Hydroxyurea, a drug widely used in therapy of several human diseases, inhibits deoxynucleotide synthesis—and, consequently, DNA synthesis—by blocking the cellular enzyme ribonucleotide reductase. Hydroxyurea inhibits human immunodeficiency virus-type 1 (HIV-1) DNA synthesis in activated peripheral blood lymphocytes by decreasing the amount of intracellular deoxynucleotides, thus suggesting that this drug has an antiviral effect. Hydroxyurea has now been shown to block HIV-1 replication in acutely infected primary human lymphocytes (quiescent and activated) and macrophages, as well as in blood cells infected in vivo obtained from individuals with acquired immunodeficiency syndrome (AIDS). The antiviral effect was achieved at nontoxic doses of hydroxyurea, lower than those currently used in human therapy. Combination of hydroxyurea with the nucleoside analog didanosine (2′,3′-dideoxyinosine, or ddl) generated a synergistic inhibitory effect without increasing toxicity. In some instances, inhibition of HIV-1 by hydroxyurea was irreversible, even several weeks after suspension of drug treatment. The indirect inhibition of HIV-1 by hydroxyurea is not expected to generate high rates of escape mutants. Hydroxyurea therefore appears to be a possible candidate for AIDS therapy.

Human immunodeficiency virus type 1 (HIV-1) viral DNA synthesis in quiescent and activated peripheral blood lymphocytes (PBLs) was studied. Incomplete viral DNA (previously demonstrated to be associated with HIV-1 virions) is carried by HIV-1 virions into quiescent and activated PBLs, contributing to the formation of an early viral DNA pool in these cells. The viral DNA is subsequently completed but only extremely slowly and inefficiently in quiescent PBLs compared to that in stimulated PBLs. We find that this correlates with significantly lower levels of dNTP substrates in quiescent compared to activated PBLs. At these low dNTP concentrations, HIV-1 reverse transcriptase acts in a partially distributive manner. Increasing dNTP concentrations from the levels of quiescent PBLs to the levels of activated PBLs increases the processive action of reverse transcriptase, which in turn stimulates rapid and efficient formation of full-length DNA. Furthermore, hydroxyurea treatment of stimulated PBLs decreases the dNTP levels and the DNA synthesis rate to levels comparable to quiescent PBLs. Our data therefore indicate that low levels of dNTP may explain why HIV-1 DNA is synthesized slowly and inefficiently in quiescent PBLs and suggest that pharmacologic induction of low dNTP levels represents a therapeutic approach for inhibition of HIV-1 replication.

Several effective SARS-CoV-2 vaccines are currently in use, but effective boosters are needed to maintain or increase immunity due to waning responses and the emergence of novel variants. Here we report that intranasal vaccinations with adenovirus 5 and 19a vectored vaccines following a systemic plasmid DNA or mRNA priming result in systemic and mucosal immunity in mice. In contrast to two intramuscular applications of an mRNA vaccine, intranasal boosts with adenoviral vectors induce high levels of mucosal IgA and lung-resident memory T cells (T_RM); mucosal neutralization of virus variants of concern is also enhanced. The mRNA prime provokes a comprehensive T cell response consisting of circulating and lung T_RM after the boost, while the plasmid DNA prime induces mostly mucosal T cells. Concomitantly, the intranasal boost strategies lead to complete protection against a SARS-CoV-2 infection in mice. Our data thus suggest that mucosal booster immunizations after mRNA priming is a promising approach to establish mucosal immunity in addition to systemic responses.

HIV-associated nephropathy (HIVAN) is now the third leading cause of end-stage renal disease in the African American population. HIV-1 infects renal tubular and glomerular epithelial cells or podocytes, cells that are a critical part of the filtration barrier. HIV-1 infection induces the loss of podocyte differentiation markers and increases podocyte proliferation. It has been previously shown that HIV-infection induces loss of contact inhibition. Here, the HIV-1 gene responsible for proliferative changes is identified by using cultured podocytes in vitro. The HIV-1 proviral construct, pNL4-3 was rendered noninfectious by replacing the HIV-1 gag/pol sequences with an EGFP reporter gene (pNL4-3: ΔG/P-EGFP). This construct was then pseudotyped with VSV.G envelope to infect podocytes that were conditionally immortalized with SV-40 T antigen. In addition, mutated constructs were engineered with premature stop codons in the HIV-1 env, vif, vpr, vpu, nef, or rev genes. The parental construct and all the other mutated constructs, with the exception of nef, induced proliferation under nonpermissive conditions and anchorage-independent growth (colony formation in soft agar) under permissive conditions. In contrast, deletion of nef markedly reduced proliferation and colony formation. Although tat alone, or tat plus rev induced marginal levels of anchorage-independent growth, coexpression with nef significantly increased colony formation. Finally, stable expression of Nef in a retroviral vector, pBabe-puro, was sufficient to induce increased proliferation and colony formation. Moreover, nef induced saturation density and loss of contact inhibition. These data indicate that Nef induces multiple proliferative effects in podocytes in culture and that nef may therefore be an important gene in the pathogenesis of HIVAN in vivo.