Therapeutic strategies for human poxvirus infections: Monkeypox (mpox), smallpox, molluscipox, and orf

Abstract

Therapeutic and vaccine development for human poxvirus infections (e.g., monkeypox (mpox) virus, variola virus, molluscum contagiosum virus, orf virus) has been largely deserted, especially after the eradication of smallpox by 1980. Human mpox is a self-limited disease confined to Central and West Africa for decades. However, since April 2022, mpox has quickly emerged as a multi-country outbreak, urgently calling for effective antiviral agents and vaccines to control mpox. Here, this review highlights possible therapeutic options (e.g., tecovirimat, brincidofovir, cidofovir) and other strategies (e.g., vaccines, intravenous vaccinia immune globulin) for the management of human poxvirus infections worldwide.

1. Introduction

Poxviruses are members of the Poxviridae family that are comprised of enveloped, double-stranded DNA viruses shaped like a brick or oval form. A well-known poxvirus is variola virus − the causative agent of smallpox which has been recognized as one of the deadliest diseases in human history [1]. Although the Poxviridae family causes a wide range of pox diseases in humans and animals, only four genera (Orthipoxvirus, Parapoxvirus, Molluscipoxvirus, and Yatapoxvirus) from the Chordopoxvirinae subfamily contain less than 20 poxviruses that cause infections in humans (Fig. 1 ). With their human-to-human transmissibility, variola virus, mpox virus, vaccinia virus, and molluscum contagiosum are responsible for several outbreaks in human history [2,3]. Other human poxviruses such as the orf virus (sore mouth infection) are often transmitted from poxvirus-infected animals to humans, whereas their human-to-human transmissions are usually rare [4,5].

Fig. 1.

The Poxviridae family tree. Human poxviruses from the Chordopoxvirinae subfamily have been highlighted. Recurrent, rare, and eradicated infections are indicated by red, blue, and green designations, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

In the past decades, antiviral research has focused on the development of antiviral agents mainly against human poxviruses from the Orthopoxvirus genus [6,7]. The other poxviruses (e.g., orf virus, sealpox virus, tanapox virus) usually cause self-limiting conditions in healthy individuals (treatment may be unnecessary) and do not trigger large outbreaks through human-to-human transmissions. Although smallpox was eradicated worldwide by 1980, the development of smallpox antiviral agents (e.g., tecovirimat, brincidofovir) and vaccines (e.g., JYNNEOS, ACAM2000) is still needed in preparing any future event of smallpox reemergence [8]. Vaccinia virus is currently studied as a popular tool for gene therapy and genetic engineering [9], but vaccinia and vaccinia-like virus infections are still documented in sporadic reports [4]. Vaccinia virus is known for its wide use in the development of smallpox vaccines, but the vaccinia virus vaccination has been mostly abandoned after the smallpox eradication worldwide. The orf virus primarily causes an infection in sheep and goats, though it can be transmitted to people and cannot be transmitted from one infected person to another. In healthy humans, the orf usually causes a benign self-limiting illness that resolves in 3–6 weeks, but in the immunocompromised patients with HIV, lupus, or cancer therapy, it can be very progressive and even life-threatening [10]. Unlike other human poxviruses that are confined to limited cases and regions, the mpox virus has suddenly spread from endemic countries to more than 100 countries in 2022, giving rise to a global concern of the 2022 mpox outbreak [11,12].

Here, we will provide an overview of the epidemiology and therapeutic strategies against human poxvirus infections, mainly focusing on mpox. Based on accumulated evidence in the literature, we will summarize the epidemiology and clinical manifestations of human poxviruses such as mpox, smallpox, vaccinia virus, molluscum contagiosum virus, and orf virus. Subsequently, potential antiviral agents with in vitro activities will be summarized. Vaccine options were also briefly mentioned for the prophylaxis of human poxvirus infections.

