The human disease-associated gene ZNFX1 controls inflammation through inhibition of the NLRP3 inflammasome, 2024, Huang et al

chillier

chillier

Senior Member (Voting Rights)

Abstract​

Inherited deficiency of zinc finger NFX1-type containing 1 (ZNFX1), a dsRNA virus sensor, is associated with severe familial immunodeficiency, multisystem inflammatory disease, increased susceptibility to viruses, and early mortality. However, limited treatments for patients with pathological variants of ZNFX1 exist due to an incomplete understanding of the diseases resulting from ZNFX1 mutations. Here, we demonstrate that ZNFX1 specifically inhibits the activation of the NLR family pyrin domain-containing protein 3 (NLRP3) inflammasome in response to NLRP3 activators both in vitro and in vivo. ZNFX1 retains NLRP3 in the cytoplasm and prevents its accumulation in the TGN38 + /TGN46+ vesicles in the resting state. Upon NLRP3 inflammasome activation, ZNFX1 is cleaved by caspase-1, establishing a feed-forward loop that promotes NLRP3 accumulation in the trans-Golgi network (TGN) and amplifies the activity of the downstream cascade. Expression of wild-type ZNFX1, but not of ZNFX1 with human pathogenic mutations, rescues the impairment of NLRP3 inflammasome inhibition. Our findings reveal a dual role of ZNFX1 in virus sensing and suppression of inflammation, which may become valuable for the development of treatments for ZNFX1 mutation-related diseases.

Link: https://www.embopress.org/doi/full/10.1038/s44318-024-00236-9
 
For their mouse work: When folks mention an effect for a particular mouse model, you can generally find info on the phenotype/symptoms for the model here: https://www.informatics.jax.org/marker/phenotypes/MGI:2138982. That link is for this gene. In this case, there are IMPC mice where they knocked out (removed the functioning version of) the gene and they see no immune defects, which they would have looked for.. And there are the mice from this group, which show a strong immune phenotype. With the mutations introduced at exon5 and exon3 in each model a similar effect might be expected (would need to dig in to that more to be sure). One difference is the background strain. The first (no immune effect) is C57BL/6J and the other (immune effect) is C57BL/6NJ. Looks like C57BL/6J may boost immune phenotypes (please an immunologist weigh in). You can also look here: https://www.proteinatlas.org/ENSG00000162711-NLRP3/single+cell to see where the genes mentioned transcripts are expressed. This gene is seen in a few different cell types, which I think make it more interesting. Just adding this to provide some additional perhaps useful context.
 
Jonathan Edwards said:
What would be the consequences of upregulating the NLRP3 pathway? Would there be more interferons or something like that? Would T cell costimuation be affected?
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It’s one part of myeloid pathogen recognition, leading to caspase and IL-1B induction plus some other cytokines I’m forgetting (though I don’t think interferon is one of the main ones)
 
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jnmaciuch said:
IL-1B
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From Wikipedia:
IL-1β, in combination with IL-23, induced expression of IL-17, IL-21 and IL-22 by γδ T cells. This induction of expression is in the absence of additional signals. That suggests IL-1β is involved in modulation of autoimmune inflammation [15]

A link to gamma delta T cells - potentially relevant? Also have we seen IL-17 pop up recently?
 
Ongoing clinical trial of an IL-1 inhibitor in Long Covid:

forestglip said:
Safety and Efficacy of Anakinra Treatment for Patients With Post Acute Covid Syndrome (PRECISION)

Description
People with COVID-19 might have sustained post-infection sequelae, known as Post-Acute Covid Syndrome (PACS). A recent consensus definition by an international panel of 265 patients, clinicians, researchers, and WHO staff suggests that post-COVID-19 condition occurs in individuals with a history of probable or confirmed SARS-CoV-2 infection, usually 3 months from the onset, with symptoms that last for at least 2 months and cannot be explained by an alternative diagnosis. Common symptoms include fatigue, shortness of breath, and cognitive dysfunction and generally have an impact on everyday functioning.

The role of immune dysregulation in PACS is indirectly supported from the findings of the SAVE-MORE randomized clinical trial, in which patients with moderate and severe COVID-19, were 1:2 randomized to treatment with placebo or anakinra once daily for 10 days. The primary endpoint was the distribution of the frequencies of patients in the 11 points of the WHO clinical progression scale (CPS) by day 28. Patients' follow-up until day 90 showed significant reduction of the incidence of PACS; this was 24.4% among placebo-treated patients and 15.7% among patients treated with anakinra.

