Suppression of Adenovirus Replication by Cardiotonic Steroids

Abstract

The dependence of adenovirus on the host pre-RNA splicing machinery for expression of its complete genome potentially makes it vulnerable to modulators of RNA splicing, such as digoxin and digitoxin. Both drugs reduced the yields of four human adenoviruses (HAdV-A31, -B35, and -C5 and a species D conjunctivitis isolate) by at least 2 to 3 logs by affecting one or more steps needed for genome replication. Immediate early E1A protein levels are unaffected by the drugs, but synthesis of the delayed protein E4orf6 and the major late capsid protein hexon is compromised. Quantitative reverse transcription-PCR (qRT-PCR) analyses revealed that both drugs altered E1A RNA splicing (favoring the production of 13S over 12S RNA) early in infection and partially blocked the transition from 12S and 13S to 9S RNA at late stages of virus replication. Expression of multiple late viral protein mRNAs was lost in the presence of either drug, consistent with the observed block in viral DNA replication. The antiviral effect was dependent on the continued presence of the drug and was rapidly reversible. RIDK34, a derivative of convallotoxin, although having more potent antiviral activity, did not show an improved selectivity index. All three drugs reduced metabolic activity to some degree without evidence of cell death. By blocking adenovirus replication at one or more steps beyond the onset of E1A expression and prior to genome replication, digoxin and digitoxin show potential as antiviral agents for treatment of serious adenovirus infections. Furthermore, understanding the mechanism(s) by which digoxin and digitoxin inhibit adenovirus replication will guide the development of novel antiviral therapies.IMPORTANCE Despite human adenoviruses being a common and, in some instances, life-threating pathogen in humans, there are few well-tolerated therapies. In this report, we demonstrate that two cardiotonic steroids already in use in humans, digoxin and digitoxin, are potent inhibitors of multiple adenovirus species. A synthetic derivative of the cardiotonic steroid convallotoxin was even more potent than digoxin and digitoxin when tested with HAdV-C5. These drugs alter the cascade of adenovirus gene expression, acting after initiation of early gene expression to block viral DNA replication and synthesis of viral structural proteins. These findings validate a novel approach to treating adenovirus infections through the modulation of host cell processes.

Elucidation of the mechanism of anti-herpes action of two novel semisynthetic cardenolide derivatives

“…The mechanism by which cardenolides affect several steps of HSV replication could be related to the inhibition of Na + / K + -ATPase in host cells. Many studies have demonstrated the modulation of Na + /K + -ATPase functions in host cells by DNA viruses (adenoviruses [18], cytomegalovirus [23,[70][71][72], and HSV [27,28]), and RNA viruses (chikungunya virus [19,73], coronaviruses [20,74,75], respiratory syncytial virus [76,77], Ebola virus [78,79], influenza virus [33,80,81], and HIV [30,31,82]). By activating signaling cascades or by altering the concentration of intracellular ions, the binding of cardenolides to Na + /K + -ATPase seems to create an unfavorable environment for viral replication.…”

“…Among them, their cytotoxic and antitumor effects recently reviewed by Cerella et al [10], De et al [11], Diederich et al [12], Schneider et al [13] and El-Seedi et al [14], as well as their anti-inflammatory [15], antiprotozoal [16], anti-oxidant and anti-aging [17] activities can be cited. Another suggested possibility is their potential antiviral action, as reported by several authors against adenovirus [18], chikungunya virus [19], coronavirus [20,21], cytomegalovirus [22][23][24], dengue virus [25], herpes virus [26][27][28], HIV [29][30][31], human papillomarivus (HPV) [32], influenza virus [33][34][35], and respiratory syncytial virus [36] replication. The effects of six well-known cardiac glycosides (digoxin, digitoxin, ouabain, convallatoxin, G-strophanthin and lanatoside C) on viral biology and the mechanisms by which they impair the replication of different RNA and DNA viruses were recently compiled [37].…”

<p>Management of Adenoviral Keratoconjunctivitis: Challenges and Solutions</p>

“…More recently, they have also shown efficacy in cancer treatments, in that treated patients demonstrated a lesser chance of relapse. 88 Hartley et al first noted its potential antiviral benefit, noting its efficacy against HAdV and herpesviruses. 89 This is due to the drugs inhibiting Na+/K+ ATPase on the cell surface, which leads to increased intracellular levels of NA+ and, subsequently, Ca++.…”

“…As such, these drugs seem to be efficacious against HAdV serotypes A-D and serve as potential treatment therapies for the treatment and prophylaxis of EKC. 88 Thus, repurposing of these cardiotonic steroids as an antiviral agent for short-term treatment of adenoviral keratoconjunctivitis as well as a prophylactic for use in individuals in close contact with patients with epidemic keratoconjunctivitis. 88 Furthermore, Marrugal-Lorenzo et al reported that mifepristone, a synthetic steroid drug, possesses anti-adenoviral properties via its interference with viral entry into the nucleus.…”

Drug repurposing for new, efficient, broad spectrum antivirals

“…The cardiac glycosides digoxin and ouabain have also been shown to impair replication of human cytomegalovirus (HCMV) and herpes simplex virus (HSV) that belong to the family of dsDNA viruses Herpesviridae (Dodson et al, 2007;Hartley et al, 2006;Kapoor et al, 2012). Inhibition happens at an early post-entry step and produces a (Grosso et al, 2017). Although the precise mechanism of action is not fully clear, these drugs altered viral mRNA processing, blocking replication before viral DNA synthesis.…”