Due to the success of a variety of organometallic polymers as anticancer agents a number of these compounds were tested for their efficacy as antiviral agents. There are a few compounds which show promising activity against vaccinia virus (dsDNA) and none against zika virus (+RNA).
Vaccinia virus is an Orthopoxvirus within the Poxviridae family of viruses . It is a double-stranded DNA virus that was previously used in the vaccination program against smallpox, which was eradicated through the World Health Organization (WHO) in 1979. Although smallpox has been eradicated, research into the disease remains invaluable, as smallpox remains a potential agent for bioterrorism. The worldwide vaccination program against smallpox was stopped in 1980, and thus, an outbreak of smallpox today would decimate the current population who lack smallpox immunity . Smallpox is caused by two viruses: variola major and variola minor, with variola major producing most of the fatal disease burden, with a case-fatality rate of 20% or more. Vaccinia virus is the vaccine strain of smallpox, and although its origins are still unclear after over 100 years of research, it is surmised that vaccinia could be the product of genetic recombination.
Vaccinia virus is a unique virus because, although it is a DNA virus, it is replicated exclusively in the cytoplasm of infected cells through a viral DNA-dependent RNA polymerase . It is also unique in that vaccinia produces two types of virions: intracellular mature virus (IMV) and extracellular enveloped virus (EEV), both of which are infectious . The IMV virions are surrounded by a single membrane, while the EEV virions possess two membranes and are critical in the dissemination of the virus. The external membrane of the EEV virions is derived from the host cell, and thus, these virions can evade both host antibody and host complement immune responses.
Upon contact with a competent host cell, vaccinia virions enter the cell through phagocytic vacuoles where they are partially stripped of their outer core, releasing the vaccinia virus dsDNA genome into the cytoplasm . Once inside the cytoplasm, the DNA-dependent RNA-polymerase of the virus transcribes the vaccinia viral genome into messenger RNAs (mRNAs) . The transcribed genes can be classified as early, intermediate, and late genes. The early vaccinia genes are needed for viral DNA replication and for assisting the virus in escaping the host’s innate immune response. The intermediate genes encode for transcriptional regulators of the late genes, and the late genes encode the structural proteins and enzymes necessary to construct the new virions. Inside cytoplasmic inclusions known as virus factories, the vaccinia virus dsDNA is packaged into an immature virion (IV), with proteolytic cleavage leading to the formation of the intracellular mature virus (IMV), which are released through cell lysis. Some virions, however, will become wrapped in Golgi-derived membranes and are known as intracellular enveloped virus (IEV). These intracellular enveloped virus particles are transported to the surface of the cell, where they can either remain at the cell surface as CEV (cell- associated enveloped virus), or they are released as extracellular enveloped virus (EEV).
Vaccinia virus is a dsDNA virus with a unique intracytoplasmic replication mechanism. Although smallpox has been eradicated worldwide, vaccinia remains relevant because of its potential use in biowarfare. Using standard plaque reduction assays, several organometallic polymers were tested against vaccinia virus for their ability to inhibit viral replication in 143 cells. Most polymers were not active; however, the derivatives of 3-AT (Table 2) showed some inhibition, protecting between 15 and 35% of cells from infection.
In Table 3, for example, all the organometallic/dicumarol polymers showed over 100% virus production. In this case, and in the case of other groups of polymers, the compounds made the cells more susceptible to lysis, resulting in an enhanced production of visible plaques compared to the number of visible plaques for the virus-infected control wells. Thus, in these instances, there was not an increase in virus production, but an increase in susceptible cells.
The organotin polymers derived from camphoric acid and lamivudine also showed the ability to inhibit vaccine replication (Table 4). The inhibition was mild at best.
Table 5 contains vaccine plaque reduction assay results for various organotin polymers. All polymers showed decent inhibition of vaccinia viruses.
In summation, a number of organotin polymers exhibit some inhibition of the vaccinia virus to merit further testing of newly synthesized polymers.
Zika virus, an enveloped, plus-stranded RNA virus belonging to the Flavivirus genus, was first discovered in Uganda in 1947 . Usually, Zika virus causes asymptomatic infection; if symptoms do occur, they include a low-grade fever, itchy rash, and arthralgia . The concern for Zika virus, however, emanates from its potential to cause severe neurological disease, such as microcephaly in newborns, as well as a handful of cases which involved development of Guillain–Barre syndrome, which causes the immune system to attack the peripheral nervous system . It gained much publicity during the recent Olympics in Brazil because of the fear by the participants, particularly women, and attenders of contracting the zika virus.
