Alirocumab therapy effectively suppresses colorectal cancer cell Growth and invasion in nude mouse liver-PCSK9 is a key driver of PI3K/Akt/p-Bad-mediated cell proliferation and antiapoptotic signaling

Background: This study assessed whether alirocumab-facilitated 5-FU therapy effectively inhibited colorectal cancer (CRC) cell proliferation/growth in the livers of nude mice. Methods and Results: In vitro results revealed that (a) cell viability at 6, 24, 48 and 72 h, (b) wound healing ability, (c) cell migratory ability, (d) protein levels of cell proliferation/antiapoptotic signalings, and (e) protein expressions of cell cyclins were significantly and progressively reduced from groups A 1 (DLD-1 cells) and A 2 (HCT116 cells), groups A 3 (siRNA-PCSK9 in DLD-1) and A 4 (siRNA-PCSK9 in HCT116), groups to A 5 (DLD-1 cells + alirocumab 10 µM) and A 6 (HCT116 cells + alirocumab 10 µM) (n = 3-6) ( all p < 0.001 ), whereas the protein expression of apoptosis exhibited contrasting effects (n = 3) ( all p < 0.001 ). These parameters, including: (a, b, c, d and e) and colony formation units (CFUs) were significantly and progressively reduced from groups B 1 (DLD-1), B 2 (DLD-1 + alirocumab 5 µM) to B 3 (DLD-1 + alirocumab 20 µM) (n = 3-6) ( all p < 0.001 ), and also significantly and progressively from groups C 1 [PCSK9 gene overexpression in DLD-1 (PCSK9 OE in DLD-1)], C 2 (PCSK9 OE in DLD-1 + alirocumab 5 µM) to C 3 (PCSK9 OE in DLD-1 + alirocumab 20 µM) (n = 3-6) ( all p < 0.001 ). Protein expression of PCSK9 was significantly and progressively increased from groups D 1 (CCD-18Co), D 2 (DLD-1) to D 3 (HCT116), and also significantly and progressively increased from groups from D 4 (THLE-2), D 5 (Huh7) to D 6 (Hep3B) (n = 3) ( all p < 0.001 ). For the animal study (n =10 for each group), two CRC cell lines were implanted into livers of nude mice and were categorized into groups G A1 (DLD1), G A2 (HCT116), G B1 (DLD1 + alirocumab), G B2 (HCT116 + alirocumab), G C1 (DLD1 + 5FU), G C2 (HCT116 + 5FU), G D1 (DLD1 + alirocumab + 5FU) and G D2 (HCT116 + alirocumab + 5FU). By day 35, the ratio of liver weight to body weight, mean bioluminescence intensity at days 21, 28 and 35, protein expressions of cell proliferation, cell cyclins, cellular levels of PCSK9 and extracellular matrices were significantly and progressively reduced from G1, G2, and G3 to G4 (n = 6-10) ( all p < 0.0001 ). Conclusion: Alirocumab-facilitated 5-FU therapy effectively suppresses PCSK9-promoted CRC proliferation/growth/invasion, highlighting that alirocumab therapy may be an alternative choice of adjuvant therapy in combination with surgery or chemotherapy.