Thread by @patientled

The PolyBio Symposium starts shortly! We'll be live-tweeting through the day - follow this thread.

1/ #LongCovid

https://twitter.com/polybioRF/status/1713940078513234303

@microbeminded2 Over my head but very cool work on respiratory profiling, and work in endometriosis!

Really cool work planned in endometriosis, spinal ligaments, etc 14/

@microbeminded2 Up next Dr. @MBVanElzakker: "Neuroinflammation and vasculature abnormalities in #LongCOVID and ME/CFS"

In looking at a brain, about half of cells are glial cells; have important functions, including immune function. 15/

@microbeminded2 @MBVanElzakker When they come in contact with pathogens or proteins from pathogens, they activate, release a protein called TSPO. TSPO is measurable with neuroimaging. 16/

@microbeminded2 @MBVanElzakker PET analysis in #LongCovid vs controls found neuroinflammation, which could implicate a bunch of things.

Really significant findings compared to controls! And not related to hospitalization or not (cohort was mostly non-hospitalized) 17/

@microbeminded2 @MBVanElzakker Their hypothesis was that cytokines wouldn't correlate with neuroinflammation, but that vascular impacts & vascular health would, and this was true!

Multiple vascular markers were correlated with neuroinflammation (more than those just on this screen). 18/ #LongCovid

@microbeminded2 @MBVanElzakker Post-doc Mark Painter filling in for John Wherry for this talk on antigen-specific T & B cells as biosensors for viral persistence and reactivation in #LongCovid! 19/

@microbeminded2 @MBVanElzakker Main two questions:

20/

Can T & B cells be used as sensitive and specific biosensors for SARS-CoV-2?

Also looking for EBV, VZV, flu, CMV, and others

This helps understand virus-specific impacts 21/

They hypothesized that not all #LongCovid patients would have these findings, but wanted to know if it was possible for them to identify subgroups with viral persistence. 22/

They did not find evidence of COVID-specific viral persistence in all #longCovid patients, but in a subcohort (27%? unsure what that number refers to). They also found evidence of EBV and VZV in a minority of LC patients. 23/

Next up, the amazing @VirusesImmunity!

She suggests 4 possible causes for #LongCovid:
viral reservoir, autoimmunity, tissue damage/dysfunction, latent virus reactivation.

24/

In autoimmune disease, patients are generating antibodies against ERV (endogenous retrovirus) 27/

Next up: Dr. Alessio Fasano, "SARS-CoV-2 persistence in MIS-C & pediatric #LongCOVID".

Early in the pandemic, there was a misconception that kids were spared (despite never being seen before). Their team thought this was counterintuitive, started looking at impact on kids 28/

Four hypotheses for #LongCovid, including persistence in the gut, chronic inflammation (including defect in zonulin-dependent barrier) (side note that MIS-C is driven by zonulin-dependent loss of gut mucosal barrier) 29/

#LongCovid is associated with increase in zonulin and oxidized LDL 30/

He suggests zonulin and oxidized-LDL could be potential biomarkers for #LongCovid. 31

3rd and 4th hypotheses are autoimmunity and microclots 32/

Next up is Dr. Locci, "Decoding Long COVID lymphoid tissue adaptive immune responses" 33/

(Hopping off, next tweeter will pick up shortly! -@ahandvanish)

34/

Hello all, @jannamoen here! Dr. Locci currently discussing her exciting approach using fine needle aspiration biopsies to study lymph tissue in #longCOVID

Her team found germinal center responses to SCoV2 appear to be more long-lasting in long COVID, suggesting immune dysregulation in the lymphatic system. /36

Highlights from the Q&A:

Akiko Iwasaki was asked whether high spike antibody levels correlate with viral persistence. Neutralization assays from the lab suggest there may not be a strong correlation between anti-spike IgG and lymphocyte responses. /37

Next up - Gene S. Tan from UCSF presenting on identification of serological biomarkers that reflect immunological status and mechanisms of viral chronic disease /38

Early, low, non-neutralizing IgG responses were observed in people who progressed to severe acute COVID-19 ("progressors"). These antibodies lacked fucosylation, enhancing their affinity for their target receptors. This correlates with worse symptoms in C19 /38

