*3.2. Human Cytomegalovirus (HCMV)*

HCMV belongs to the β-herpesvirus family, with a high prevalence, infecting up to 80% of the general population usually asymptomatic in healthy people [45]. Diseases associated with HCMV include glandular fever and pneumonia. HCMV is also an important pathogen in organ transplant patients responsible for significant morbidity and mortality in organ transplant recipients, and a major cause of disease in patients with HIV infection [46]. HCMV infections in newborns may result in hearing loss, mental retardation and palsy [47]. The available FDA-approved therapeutic options for HCMV infection include ganciclovir, foscarnet, cidoforvir, and fomivirsen [48]. These drugs have different mechanisms of actions or applications, and represent the successes that had been made against the challenges of HCMV [48–50]. Several anti-HCMV drugs were reported to have low potency, poor oral bioavailability, and adverse side effects [50]. Moreover, drug resistance strains were reported to emerge [51]. Hence, there has been a renewed interest in search of new inhibitors of HCVM [50,51]. Of greater concern is the increase in the number of people living with transplanted organs, and the increase in HIV infected people [52]. Technological advancements have enabled organ transplants to be more accessible while the increase in HIV-infected individuals is due to new retroviral therapies that have converted HIV infection to chronic disease as infected people live longer, leading to increased cases of HCMV infection [52]. In 2011, the first case of HCMV-treated with AIC246, a novel anti-CMV compound that targets the viral terminase complex and remains active against virus resistant to DNA polymerase inheritors was reported [53], which represents a good example of the renewed interest for HCMV inhibitors [53]. Almost 60 patents claiming novel agents for the treatment of HCMV were launched from January 1996 to 2000, but so far none of these projects has led to the approval of an anti-HCMV drug [49]. However, the recent FDA approval of letermovir (Figure 3), providing a long-awaited alternative for preventing cytomegalovirus infection in allogeneic hematopoietic stem cell transplant recipients [54] is very encouraging.

**Figure 3.** PREVYMIS™ (letermovir) a recently (2017) FDA-approved drug for the prevention of *Human cytomegalovirus* (HCMV) infection and disease in organ transplant patients.

A class of myxobacterial compounds, myxochelin, belonging to a larger group of natural products, siderophores, were isolated from several strains of myxobacteria [53,55]. Siderophores are secondary metabolites produced by some microorganisms under iron-limiting conditions, and enhance the uptake of iron [56]. Other siderophores isolated from myxobacteria includes nannochelins and hylachelins. Various studies have revealed myxochelins to be potent antitumour agents [55,57–59]. The antitumour activity was demonstrated to be caused by inhibition of human 5-lipoxygenase (5-LO) [58]. Surprisingly the inhibition of 5-LO by myxochelins was found not correlating with the iron affinities [58]. The enzyme 5-LO is responsible for the catalysis of two initial steps in the biosynthesis of leukotriens, starting from arachidonic acid [58]. Leukotriens are well-known mediators of a variety of allergic reactions such as inflammatory, rheumatic arthritis, allergic rhinitis and cardiovascular diseases [58]. Importantly, 5-LO pathways were associated with cancer proliferation, hence explaining the observed strong anticancer activity of myxochelin [58,60]. Nannochelins are reported to have no significant antimicrobial activity [24].

Myxochelin A (**12a**) was initially isolated from the culture broth of *Angiococcus disciformis* (strain An d30). Later on, myxochelins B (**12b**), C (**12c**), D (**12d**), E (**12e**), and F (**12f**) were isolated and also synthesised (Figure 4) [60,61]. The corresponding biosynthetic gene clusters have been identified in *Stigmatella aurantiaca* (Sga 15), and *Sorangium cellulosum* (Soce 56) [62]. Additional siderophores have been isolated from *Nannocystis exedens* (**21a**–**21c**) and *Hyalangium minutum* (**20a**–**20c**) [63,64]. Myxochelin C (**12c**) inhibited HCMV with an IC50 value of 0.7 μg/mL [46,53]. It could in future become feasible to test others among the over 500 different siderophores that are known to science [65]. In particular, the known myxobacterial-derived siderophores, such as nannochelins (**21a**–**21c**), hylachelins (**20a**–**20c**), and all the other myxochelin analogues (Figure 4) should be screened for various antiviral activities, especially anti-HCMV, and should be studied for structure activity relationship for possible discovery of more potent antivirals.

#### *3.3. Ebola Virus Disease (EVD)*

Ebola haemorrhagic fever is caused by the Ebola virus (EBOV), a single stranded RNA enveloped virus belonging to the family *Filoviridae*. EVD first appeared in 1976 in two simultaneous outbreaks, one in Nzara, South Sudan, and the other in Yambuku, the Democratic Republic of Congo. The latter occurred in a village near the Ebola River, from which the disease takes its name [65,66]. EVD case fatality rate is around 50%, with different cases from 25% to 90% fatality in past outbreaks reported [65]. Furthermore, EBOV is known to persist in immune-privileged sites, such as testicles, inside of the eye, and central nervous system, and in some people who have recovered from EVD [65]. The effect of the persistence is yet to be known [66].