Zika virus particles contain an inner nucleocapsid surrounding the genomic RNA, and the nucleocapsid is wrapped in an envelope that contains the viral membrane protein (M) and the viral envelope protein (E) . The RNA of Zika virus encodes 3,423 amino acids which are translated as a large polyprotein, which is subsequently cleaved into 10 + individual viral proteins. The viral nonstructural protein NS3 has helicase and nucleoside triphosphatase activities, while NS5 is the viral RNA-dependent RNA polymerase which is required for viral genome replication. Zika virus infects human neural progenitor cells (hNPCs) through clathrin-mediated endocytosis. The acidic environment of the endosome induces conformational changes in the viral envelope (E) glycoprotein leading to fusion between the viral and endosomal membrane and subsequent release of the zika virus RNA into the cellular cytoplasm. The viral RNA-dependent RNA polymerase is responsible for replication and translation of the viral genome, along with currently unidentified cellular factors. Immature virions bud into the endoplasmic reticulum, where they receive their cellular-derived envelope with embedded viral prM (membrane) and envelope proteins. Immature virions complete the process of maturation as they proceed through the trans-Golgi network, and the virions are eventually released from the cell through exocytosis (Figs. 9, 10, 11, 12, 13, 14).
Zika virus is a single-stranded, plus-sense RNA virus that has garnered worldwide attention recently due to its connection to neurological birth defects. Table 6 shows the results for the compound assays for zika virus. Unlike vaccinia virus, in which a plaque assay technique was used to assess the efficacy of the compounds, we could not get zika virus to produce defined plaques, so we used a cytopathic effect assay. For the organotin polymers, results indicate that two groups of compounds show promise as antiviral agents against zika virus. These compounds are derived from camphoric acid and lamivudine.
Lamivudine (Fig. 15; also called 3TC) is a potent reverse transcriptase prodrug antivirial molecule employed in the treatment of AIDS [26,27,28,29,30]. Structurally, lamivudine is a nucleoside analogue. It is administered several times daily because of its short half-life of 5–7 h. It has additional problems including negative effects from the accumulation of the drug, high cost, and lack of patient compliance. Further, there is an increased incidence of co-infection of HIV with such diseases as tuberculosis. Co-treatments are being investigated. For instance, co-loaded polymer microspheres containing lamivudine and an anti-tuberculosis drug such as isoniazid have been described that allow the treatment of both the HIV and tuberculosis .
All the organotin polymers derived from lamivudine and camphoric acid exhibit total inhibition of the zika virus, strain 502. They all inhibited infection of Vero cells at 0.025 µg/mL. Against Zika strain 502, the products from diethyltin dichloride and camphoric acid and diphenyltin dichloride and camphoric acid inhibited infection of Vero cells at concentrations of 0.000391 and 0.000781 μg/mL, respectively. Thus, it appears that the organotin and camphoric acid-derived compounds merit further testing against zika virus as potential novel antiviral agents. These concentrations are in the nano or near nanogram region (Fig. 16).
In hopes of determining additional products that exhibit good inhibition of the zika virus several polymers that had structural characteristics like camphoric acid were tested for their ability to inhibit the zika virus. We identified possible structural characteristics that we have begun to explore. Camphoric acid has two acid groups and further a ring system from which the acid groups are attached. We have synthesized many polymers that exhibit good anticancer activity. Table 6 contains results for some of these. Figure 17 contains the structures for some of these. In no case was any inhibition of the zika virus found. Additional structures were studied but thus far with no success. We continue to seek further structures that will inhibit the zika virus.
The influence of the polymeric nature was briefly studied. A dimer of the dibutyltin camphoric acid was synthesized through simply reacting the camphoric acid with tributyltin chloride. Because the organotin reactant has only one reactive group, it can only react at the ends of the camphoric acid without chain extension (Fig. 18).
The dimer showed the ability to inhibit the zika virus, Table 6, but inhibition was significantly less compared with the dibutyltin/camphoric acid polymer. Thus, the polymeric nature is positive in this case.