A few additional treatment possibilities in COVID19 (SARS Cov-2) addressing furin-like cleavage and pyroptosis (caspacin-1 activation of inflammasome NLRP3)

*stay tuned for a blog looking at flavonoids and SARS Cov-1 and Cov-2

It looks like pyroptosis, a novel form of inflammatory cell death, may be a cause of the increased virulence of COVID19 (although it was seen in SARS Cov 1, too). Pyroptosis initiates upregulation of inflammasome NLRP3 via caspase-1 proteases in the canonical pathway. Other caspases can also trigger the inflammasome initiated by a variety of triggers (see figures 1, 2 below). The end results of NLRP3 upregulation includes IL-1beta and IL-18 production, and cell death. In COVID19 patients, higher serum IL-1beta (a surrogate marker for NLRP3 activation) has been noted. And lymphocytopenia in COVID19 is speculated to be due to chronic pyroptosis activation. On autopsy in four patients who died from  SARS-Cov1, it looks like pyroptosis-induced damage was present in multiple tissues including lungs, leading to significant organ damage. It’s thought that in SARS Cov1, its 3a protein that activated the inflammasome in LPS-primed macrophages. 

Of course, pyroptosis can be a protective antimicrobial response (damaged, infected cells are aggressively killed and Th1 gets turned on via the cytokines initiated by NLRP3) but it can be chronically, inappropriately activated by when hijacked by the virus. In fact, observed in some viral infections is an inhibition of cell death (normally triggered by gasdermin D, GSDMD) allowing inflammation and viral propagation to continue unabated. 

Not surprisingly, chronic activation of pyroptosis can occur with other DAMP/PAMP exposures (figure 2) think proinflammatory/high sugar & processed food diet, dysbiosis/ LPS, stress, toxins, drugs, ROS, mitochondrial damage, etc. Chronic activation has been noted in inflammatory diseases such as type II diabetes, obesity, autoimmune disease, cardiovascular disease, cancer. Interestingly, it might be that CD4 T-cell depletion in HIV is mediated by caspase-1 pyroptosis. 

The cytokine storm seen in SARS Cov1 and Cov2 might be due to chronic pyroptosis activation. However, known activators of NLRP3 in SARS-CoV differ from CoV2.  While SARS Cov1 and COVID19 are genetically very similar, the extraordinarily high rate of infectivity of COVID19 is unique. Prepublication research suggests the cause might be a unique furin-like cleavage site on the spike protein of Cov2 that was absent in Cov1.  

  

Understanding that treatments for COVID19 are speculative, here are some considerations based on what we’re learning about pathophysiology.

1. Essential, powerful basics for all of us

In Functional Medicine, we talk about the “MLF” or modifiable lifestyle factors. We want to get on that now (and for future prep): Resolve the baseline inflammation that could contribute to COVID19-driven inflammation. FxMed providers are great at thinking through reducing PAMP/DAMP exposure (and, thanks to Dr. Rountree’s teaching for 10+ years in the Immune Module, most of us know his refrain on possible issues: Toxins, bugs, drugs, foods) 

• Clean, anti-inflammatory diet (see below- consider a keto leaning diet or caloric restriction as a ramped up anti-inflammatory approach)
• Tend to gut as needed (gut permeability, LPS-driven endotoxemia, dysbiosis, inflammation, etc)
• Reduce toxin exposures
• Sufficient nutrients
• Adequate sleep
• Exercise
• De-stress
• Connect with your fellow humans.