Now they are asking how antibody fucosylation is impacted in long COVID. LC phenotype is consistent with high inflammatory state without neutralization activity, not ideal for long-term health. /39

Exosomes are an interesting target for the group - these are basically little envelopes released by cells that could shield the contents from detection. /40

Next up - Dr. David Price discusses SARS-CoV-2 resevoirs in the lung as potential drivers of #longCOVID. His group found that peripheral virus-specific T cell frequencies were approximately the same between healthy controls and LC patients. /41

This implies the immune response in #longCOVID patients may be blunted - specific T cell frequencies should be increased after encountering a virus. (Sorry, missed the screenshot for this slide!) /42

Prev slides were discussing frequency - but further analysis found the /behavior/ of SCoV2 specific T cells were also different. /43

Activation of the alternative complement pathway, which is triggered by direct exposure to damaged cells and/or pathogens, is a predictive feature of long COVID. These data highlight ongoing tissue damage consistent with localized reservoirs of SCoV2 /43

New study aims to elucidate the SCoV2 resevoir and related pathology in lung tissue with a variety of next-generation genomic technologies. Keep an eye out for this in the future! /44

We are now hearing from Dr. Max Qian from J. Craig Venter Institute on the use of AI approaches to identify #longCOVID endotypes, a very popular methodological approach for discovery-driven research. /45

Machine learning is often applied in long COVID - it usually involves using an unsupervised process to split the population into clusters based on dimensionality reduction (or similar) methods. This can be paired with supervised learning classification /46

Machine learning has been successfully applied to long COVID. Clustering can be done based on symptom group presentations. Current studies suggest at least 10 sub-phenotypes (probably not the correct phrase) in #longCOVID /47

Ongoing work is being performed on semi-supervised topic modeling incorporating important events like vaccination and reinfection. These tools can help us understand divergent presentations and mechanisms in such a heterogeneous condition /48

Some interesting data suggesting common demographic variables like race, BMI, smoking status, and sex are not correlated with #longCOVID - it impacts these groups equally. Variables that DID correlate with LC included HIV status, fatigue, and diabetes /49

Next up is Dr. @resiapretorius discussing biofilms and endothelial debris contributions to #microclots.

Importantly, latest studies includes both LC and vaccine injury groups!

They have published a lot of work showing platelet hyperactivation in #longCOVID /50

@resiapretorius Their first paper describing flow cytometry to systematically measure microclots was recently published and will hopefully facilitate microclots research around the world /51

@resiapretorius More recent work in the lab is focused on novel methods to analyze and trace biofilms, amyloid microclots using fluorescent microscopy. These could easily be automated! /52

@resiapretorius Final points - microclotting and endothelial damage point to heterogenous and complex disease pathology. Size, number of clots alone will not be enough to explain the pathology in #longCOVID, framework needs to include platelet activation and endothelialitis /53

@resiapretorius Next on the podium is Dr. Marcelo Freire @drmfreire discussing immune, clotting, and infectious biomarkers of #longCOVID blood and saliva.

The mouth is a portal to the body, and the oral cavity can be an important site for pathogenesis of chronic diseases /54

@resiapretorius @drmfreire Fun fact - your saliva contains a lot of immune cells, which are alive and active fighting off pathogens in your mouth. These leukocytes are highly heterogeneous and correspond to specific pathogens /55

@resiapretorius @drmfreire Proteomics analysis found similar disruptions in protein expression from saliva compared with plasma (from blood). Saliva is a much easier sample to collect!

