NUCLEAR-LOCALIZATION SIGNAL OF HIV-1 AS A NOVEL TARGET FOR THERAPEUTIC INTERVENTION

Background: Human immunodeficiency virus type 1 (HN-1) is a lentivirus and shares with other members of this retroviral subfamily the abilit y to replicate in nondividing cells, in particular, cells of the monoc yte/macrophage lineage. This feature relies on the presence of a speci fic nuclear localization signal (NLS) within the viral matrix protein (MA p17), which to some degree can be complemented by the activity of the viral vpr gene product. The MA p17 NLS ensures efficient transport ation of the viral preintegration complex into the nucleus of an infec ted macrophage and confers persistence of HIV-1 in quiescent T cells, and therefore presents an attractive target for therapeutic interventi on. Materials and Methods: Nuclear localization signals (NLS) in gener al and the HIV-1 MA p17 NLS in particular are characterized by a stret ch of positively charged amino acids including one or more lysine resi dues. A series of compounds potentially capable of binding and reactin g with lysine by forming Schiff base adducts was synthesized. Our spec ial consideration was to make compounds that would preferentially bind to two closely contiguous amino functions, as opposed to isolated sin gle lysine residues. We assumed that this approach might specifically target the compound to NLS while affecting other regions less, thus re ducing nonspecific cytotoxicity. Antiviral activity was assessed in pr imary monocytes and in peripheral blood lymphocytes (PBL) infected wit h HIV-1(ADA) strain. Viral replication was monitored by reverse transc riptase (RT) activity in the supernatant. Efficiency of nuclear import ation of the viral preintegration complex was estimated by the formati on of 2-LTR circle forms of HIV-1 DNA and also by in situ PCR techniqu es. Results: Arylene bis(methyl ketone) compounds with a nitrogenous t hird substituent, especially a pyrimidinic side-chain, inhibited HIV-1 replication in human monocytes at an IC50 as low as 1 nM. These compo unds did not block HIV-1 replication in peripheral blood lymphocyte cu ltures. The inhibitory effect observed in monocyte cultures appeared i n the context of markedly reduced nuclear importation of viral DNA in the presence of the drug. No cytotoxic effects of the compounds was ob served in vitro at concentrations as high as 10 mu M. An amidinohydraz one derivative of the most active compound was about 100 times less ac tive than the parent, indicating that carbonyl,groups were instrumenta l in the antiviral effect. Conclusions: These early results suggest th at retroviral replication in nondividing cells is susceptible to pharm aceutical intervention targeted against the NLS activity of HIV-1 prot eins in the viral preintegration complex. The compounds described effi ciently block translocation of viral DNA to the nuclei of infected pri mary monocytes, and inhibit viral replication. This inhibition is effe ctive only in nondividing cells and is not seen in proliferating cultu res, such as activated PBLs. Thus, drugs that target HIV-1 NLS may be useful to specifically block the macrophage arm of HIV infection and c ould thereby be of value in treating macrophage-specific manifestation s of HN disease, such as HIV-1 dementia. In combination with other dru gs, potential therapeutics exploiting this target may also help to con trol the progression of HIV-1 infection and disease.