Poly(ADP-ribose) polymerase-1 (PARP-1) plays a crucial role in DNA repair, mediating approximately 90 % of ADP-ribosylation processes associated with DNA damage response. Consequently, inhibiting PARP-1 with small molecules represents a promising strategy for cancer therapy. Utilizing a structure-based design and molecular hybridization approach, we developed three novel series of spirobenzoxazinone-piperdine/salicylamide-based derivatives. These compounds were evaluated for their in vitro PARP-1 inhibitory activity, and their structure–activity relationships were analyzed. At 10 µM concentration, derivatives (18a-d) demonstrated nearly complete inhibition, and the spirocyclic derivative (7c) also achieved a considerable inhibitory effect, with IC50 values in the low micromolar range. The most promising compounds (7c, 18a-d) were tested for their antiproliferative activity against six cancer cell lines. Notably, compounds (7c) and (18d) exhibited significant antiproliferative effects against H1299 and FaDu cells, which correlated with their calculated logP values. These compounds were also tested against normal human skin fibroblasts (HSF), revealing a favorable safety profile compared to cancer cells. Basal anti-PARP-1 activity of the most promising compounds was validated in the HCT116 colorectal cancer cell line. Western blot analysis confirmed robust cleavage of PARP-1, indicating enzymatic inhibition and loss of PARP-1 activity. Combining these inhibitors with doxorubicin showed synergistic lethality in colony-formation assay. Finally, a molecular docking study was conducted to examine the binding modes of these compounds within the PARP-1 active site. The results demonstrated binding modes comparable to those of olaparib and other approved PARP-1 inhibitors, maintaining the key interactions necessary for activity. Based on these findings, compounds (7c) and (18d) emerge as promising candidates for further development in targeting anti-cancer drug resistance through PARP-1 inhibition.