This indicates that salivary proteins could be used as biomarkers, and could also be driving pathology /56

@resiapretorius @drmfreire Some very beautiful images showing transcriptional expression in situ (within tissue), in this case in samples from patients with diabetes. The goal is to apply this to #longCOVID /57

@resiapretorius @drmfreire Now we have Dr. Annie Antar discussing evidence of SCoV2 persistence via evaluation of Long COVID B cell responses at Johns Hopkins

Measles is another RNA virus that causes post-acute sequelae, the most severe being paralyzing polio, which results from replication in the CSF /58

@resiapretorius @drmfreire Measles results in long-term disruptions of the adaptive immune system, this is previous work serves as inspiration for the SARS-CoV-2 studies /59

@resiapretorius @drmfreire Cool study design - patients enroll at acute symptom onset, samples can then be correlated with reported long COVID symptoms. Delayed viral RNA clearance from nose/saliva was associated with brain fog, and elevated IL-6 associated with long COVID at 90 days /60

@resiapretorius @drmfreire The survey team worked closely with @ahandvanish to ensure their questionairres accurately captured #longCOVID. Future studies include TCRseq - T-cell receptor sequencing /61

@resiapretorius @drmfreire @ahandvanish Now is Dr. Maayan Levy discussing serotonin reduction and viral persistence in post-acute sequelae of viral infection. Serum metabolomics combined with machine learning identified several clusters, but serotonin reduction was strongest predictor /62

@resiapretorius @drmfreire @ahandvanish Interestingly, serotonin reduction was not unique to SCoV2 - it was observed in mice following VSV, ARM, etc. By mimicking viral RNA with poly(I:C), they showed viral RNA activated interferon responses to reduce serotonin. This reduction is caused by disrupted gut absoption /63

@resiapretorius @drmfreire @ahandvanish This study highlights the importance (and complexities) around serotonin signaling following viral infections!

Now we have Dr. Lael Yonker on her pediatric #longCOVID #longCOVIDkids work.

Multiple studies show SCoV2 in the gut, which causes zonulin release and can lead to MIS-C/PIMS in children. Can blocking zonulin release put the brakes on inflammation onset?
/65

Open-label trial of zonulin receptor antagonist expedited the resolution of GI symptoms and spike clearance in children with MIS-C. Really promising results, currently in phase 2 DBPC trial! /66

Can circulating spike act as a surrogate marker for larazotide? This is a case study showing several spikes and rebounds in CRP until given larazotide (zonulin receptor blocker), which quickly stabilized CRP and spike at low levels /67

Alright, @jannamoen signing off! Tweets will continue below from other volunteers for the rest of the session! Thanks for tuning in. /68

@jannamoen @ahandvanish checking back in for the next few! 69/

@jannamoen @ahandvanish Next up is Dr. Tim Henrich "Total body PET imaging for T cell activity & deep tissue viral reservoirs in Long COVID".

There's lots of inflammation and immune dysregulation. "It's likely a tissue-based phenomenon." But tissue is hard to get!

#LongCovid 70/

So new imaging - in this case non-invasive nuclear imaging - may be helpful.

They are using a tracer called F-ARG, which is taken up by T cells, so it's a marker for activated T cells. 71/

Cohort was at a median of 195 days with #LongCovid, max was 2.5 years. Mostly non-hospitalized. Many had lost COVID antibodies!

Also had recovered COVID and pre-COVID healthy controls. 72/

Increased T cell activation in #longCovid patients compared to controls - curious places, cervical spine, thoracic spine, bone marrow, cauda equina. 73/

Back to LIINC study - found spike RNA in some areas.

Plans to do whole-body imaging through immunoPET imaging of the spike protein! 74/

A critical point - everyone having COVID just generally changes research baselines on inflammation, brain, cardiovascular health etc! 75/

Next is Dr. Matthew Frank, "A multi-hit model of Long COVID development".

Successive inflammatory insults = neuroinflammatory and brain changes in #LongCovid. 76/

He is proposing that #LongCovid comes from two hits: first, the spike hit from COVID, and then a second inflammatory hit (could be bacterial, another virus, environmental hit. potentially reinfection?)

This second hit causes the neuro impacts on the brain 77/

They ran an experiment on this hypothesis with animals. NLRP3 (gatekeeper of neuroinflammation) became elevated 9 days after the initial infection. Wonder if this is why there is a delay in neurocog symptoms?