2. Epidemiology

Before 1970, the mpox virus was initially recognized as a nonhuman primate pathogen, until human mpox cases were reported in the Democratic Republic of Congo [13]. It was then reported in the Tshuapa Province of the Democratic Republic of Congo [14,15]. Although the natural host reservoir of mpox is still not known, the mpox virus had been found in almost 14 species of African rodents [16]. From 1971 to 2017, the mpox virus was originally endemic in the tropical rainforest areas of West and Central Africa (Fig. 2 A), where the mpox virus has animal reservoirs [17]. In 2022, however, mpox has unexpectedly spread to non-endemic regions outside of Africa [18], triggering a multi-country outbreak (Fig. 2B). The first case of mpox in the UK was imported from Nigeria on May 4, 2022 [19]. This expanded to a family cluster of three cases of mpox in May 2021 [20]. From April to June 2022, the mpox virus spread over 16 countries [21]. This made Kozlov wonder whether the mpox outbreak could be contained [22]. Palmore and Henderson correctly wondered whether we are now facing a new infection when the clinical and public health resources have been stretched to the limit by COVID-19 pandemic [23].

Fig. 2.

A geographical map of mpox cases

(A) The total number of mpox cases (years: 1970 to 2017) in West and Central Africa was collected from the WHO survey [17]. (B) The geographical distribution of total mpox cases from 2022/01/01 to 2022/11/30. The data was collected from the US CDC (https://www.cdc.gov/poxvirus/monkeypox).

On July 23, 2022, the World Health Organization (WHO) Director-General Tedros Adhanom Ghebreyesus, announced the 2022 mpox outbreak as a Public Health Emergency of International Concern [24]. As of November 30, 2022, mpox has reached >100 countries with a total number of >81,000 mpox cases worldwide (Fig. 2B), but the case-fatality rate is rather low (<0.1%). In the current outbreak, >29,000 mpox cases have been documented in the United States. According to a survey by the US CDC, a majority of these mpox cases were identified among the high-risk population of gay, bisexual, and other men who have sex with men (MSM) [25]. This MSM community was also the principal target of mpox in the UK from April to May 2022 [26].

Anyone can get mpox through close skin-to-skin contact and respiratory droplets from mpox-infected patients, but why the 2022 mpox outbreak is primarily hitting MSM in the United States, the UK, and many other countries? Although the mechanisms are still not fully understood, highly interconnected sexual networks among the MSM community might have established a high opportunity for the mpox spread, especially among those with many sexual partners [27]. Apart from intimate contact during the oral, anal, or vaginal sex or touching the genitals or anus of mpox-infected patients, mpox can be also transmitted by prolonged face-to-face contact, massage, and kissing (https://www.cdc.gov).

3. Clinical manifestations and symptoms

Clinical features of five human poxviruses (mpox virus, variola virus, vaccinia virus, orf virus, and molluscum contagiosum virus) have been summarized in Table 1 . After a certain period of incubation (e.g., mpox: 3–34 days, smallpox: 7–17 days, molluscum contagiosum virus: 2 weeks to 6 months), human poxvirus infections often lead to localized or generalized skin lesions, progressing from papules, to vesicles and scabs. Depending on the species and strains of poxviruses involved, skin lesions often have various features such as lesion areas (e.g., hands, feet, face, genitals), varying sizes, and different shapes (Fig. 3 ). Apart from skin lesions, patients may also experience common symptoms and signs such as rash, fever, lymphadenopathy, headache, and body aches (Table 1).

Table 1.

Overview of five human poxvirus infections.

Poxvirus	Mpox virus	Smallpox (variola virus)	Vaccinia virus	Orf virus	Molluscum contagiosum virus
Discovered year	1958	∼1500 BC	1796	1934	1817
Total cases in 2022	>80,000	No	Some	Many	Many
Incubation period	3–34 days	7–17 days	5–19 days	3–7 days	2 weeks to 6 months
Transmission route	Close contact with mpox lesions, rash, and scabs	Person-to-person contact, virus-containing fluids or contaminated fomites	Close contact with vaccinia-containing lesions, milk or contaminated fomites	Contact with orf-infected animals or contaminated fomites	Close contact with infected skin, contaminated fomites, or sexual contact
Common symptoms	Skin or anogenital lesion, rash, fever, lymphadenopathy, pharyngitis, headache, lethargy, myalgia [21]	Skin lesions, fever, headache, fatigue, rash, facial scarring, pneumonia, blindness, encephalitis [93]	Skin lesions, rash, fever, headache, body aches, nausea, lymphadenopathy [94]	Skin lesions mostly on the fingers, hands, or forearms; mild fever, fatigue, swollen lymph nodesb	Skin lesions and bumpsb
Disease type	A self-limited disease in most cases	A serious and often deadly disease	A self-limited disease in most cases	A self-limited disease in most cases	A self-limited disease in most cases
Approved/repurposed antivirals	Tecovirimata, Brincidofovira, Cidofovira	Tecovirimat, Brincidofovir	VIGIV (Vaccinia Immune Globulin Intravenous, Human); Trifluridine eye drops for vaccinia virus keratitisa [95]	–	–
Approved vaccines	JYNNEOS, ACAM2000a	JYNNEOS, ACAM2000, Dryvax (discontinued)	ACAM2000	–	–

