Exploration of the treating the host concept, using repurposed agents acting on the renin angiotensin system – ATC C09.
From the preceding discussion, a framework for treating the host and repurposing can be employed to explore the eligibility of drugs that influence the renin angiotensin system as candidates for clinical trials. In particular to identify what is known about these agents in regard to:
1) Pathogenesis 2) Target homology of COVID-19 3) Prevention/ Treatment and 4) Dose-Response curve in relation to safety.
1. Pathogenesis. To treat the host requires an understanding of the physiology of this response to that condition (in this case COVID). In addition, knowledge is needed of chemical, structural and immunological homology with the novel action of the repurposed drug and the molecular events that drive the consequences of viral pathogenesis in COVID-19 disease. As the main causes of death in COVID are pulmonary vasoconstriction, pulmonary oedema, thrombosis, and fibrosis, a focus on RAS inhibition, a key driver of (a) Pulmonary Complications – pro-thrombotic and inflammation in the lung (b) Innate Immune Response and (c) Vascular Response - vasoconstriction, inflammation and coagulation is key.
1a. Pulmonary Complications – pro-thrombotic and inflammatory lung disease. During early 2020, an enormous amount was written on the pathogenesis of COVID-19, including comprehensive summary review articles with strong emphasis on pulmonary pathophysiology in this condition (11-13, inter alia ). The relationship between the dysregulation of the pulmonary renin angiotensin system (RAS) and the progression to severe acute respiratory syndrome (SARS) including the intermediate stages of infections and pneumonia has been discussed at length. By way of summary the fundamental initiation is the role of ACE2 as the receptor for SARS coronavirus infection (14). It is the impact of this initiation that drives a pulmonary dysregulation of the finely balanced RAS. This balance involves the generation of angiotensin II by ACE and the inactivation of that peptide by ACE2. The functional role of ACE2, its catalytic product angiotensin1-7 and the MAS receptor are the counterbalance moieties for angiotensin II and its AT1R receptor (11). The interaction of angiotensin 1-7 with MAS receptors promotes an anti-inflammatory response, vasodilation an anti-fibrotic response and is seen as protective. Down regulation of ACE2 in acute lung injury models with SARSCoV infection supports a protective role for RAS blockade in this form of pulmonary infection (11). The core hypothesis is that a dysregulated RAS is core to the pathophysiology of COVID-19 driven lung damage and acute respiratory distress syndrome (15). This builds upon much earlier experimentation showing the angiotensin II receptor blocker (ARB) losartan could effectively reduce acute lung injury induced by SARS-CoV (16).The summary statement is most apt:“Therefore, in acute lung injury, ACE, Ang II, and AT1R function as lung-injury-promoting factors, while ACE2 protects from lung injury” (17).
It follows that from a repurposing standpoint that there exists a strong target homology with agents that promote RAS inhibition and the pathogenesis of COVID-19 disease in the lung in terms of pathophysiology and pharmacology.
1b. Innate Immune Response. Angiotensin II, via activation of the AT1R receptor, is pivotal in modulating immune responses. As highlighted by Veerappan et al 2008 (18) mast cells are found in the submucosal tissue in human bronchi in the upper and lower respiratory tree. Mast cells rapidly recognise pathogens and mount an immune response that includes plasma extravasation, immunological and cytokine modulation, leukocyte recruitment and ultimately cellular inflammation (19). What is critical is the role of mast cells in the bronchi in the release of renin and the initiation of the production of the angiotensin II that acts through AT1R receptors (18). Moreover, in an experimental model of hypersensitivity, anaphylactic mast cell degranulation has been shown to result in angiotensin bronchoconstriction (18). Of significance is the observation that mast cell chymase can generate angiotensin II independently of ACE activity (20), the salient point being that ACE is membrane bound and chymase upon release moves within interstitial spaces. ACE also modulates macrophage and neutrophil function in its role in the innate and adaptive response (21).
Angiotensin II mediates its pro-inflammatory responses by signalling through AT1R receptor.
These responses include, leukocyte recruitment, increased production of reactive oxygen species by receptor (AT1R) activation of NADPH oxidase, contributing to inflammation and the stimulation of the innate immune responses (summarised in 21). As mentioned above angiotensin 1-7 serves as a counter to the pro-inflammatory influence of angiotensin II. Angiotensin 1-7 achieves this by acting through the MAS receptors expressed at the surface of the bronchial smooth muscle and alveolar epithelium (12). With the viral infection in COVID-19 the cytokine storm is a key characteristic. The release of large amounts of pro-inflammatory cytokines and chemokines (IFNα, IFNγ, IL-1β, IL-6, IL-12, IL-18, IL-33, TNFα, TGFβ, CXCL10, CXCL8, CXCL9, CCL2, CCL3,CCL5) by immune effector cells drives the aberrant inflammatory response (22).
