The human polyomavirus, JCV, is the etiologic agent of a fatal central nervous system (CNS) demyelinating disease known as progressive multifocal leukoencephalopathy (PML). PML occurs predominantly in immunosuppressed patients and remains an intractable complication in AIDS. To date, there are no effective therapies to treat PML. We previously demonstrated that the neuroleptic drug, chlorpromazine, inhibits the endocytic pathway used by JCV to infect glial cells. In this paper, we demonstrate that nontoxic doses of chlorpromazine are effective at inhibiting JCV multiplication and spread in a tissue culture model. The clinical efficacy of this drug or related compounds in treating PML has not been evaluated.