-: No antivirals/vaccines have been approved.

^aOff-label use, which still requires further evidence from clinical trials.

^bInformation was collected from the US CDC (https://www.cdc.gov/poxvirus/).

Fig. 3.

Electron micrograph images (A) and clinical images (B) of mpox, smallpox, molluscum contagiosum, and orf in humans. The micrograph and clinical images for mpox [by Charles D. Humphrey, Tiara Morehead, Russell Regnery, Brian W.J. Mahy], smallpox [by Cynthia S. Goldsmith, Russell Regnery], vaccinia virus [by Moses Grossman], orf virus [by Maureen Metcalfe, Tom Hodge, James H. Nakano], and molluscum contagiosum [by James H. Nakano, Susan Lindsley] were obtained from the Centers for Disease Control and Prevention [http://phil.cdc.gov/phil/home.asp] with the permission for free publication.

During the 2022 mpox outbreak, the clinical presentation of mpox is characterized by anogenital lesions and rashes that relatively spare the face and extremities. The most prevalent sign/symptom is fever followed by inguinal lymphadenopathy and exanthema. Asthenia, fatigue, and headache were described in 22.86% and 25.71% of the subjects, respectively. Myalgia was present in 17.14% of the cases [28]. In another study [29], fever, rash, and lymphadenopathies were reported as possible clinical manifestations of mpox. The appearance of mpox cases with exclusive genital lesions [30], elicited the hypothesis that mpox could be another sexually transmitted infection [31,32]. That mpox can cause genital lesions was documented by Patrocinio-Jesus and Peruzzu [33]. An ulcerating genital rash was also observed in the majority of mpox cases [34].

4. Therapeutic strategies

In 2006, our team reported that antiviral treatment was more effective than smallpox vaccination upon lethal monkeypox (mpox) virus infection [35]. This study was done in cynomolgus monkeys (Macaca fascicularis) after lethal intratracheal infection with mpox virus (Fig. 4 A). The smallpox vaccine used was the Elstree strain, and as the antivirals served cidofovir and HPMPO-DAPy (Fig. 4B).

Fig. 4.

Antiviral agents for human poxvirus infections.

(A) Kaplan–Meier plot of vaccine versus cidofovir and HPMPO-DAPy with the post-exposure effect on the survival of macaques (this subfigure was copied from our previous publication [35]). Arrowheads show the time points of vaccination or antiviral treatment. After intratracheal mpox infection, macaques were sham-treated (group I), vaccinated with smallpox vaccine Elstree-RIVM (group II), treated with five-dose cidofovir (group III), with six-dose cidofovir (group IV), with five-dose HPMPO-DAPy (group V) or with six-dose HPMPO-DAPy (group VI) [35]. Chemical structures of cidofovir (B), HPMPO-DAPy (C), tecovirimat (D), and brincidofovir (E).

Apart from cidofovir and HPMPO-DAPy, tecovirimat and brincidofovir are small-molecule inhibitors that have been approved by the US FDA for smallpox treatment (Table 2 ), though their efficacy has not been evaluated in randomized clinical trials of patients with smallpox. This section will discuss their potential for controlling human poxviruses in some scenarios.

Table 2.

Summary of tecovirimat, brincidofovir, and cidofovir^a.

