Identification of soluble biomarkers that associate with distinct manifestations of long COVID, 2025, Gao, Buggert, Peluso et al

[forestglip]

Now published: post with link here

*******
Identification of soluble biomarkers that associate with distinct manifestations of long COVID

Marcus Buggert, Yu Gao, Curtis Cai, Sarah Adamo, Elsa Biteus, Habiba Kamal, Lena Dager, Kelly Miners, Sian Llewellyn-Lacey, Kristin Ladell, Pragati Sabberwal, Kirsten Bentley, Jinghua Wu, Mily Akhirunnesa, Samantha Jones, Per Julin, Christer Lidman, Richard Stanton, Helen Davies, Soo Aleman, David Price, Paul Goepfert, Steven Deeks, Michael Peluso

Abstract
Long COVID is a heterogeneous clinical syndrome of uncertain etiology triggered by infection with SARS-CoV-2. We employed ultrasensitive approaches to profile the immune system and plasma proteome in healthy convalescent individuals and patients with long COVID.

Symptomatic disease was not consistently associated with quantitative differences in immune cell lineage composition or antiviral T cell immunity. Healthy convalescent individuals nonetheless exhibited higher titers of neutralizing antibodies against SARS-CoV-2 than patients with long COVID, and extensive phenotypic analyses revealed a subtle increase in the expression of some coinhibitory receptors, most notably PD-1 and TIM-3, among SARS-CoV-2 nonspike-specific CD8+ T cells in patients with long COVID.

We further identified a plasma biomarker signature of disease linking breathlessness with apoptotic inflammatory networks centered on the hub protein TRAF2 and dysregulated pathways associated with lung injury, cell cycle progression, and platelet activation, which could potentially inform the diagnosis and treatment of long COVID.

Link | PDF (Preprint)

 

[Hutan]

From another study:
Improvement of immune dysregulation in individuals with long COVID at 24-months following SARS-CoV-2 infection, Matthews et al, 2024

LC participants show elevated nucleocapsid IgG levels at 3 months, and higher neutralizing capacity up to 8 months post-infection. Increased spike-specific and nucleocapsid-specific CD4+ T cells, PD-1, and TIM-3 expression on CD4+ and CD8+ T cells were observed at 3 and 8 months, but these differences do not persist at 24 months.

It also found increased PD-1 and TIM3 expression (but higher rather than lower specific antibody levels) in the Long Covid group.

It looks like heterogeneity was a problem in this study. I wish researchers would do more sorting of potential participants on obvious symptom differences before they do their studies.

 

[forestglip]

HAVCR2 (TIM-3) on Wikipedia

Hepatitis A virus cellular receptor 2 (HAVCR2), also known as T-cell immunoglobulin and mucin-domain containing-3 (TIM-3), is a protein that in humans is encoded by the HAVCR2 (TIM-3)gene. HAVCR2 was first described in 2002 as a cell surface molecule expressed on IFNγ producing CD4+ Th1 and CD8+ Tc1 cells.[5][6] Later, the expression was detected in Th17 cells,[7] regulatory T-cells,[8] and innate immune cells (dendritic cells, NK cells, monocytes, macrophages).[9][10] HAVCR2 receptor is a regulator of the immune response.

Clinical significance
HAVCR2 expression is up regulated in tumor-infiltrating lymphocytes in lung,[8] gastric,[30] head and neck cancer,[31] schwannoma,[32] melanoma[33] and follicular B-cell non-Hodgkin lymphoma.[34] It is also up-regulated in tumour-associated macrophages in various malignancies, including melanoma, especially in immunotherapy-resistant context.[9]

The HAVCR2 pathway may interact with the PD-1 pathway in the dysfunctional CD8+ T cells and Tregs in cancer.[35][8] HAVCR2 is mainly expressed on activated CD8+ T cells and suppresses macrophage activation following PD-1 inhibition.[36] Upregulation was observed in tumors progressing after anti-PD-1 therapy.[37] This seems to be a form of adaptive resistance to immunotherapy. Multiple phase 1/2 clinical trials with anti-HAVCR2 monoclonal antibodies (LY3321367,[38] Eli Lilly and Company; MBG453,[39] Novartis Pharmaceuticals; TSR-022,[40] Tesaro, Inc.) in combination with anti-PD-1 or anti-PD-L1 therapies are ongoing.

