The histone deacetylase inhibitor valproic acid (VPA) is a potential drug that could be adapted to prevent the progression and complications of SARS-CoV-2 infection. VPA has a history of research in the treatment of various viral infections. VPA inhibits SARS-CoV-2 virus entry, suppresses the pro-inflammatory immune cell and cytokine response to infection, and reduces inflammatory tissue and organ damage by mechanisms that may appear to be sex-related. The antithrombotic, antiplatelet, anti-inflammatory, immunomodulatory, glucose- and testosterone-lowering in blood serum effects of VPA suggest that the drug could be promising for therapy of COVID-19. Sex-related differences in the efficacy of VPA treatment may be significant in developing a personalised treatment strategy for COVID-19.
1. SARS-CoV-2 Virus and VPA
The docking, binding energy calculation determines that VPA metabolite 4-ene-VPA-CoA creates a stable interaction with nsP12 of SARS-CoV2 RNA polymerase and VPA-CoA could specifically inhibit the target. SARS-CoV-2 RNA polymerase is an enzyme playing in viral RNA replication and the virus’s survival in a host [
51,
52]. The SARS-CoV-2 virus x-ray crystal structure of a critical protein in the virus’s life cycle is the central protease (M
pro, 3CL
pro) [
53,
54,
55]. The M
pro importance recognises M
pro as a target for antiviral drugs, designed as a virus 3CL
pro inhibitor, for COVID-19 therapy [
56,
57]. HDACs’ inhibitors are tightly bound into the active site of the crystallographic virus M
pro structure [
58]. The SARS-CoV-2 protease NSP5 interacts with the HDAC2. Researchers predict that NSP5 may inhibit the transportation of HDAC2 into a nucleus, and could affect the HDAC2 strength to interfere with the interferon response and inflammation [
59,
60]. Experimental studies show that the binding of HDAC2 to the promoters was lower in females than in males [
61]. The HDAC2 activity can not only be modulated by VPA binding to the catalytic center, but the HDAC2 protein level is susceptible to selective regulation by VPA [
62]. VPA blocks the zinc-containing catalytic domain of HDACs [
63]. VPA treatment reduced HDAC2 level in male rats’ brain frontal cortex tissue, but no VPA effect was for HDAC2 protein in females [
64].
2. Sex-Related COVID-19 Infection Progression Mechanisms and VPA
Due to the high viral infection load, membrane ACE2 and its mRNA expression are significantly diminished in COVID-19 patients [
69,
94,
95,
96]. At a later COVID-19 infection stage, down-regulated ACE2 in tissues may worsen the imbalance in the renin-angiotensin system (RAS). ACE2 has a protective effect through RAS regulation [
97], and protects against RAS-mediated activations of harmful effects [
69,
98]. Depleting ACE2 in tissue cells leads to an increase in angiotensin II (Ang II) blood serum level and the activation of the AT
1 receptors, which would activate ADAM17 more. The ACE2 expression is transcriptionally suppressed due to AT
1 activation [
99]. Increased Ang II levels act as a vasoconstrictor and a pro-inflammatory molecule through AT
1 [
100]. ACE2 knockout results in pathology similar to the acute respiratory distress syndrome in mice [
101]. The ACE2 molecule reduces RAS activity by converting Ang II into Ang 1–7 [
102,
103,
104], decreasing Ang II level and the AT1 activation, which manages reduced pathological inflammation effects in tissues [
69,
105,
106]. Sex distinctions of the RAS in response to stimulation and inhibition of the system have been reviewed [
81,
107,
108]. Higher levels of ANG (1–7) in women may inhibit the harmful effects of ANG II and its activation [
109]. Compared with female rats, males have higher AT1 receptor RNA, higher AT1 protein levels, higher receptor density in kidneys and ∼40% higher specific AT1 binding in the glomeruli than females. These differences are 17β-estradiol (E2) dependent [
81,
108,
110]. Activation of AT1 mediates ANG II’s biological functions, such as sodium reabsorption, vasoconstriction, increased oxidative stress and inflammation [
111,
112]. VPA, inhibiting HDAC1 and HDAC2, down-regulates Ang II and AT1 activity [
113,
114]. VPA reverses the ANG II-induced increment of HDAC2 RNA and protein levels in cardiomyocytes [
115]. Anti-hypertensive action of VPA is mediated by the inhibition of HDAC1 via acetylation processes [
113,
116].