Many cells of the innate immune system have been shown to have an important role in the development of human COVID lung. Pulmonary RAS has a key role in bronchoconstriction consistent with the release of renin and subsequent angiotensin II formation with mast cell degranulation (18). Moreover, in an animal model of hypersensitivity mast cell degranulation was associated with angiotensin II mediated constriction, inhibited with an AT1R blocker or an inhibitor of renin (18). From a repurposing standpoint that there exists a strong target homology with agents that promote RAS inhibition and the modulation of innate immune responses in the lung.
1c. Vascular; vasoconstriction, inflammation and coagulation.
There is growing evidence that there exists a confluence of vascular and diffuse alveolar damage with SARS and possibly also COVID-19 disease. In some of the data with COVID-19 pneumonia, McGonagle et al (23) have drawn attention to the similarities of blood vessel wall oedema, modest vessel wall immune cell infiltration, hyaline thrombosis and haemorrhagic change described earlier for patients with SARS, consistent with the observation that disseminated intravascular coagulation may develop late in COVID-19 disease. Consistent with this view Teuwen et al 2020 (24) suggested that pulmonary endothelial cells (ECs) have been largely overlooked as a target in COVID-19, as the attending ECs altered vessel barrier function promotes a pro-coagulative vascular inflammatory state and an inflammatory cell infiltration. They also highlight the fact that SARS-CoV-2 binds to the ACE2 receptor impairing the enzyme that is the counterbalance for the vasoactive angiotensin II. The lung pathology COVID-19 disease displays microvascular thrombosis and haemorrhage with extensive alveolar and interstitial inflammation termed lung-restricted vascular immunopathology (23). In the early stage is distinct from diffuse pulmonary intravascular coagulopathy. Cao and Li (25) also similarly highlighted the two hallmarks of critical patients with COVID-19 disease namely progressive inflammation and the trend to hypercoagulation. Furthermore, they draw attention not only to the inflammatory state as a trigger for coagulation, but endothelial cell disseminated intravascular coagulation, characterising the COVID-19 hypercoagulable state as displaying elevated D-dimer (a fibrin degradation product) and fibrinogen concentrations and prolonged prothrombin time (25).
Previous studies have shown that in pulmonary hypertension there is a reduced ACE2 activity and that augmentation of ACE2 reduced oxidant and inflammatory markers as well as improving pulmonary hemodynamics (26). Thirty years ago, our team established that in hypertensive blood vessels the endothelial cell mediated relaxation was impaired and this could be reversed with impairment of prostanoid system or treatment with the ACEI captopril (27). Collectively demonstrating a key role of the vascular endothelium in coagulation, inflammation and vasoconstriction and the capacity for abnormal function driven by the RAS. The RAS influence on the vasculature also extends to trophic factors. The neurotrophic peptide NGF is believed to be a pro inflammatory mediator in airways and promotes bronchial hyperresponsiveness, Freund-Michel et al. (28). Of significance was our demonstration that the AT1R blocking agent losartan reversed the abnormal concentrations of NGF in blood vessels in hypertension (29).
As we indicated earlier there is growing evidence that there exists a confluence of vascular and diffuse alveolar damage with SARS and possibly COVID-19. Gonzalez‑Jaramillo et al 2020 in a commentary on the double burden of COVID-19, also implicates the RAS in chronic pulmonary disease such as pulmonary fibrosis and pulmonary arterial hypertension (30). Similarly, PAH is characterized by reduced ACE2 activity and the production of Angiotensin-(1–7) by ACE2 activating Mas receptors activation improves experimental models of PAH (26).
As pointed out by Cao and Li 2020 (25), SARS-CoV-2 can initially replicate in the upper respiratory track and later in the disease course replicate in the lower respiratory tract generating a secondary viremia with a subsequent attack on organ targets expressing ACE2. A dynamic consistent with the possibility outlined by Kreutz et al 2020 (11) and other groups that ACEIs and ARBs may be associated with lower incidence and/or improved outcome in patients with lower respiratory tract infections. From a repurposing standpoint that there exists a strong target homology with agents that promote RAS inhibition that involves blood vessel and vascular endothelial cell damage.