HAVCR2 is also an exhaustion maker for NK cells. Blockade of this receptor can improve the NK cells antitumor activity in esophageal cancer, melanoma and lung adenocarcinoma.[24]

The role of HAVCR2 in the T-cell dysfunction has been investigated in chronic viral infections. Together with PD-1, HAVCR2 negatively regulate CD8+ T-cells and thus, in vivo blockade of HAVCR2 and PD-1 led to the restoring of antiviral immunity.[41]

A recent genome-wide association study (GWAS) has found that genetic variations in HAVCR2 are associated with late-onset sporadic Alzheimer's disease (LOAD). HARVC2 is capable of interacting with amyloid-β precursor protein.[42]

The page also lists some ligands that bind to TIM-3, a couple of which rang a bell.

Gal-9
HAVCR2 is primarily activated by soluble galectin-9.[25] The engagement leads to stimulation of an influx of calcium to intracellular space and induction of programmed cell death, apoptosis, cell necrosis or T cell anergy.[26][17][18] As a consequence, a suppression of Th1 and Th17 responses and induction of immune tolerance occurs, gal-9/HAVCR2 increases the immunosuppressive activity of Treg cells.[14]

Now published as —

Exploring the role of galectin-9 and artemin as biomarkers in long COVID with chronic fatigue syndrome: links to inflammation and cognitive function
Elahi, Shokrollah; Rezaeifar, Maryam; Osman, Mohammed; Shahbaz, Shima

This study aimed to assess plasma galectin-9 (Gal-9) and artemin (ARTN) concentrations as potential biomarkers to differentiate individuals with Long COVID (LC) patients with myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) from SARS-CoV-2 recovered (R) and healthy controls (HCs).

Receiver operating characteristic (ROC) curve analysis determined a cut-off value of plasma Gal-9 and ARTN to differentiate LC patients from the R group and HCs in two independent cohorts. Positive correlations were observed between elevated plasma Gal-9 levels and inflammatory markers (e.g. SAA and IP-10), as well as sCD14 and I-FABP in LC patients. Gal-9 also exhibited a positive correlation with cognitive failure scores, suggesting its potential role in cognitive impairment in LC patients with ME/CFS.

This study highlights plasma Gal-9 and/or ARTN as sensitive screening biomarkers for discriminating LC patients from controls. Notably, the elevation of LPS-binding protein in LC patients, as has been observed in HIV infected individuals, suggests microbial translocation. However, despite elevated Gal-9, we found a significant decline in ARTN levels in the plasma of people living with HIV (PLWH). Our study provides a novel and important role for Gal-9/ARTN in LC pathogenesis.


Link | PDF (Frontiers in Immunology) [Open Access]

It lists CEACAM1. We've seen upregulation of CEACAM3, though I'm not sure how related they are.

CEACAM1
The last known TIM3 receptor ligand is CEACAM1 glycoprotein. It is co-expressed with TIM3 T cells but also monocytes, macrophages, dendritic cells. It binds to the CC´ and FG loops of the TIM3 protein. CEACAM1 can also bind to TIM3 intracellularly (cis presentation) and is likely to be important for TIM-3 maturation on cell surface. The CEACAM1 binding contributes to the development of T cell tolerance, triggers the release of BAT3 from TIM-3 leading to inhibition of TCR signaling, and also inhibits the immune response of myeloid cells.[14]

Upregulated
CEACAM3

Edit: CEACAM1 was upregulated in long COVID but not ME/CFS in the quoted post. CEACAM3 was upregulated in both conditions.

 

[forestglip]

From reference 41 in Wikipedia page above:

Cooperation of Tim-3 and PD-1 in CD8 T-cell exhaustion during chronic viral infection, 2010, Hyun-Tak Jin et al

Our results show a correlation between coexpression of Tim-3 with PD-1 and deeper CD8 T cell exhaustion. However, it is not clear whether the Tim-3 and PD-1 pathways have overlapping functions or contribute independently to T cell dysfunction during chronic viral infection. To address this issue, we blocked the Tim-3 and PD-1 pathways with Tim-3-Ig fusion protein (Tim3Ig) and blocking antibody to PD-L1 (αPDL1), either alone or in combination during chronic LCMV [lymphocytic choriomeningitis virus] infection.

