Knockout of PAK1 and PAK4 supresses tumour growth associated with vasculogenic mimicry inhibition through EphA2-VE-cadherin-MCAM pathway

Pancreatic ductal adenocarcinoma (PDA) remains largely refractory to anti-angiogenic strategies, and non-endothelial perfusion mechanisms such as vasculogenic mimicry (VM, endothelial-like channels formed by tumour cells) may sustain tumour progression. Here, we examined whether the combined knockout of p21-activated kinase 1 and 4 (PAK1&4) affects vascular mimicry (VM) programmes in pancreatic cancer. KPC wild-type or PAK1&4 knockout cells were injected subcutaneously into immunodeficient mice. Knockout of PAK1& 4 suppressed tumour growth, associated with VM inhibition, but not endothelial angiogenesis. Knockout of PAK1& 4 reduced tumour expression of VM markers EphA2, VE-cadherin and MCAM, and decreased EphA2⁺VE-cadherin⁺ and EphA2⁺MCAM⁺, CD31⁺VE-cadherin⁺ and CD31⁺MCAM⁺ cells. In vitro, PAK1 knockout and PAK1& 4 knockout suppressed VM-like tube formation and migration, whereas PAK4 knockout enhanced tube formation. Global proteomics linked PAK1 knockout to downregulation of EPH-Ephrin signalling and reduced EphA2-MCAM-RhoA abundance, while PAK4 knockout enriched blood-vessel morphogenesis molecules. These findings identify a PAK-dependent VM programme and suggest that dual PAK targeting inhibits tumour growth by VM inhibition.