genus of the family Poxviridae
contains a number of pathogens known to infect humans, including variola virus (VARV, the causative agent of smallpox), cowpox virus, camelpox virus, vaccinia virus, and monkeypox virus (MPXV). Human infection with members of this genus results in varying degrees of morbidity and mortality. Virions are enveloped and brick-shaped, with a dumbbell shaped core containing the genetic material [1
contain a single, linear piece of double-stranded DNA with highly conserved central regions and more variable terminal ends [1
]. The proteins expressed from the terminal ends are predominantly involved in immunomodulation and/or host range determination [2
VARV, the etiological agent of smallpox, causes an acute, systemic lesional disease with a mortality rate of approximately 30% [5
]. Eradicated in 1977, smallpox remains a constant threat due to its potential use as a biological weapon for mass dissemination to a largely unprotected worldwide population. Unfortunately, VARV is not the only member of the Orthopoxvirus
genus that causes severe disease in humans and has the potential for development as a biological weapon. The global eradication of smallpox and the subsequent cessation of smallpox vaccination in 1980 allowed for the emergence of another lethal zoonotic disease, monkeypox.
Similar to smallpox, monkeypox is a systemic lesional disease with a prodrome period of flu-like symptoms (fever, malaise, chills, headache) followed by the development of a progressive maculopapular rash which expands in a centrifugal pattern and progresses from papules to vesicles to pustules and finally to crusts [7
]. MPXV is a zoonotic virus endemic in the Democratic Republic of the Congo (DRC) where it regularly emerges from reservoir species, including squirrels and other rodents [12
], to cause serious disease outbreaks in humans. The best estimate of mortality rate is approximately 10%; however, this is likely an underrepresentation due to sporadic reporting since 1986 and a lack of information concerning the complete geographic range of human monkeypox disease [9
There are 2 distinct clades of MPXV, West African and Central African. MPXV strains belonging to the West African clade are far less virulent than Central African strains in both humans and non-human primates, with diminished morbidity and human-to-human transmissibility [19
]. The MPXV outbreak that occurred in the Midwestern United States in 2003 was caused by a West African strain of MPXV and thus resulted in less severe disease than what is typically seen in outbreaks in Central Africa [21
]. This outbreak did, however, demonstrate the ability of MPXV to reach beyond the African continent and cause disease in MPXV-naïve populations. Although outbreaks of Central African monkeypox have not been seen outside of Africa, predictions based on an ongoing active disease surveillance study in the DRC suggest that spread to a MPXV-naïve population could have significant public health impacts. This study was conducted in nine health zones in the DRC and revealed a dramatic increase in monkeypox cases, with 760 laboratory confirmed cases identified from 2005 to 2007 [18
]. Although previous vaccination against smallpox was found to still confer significant protection, only approximately 25% of the population in the sampled health zones had evidence of past vaccination [18
]. Data suggesting that the incidence of human-to-human transmission of MPXV is on the rise in this region is also concerning [18
] and could suggest that fading herd immunity coincident with a rise in the number of unvaccinated persons is allowing for more efficient spread. Additionally, it is possible that genetic variants are emerging that are more highly adapted to humans. Taken together with a long incubation period, which allows for a significant period of time during which a person is potentially contagious but asymptomatic, and its potential use as a biological weapon, it is evident that the development of therapeutic methods to treat active MPXV infections is critical.
In this paper, we investigate the potential use of interferon (IFN)-β as an anti-MPXV therapeutic. IFN-β is already US Food and Drug Administration (FDA) approved for the treatment of multiple sclerosis in four forms: Betaseron, Rebif, Avonex, and Extavia. All of these products have well-defined safety records for human use (FDA).
IFN-β is a type I IFN that plays a key role in the innate immune response by promoting the production of IFN-stimulated genes that inhibit protein synthesis, induce apoptosis, and activate macrophages and natural killer cells [23
]. Additionally, type I IFNs enhances the adaptive immune response by upregulating major histocompatibility complex-I/II expression on the surface of antigen-presenting cells [23
IFNs have been generally overlooked as anti-Orthopoxvirus
agents due to the large number of immunomodulatory proteins expressed by viruses belonging to this genus. To date, 13 Orthopoxvirus
proteins have been shown to have anti-IFN activity: A46, A52, K7, N1, B14, K1, M2, COP-B19, B8, H1, E3, K3, and C7 [26
]. Recently, a 14th protein was identified, VARV-G1R, which binds to NF-κB and inhibits NF-κB regulated gene expression [27
]. Some of these proteins have also been shown to play key roles in host range determination and virulence during vaccinia virus infection. Unfortunately, most of these proteins have not been fully characterized, and the activity of orthologs expressed by MPXV and VARV has not been extensively investigated at this time. One of the best characterized of these proteins is E3. E3 has been studied in vaccinia virus and is known to block the activation of PKR [28
]. Although VARV contains a full length and fully functional E3L, MPXV lacks the N-terminal domain responsible for binding Z-DNA and PKR [11
]. Removal of this domain results in a decreased virulence in murine models of vaccinia infection [32
]. K3 is a homolog of eIF-2α that sequesters PKR thereby preventing phosphorylation of native eIF-2α by PKR [33
]. It is a host range gene that is expressed by both VARV and vaccinia virus but not by MPXV [11
]. C7 and K1 have also been shown to affect the cell tropism of vaccinia virus [37
]. Although their exact functions are less well understood, it is believed that they employ a novel mechanism to antagonize IFN [37
]. While the role of VARV-G1 in host range restriction has not been explicitly demonstrated, G1 orthologs are present in some of the most highly pathogenic Orthopoxviruses
, including VARV and MPXV, but not in vaccinia virus, suggesting that this protein may be a key virulence factor [27
The detailed comparative study of COP-B19 orthologs from MPXV and VARV represents the first cross-species functional analysis of any of the anti-IFN immunomodulators [42
]. In this study, COP-B19 (aka B18 or IFN α/βR) from vaccinia virus was found to react very strongly with human and murine IFN-α and IFN-β. In contrast, the VARV ortholog, B17, bound to murine IFN-β very poorly but bound to human IFN-β better than vaccinia B18. Although this study didn't give as detailed of a description of the binding properties of MPXV B16, it did suggest that immunomodulatory proteins such as COP-B19 may play significant roles in host range restriction. Additionally, it showed that the analysis of Orthopoxvirus
immunomodulatory proteins cannot be limited to vaccinia virus but must be carried out for all Orthopoxviruses
as orthologs may function differently and/or be affected to varying degrees by the host immune response. The genetic and functional variability of the immunomodulatory proteins necessitate that prophylactic or therapeutic agents that are intended to overcome the action of these proteins be tested for efficacy with all Orthopoxviruses
as the susceptibility of these viruses may vary significantly.
Although IFN-β has been shown to substantially diminish vaccinia virus pathogenesis in vivo
], the susceptibility of MPXV to IFN-β is uncertain. In this study, we found that MPXV production and release were significantly reduced in the presence of IFN-β. Additionally, IFN-β was able to inhibit MPXV when introduced 6-8 h after infection, revealing its potential for use as a therapeutic against established infections. IFN-β treatment was able to induce the expression of the antiviral protein MxA during MPXV infection, and constitutive expression of MxA was able to inhibit virus production. Collectively, the data show that IFN-β is a strong novel candidate for further investigation as a prophylactic and therapeutic against MPXV.