The viral load of mice treated with isotype control or Tim3Ig alone in the serum was not significantly altered after treatment (Fig. 5). In contrast, αPDL1 alone or dual blockade (Tim3Ig + αPDL1) treatments led to 4.2-fold and a 6.3-fold reductions, respectively, in virus titer of serum (Fig. 5).

Similarly, treatment with Tim3Ig alone resulted in slightly lower viral loads in the spleen, liver, and lung, whereas treatment with αPDL1 alone showed significant reduction of viral load in all tissues compared with the isotype control group (Fig. 5). More impressively, dual blockade with Tim3Ig and αPDL1 led to further reduction in viral loads in all tissues compared with blockade with αPDL1 alone (Fig. 5).

 

[Sean]

It looks like heterogeneity was a problem in this study. I wish researchers would do more sorting of potential participants on obvious symptom differences before they do their studies.

Heterogeneity is going to remain a significant confounder until we get some good biomarkers to help reveal any sub-groups and generally clarify diagnostic criteria.

 

[Hutan]

Heterogeneity is going to remain a significant confounder until we get some good biomarkers to help reveal any sub-groups and generally clarify diagnostic criteria.

For sure, but there are things that can be done to make more homogeneous cohorts e.g. people whose collection of symptoms meet ME/CFS criteria and who have had the symptoms for over one year; people with observable lung damage; people who were on mechanical ventilation and have observable lung damage...

 

[SNT Gatchaman]

It doesn't read as if these particular cohorts were hospitalised / ventilated or had observable lung damage.

All participants had a clearly defined episode of acute COVID-19 con¦rmed via direct molecular evidence of infection with SARS-CoV-2.

Eligible patients were men and non-pregnant women over the age of 18 years with no alternative explanatory disease and symptoms that persisted for at least 12 weeks after the initial diagnosis of acute COVID-19. One persistent symptom was sufficient for the diagnosis of long COVID. All patients underwent a comprehensive medical evaluation, including chest radiography, electrocardiography, lung function tests (spirometry with gas transfer as indicated and measurement of exhaled nitric oxide), and standard blood tests (autoantibody screens, bone, liver, and kidney function, coagulation screens, full blood count, and markers of nutrition).

All baseline medical evaluations were normal in patients with long COVID.

 

[Kalliope]

The paper has now been published

https://www.nature.com/articles/s41590-025-02135-5

 

[Kalliope]

Press release from Karolinska Institutet:
Long COVID biomarkers found - associated with respiratory problems

Quote:
“The proteins were mainly found in patients with Long COVID and severe respiratory problems,” says Dr Buggert. “This is a biomarker pattern that we know to be linked to inflammatory signal pathways involved in cell death and lung damage and that has also been observed in other patient groups with severe pulmonary disorders.”

https://via.tt.se/pressmeddelande/3...piratory-problems?publisherId=3236933&lang=sv

 

[Dolphin]

https://www.nature.com/articles/s41590-025-02135-5

• Article
• Open access
• Published: 30 April 2025

Identification of soluble biomarkers that associate with distinct manifestations of long COVID

• Yu Gao,
• Curtis Cai,
• Sarah Adamo,
• Elsa Biteus,
• Habiba Kamal,
• Lena Dager,
• Kelly L. Miners,
• Sian Llewellyn-Lacey,
• Kristin Ladell,
• Pragati S. Amratia,
• Kirsten Bentley,
• Simon Kollnberger,
• Jinghua Wu,
• Mily Akhirunnesa,
• Samantha A. Jones,
• Per Julin,
• Christer Lidman,
• Richard J. Stanton,
• Paul A. Goepfert,
• Michael J. Peluso,
• Steven G. Deeks,
• Helen E. Davies,
• Soo Aleman,
• Marcus Buggert &
• David A. Price

Nature Immunology volume 26, pages692–705 (2025)Cite this article

• Metricsdetails
  

Abstract

Long coronavirus disease (COVID) is a heterogeneous clinical condition of uncertain etiology triggered by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

Here we used ultrasensitive approaches to profile the immune system and the plasma proteome in healthy convalescent individuals and individuals with long COVID, spanning geographically independent cohorts from Sweden and the United Kingdom.

Symptomatic disease was not consistently associated with quantitative differences in immune cell lineage composition or antiviral T cell immunity.

