Galectin-3 causes CD8 dysfunction in severe Covid-19 cases

And we finally have a proper inhibitor to it

Both of the following papers are directly correlated with the following substack, while I often don’t repeat myself and left for the readers to “dig”, this is one of the very few substacks I break this unofficial rule, it is one of the most important dynamics of the virus and the Spike Protein, therefore the overall pathophysiology, and it will influence the long-term effects of the virus.

Chimeric nature of Galectin-3 and SARS-CoV-2

You must read the substack above to be able to comprehend what we are about to cover entails.

Epithelial galectin-3 induces mitochondrial complex inhibition and cell cycle arrest of CD8⁺ T Cells in severe/critical ill COVID-19

During virus infection, pathogen-specific CD8+ T cells are activated, proliferated and expanded to against virus infection, and cellular metabolism and mitochondrial function are the main determinants of this process. CD8+ T cell activation is accompanied by extensive metabolic reprogramming with increased expression of nutrient transporter proteins and enzymes involved in glycolysis. In addition, effector CD8+ T cells rely on enormous energy generated by oxidative phosphorylation and glycolysis to complete the cell cycle.

The mitochondrial oxidative phosphorylation system, a key system for energy supply in eukaryotic cells, comprises five enzymatic complexes (mitochondrial complex I to V) and two mobile electron carriers. Mitochondrial complexes I (NADH-CoQ reductase), II (succinate-CoQ reductase), III (CoQH2-c reductase), IV (cytochrome c oxidase), and V (oligomycin sensitive ATPase) catalyze ATP production through a series of redox reactions, and adequate energy supply is of vital importance for CD8+ T cell proliferation and expansion. Therefore, insufficient cellular metabolism and mitochondrial energy generation might be responsible for the impaired functions and limited expansion of CD8+ T cells in severe/critical COVID-19.

Over the past three years, a contentious debate has ensued regarding the CD8+ T cells' malfunctioning (the cells responsible for battling viruses, intracellular bacteria, and cancer surveillance) and the precise cause of said malfunction. Although no single causative agent has been identified, a plethora of papers, some of which I have previously discussed in this substack, demonstrate a consistent outcome: CD8+ T cells are of paramount importance in effectively combating and eradicating diseases, including SARS-CoV-2. In this particular study, a subset of CD8+ T cells was found to have undergone proliferation and depletion, rendering them exhausted and incapable of producing more of these cells, all while demonstrating impaired mitochondrial function in severe and critical patients. Subsequent experiments revealed that the ORF3a protein from SARS-CoV-2 induced Galectin-3 expression in epithelial cells, leading to the inhibition of mitochondrial function and loss of proliferating ability, ultimately causing low CD8+ expansion.

The authors of this study sought to understand precisely how and why this dysfunction manifests. They examined CD8+ T cells from severe and critical cases, dividing them into 13 clusters, and found Cluster 10 to be the most severely compromised. Cluster 3, while not demonstrating any obvious differences in the expression of certain pathways/genes, may be correlated with the development of dysfunctional cells in Cluster 10. The same dynamic as further observed in regard to mitochondrial function, such observation is found below. Cluster 10 had significant downregulation of mitochondrial-related pathways. We find ourselves back at a point I raise almost every other week at this point, at the core of this issue (SARS-CoV-2 pathogenesis) lies improper mitochondrial function.

Impairment of mitochondrial function in the cell-cycle-arrest cluster

Enrichment results of C10 showed plenty of downregulation of mitochondrial-related pathways, while enriched terms in C3 had little relationship with mitochondrial function. This strong decrease of respiratory chain-related genes and mitochondrial-related signaling pathways revealed that the mitochondrial biogenesis of C10 was suppressed, and its paramount function — supply energy for life activities — was impaired.

The entire family of Galectins plays important roles in various stages of cell function, including immune cells, in T cells they often negatively impact their function by directly impacting the proliferation and survival of these cells.

The analysis revealed that two galectins, galectin-3 and galectin-9, were expressed at high levels in certain epithelial cells. Additionally, six ligand-receptor interactions involving these galectins and specific proteins on the surface of CD8+ T cells were identified. The analysis suggested that the epithelial cells expressing high levels of galectins had a greater negative effect on Cluster 10 (a compromised group of CD8+ T cells) than on Cluster 3. Furthermore, certain receptors that galectins bind to were found to be more highly expressed in Cluster 10 compared to Cluster 3, indicating that the binding of galectins to these receptors may contribute to the dysfunctional state of the CD8+ T cells in Cluster 10.