After the end of the SAVE-MORE trial, the understanding of the immune activation of PACS and the development of tools for the evaluation of patients have become the main aims of the Hellenic Institute for the study of sepsis (HISS) group. More precisely, patients with medical history of COVID-19 pneumonia during three separate time periods and matched comparators for age, sex, comorbidities, and state of vaccination were followed up and evaluated for PACS. Main findings can be summarized as follows:
  1. For at least one year after acute COVID-19 there is considerable immune dysregulation involving both the innate and the adaptive responses.
  2. Patients with PACS may be classified into four main phenotype clusters: fatigue involving 70.8%, respiratory cluster involving 33.2%, systemic symptoms involving 17.7% and other symptoms involving 26.1%.
  3. The risk for progression into PACS was significantly lower among patients treated with anakinra in the acute stage (odds ratio 0.59, p: 0.017) showing a role of IL-1 for the progression into PACS.
  4. Patients with fatigue bring distinct immunotype compared to the respiratory cluster.
  5. IP-10 (interferon-gamma-induced protein-10) at levels more than 250 pg/ml has sensitivity 99.3%, specificity 90.9%, positive predictive value (PPV) 97.9% and negative predictive value (NPV) 97.6% for the diagnosis of the post acute COVID immune dysregulation.
PRECISION is a proof-of-concept, randomized clinical trial (RCT) aiming to evaluate the efficacy and safety of anakinra in patients with PACS in improving the clinical and immunological state over 4 to 8 weeks as measured by a composite endpoint, namely, the "Score of PACS progression reversal".

Eligibility


Link (with contact info)

Only the Greece sites have started recruiting at this point.
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Derya Unutmaz reports an N=1 improvement from Ankinra (suggested by Grok, of all things [not an endorsement by me]):

Mij said:
My close physician friend just messaged me-this is amazing: He had been following a patient with a chronic disease for many years, trying various treatments to no avail. He recently uploaded all the data to Grok, which suggested a specific treatment. Now, the patient seems cured!
https://twitter.com/user/status/1902381963111014819
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According to OMIM loss of function of ZNFX1 leads to hyperinflammation and immunodeficiency. I remember reading a paper which I can't find at the moment which suggested these people usually die in childhood of severe viral infections of various kinds, or even without evidence of a virus seem to have a lot of sterile inflammation and tissue damage. If the gtex expression data from the Decode hit is correct though this would be the opposite of the expected effect in ME/CFS.

The logic of what they set out in this thread's paper, combined with an increased ZNFX1 expression from the gtex data (as much as you can trust that data) - would be a downregulation of the NLRP3 inflammasome pathway, not an increase, as the increased levels of ZNFX1 would bind to the NLRP3 protein therefore preventing inflammasome assembly.

I think maybe ZNFX1 could intersect with other virus sensing pathways, so I thought if this gene is relevant in ME then it could represent a move away from an IL-1 phenotype and towards some other less inflammatory viral response (like interferon related?).
 
chillier said:
The logic of what they set out in this thread's paper, combined with an increased ZNFX1 expression from the gtex data (as much as you can trust that data) - would be a downregulation of the NLRP3 inflammasome pathway, not an increase, as the increased levels of ZNFX1 would bind to the NLRP3 protein therefore preventing inflammasome assembly.
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Ah, my mistake, apologies for flooding the thread with irrelevant speculation then!
 
chillier said:
Not necessarily! It depends on how much we can trust the eQTL data, and from what I've gathered it may not be all that reliable.
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I had this paper from yesterday in mind when posting
Chandelier said:
link.springer.com

The NLRP3 inflammasome as a key pathway in the affective and chronic fatigue symptoms of Long COVID - Journal of Translational Medicine

The neuropsychiatric and somatic manifestations (physio-affective phenome) of Long COVID are substantially predicted by elevated peak body temperature (PBT
link.springer.com link.springer.com

Authors: Yingqian Zhang, Hussein Kadhem Al-Hakeim, Hawraa Kadhem Al-Jassas & Michael Maes

Abstract​

Background​

The neuropsychiatric and somatic manifestations (physio-affective phenome) of Long COVID are substantially predicted by elevated peak body temperature (PBT) and diminished oxygen saturation (SpO2) during the acute infectious stage.
The latter is linked to the immune pathophysiology of Long COVID involving activation of the immune-inflammatory response system (IRS) and the nucleotide-binding domain leucine-rich repeat and pyrin domain-containing receptor 3 (NLRP3) inflammasome.
Nevertheless, there is a lack of data indicating whether NLRP3 and its components are implicated in the physio-affective phenome of Long COVID.

Method​

We enrolled 161 Long COVID patients 6 to 9 months after the acute phase and divided them into two groups based on their baseline PBT and SpO2 levels, corresponding to mild and severe acute COVID-19.
We assessed serum NLRP3, caspase-1, C-reactive protein (CRP), interleukin (IL)-18, IL-1β, IL-10, fibronectin, and Gasdermin D (GSDMD) during Long COVID.