2. A few considerations for taming inflammasome activation

• Melatonin (see figure 3 below)
• • • Consider 3mg- 30mg (the latter is used in oncology)
    • Effective inhibitor of NLRP3 activation via a variety of mechanisms
• The chief ketone beta hydroxybutyrate has been shown to inhibit inflammasome activation. *We will discuss soon how to support modest ketone production safely, without promoting inflammation.
• • • Consider caloric restriction (this can be as simple as a 7-7 eating schedule)
    • Consider low carbohydrate diet
    • Consider MCT oil
    • Consider ketone esters (I don’t personally have experience with these outside of oncology nutrition)
    • Exercise will fuel ketone production, too
• Specialized pro-resolving lipid mediators
• • • Metagenics SPM Active: 2 caps daily; increase to 2 caps BID or perhaps TID with active infection 
      • SPMs inhibit priming and activation of macrophage NLRP3 inflammasome (in vivo, in vitro) Details: SPMs, specifically D2, suppressed IL-1 beta production and secretion in LPS and ATP challenged macrophages. D2 was also shown to reduce inflammasome assembly and caspase-1 activity (cell studies). In vivo, D2 blocked the iinflammasome as shown by reduced IL-1beta release and increase M2 markers of inflammation resolution. 
      • SPMs really require a blog unto themselves. There are a host of studies on SPMs and inflammation, including inhibition of the inflammasome. While we need more human in vivo data, the research to date is very interesting to me. I am bullish on the SPM benefit. Note that we CAN make SPMs from Omega 3 and arachidonic acid, but not everyone makes SPMs equally well (it’s thought that the failure of inflammation resolution, ie- chronic inflammation, is the inability to kick in SPM production). Low dose aspirin can support aspirin-triggered synthesis of some SPMs, whereas a full  ASA dose and other NSAIDs can inhibit production. 
• Palmitoylethanolamide (PEA)
• • I referenced this little molecule here and this is a great review
  • Take 600mg/day as prevention; 600mg TID during active infection
  • Shown to inhibit inflammasome
  • We are using Metagenics Hemp oil plus (with PEA) and Mirica (PEA with luteolin)

Add your own list here- there are MANY options; share with us those you like. 

 

Furin-like cleavage site. Can we slow infectivity down?

Luteolin has been studied for its inhibition of pro protein convertase furin. Dengue fever, which also infects human cells via the furin cleavage system, was inhibited in vitro and in vivo by the application of luteolin – it appears that luteolin interrupted an efficient cleavage process rather than though competitive inhibition.  Chinese herb Viola Yedoensis Makino  

In vitro assays also show oroxylin A, chrysin and baicalin are able to inhibit furin, with oroxylin A being the most potent of the three. Oroxylin A and chrysin used in this study were isolated from the stem bark of oroxylum indicum. 

[As I mentioned above, we are using a PEA product with luteolin- KF]

 

 Figure 1: A hypothesis of the relationship between SARS-CoV-2 and cell pyroptosis. The COVID-19 may be linked to cell pyroptosis, especially in lymphocytes through the activation of the NLRP3 inflammasome. Morphological changes in lymphocytes and macrophages, nucleic acid and protein levels in classical and non-classical cells, detection of NLRP3 and GSDMD, and the role of inflammatory cytokines IL-1β and IL-18 requires further research

 

Figure 2 A schematic diagram of Pyroptosis pathways. Pyroptosis is initiated when DAMPs or PAMPs activate the inflammasomes. Activated inflammasomes can lead to the cleavage of caspase-1. The activated caspase-1 cleaves Gasdermin D (GSDMD), in turn to form the N-fragment of GSDMD and cell membrane pores, resulting in pyroptosis. Activated caspase-1 also promotes the maturation and secretion of IL-1β and IL-18, which is also an important molecular event during the pyroptosis procession. When LPS binds to the precursor of caspase-4/5/11, it can also cause pyroptosis. The other way to activate pyroptosis is caspase-3/ Gasdermin E (GSDME). Caspase-3 can be activated by mitochondrial and death receptor pathway. The activated caspase-3 then cleaves GSDME, to produce GSDME N-fragments, forming pores in the plasma membrane, causing cell swelling and pyroptosis. DAMPs, danger-associated molecular patterns; PAMPs, pathogen-associated molecular patterns; IL-1β/IL-18, interleukin-1β/interleukin-18; LPS, lipopolysaccharide.

 

Figure 3: Schematic representation of melatonin’s protective effects against the inflammatory process involving NF-κB and NLRP3 inflammasome activation.