3. COVID-19 Thrombotic Complications and VPA
3.1. SARS-CoV-2 and Sex-Related Thrombotic Complications
The pathophysiology of COVID-19 complications is characterised by clinical features of thrombosis and disseminated intravascular coagulopathy in the airways, myocardium, kidneys, brain and other organs [
179]. Thrombosis is found in approximately 30% of COVID-19 hospitalised patients [
180]. The incidence of thrombosis in COVID-19 is higher in men than in women and explains the higher mortality in men [
181]. In an analysis of 29 studies, 70% of all thromboembolic events occurred in men and 30% in women [
182]. Viral invasion due to severe vascular endothelial damage triggers the coagulation cascade, impairs fibrinolytic activity, releases von-Willebrand factor [
13], increases total cytokine release, activates platelets and the complement system and generates thromboxane [
183,
184]. SARS-CoV-2 can directly activate coagulation via the viral M
pro; the active site of M
pro is structurally similar to the active site of FXa and thrombin and can therefore activate coagulation [
185]. The development of thrombosis has been attributed to the direct effects of the virus by increasing the levels of pro-inflammatory cytokines and pro-inflammatory M1 macrophages, by activation of the complement system and by endothelial dysfunction, leading to disseminated intravascular coagulopathy [
186,
187,
188,
189]. Endothelial dysfunction and its association with thrombosis have been implicated in SARS-CoV-2-induced target organ damage [
190]. VPA binding to SARS-CoV-2 M
pro is expected to inhibit M
pro pathways [
191]. Older men with hypertension, chronic kidney disease, coronary disease, diabetes mellitus and obesity are at increased risk of thrombotic complications [
192,
193,
194]. Changes in plasma levels of D-dimer, von Willibrand factor (vWF), fibrinogen, tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1 antigen (PAI-1) antigen are associated with poorer outcomes in COVID-19 patients [
195,
196,
197,
198].
3.2. VPA Effect on Thrombosis Mechanisms, COVID-19
The VPA effects on thrombogenesis have been explored in pre-clinical studies and during the treatment of patients with VPA (
Table 2). HDAC inhibitors have reduced platelet counts and inhibit platelet function [
199,
200], while other VPA experimental and clinical trials did not find such effects [
201,
202]. The baseline platelet count was similar in women and men. A causal relationship between prolonged use and rising plasma VPA levels and reduced platelet counts, with reversal of thrombocytopenia after reduction of VPA dosage, was reported: that of thrombocytopenia substantially increased at VPA levels above 100 ug/mL in women and above 130 ug/mL in men; women were significantly more likely to develop thrombocytopenia [
203]. There is a significantly higher female overrepresentation in heparin-induced thrombocytopenia, with females at approximately twice the risk of thrombocytopenia than males. However, the underlying mechanism for this sex difference is unclear [
204]. VPA may affect several different coagulation factors: decrease in von Willebrand factor:antigen (vWF:Ag) concentration [
205,
206]; protein C level [
205,
207], protein S level [
207,
208], antithrombin III level, decrease prothrombin time [
205,
206] and increase activated partial thromboplastin time [
205,
206,
207].
Table 2. VPA treatment effect on thrombogenesis.