Galectin-3 inhibits mitochondrial complex III/IV genes transcription by suppressing NRF-1

Our single-cell transcriptome analysis revealed that galectin-3-related ligand-receptor interactions between lung epithelial cell clusters and CD8+ T cell clusters were significantly enhanced in CD8+ T cell subsets with impaired mitochondrial function and cell cycle arrest.

Taken together, our findings suggest that galectin-3 mediates the down-regulation of mitochondrial complex III/IV genes by suppressing NRF1 nuclear translocation and transcriptional activation.

The down-regulation of mitochondrial function comes from the higher expression of Galectin-3 and its interaction with many cells, and this is a byproduct of infection of epithelial cells, and ORF3a plays an impactful role in this dynamic. This protein is completely different from the NTD present in the Spike Protein, but the pathogenic effects describe here are significant because the Spike Protein induces the production of Gal-3 everywhere, not just in epithelial cells.

This also will serve as a reminder of why so many people treated with Paxlovid suffer rebounds, have poor health after the treatment, and become little mutation machines since Paxlovid directly affects both types of CD cells you need to properly fight the infection.

Paxlovid use becomes dangerous, HCQ works

As I stated to the degree of becoming repetitive and a nuisance, Galectins play a massive role in many of the different aspects of what SARS-CoV-2 is capable of inducing, so I will demonstrate and cite merely a few of the most significant ones in my opinion (new ones btw, not merely repeating myself endlessly). The following paper is obligatory reading for anyone who wants to understand both galectin and autoimmunity (therefore autoimmunity caused by infections).

Galectin-1 and -3 are expressed in nearly every tissue and by most cell types examined^7,8, whereas other galectins appear to have a more restricted expression profile: galectin-7 is mainly expressed by stratified epithelial cells; galectin-9 is mainly expressed by gastrointestinal epithelial cells, the thymus and endothelial cells; and galectin-12 is mainly expressed by adipocytes^9,10,11,12.

Fig. 2: Regulation of adaptive immunity by galectins.

Intracellular galectin-3 was also found to be localized in the immunological synapse in both naive and memory CD8⁺ T cells when activated by γ-herpesvirus (MHV68)⁴⁰. Galectin-3 knockout mice mount a stronger MHV68-specific CD8⁺ T cell response compared with wild-type mice⁴⁰, indicating that intracellular galectin-3 can upregulate the threshold for T cell activation. Overall, these studies suggest that both extracellular and intracellular galectin-3 can dampen T cell activation by altering the distribution and half-life of distinct cell surface glycoproteins.

For example, galectin-1, -8 or -9 knockouts have an exaggerated clinical score and increased numbers of T_H1 cells and T_H17 cells in the EAE model⁴⁵. Of note, increases in the number of CD8⁺ T cells have also been observed in galectin-9 knockout mice following challenge with herpes simplex virus (HSV)⁵², suggesting that the regulation of T cell fate is not limited to CD4⁺ T cells or autoimmunity. Together, these studies suggest that several different types of galectins have a role in the contraction of T cell populations and raise the possibility that systemic administration of galectins may be of therapeutic value in autoimmune diseases.

Given the contextual, intricated, double-edge nature of Galectins, and their fairly complex modulatory roles in many cell types, states, and in the presence of sugar molecules, decyphering their roles is a colossal task, yet in the last decade and a half scientists made strides. The first paragraph in the quote above is the one that bears more significance when we talk about SARS-CoV-2 infection and its long-term sequelae and impact. γ-herpesvirus is a subfamily of herpes viruses, with major significance to long-term damage from acute (bystander effect) and latent infections, belonging to this family are HHV-8 called Kaposi sarcoma Herpesvirus (KSHV), HHV-4 formally known everywhere as Epstein-Barr Virus, both known oncogenic viruses, and Epstein-Barr being closely correlated with Long-Covid. MHV68 is a closely related virus to human gamma herpesvirus and a common animal model to study their human counterpart.

