Two-, three-, and four-drug antiretroviral combinations in either simultaneous or sequential regimens were evaluated for their ability to suppress human immunodeficiency virus (HIV) type 1 replication in vitro. Zidovudine, lamivudine, saquinavir, and nevirapine were used at IC(90)s, IC(99)s, or IC(> or = 99)s in a CD4-positive human lymphoblastoid cell line (H9 cells) acutely infected with HIV-1. In sequential regimens, drugs were added at weekly intervals. In simultaneous regimens, all drugs were added on day 0. Increasing the number of drugs in a combination regimen both increased the degree of viral inhibition and delayed the time of breakthrough viral replication. Simultaneous regimens provided more profound and earlier viral inhibition than did sequential regimens. However, sequential addition provided relatively more durable viral inhibition than did simultaneous regimens when drug concentrations were low. The relative effectiveness of different HIV-1 therapeutic strategies depends on both the numbers and concentrations of the drugs used.