Healthy convalescent individuals nonetheless exhibited higher titers of neutralizing antibodies against SARS-CoV-2 than individuals with long COVID, and extensive phenotypic analyses revealed a subtle increase in the expression of some co-inhibitory receptors, most notably PD-1 and TIM-3, among SARS-CoV-2 nonspike-specific CD8+ T cells in individuals with long COVID.

We further identified a shared plasma biomarker signature of disease linking breathlessness with apoptotic inflammatory networks centered on various proteins, including CCL3, CD40, IKBKG, IL-18 and IRAK1, and dysregulated pathways associated with cell cycle progression, lung injury and platelet activation, which could potentially inform the diagnosis and treatment of long COVID.

 

[Dolphin]

https://www.eurekalert.org/news-releases/1082365

News Release 30-Apr-2025
Long COVID biomarkers found – associated with respiratory problems
Peer-Reviewed Publication

Karolinska Institutet


Researchers at Karolinska Institutet have identified biomarkers in the blood associated with symptoms of long COVID, particularly severe respiratory disorders. The discovery can pave the way for future diagnosis and treatment. The results are published in the top-ranking scientific journal Nature Immunology.

Long COVID, also known as Post COVID, is a condition characterised by persistent symptoms of previous COVID-19. A new study from Karolinska Institutet, Sweden, and Cardiff University, UK, led by Marcus Buggert, docent at the Department of Medicine, Karolinska Institutet (Huddinge), has identified a set of proteins in the blood of people with Long COVID.

“The proteins were mainly found in patients with Long COVID and severe respiratory problems,” says Dr Buggert. “This is a biomarker pattern that we know to be linked to inflammatory signal pathways involved in cell death and lung damage and that has also been observed in other patient groups with severe pulmonary disorders.”

Detailed analysis of blood samples

Severe and permanent symptoms of acute breathlessness is one of the most common and most typical symptoms of Long COVID. The researchers also studied samples from a group of patients that had recovered from their previous COVID-19 and who had, interestingly, none of these proteins in their blood.

The finding was based on a detailed analysis of blood samples from independent patient groups in Sweden and the UK, something that had not been done before.

All 265 patients who participated in the study had contracted COVID-19 during the early days of the pandemic when no vaccine was yet available. Using advanced techniques, the researchers measured thousands of proteins in the blood plasma, which they related to the patients’ symptoms. They also used flow cytometry to conduct immunological analyses.

“By identifying the proteins that are elevated in affected patients, we’re creating a platform from which to develop diagnostic tools and new targeted therapies,” Dr Buggert says. “This is especially important since there are no specific biomarkers and treatments for Long COVID.”

Symptom biology

The results of the study expose the underlying biological processes that can cause certain patients to experience severe symptoms long after previous COVID-19.

The next step in the research is to understand what underpins this pattern by studying lung and gastrointestinal tissue. In doing so, the researchers hope to locate the source of the identified proteins and find if there is any remaining inflammation or tissue damage in specific organs of patients with Long COVID.

The study was funded by the PolyBio Research Foundation, the Swedish Research Council, SciLifeLab/KAW National COVID-19 Research Program, the Knut and Alice Wallenberg Foundation and Karolinska Institutet. Some of the co-authors receive consultancy and lecture fees from pharmaceutical companies, but they are unconnected to this study.

Publication: “Identification of soluble biomarkers that associate with distinct manifestations of long COVID”, Yu Gao, Curtis Cai, Sarah Adamo, Elsa Biteus, Habiba Kamal, Lena Dager, Kelly L. Miners, Sian Llewellyn-Lacey, Kristin Ladell, Pragati S. Amratia, Kirsten Bentley, Simon Kollnberger, Jinghua Wu, Mily Akhirunnesa, Samantha A. Jones, Per Julin, Christer Lidman, Richard J. Stanton, Paul A. Goepfert, Michael J. Peluso, Steven G. Deeks, Helen E. Davies, Soo Aleman, Marcus Buggert & David A. Price, Nature Immunology, online 30 April 2025, doi: 10.1038/s41590-025-02135-5.

Journal
Nature Immunology

DOI
10.1038/s41590-025-02135-5

Method of Research
Observational study

Subject of Research
People

Article Title
Identification of soluble biomarkers that associate with distinct manifestations of long COVID

Article Publication Date
30-Apr-2025