate for further development as an anticancer and antimicrobial agent, with potential applications in the treatment of various cancers and bacterial infections. The study highlights the importance of combining experimental and computational approaches in the discovery and development of novel therapeutics.This study reports the synthesis, characterization, and evaluation of 18 novel carbazole derivatives containing thiosemicarbazide functional groups, which exhibit potent antioxidant, anticancer, and antimicrobial activities. The compounds were synthesized through a series of chemical reactions, including esterification, hydrazination, and coupling with various isothiocyanates. The synthesized compounds were characterized using NMR, mass spectrometry, and other analytical techniques. Antioxidant activity was assessed using the DPPH radical scavenging assay, revealing that compounds 4h and 4o showed the highest antioxidant activity with IC50 values of 0.73 and 0.38 μM, respectively. Anticancer activity was evaluated using MTT assays on MCF-7 cancer cells, with compound 4o showing the highest potency (IC50 = 2.02 μM). Compound 4o also exhibited significant antimicrobial activity against S. aureus, E. coli, and C. albicans, with MIC values of 1.56 μM, 1.56 μM, and 0.39 μM, respectively. The study further investigated the molecular mechanisms underlying the anticancer activity of compound 4o, revealing its ability to target the PI3K/AKT/mTOR signaling pathway, induce apoptosis, and cause cell cycle arrest in MCF-7 cells. Molecular docking simulations supported these findings, showing favorable binding interactions between compound 4o and key enzymes in the PI3K/AKT/mTOR pathway. Overall, the results indicate that compound 4o is a promising candidate for further development as an anticancer and antimicrobial agent, with potential applications in the treatment of various cancers and bacterial infections. The study highlights the importance of combining experimental and computational approaches in the discovery and development of novel therapeutics.