Antiviral inhibitors	Tecovirimat (TPOXX)	Brincidofovir (Tembexa)	Cidofovir (Vistide)
Approval date	July 2018	June 2021	June 1996
Approved indication	Human smallpox disease in adults and pediatric patients weighing at least 13 kg	Human smallpox disease in adult and pediatric patients, including neonates	Human cytomegalovirus retinitis in AIDS patients
Mechanism of drug action	Tecovirimat targets and inhibits the orthopoxvirus VP37 protein to block its interaction with cellular Rab9 GTPase and TIP47	Brincidofovir inhibits the viral DNA synthesis mediated by the viral DNA polymerase.	Cidofovir inhibits the DNA synthesis mediated by the viral DNA polymerase.
Pharmaceutical form	Capsules for oral use	Tablets or oral suspension for oral use	Solution for infusion
Recommended doseb	•Patients weighting ≥120 kg: 600 mg three times daily for 14 days; •Patients weighting <120 kg and ≥40 kg: 600 mg twice daily for 14 days; •Patients weighting <40 kg and ≥24 kg: 400 mg twice daily for 14 days; •Patients weighting <25 kg and ≥13 kg: 200 mg twice daily for 14 days;	•Patients weighting ≥48 kg: two 100 mg tablets or 20 mL oral suspension (for patients who cannot swallow tablets) once weekly for 2 doses; •Patients weighting ≥10 kg and <48 kg: 4 mg/kg oral suspension once weekly for 2 doses; •Pediatric patients weighting <10 kg: 6 mg/kg oral suspension once weekly for 2 doses	•Induction treatment: 5 mg/kg body weight (given as an intravenous infusion at a constant rate over 1 h) administered once weekly for two consecutive weeks; •Maintenance treatment after the induction treatment: 5 mg/kg body weight (given as an intravenous infusion at a constant rate over 1 h) administered once every two weeks.
Adverse effectsb	Headache, nausea, abdominal pain, vomiting	Diarrhea, nausea, vomiting, abdominal pain	Headache, nausea, vomiting, alopecia, rash, chills, fever, asthenia, neutropenia, renal failure
Drug interactions	Tecovirimat is a weak inducer of cytochrome P450 (CYP)3A and a weak inhibitor of CYP2C8 and CYP2C19. Repaglinide and midazolam should not be administered to avoid drug interactions.	Brincidofovir should not be administered with inhibitors of organic anion transporting polypeptide 1b1 and 1b3	Cidofovir should not be administered with probenecid and tenofovir.

^aInformation was collected from the drug label.

^bEvaluation of drug dosage and adverse reactions in patients with human poxvirus infections has not been reported.

4.1. Tecovirimat (alternative name: ST-246, brand name: TPOXX®)

Tecovirimat has been documented by Grosenbach et al. [36] for the oral treatment of smallpox. It was further hailed as the first-in-class therapy for the emerging mpox outbreak [37]. Delaune and Iseni [38] highlighted the difficulties to develop new drugs targeting an eradicated disease under the FDA “animal efficacy rule” with the few, and imperfect, animal models available. According to this review, the study of Grosenbach et al. [36], allowed the validation of a dosing regimen for oral tecovirimat (600 mg twice daily). Almost a decade before tecovirimat was finally approved by the US FDA on July 13, 2018, it had been the subject of several review articles pointing to its potential in the treatment of orthopoxvirus infections, i.e., smallpox [[39], [40], [41]]. Tecovirimat, as well as cidofovir and brincidofovir, featured (again) among the antiviral agents active against mpox in a 2022 clinically oriented review article by Siegrist and Sassine [42]. The combination of tecovirimat with brincidofovir resulted in synergistic efficacy against orthopoxvirus infections in vitro and in vivo [43]. For the treatment of mpox virus infection, tecovirimat can also be successfully combined with the smallpox vaccine ACAM2000 [44,45].

