Cancer's global impact necessitates innovative and less toxic treatments. Thiosemicarbazones (TSCs), adaptable metal chelators, offer such potential. In this study, we have synthesized N (4) -substituted heterocyclic TSCs from syringaldehyde (TSL1, TSL2), and also report the unexpected copper -mediated cyclization of the TSCs to form thiadiazoles (TSL3, TSL4), expanding research avenues. This work includes extensive characterization and studies such as DNA/protein binding, molecular docking, and theoretical analyses to demonstrate the potential of the as -prepared TSCs and thiadiazoles against different cancer cells. The DFT results depict that the thiadiazoles exhibit greater structural stability and reduced reactivity compared to the corresponding TSCs. The docking results suggest superior EGFR inhibition for TSL3 with a binding constant value of - 6.99 Kcal/mol. According to molecular dynamics studies, the TSL3-EGFR complex exhibits a lower average RMSD (1.39 nm) as compared to the TSL1-EGFR complex (3.29 nm) suggesting that both the thiadiazole and thiosemicarbazone examined here can be good inhibitors of EGFR protein, also that TSL3 can inhibit EGFR better than TSL1. ADME analysis indicates drug -likeness and oral availability of the thiadiazole-based drugs. The DNA binding experiment through absorption and emission spectroscopy discovered that TSL3 is more active towards DNA which is quantitatively calculated with a Kb value of 4.74 x 106 M-1, Kq value of 4.04 x 104 M-1and Kapp value of 5 x 106 M-1. Furthermore, the BSA binding studies carried out with fluorescence spectroscopy showed that TSL3 shows better binding capacity (1.64 x 105 M-1) with BSA protein. All the compounds show significant cytotoxicity against A459 -lung, MCF-7-breast, and HepG2-liver cancer cell lines; TSL3 exhibits the best cytotoxicity, albeit less effective than cisplatin. Thiadiazoles demonstrate greater cytotoxicity than the TSCs. Overall, the promise of TSCs and thiadiazoles in cancer research is highlighted by this study. Furthermore, it unveils unexpected coppermediated cyclization of the TSCs to thiadiazoles. C1 [Manakkadan, Vipin; Palakkeezhillam, Vishnunarayanan Namboothiri Vadakkedathu; Rasin, Puthiyavalappil; Kumar, Vaishnu Suresh; Sreekanth, Anandaram] Natl Inst Technol Tiruchirappalli, Dept Chem, Tiruchirappalli 620015, Tamil Nadu, India. [Haribabu, Jebiti] Univ Atacama, Fac Med, Carreras 1579, Copiapo 1532502, Chile. [Haribabu, Jebiti] Chennai Inst Technol CIT, Chennai 600069, India. [Vediyappan, Ramesh] Madurai Kamaraj Univ, Sch Chem, Dept Organ Chem, Madurai 625021, Tamil Nadu, India. [Kumar, Vaishnu Suresh; Garg, Mohit] Birla Inst Technol & Sci, Dept Chem Engn, Pilani 333031, Rajasthan, India. [Bhuvanesh, Nattamai] Texas A&M Univ, Dept Chem, College Stn, TX 77842 USA. C3 National Institute of Technology (NIT System); National Institute of Technology Tiruchirappalli; Universidad de Atacama; Madurai Kamaraj University; Birla Institute of Technology & Science Pilani (BITS Pilani); Texas A&M University System; Texas A&M University College Station