"There is a persistent cellular change in the brain" 78/

Switching off to another volunteer now! 79/

VanElzakker in Q&A on questions about serotonin paper: underscores SSRIs are reuptake inhibitors; this is not the same as adding it or producing more of it

VanElzakker now summarizing research on detecting fibrin-based clots using radioligands; PolyBio interested in pursuing this in LC

VanElzakker discussing images of micro clots (notes amyloid here differs from amyloid in Alzheimer's). /81

VanElzakker contrasting images of neutrophil movement in patients & controls/82

"platelets and neutrophils are responding to the same thing" (image on left is neutrophils, right shows platelet overlay)/83

the hope is this will lead to PET imaging that can provide information /84

VanElzakker now onto a second study!/85

Very interesting research drawing on CCI, instability, with potential to do vagus nerve stimulation in scanner to observe results/86

Next up: Sara Cherry on tissue models in SARS-COV-2, especially viral persistence in the GI tract /87

Where we're at right now: approaches include direct acting antivirals and immunomodulators /88

We know that antivirals can reduce viral load during acute infection, but what about viral persistence /89

Cherry working here to ask questions about the difference between virus in airways during acute infection and in the gut later on/90

Noting high and persistent levels of virus after acute phase of infection, but questions outstanding about when and how to dose antivirals /91

Questions guiding Cherry's work / 92

Next up: Steven Deeks on experimental medical studies for infection-associated chronic illnesses, including LC. Noting need to define a mechanism in which we can interest pharma, industry partners to invest/ 93

Deeks noting the pileup of multiple pathways, all of which are actionable / 94

Deeks notes LIINC cohort shows emerging relationships between viral persistence and symptoms, though this research is ongoing / 95

One approach: antivirals targeting viral persistence (noting Stanford, Yale Paxlovid studies), but noting limits of antivirals that target replication of virus only / 96

Another approach: study of monoclonal antibodies that clear the viral reservoir itself / 97

Other pathways to intervene: baricitinib (JAK inhibitor) / 98

David Putrino up next on "how to be an innovator in the space of infection-associated chronic illnesses": noting that it takes 17 years to "go from bench to bedside" because funding rewards incremental research / 99

Putrino: hyperspecialization and siloing creates competition among departments, makes it difficult to approach infection-associated chronic illnesses b/c of the need for interdisciplinary knowledge and care / 100

Putrino: his work at Mount Sinai focused on adopting care models from pragmatic clinical trials. Importance of moving quickly, partnering with industry /101

"Single guiding principle" for Putrino lab's work: "Never let a patient be told 'there's nothing more we can do'". But clinical adoption is the hard part: barriers in time, funding, difficulty of behavioral change / 102

Putrino's model: building a center to address infection-associated chronic conditions; essential to address and treat together / 103

Putrino: importance of studying multiple therapies, getting at combinations that work for different endotypes (noting that adaptive platform trials are "not your grandpa's monotherapy trials") / 104

Q. to Dr. Cherry: is the gut/lung primary source of viral persistence, but what about other sites? A: thinks other tissues are important and they are expanding to this, hoping for more primary samples / 105

Q. for Peluso: how is the monoclonal he's studying different from Regeneron or others used for acute COVID? A. This is an investigational monoclonal--was the last monoclonal standing at end of 2022.

Q. will these need to be combined with other drugs? A. combination therapeutics used in HIV to avoid resistance; interesting to look at multiple mechanisms contributing to resistance and target those. Cherry adds that combining multiples should be studied / 107

Q. to VanElzakker on vagus nerve and muscarinic receptors. A. Notes that the idea that the vagus nerve is infected is a hypothesis at this point, likely we need multi pronged approaches. Proal notes recent finding from autopsy study of SARS-COV-2 in vagus nerve / 108

Q. on sleep and reduced parasympathetic activity at night. A. VanElzakker: sleep one of multiple areas of dysfunction. They give a take-home wearable sleep device to study this in days prior to scans to capture data on this. Probs with sleep likely snowball w/other symptoms / 109

Q. on trial construction, being patient-centered, and including housebound patients. A. Iwasaki: now can enroll ppl across US (except Alaska/Hawaii) and deliver meds, take samples from home. Peluso notes PLRC input, patient advocates, community board / 110