**Figure 4.** Myxobacterial-derived siderophores.

The re-emergence of Ebola occurred in West African countries causing 11,308 deaths leading to the WHO on 8 August 2014, declaring the epidemic to be an international public health emergency [66]. An experimental Ebola vaccine called rVSV-ZEBOV has been reported to show high protection against EVD [67]. No drug or licensed vaccine currently exists; hence there is an urgent need for drugs that inhibit entry or multiplication of EDV [62,67]. Developing an assay to test compounds for anti-EBOV poses a significant challenge because of the cost of equipment for the high risk, Biosafety S4 organism,

involved [66]. However, various metabolites from myxobacteria were screened for EBOV inhibition by an assay with a surrogate system using Ebola envelope glycoprotein GP-pseudotyped lentiviral vectors (Figure 5) [67]. GP-pseudotype lentiviral vectors were used as tools to investigate the entry process of the viruses, enabling studies without the need of using the native Ebola virus reducing the safety level from the highest level 4 to level 2 [67]. The same analysis was conducted with vesicular stomatisis virus (VSV)-G-pseudotyped vectors to determine the EBOV-specificity of the inhibitory function of the compounds. Chondramides (**13a**–**13d**) were reported to inhibit EBOV-GP-mediated transduction with impressive IC50 values of 24–42 nM. The VSV-G-mediated transduction was less efficient, with an IC50 value of 55–111 nM [66]. Chondramides (**13a**–**13d**), a class of compounds known to interfere with actin, were isolated from a myxobacterium belonging to the genus *Chondromyces* [68]. Members of the genus *Chondromyces* belong to those myxobacteria known to synthesise two or more chemically unrelated secondary metabolites with different mechanisms of action [69]. Other promising hits were the noricumazoles, a family of polyketides from *Sorangium cellulosum*. Noricumazole A (**14a**) was found to inhibit EBOV-GP with an IC50 value of 0.33 μM. In fact, **14a** was found to be EBOV-GP specific and showed no significant inhibition against (VSV)-G-pseudotyped vectors [67]. Noricumazoles are known to be potassium channel blockers with **14a** known to be highly toxic while the derivatives, **14b** and **14c**, are equally active with lower toxicity [67,70]. The screening of myxobacterial natural compounds library resulted in the identification of inhibitors of EBOV-GP pseudotyped vectors, chondramides and noricumazole, whose mechanism of action is actin-stabilising and the channel blockers respectively [67]. These metabolites will give insights into the EBOV infection mechanism, rather than being used as drugs, because the modes of actions are expected to have side effects. However, the lower toxicity of **14b** and **14c**, which are derivatives of **14a**, is exciting and qualifies **14a** to be considered as a lead structure for the development of EBOV inhibitors.

**Figure 5.** Structures of myxobacterial-derived compounds with activity against Ebola virus (EBOV). Chondramides (**13a**–**d**) are known to be actin inhibitors, while noricumazols (**14a**–**c**) are known potassium channel inhibitors [70,71].

#### *3.4. Hepatitis C Virus*

HCV is an enveloped, single-stranded RNA virus with positive polarity (ss (+) RNA). HCV is transmitted by blood-to-blood contacts, such as through intravenous injections, blood transfusion, and various exposures to blood contaminants. It can also be transmitted by contact with bodily fluids including saliva or semen of an infected person [72]. By 2015, there were 71 million people infected with HCV globally [73]. HCV and hepatitis B virus (HBV) infection are the major causes of hepatocellular

carcinoma (HCC), associated with cirrhosis [74]. Currently, no products are available to prevent HCV infection. There are some drugs available that can cure HCV infection [75]. However, treatment is complicated by HIV-HCV/HBC co-infections with drug-drug interactions between anti-HIV and anti-HCV drugs, resulting in serious side effects and can lead to the death of patient [8]. The discovery of broad-spectrum antivirals may play an essential role in overcoming this challenge.

The recently isolated compounds from *Labilithrix luteola*, labindoles A (**15a**), and B (**15b**) have been reported to have moderately inhibited HCV (Figure 6) [75]. Interestingly the labindoles were said to have no cytotoxicity, anti-bacterial or antifungal activities. 3-chloro-9H-carbazole (**17**) and 4-hydroxymethyl-quinoline (**18**) also isolated from *Labilithrix luteola* were reported to have a strong inhibition of HCV [75]. Soraphens are a family of polyketide-derived macrolactones comprising over 50 metabolites known for strong antifungal activity [76]. However, recent studies have suggested that soraphen A (**16**) inhibits HCV replication in HCV cell culture models expressing subgenomic and full-length replicons as well as a cell culture-adapted virus with an IC50 value of 5 nM [11,77]. The HCV assay involved the development of subgenomic replicons that replicate autonomously in the human hepatoma cell line Huh-7 to be able to screen for anti-HCV activity. The subgenomic replicons are genetic materials from HCV, which represent the actual invasion and replication of HCV on the liver cells [75]. Furthermore, **16** is known to depolymerise the acetyl-CoA carboxylase (ACC) complexes into less active dimers [77]. The mechanism of action of **16** is a valuable probe to study the roles of ACC polymerisation and enzymatic activity in viral pathogenesis [77]. Various minor structure alterations of **16** did not affect the antiviral activity [11]. Owing to the fact that soraphens inhibit both HIV and HCV, it has been proposed that the broad-spectrum activity of **16** could be due to targeting commonly used host factors or pathways necessary for viral replication [78]. Another recently isolated myxobacteria-derived secondary metabolite, lanyamycin (**22**) from *Sorangium cellulosum* (strain Soce 481) moderately inhibited HCV with IC50 value of 11.8 μM [78]. The macrolide, **22**, is closely related to the bafilomycins, a class of secondary metabolites from actinobacteria [79]. Interestingly bafilomycin A was previously shown to possess good activity (IC50 value of 0.1 nM) against influenza A virus, which is below its cytotoxic levels [80]. Screening of **22** for activity against influenza A virus and other pathogenic viruses, and studying the mode of action would be interesting.

**Figure 6.** Compounds with activity against Hepatitis C Virus.