2. Target homology of COVID-19 and therapeutic agents .
SARS-CoV-2 is an RNA virus and they have high mutation rates up to a million times higher than their hosts, and it is this high rate that enables the viruses to avoid host immunity and to achieve drug resistance (Pachetti et al 2020- (31). From the standpoint of drug repurposing the selected drug should target, not the virus, but that aspect of the pathophysiology of COVID-19 that is tightly linked to the conserved mutational characteristics of the virus and its class.
2a. Conserved target for SARS-CoV-2. It is now generally accepted that ACE2 is the receptor for SARS-CoV and SARS-CoV-2 to enable entry to the host (32). Moreover, South et al 2020 (33) suggested that angiotensin may promote both lung injury and ARDS in SARS-CoV, SARS-CoV-2/COVID-19 and possibly in MERS.
Using crystal structures for NL63 coronavirus (NL63-CoV) and SARS coronavirus (SARS-CoV) receptor binding domains Wu et al 2011 demonstrated virus binding hot spots on their common receptor, the human angiotensin-converting enzyme 2 (ACE2). From that experimentation Wu et al 2011 (34) suggested that the hot spot features were amongst the driving force for the convergent evolution of these two viruses. It follows that this would infer multiple binding opportunities for differing or mutated viruses but restricted to a common protein target. The targeted physiological process for driving pulmonary inflammation will be hard for a virus to abandon without significant penalty and can be viewed as conserved. From a repurposing standpoint that there exists a strong target homology with agents that promote RAS inhibition and the common conserved site of the pathogenesis for COVID-19 and related viruses.
3. Prevention to Treatment. An understanding of how the repurposed drug provides benefit in treating the host across the cascade from prevention to the life-threatening consequences of other viral pathogenesis in COVID-19 disease. Cao and Li 2020 importantly summarise the clinical course of SARS-CoV-2 infection by dividing it into three phases: viremia phase, acute phase (e.g. with pneumonia) and severe or recovery phase (25). From the standpoint of drug repurposing in COVID we have adopted a similar approach, with particular focus on the severe phase, where the host system has switched on innate immune system, affecting inflammation. oedema, and vasconstriction. Our treatment focus in this phase is on RAS inhibition and (a) predisposition and age (b) predisposition with hypertension and co-morbidities (c) treatment and secondary infection and (d) intubation.
3a. Predisposition and age . Increasing age has been shown to be a strong correlative factor with morbidity and mortality with COVID. As summarized by Shahid et al. studies have highlighted the relatively higher mortality rates in older adults (35). While a variety of reasons may underpin this predisposition, it would appear that the RAS has a strong input to aging by way of mitochondrial function, specifically a strong relationship between free radicals, the mitochondria and aging, with excessive free radical production linked to dysregulation of RAS (36). Key in this dysregulation is the angiotensin receptor AT1R and its linkage to NADPH and increased production of reactive oxygen species (ROS), and a role of RAS in decreasing ROS scavenging enzymes. As highlighted by Wilson et al 2016 (37) in mitochondria the receptors for angiotensin (AT1R and AT2R) are linked to respiration and nitric oxide production. Furthermore the presence of angiotensin 1-7 in purified mitochondria and the conversion of angiotensin I to angiotensin 1-7 by endopeptidases suggests a intramitochondrial pathway for formation of angiotensin 1-7 (37). Recently Wang et al (38) have shown that overexpression of ACE2 through mitochondrial function enhances the protective effects of endothelial progenitor cells (EPC) on endothelial cell injury. It remains to be determined whether the dysregulation of RAS described in COVID-19 also similarly has a deleterious effect on mitochondrial function, as there appears to be an emerging possibility that a complex intersection between aging, mitochondrial function, apoptosis, the RAS and coronaviruses exist. Lai et al conducted a proteome analysis on human promonocyte HL-CZ cells expressing SARS CoV3CL., finding that 36% of the up regulated proteins were located in the mitochondria, including the apoptosis-inducing factor ATP synthase beta chain and cytochrome c oxidase. They suggested that SARS CoV3CLpro could induce mitochondrial-mediated apoptosis (39). Additionally, it is possible that mitochondrial dysregulated RAS and free radical production in aging acts in concert with viral RAS dysregulation in COVID-19 disease. From the standpoint of repurposing what is important is the observation that inhibition of RAS with enalapril and losartan preserve mitochondrial function from the effects of aging (40).