Tecovirimat was first identified as an egress inhibitor that targets the highly conserved orthopoxvirus VP37 protein to block its interaction with host Rab9 GTPase and TIP47 to prevent the virus from egress [46]. Subsequently, its oral dosage (400 mg once daily for 14 days) was considered effective for the prevention or treatment of smallpox or mpox virus infections in humans [47]. It was then found effective in protecting nonhuman primates from smallpox and mpox virus challenges [48]. Tecovirimat has also proven effective in the postexposure prophylaxis and therapy of rabbitpox in rabbit models [49]. From a pharmacokinetic viewpoint, the dog showed the lowest, and the monkey the highest half-life time in an interspecies allometric scaling of tecovirimat [50]. Still, in 2013 it became also known that tecovirimat was effective against vaccinia virus infection in immunodeficient mice with partial T-cell reconstitution [51]. Tecovirimat was also active against Cantagalo virus, the etiologic agent of a pustular disease in dairy cows and dairy workers in Brazil [52]. A single daily oral dose (400 or 600 mg) of tecovirimat for 14 days was found safe and well tolerated in human volunteers [53], while its efficacy in the treatment of nonhuman primates infected with variola virus (smallpox) was confirmed [54]. Oral administration of tecovirimat at 400 mg daily can be expected to provide plasma concentrations above those effective against orthopoxvirus infections in humans [55]. In cynomolgus macaques, tecovirimat proved effective against a lethal aerosol mpox virus challenge [56].

A recent observational study reported that tecovirimat reduced viral loads and shortened the recovery time in one mpox-infected female who had no HIV, HBV, or HCV infection [57]. A case report described the preemptive use of tecovirimat in a patient with acute myeloid leukemia after smallpox vaccination [58]. To evaluate the safety and efficacy of tecovirimat in mpox-infected patients, randomized placebo-controlled double-blinded trials have been established, including the STOMP study (NCT05534984), the PLATINUM-CAN study (NCT05534165), the UNITY study (NCT05597735), and the study (NCT05559099). These trials are still ongoing and clinical evidence of tecovirimat efficacy will be likely available in 2023.

4.2. Brincidofovir (alternative name: CMX001, brand name: Tembexa®)

Alkoxyalkyl esters of (S)-HPMPA and (S)-HPMPC, i.e. the hexadecyloxypropan-1-ol ester of cidofovir (HDP-cidofovir) are active against orthopoxviruses, i.e. vaccinia virus in cell monolayers and organotypic cultures [59]. They have also been found effective against the orf (contagious ecthyma) virus in vitro [60], and various other orthopoxviruses [61], including ectromelia (mousepox) [62,63]. HDP-cidofovir was subsequently renamed brincidofovir (CMX001) and found effective in the treatment of rabbitpox virus infections in New Zealand White Rabbits [[64], [65], [66]].

In June 2021, brincidofovir - the prodrug of cidofovir - was approved for the treatment of human smallpox disease caused by the variola virus in adults and children. Brincidofovir is now evaluated for its efficacy in the treatment of mpox virus infection in prairie dogs [67]. Given the devastating potential of smallpox as a bioweapon, brincidofovir might be advocated as imperative in a multipronged defense strategy [68] to combat orthopoxviruses, including mpox, at large. Recently, a retrospective observational study indicated that brincidofovir offered no convincing clinical benefit and was associated with a recognized adverse event of deranged liver enzymes in a small cohort of three mpox-infected patients [57]. Nevertheless, the clinical efficacy and safety of brincidofovir in mpox-infected patients should be further evaluated by randomized clinical trials (e.g., NCT01143181).

4.3. Cidofovir (brand name: Vistide®)

In June 1996, cidofovir was approved by the US FDA for the treatment of cytomegalovirus retinitis in AIDS patients. Similar to the mechanisms of other nucleotide analogues, cidofovir is an acyclic cytidine nucleoside phosphonate that blocks the replication of DNA viruses [69,70]. Cidofovir has been well-documented in the therapy and short-term prophylaxis of poxvirus infections [[71], [72], [73]]. Among the poxviruses, cidofovir has shown in vitro activity against orthopox (e.g., vaccinia, variola (smallpox), cowpox, mpox, camelpox, ectromelia (mousepox)], molluscipox (molluscum contagiosum) and parapox (orf) viruses [[71], [72], [73]]. In animal models, cidofovir could reduce the development of orf virus lesions in lambs [74] and rescue mice from lethal mousepox infection [75]. In humans, cidofovir has been successfully used for the treatment of recalcitrant molluscum contagiosum virus and orf virus immunocompromised patients [76].

