Stable ASCT2 and LAT1 Suppression in Human Hepatocellular Carcinoma Cells Fails to Measurably Affect Proliferation, Anchorage-independent Growth and mTORC1 Signaling

Amino acid transporters ASCT2 and LAT1 are coordinately enhanced in human cancers where, among other roles, they are thought to drive mammalian target-of-rapamycin (mTOR) growth signaling. To assess their value as therapeutic targets in hepatocellular carcinoma cells (HCC), both transporters were targeted and suppressed with lentiviral vectors producing specific short hairpin RNA in SK-Hep1 cells. Five ASCT2 and four LAT1 stable knockdown cell lines were produced (each with a unique shRNA sequence) with proportionally suppressed glutamine and leucine transport, respectively, and suppressed transporter expression at the mRNA and protein level by 20% to 50% relative to controls. Suppression of each transporter failed to affect cell proliferation. In addition, soft agar colony formation was not compromised in transporter-suppressed cells; neither was basal or amino acid-stimulated mTOR complex 1 (mTORC1) signaling as assessed by 4E-BP1 phosphorylation. No evidence was found for an enhanced reliance on autoph...