Hepatitis C virus (HCV) is a major cause of liver cirrhosis and hepatocellular carcinoma worldwide. We previously reported that cyclosporin A (CsA) inhibits HCV-1b replication. However, its inhibition of JFH-1 (HCV-2a) was much less. Since HCV genotype clearly affects the in vitro and in vivo response to anti-viral therapy, we wished to examine the effect of CsA and its non-immunosuppressive derivative NIM811 on HCV genotype 4a replication. We first established an in vitro system supporting HCV-4a infection and replication using immortalized human hepatocytes, HuS-E7/DN24 (HuS) cells, and these cells were infected with sera obtained from Egyptian patients with chronic HCV-4a infection. HuS cells supported more robust HCV-4a replication than both HuH-7.5 and PH5CH8 cells, and HCV-4a infection and replication were completely inhibited by 3 μg/ml CsA and 0.5 μg/ml NIM811. Thus, HuS cells are a good model system supporting the infection and high-level replication of HCV-4a, and both CsA and NIM811 effectively inhibit HCV-4a replication in this system.