Novel 3‐Substituted‐2<i>H</i>‐Chromene Scaffold Based Fluorinated Hydrophobic Fragment as In‐Vitro Antiproliferative Agents and Apoptosis Inducers Targeting Both VEGFR‐2/BRAF^V600E and <i>h</i>‐DHFR With Molecular Docking Simulation

Recently, there has been an increasing interest in the use of protein kinase inhibitors as a therapeutic strategy for the treatment of cancer. In this study, a new series of 2H-chromene derivatives (2-5 and 6-8) and 3H-benzo[f]chromene carbohydrazide derivative (9) were synthesized. The structure of the designed derivatives was characterized by IR, ¹H/¹³C NMR, and elemental analysis. Moreover, the cytotoxic activity of the newly synthesized chromenes was evaluated against breast cancer cell lines (MDA-MB-231 and MCF-7) and a cervical cancer cell line (HeLa). The results of these evaluations demonstrated promising activity, ranging from good to moderate. Additionally, the lung fibroblast cell line (WI-38), as a normal cell line, was also utilized to assess the active derivatives' selectivity. Among the compounds tested, chromene derivative 3 demonstrated the highest potency, exhibiting IC₅₀ values of 5.36 ± 0.50, 7.82 ± 0.60, and 9.28 ± 0.70 µM against the MDA-MB 231, MCF-7, and HeLa cell lines, respectively. The potential of chromone 3 as a multi-targeted anticancer agent was assessed by evaluating its activity against BRAF and VEGFR-2. Notably, the most promising chromene derivative 3 demonstrated significant VEGFR2 activity with an IC₅₀ value of 0.224 µM compared to sorafenib's 0.045 µM, while exhibiting inhibitory activity against BRAF with an IC₅₀ value of 1.695 µM relative to Vemurafenib's IC₅₀ value of 0.468 µM. In addition, compound 3 inhibits the DHFR enzyme with an IC₅₀ value of 2.217 ± 0.014 µM, compared to methotrexate (IC₅₀ = 0.4315 ± 0.019 µM). These results revealed that the compound has multifaceted mechanisms of action that may augment its therapeutic effectiveness. In addition, compound 3 causes overexpression of caspase-3 and Bax by 6.13 and 8.85-fold, respectively. It also downregulates the antiapoptotic Bcl-2 level by 0.4775-fold compared to the untreated MDA-MB 231 cells. Flow cytometry analysis of MDA-MB-231 cells indicates that compound 3 induces cell cycle arrest in the G0-G1 phase, with an observed percentage of 73.15%. The in-silico toxicity prediction was evaluated and demonstrated a good toxicity profile. Finally, molecular docking studies supported these findings by confirming strong binding affinities of the derivatives to VEGFR-2, BRAF, and DHFR.