Taken together with what we covered so far, now we have another mechanistic understanding of how, where, and why severe and critically ill Covid patients have months-long dysfunctional immunity, and how certain viruses are escaping latency, with the multiple front Galectin production. This also gives hint (insights if you know where to look) into why a non-inflammatory and tolerant immune state is becoming more predominant in those vaccinated against SARS-CoV-2.

Coming to the second section of this substack, the good news part.

An Oral Galectin Inhibitor in COVID-19—A Phase II Randomized Controlled Trial

SARS-CoV-2 vaccines play an important role in reducing disease severity, hospitalization, and death, although they failed to prevent the transmission of SARS-CoV-2 variants.Therefore, an effective inhibitor of galectin-3 (Gal-3) could be used to treat and prevent the transmission of COVID-19.ProLectin-M (PL-M), a Gal-3 antagonist, was shown to interact with Gal-3 and thereby prevent cellular entry of SARS-CoV-2 in previous studies. Aim: The present study aimed to further evaluate the therapeutic effect of PL-M tablets in 34 subjects with COVID-19. Methods: The efficacy of PL-M was evaluated in a randomized, double-blind, placebo-controlled clinical study in patients with mild to moderately severe COVID-19. Primary endpoints included changes in the absolute RT-PCR Ct values of the nucleocapsid and open reading frame (ORF) genes from baseline to days 3 and 7. The incidence of adverse events, changes in blood biochemistry, inflammatory biomarkers, and levels of antibodies against COVID-19 were also evaluated as part of the safety evaluation. Results: PL-M treatment significantly (p = 0.001) increased RT-PCR cycle counts for N and ORF genes on days 3 (Ct values 32.09 ± 2.39 and 30.69 ± 3.38, respectively) and 7 (Ct values 34.91 ± 0.39 and 34.85 ± 0.61, respectively) compared to a placebo treatment. On day 3, 14 subjects in the PL-M group had cycle counts for the N gene above the cut-off value of 29 (target cycle count 29), whereas on day 7, all subjects had cycle counts above the cut-off value. Ct values in placebo subjects were consistently less than 29, and no placebo subjects were RT-PCR-negative until day 7. Most of the symptoms disappeared completely after receiving PL-M treatment for 7 days in more patients compared to the placebo group. Conclusion: PL-M is safe and effective for clinical use in reducing viral loads and promoting rapid viral clearance in COVID-19 patients by inhibiting SARS-CoV-2 entry into cells through the inhibition of Gal-3.

In relation to Gal-3 inhibitors, it is plausible that PL-M interacts with Gal-3 in the same way as the structurally similar NTD of the S1 subunit of SARS-CoV-2. Overall, the inhibition of Gal-3 and blockade of the NTD of the S1 subunit and ACE2 receptors may be the anti-SARS-CoV-2 actions of PL-M. Gal-3 is upregulated in influenza A virus infection and supports replication.

If you read so far, and especially if you read the previous Galectin-3 piece, you are now fully aware of the significance, and impact Gal-3 has not only in regards to SARS-CoV-2 infection but to many diseases, from chronic to latent viruses, autoimmunity to inflammatory conditions, a true sugary chimera. Being such a major player at the very start of many molecular cascades, it is no surprise that finally attention to the modulation and inhibition of Galectins has finally been achieved.

In regards to its pathogenic role in a sizable portion of many suffering from IgG4-Related Diseases, among many other conditions such as Long Covid (with its interplay with many latent infections and chronic conditions), it is safe to say that I would basically cut the red tape, scale production to hundreds of millions, and start prescribing this inhibitor (For the curious and anyone else reading this, here is the site of the company responsible for this inhibitor, BioXyTran).

Save a very specific molecular pathway that Pfizer has thrown millions (at this point most likely 10+) into solving, with no solution proposed so far, this is effectively “the next best thing”, and one that is within reach of most countries in need.

Shortly after writing the first Galectin-3 substack, I wrote this short guide on how to modulate Galectin-3, and once again it will read “repetitive” by the simple fact that a lot of the supplements I suggest do influence Gal3 in a positive way. FYI modulation is fairly different than inhibition, modulation has a lot of off-target effects, often positive, and this is no substitute for a designer inhibitor, it was just what we had at the time.

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