Lupeol and Paclitaxel cooperate in hindering hypoxia induced vasculogenic mimicry via suppression of HIF-1α-EphA2-Laminin-5γ2 network in human oral cancer

Abstract

Vasculogenic mimicry (VM), defined as an endothelial cell independent alternative mechanism of blood and nutrient supply by dysregulated tumor cells, is associated with poor prognosis in oral squamous cell carcinoma (OSCC). Here we aim to investigate the underlying molecular mechanism of the synergistic effect of phytochemical Lupeol and standard microtubule inhibitor Paclitaxel in reversing the hypoxia induced VM formation in OSCC. The results demonstrated that the hypoxia induced upregulation of HIF‐1α led to augmentation of signaling cascade associated with extracellular matrix remodeling and EMT phenotypes that are mechanistically linked to VM. Induction of HIF‐1α altered the expression of EMT/CSC markers (E‐Cadherin, Vimentin, Snail, Twist and CD133) and enhanced the ability of cell migration/invasion and spheroid formation. Subsequently, the targeted knockdown of HIF‐1α by siRNA led to the perturbation of matrigel mediated tube formation as well as of Laminin‐5γ2 expression with the down‐regulation of VE‐Cadherin, total and phosphorylated (S‐897) EphA2, pERK1/2 and MMP2. We also observed that Lupeol in association with Paclitaxel resulted to apoptosis and the disruption of VM associated phenotypes in vitro. We further validated the impact of this novel interventional approach in a patient derived tumor explant culture model of oral malignancy. The ex vivo tumor model mimicked the in vitro anti‐VM potential of Lupeol‐Paclitaxel combination through down‐regulating HIF‐1α/EphA2/Laminin‐5γ2 cascade. Together, our findings elucidated mechanistic underpinning of hypoxia induced Laminin‐5γ2 driven VM formation highlighting that Lupeol‐Paclitaxel combination may serve as novel therapeutic intervention in perturbation of VM in human OSCC.

Lupeol synergizes with 5-fluorouracil to combat c-MET/EphA2 mediated chemoresistance in triple negative breast cancer

“…Western blotting was performed as previously described. 64 Total cell lysates (50 μg) were resolved by SDS-PAGE and transferred to Polyvinyldene di-fluoride (PVDF) membrane by semi-dry electroblotting (BIO-RAD). After non-specific protein blocking with 5% non-fat dry milk for 1 h at room temperature, the membranes were incubated with primary antibodies overnight at 4°C followed by incubation with appropriate horseradish peroxidase (HRP) tagged secondary antibodies for 1 h at room temperature.…”

Advances in Small Molecular Agents against Oral Cancer

“…In addition, paclitaxel inhibits the growth of oral cancer cell lines by inhibiting the EGFR signalling pathway [90]. Paclitaxel combined with lupeol inhibits the simulation of hypoxia-induced angiogenesis [91]. Ursolic acid (57) induces caspase-dependent cell apoptosis by down-regulating the expression of multiple biomarkers, including Akt/mTOR/NF-xB signalling [92].…”

“…Betulinic acid (61) inhibits the proliferation of OSCC cells by regulating ROS and p53 signalling, making it a potential drug for the treatment of oral cancer [96]. Lupeol (62) can promote the apoptosis and inhibit the proliferation of OSCC cells by inducing the phosphorylation of EGFR and inhibiting the activation of downstream molecules, such as protein kinase B (or AKT) and NF-κB [91,97]. Zhang et al [98] found that 20(S)-ginsenoside Rh2 (63) induces the apoptosis and inhibits the growth of OCC by inducing G0/G phase arrest and significantly down-regulating the levels of p-Src, p-B-Raf, and p-ERK1/2 proteins.…”

Exploring the dynamic interplay between cancer stem cells and the tumor microenvironment: implications for novel therapeutic strategies

“…CSCs play a role in this process through vasculogenic mimicry (VM), where they transform into endothelial-like cells, forming tube-like structures without true endothelial cells [ 233 ]. Hypoxia in the tumor environment activates various signaling pathways (e.g., HIF, c-Myc, Sox-2, Oct-4, adenosine/STAT3/IL-6 pathway, MAPK/ERK pathway, Notch, Wnt, Hedgehog, and Hippo signaling) [ 234 ], promoting the multi-directional differentiation potential of CSCs and influencing CSC transformation into incomplete endothelial cells that exhibit VM in different tumor types (e.g., oral squamous cell carcinoma, breast cancer, liver cancer, colorectal cancer, and cholangiocarcinoma) [ 235 – 239 ].…”