The era of the acyclic nucleoside phosphonates (ANPs) was born with the advent of (S)-HPMPA [(S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine]. This compound was shown to be effective against various DNA viruses such as the vaccinia virus [77]. Its cytosine counterpart, (S)-HPMPC [(S)--1-(3-hydroxy-2-phosphonomethoxypropyl)cytosine, cidofovir, Vistide®] was shown to be active against vaccinia virus by De Clercq et al. [78]. Its active metabolite, the diphosphate, proved inhibitory to the vaccinia virus DNA polymerase [79]. Although it has been approved only for anti-cytomegalovirus treatment, cidofovir can be possibly repurposed for treating other DNA viruses (e.g., vaccinia virus, molluscum contagiosum, Epstein-Barr virus, adenovirus infection, and human polyoma infections) under certain conditions [[80], [81], [82]].

4.4. (R)-HPMPO-DAPy

(R)-HPMPO-DAPy offers the same antiviral spectrum as cidofovir [35], as further reviewed by E. De Clercq [83]. This means herpes-, polyoma, papilloma-, adeno- and all poxvirus infections, and among the latter, orthopox, parapox and molluscipox, and among the orthopox, smallpox and mpox viruses. This also extends for the latter to the alkoxyalkyl esters, i.e. the hexadecyloxypropyl derivatives, which have so far not been explored and offer unique opportunities for further investigation and possible development to combat poxvirus infections including smallpox and mpox.

5. Perspectives

In addition to antiviral options, vaccines (e.g., JYNNEOS, ACAM2000) and intravenous vaccinia immune globulin are also available for the management of mpox virus infection [84]. In September 2022, JYNNEOS was approved by the US FDA for the prevention of mpox in adults (≥18 years). ACAM2000 has also been recommended by the US CDC for mpox prophylaxis. In principle, smallpox vaccines likely protect people exposed to mpox [85]. An example in point of a such vaccine is the Modified Vaccinia Ankara (MVA) strain [86]. In addition to vaccines and therapeutics, nonpharmaceutical interventions (e.g., safe sex, good hygiene, avoiding contact with infected subjects) are also important for the prevention of human poxvirus infections in current and future outbreaks.

To date, the US FDA has advised in favor of two inhibitors, tecovirimat and brincidofovir, for the treatment of smallpox (Table 2). Although tecovirimat and brincidofovir have not yet been approved by the US FDA for mpox treatment, they are currently under evaluation for those patients at risk of severe mpox disease. Cidofovir is not FDA-approved for the treatment of variola virus infections but could be used during an outbreak under an appropriate regulatory mechanism (such as an investigational new drug protocol or Emergency Use Authorization).

In addition to mpox, progressive vaccinia virus infection which might develop as a potentially lethal complication in smallpox vaccine recipients with severely impaired cellular immunity may also be considered an important indication for the therapy with vaccinia immune globulin, tecovirimat, and brincidofovir [87]. Future novel anti-orthopoxviral strategies to combat mpox virus infection may be based, for example, on methylene blue derivatives [88] and adamantane derivatives [89]. The ternary inclusion complex containing hydroxypropyl-β-cyclodextrin and meglumine might be a potential formulation strategy for enhancing the solubility and bioavailability of tecovirimat [90].

The mpox strains in 2022 have segregated in a divergent phylogenetic branch, likely reflecting continuous accelerated evolution [91]. Furthermore, multiple lineages of the mpox virus have been detected [92], giving rise to the concern about drug resistance and compromised vaccine effectiveness. Future drug development may focus on pan-poxvirus small-molecule inhibitors and their combinations, ideally delivered orally, to control current and future outbreaks of evolving human poxviruses.

Funding

This study was supported by the National Nature Science Foundation of China (31871324, 81730064, 31571368) and National Science and Technology Major Project of China (2018ZX10715004). The funders had no role in study design, data collection, data analysis, data interpretation, or writing of the report.

CRediT authorship contribution statement

Erik De Clercq: Conceptualization, Writing – original draft, Supervision. Yike Jiang: Data collection, Data curation, Visualization. Guangdi Li: Visualization, Conceptualization, Writing – review & editing, Validation, Funding acquisition.

Declaration of competing interest

The authors disclose no conflicts.