Pharmacological Cyclin-Dependent Kinase Inhibitors Inhibit Replication of Wild-Type and Drug-Resistant Strains of Herpes Simplex Virus and Human Immunodeficiency Virus Type 1 by Targeting Cellular, Not Viral, Proteins

Abstract

Pharmacological cyclin-dependent kinase (cdk) inhibitors (PCIs) block replication of several viruses, including herpes simplex virus type 1 (HSV-1) and human immunodeficiency virus type 1 (HIV-1). Yet, these antiviral effects could result from inhibition of either cellular cdks or viral enzymes. For example, in addition to cellular cdks, PCIs could inhibit any of the herpesvirus-encoded kinases, DNA replication proteins, or proteins involved in nucleotide metabolism. To address this issue, we asked whether purine-derived PCIs (P-PCIs) inhibit HSV and HIV-1 replication by targeting cellular or viral proteins. P-PCIs inhibited replication of HSV-1 and -2 and HIV-1, which require cellular cdks to replicate, but not vaccinia virus or lymphocytic choriomeningitis virus, which are not known to require cdks to replicate. P-PCIs also inhibited strains of HSV-1 and HIV-1 that are resistant to conventional antiviral drugs, which target viral proteins. In addition, the anti-HSV effects of P-PCIs and a conventional antiherpesvirus drug, acyclovir, were additive, demonstrating that the two drugs act by distinct mechanisms. Lastly, the spectrum of proteins that bound to P-PCIs in extracts of mock-and HSV-infected cells was the same. Based on these observations, we conclude that P-PCIs inhibit virus replication by targeting cellular, not viral, proteins.To ensure specificity and avoid toxicity, most antiviral drugs are designed to target viral proteins. Such drugs, however, select for drug-resistant viral mutants. Moreover, these drugs exhibit activity against only a few closely related viruses. In contrast, antiviral drugs that target cellular proteins required for viral replication would not be constrained by these limitations. In the past several years, pharmacological cyclin-dependent kinase inhibitors (PCIs) have been shown to inhibit the replication of four clinically important viruses: human cytomegalovirus (HCMV) (6), herpes simplex virus type 1 (HSV-1) (56-58), human immunodeficiency virus type 1 (HIV-1) (9, 47, 69), and varicella-zoster virus (J. Moffat, State University of New York, Upstate Medical University, personal communication). However, it is as yet unclear whether the antiviral effects of these drugs are mediated exclusively by inhibition of their known cellular targets, or by inhibition of yet-unknown viral targets.Of the PCIs developed to date, the 2,6,9-trisubstituted purines (P-PCIs), such as Roscovitine (Rosco) (45) and Purvalanol (Purv) (26), are the most specific and best characterized. Rosco and Purv differ in potency (Purv is more potent than Rosco [26,45]) but not in selectivity or mechanism of action. Both drugs inhibit cdk1, -2, and -5 and erk1 and -2 (at Ϸ50-to 1,000-fold higher concentrations than are needed to inhibit cdks), but they do not inhibit cdk4 or -6 or a large number of other kinases (26,36,45). Mechanistically, Rosco and Purv compete with ATP for binding to the ATP-binding pocket of the target cdks (16,26,45,68). All known effects of Rosco and Purv on cells can be attributed to inhi...

Herpes simplex virus-type 2 infectivity and agents that block gap junctional intercellular communication

“…The antiviral xanthate D609 prevented HSV-1 replication by inhibiting the viral US3 protein kinase, protein kinase C, and viral protein phosphorylation (Walro and Rosenthal, 1997). Conversely, pharmacological cyclin-dependent kinase inhibitors exert their antiviral effects on herpes viruses by blocking cellular rather than viral proteins (Schang et al, 2002). Clearly the phosphorylation of both viral and cellular proteins is important for virus replication and protein kinase inhibitors exert their anti-viral effects by preventing phosphorylation of cellular and/or viral proteins.…”

p42/p44 MAPKs are intracellular targets of the CDK inhibitor purvalanol

“…In addition to CDKs, p42/p44 MAPKs were found to interact with the purvalanol matrix, in most models: lugworm oocytes, Xenopus oocytes, porcine and human brains (Knockaert et al, 2000; data not shown), rat tissues (Knockaert et al, 2000; data not shown), rat hepatocyte primary cultures (data not shown) and in seven different mammalian cell lines (Figure 5b; Knockaert et al, 2000;Schang et al, 2002). These observations are quite striking, given the large difference between the IC 50 values of purvalanol for CDKs and MAPKs in vitro (Figure 4).…”

An inhibitor of cyclin-dependent kinases suppresses TLR signaling and increases the susceptibility of cancer patients to herpes viridae

“…After written informed consent was obtained, peripheral blood (PB) samples were obtained from patients enrolled in the Phase I dose-escalation study, as an ancillary study to better characterize the toxicity observed in the clinical study. 13 Patients bearing advanced/metastatic solid tumors were treated with repeated cycles of PHA-793887 administered as a 1 hour IV infusion on day 1, 8, 15 of a 4-week cycle, i.e., with treatment on days 1, 8, 15 followed by a 2 week resting period (day [16][17][18][19][20][21][22][23][24][25][26][27][28]. In 3 patients, before and after each injection, heparinized blood was taken on days 0, 2, 14 and 16 ( Fig.…”

“…These observations could appear paradoxical in as much as recent studies have shown that replication of Herpes Simplex Virus (and other large DNA viruses) requires cellular proteins including CDKs. [14][15][16][17][18][19][20][21] One of the potential mechanisms accounting for these adverse effects might be the negative regulation of TLR signaling pathways by CDKi and/or GSK3β inhibition, as suggested by previous in vitro studies.…”

“…Tumor associated-mutations deregulate distinct CDKcyclin complexes causing unscheduled proliferation. Deregulation of CDK4 and CDK6 activities have been implicated in a variety of human malignancies, CDK4 being hyperactive in epithelial a 4-week cycle, i.e., with treatment on days 1, 8, 15 followed by a 2-week resting period (day [16][17][18][19][20][21][22][23][24][25][26][27][28] (Fig. 1A).…”