Despite the huge efforts employed to implement novel chemotherapeutic paradigms for lung cancer, the disease still remains a major concern worldwide. Targeting molecular pathways as Hedgehog (Hh) and Mitogen-activated protein kinase (MAPK) represent a new hope in lung cancer treatment. This work was undertaken to evaluate the antitumor effects of GANT61 (5 μM), BI-847325(30 μM), and GANT61 (5 μM)/BI-847325(30 μM) combination on A549 adenocarcinoma lung cancer cell line. The growth inhibition 50 (GI50) for both drugs was performed using MTT. The protein levels of Caspase-3, Bcl-2-associated X protein (Bax), Myeloid cell leukemia sequence 1 (MCL-1), cyclin D1, vascular endothelial growth factor (VEGF), extracellular signal-regulated kinases (ERK), p-Akt, and phosphohistone H3 (pHH3) were measured using ELISA. Glioma-associated oncogene homolog 1(Gli1) gene expression was assessed by quantitative real-time PCR. The GI50 for GANT61 and BI-8473255 were 5 µM and 30 µM, respectively. Caspase-3 and Bax protein levels were significantly elevated while MCL-1, cyclin D1, VEGF, ERK 1/2, p-Akt, and pHH3 levels were significantly reduced by both drugs and their combination relative to the control group. Gli1 gene expression was down-regulated in all groups relative to the control group. GANT61, BI-847325 and their combination inhibited proliferation and angiogenesis but activated the apoptotic pathway. Both drugs conferred a profound negative impact on the crosstalk between each of Hh and MAPK pathways and Phosphoinositide 3 -kinases (PI3K)/Akt/Mammalian target of Rapamycin (mTOR). To the best of our knowledge, the antitumor effects of BI-847325/GANT61 combination have not been tested before. Further in-vitro and in-vivo studies are warranted to support the findings.