# |
Thrombogenesis Related Factor |
Cells/Animals/Human |
Sex |
VPA Treatment Effect |
Ref. |
1. |
Complement C3 |
HepG2 cells |
unknown |
↓ C3 gene expression |
[209] |
2. |
t-PA |
Human umbilical vein endothelial cells |
unknown |
↑ t-PA production |
[210] |
3. |
ICAM-1 expression |
Human umbilical vein ECs and human coronary artery EC |
unknown |
↓ ICAM-1 expression |
[83] |
4. |
Platelets number |
C57BL/6 mice |
unknown |
↓ platelets count |
[200] |
5. |
Vascular t-PA |
C57BL/6 mice |
males |
↑ endothelial vascular t-PA production; ↓ fibrin accumulation in response to vascular injury |
[201] |
6. |
E-selectin and ICAM-1 |
Sprague–Dawley rats with subarachnoid hemorrhage induced vasospasm |
males |
↓ the E-selectin and ICAM-1 level |
[211] |
7. |
Platelets number |
Epileptic adult patients and healthy control |
men and women |
relationship between rising plasma VPA level and reduced platelet counts, with female sex additional risk factor |
[203] |
8. |
Arachidonate cascade thromboxane A2 in platelets |
Epileptic adult patients and healthy control |
men |
↓ activity of the arachidonate cascade in platelets; ↓ the cyclooxygenase pathway; ↓ synthesis thromboxane A2 |
[199] |
9. |
Von Willebrand factor:antigen |
Epileptic children patients and healthy control |
male + female (combined) |
↓ concentration in blood serum |
[205,206] |
10. |
Protein C |
Epileptic children patients and healthy control |
male + female (combined) |
↓ concentration in blood serum |
[205,207] |
11. |
Protein S |
Epileptic children patients and healthy control |
male + female (combined) |
↓ concentration in blood serum |
[207,208] |
12. |
Antithrombin III |
Epileptic children patients and healthy control |
male + female (combined) |
↓ concentration in blood serum |
[205,206] |
13. |
Prothrombin time |
Epileptic children patients and healthy control |
male + female (combined) |
↓ concentration in blood serum |
[205,206] |
14. |
Activated partial thromboplastin time |
Epileptic children patients and healthy control |
male + female (combined) |
↓ concentration in blood serum |
[205,206,207] |
↓ decreased; ↑ increased.
SARS-CoV-2 activates the complement system, either directly or through an immune response. Activated complement promotes inflammation [
212]. Complement activation is increased and constant in severely ill COVID-19 patients, and complement activation is via the alternative pathway (AP) [
213]. The anaphylatoxins C3a and C5a are significant contributors to the cytokine storm syndrome [
214]. The healthy adult population is characterised by substantial sex-related differences in complement levels and function: significantly lower AP activity was in females than males. AP revealed lower C3 levels in women [
215]. In experimental intestinal ischemia with an acute inflammatory response, complement activity was sex-dependent: female MBL-/- and P-/- mice had significantly less C5a in their serum than males [
216]. Experimental results indicate that lysine acetylation by VPA is associated with attenuated C3 gene expression. VPA-associated reductions in circulating complement and clotting factors result from changes in liver-specific gene expression [
209]. VPA inhibits intercellular adhesion molecule-1 (ICAM-1) and E-selectin [
83,
211]. Analysis from patients hospitalised with COVID-19 showed higher circulating VCAM-1 and E-selectin levels in men than women [
217]. The endothelial cell adhesion molecules elevated levels promote tissue infiltration of circulating leukocytes and are associated with inflammation and thrombosis, which occur at a higher frequency in males [
218]. VPA reduced endothelial cell dysfunction through the mechanisms of action of transforming growth factor-β (TGF-β) and vascular endothelial growth factor (VEGF) in a porcine model of ischemia-reperfusion of hemorrhagic shock [
219]. Inhibiting TGF-β activity, VPA alleviates pulmonary fibrosis through epithelial-mesenchymal transition inhibition in vitro and in vivo [
132]. Estradiol has been shown to decrease TGF-β1 synthesis [
220]. VPA inhibiting IL-12 and TNF-α, reversing macrophage polarisation from pro-inflammatory to the anti-inflammatory type, and reducing macrophage infiltration reduces the risk of thrombosis [
117,
137].
3.3. VPA and Fibrinolysis
Treatment with VPA in a rat thrombosis model reduced thrombus formation and did not increase bleeding tendency [
201]. VPA can selectively manipulate the fibrinolytic system to reduce thrombus formation in blood vessels in vivo. In a murine model of thrombosis induced by intravascular injury, VPA treatment increased t-PA production in blood vessels [
201,
210,
221,
222,
223] was associated with less fibrin accumulation and fewer thrombi [
201,
224]. Impaired fibrinolysis, due to reduced t-PA production and depleted storage or increased expression of a significant inhibitor of fibrinolysis PAI-1 [
225,
226], has been reported in coronary heart disease patients with cardiovascular risk factors, such as hypertension and obesity [
227,
228,
229,
230,
231]. A clinical trial of VPA treatment showed a significant reduction in PAI-1 and signs of improvement in fibrinolysis, favourably altered the balance between t-PA and PAI-1, and the dose of VPA treatment was significantly lower than the usual dose of VPA for epilepsy [
202,
223,
224]. Thus, VPA could be a potential alternative for preventing thrombotic events based on improved endogenous fibrinolysis [
202].
This entry is adapted from the peer-reviewed paper 10.3390/biomedicines10050962