Results​

All of the aforementioned indicators (except IL-10) were substantially higher in Long COVID patients who had previously experienced severe COVID-19 than in those who had mild acute COVID-19.
The physio-affective phenome of Long COVID and the severity of the acute COVID-19 were significantly correlated with these IRS biomarkers, with the exception of fibronectin.
The variance in the overall severity of Long COVID is accounted for (49.5%) by the combined influence of fibronectin, IL-10, SpO2, and PBT.

Conclusion​

The severity of Long COVID is strongly associated with IRS and NLRP3 activation and the severity of the inflammatory response during the acute infectious phase.
NLRP3 activation is a drug target to treat Long COVID.


DOI: 10.1186/s12967-026-07703-3
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chillier said:
I think maybe ZNFX1 could intersect with other virus sensing pathways, so I thought if this gene is relevant in ME then it could represent a move away from an IL-1 phenotype and towards some other less inflammatory viral response (like interferon related?).
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Yeah that's a potential way to look at it--virus would be detected by and induce different facets of the immune response simultaneously (through intracellular RNA/DNA sensors, TLRs, inflammasome, adaptive recognition, etc.), each of which trigger different combinations and strengths of cytokines, so less efficacy of one of those arms could very well shift the proportion of the local and systemic cytokine cocktail.

[Edit: if inflammasome was suppressed but intracellular sensors were still active then my guess is that leads to more interferon--I remember reading that after viral DNA/RNA in a host cell triggers interferon from the parenchymal cGAS or RIG-I response, the DNA/RNA can then get packaged with the inflammasome and shunted into the extracellular space, where it stimulates macrophages and triggers other cytokines besides interferon]
 
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Found one of the papers I was thinking of:
www.nature.com

Cytosolic escape of mitochondrial DNA triggers cGAS-STING-NLRP3 axis-dependent nucleus pulposus cell pyroptosis - Experimental & Molecular Medicine

An investigation of the mechanisms responsible for intervertebral disc degeneration (IVD) reveals a potential treatment to limit the progression of low back pain. IVD involves widespread inflammation and cell death inside discs, but exactly how the condition progresses is unclear. Cao Yang, Kun...
www.nature.com www.nature.com

I landed on it while following up on my interest in mtDNA/interferon, but viral DNA activates the same sensor.

Though selfishly I’m now wondering if a ZNFX1 mutation could be relevant in both contexts—skew towards interferon during initial infection, and then afterwards mtDNA released into the cytosol during normal cell stress fails to elicit the same NLRP3 activation and pyroptosis, so those cells end up maintaining an interferon response for longer than they would if they were properly cleared out (and it doesn’t get locally counteracted by IL-1B signaling as much)
 
jnmaciuch said:
so those cells end up maintaining an interferon response for longer than they would
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When I was digging around this before I wondered about something not being as efficient around cleanup meaning the response goes on for longer and potentially amplifies more than normal. Ubiquitin proteasome area. Not sure we’ve really got any evidence for this though. But I still like your interferon/cGAS-STING/NLRP3 angle. Could the threshold for activation of these pathways just be lower in the context of the B-cell/lipid findings? Another contributory factor rather than anything?
 
hotblack said:
Could the threshold for activation of these pathways just be lower in the context of the B-cell/lipid findings? Another contributory factor rather than anything?
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I am not sure! I assume that NLRP3 related processes happen somewhat independently from B cell responses, though there may be points where they overlap and interact
 
jnmaciuch said:
I am not sure! I assume that NLRP3 related processes happen somewhat independently from B cell responses, though there may be points where they overlap and interact
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Usually I guess. But could this all happen, within B cells themselves? Sorry I’m crossing threads as well as streams of thought a bit here.. But if B cells are in his lipid altered state, could that push pathways like TLR, NLRP3, cGAS–STING within them closer to thresholds for triggering/signalling?

I was thinking if the B cells have this hangover or whatever which leaves them in this state, floating around ready to kick off at the slightest stimulation, it could be another way of having the sensitisation and innate/adaptive crossover we talked about from FCyRI and interferon gamma? And why knocking a load of certain B cells out seems to help some people. Or is this a bit if a stretch?
 
hotblack said:
Or is this a bit if a stretch?
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maybe! I think when the questions become “is it possible that X part of the immune system could have an impact on Y part of the immune system?” the answer will pretty much always be yes—you just need specific observations to determine whether that interaction has enough strength to really matter.

Which isnt to say there’s no merit to making the connections, just that we don’t have specific observations to support the importance of all the intermediate steps (that I know of, I definitely don’t have exhaustive knowledge on the subject)
 
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