Please can you give a brief introduction to type 3 interferons (IFN)?
The type 3 interferons - usually called the “lambda” interferons, or “IFNL” - are the most recently identified IFNs. We (myself and Sergei Kotenko) originally identified the receptor and then the three ligands (IFNL1,2,3; very recently a fourth, IFNL4, was discovered); the key paper was published in 2003.
The Lambdas signal very similarly to the Type 1s and so Type 1 and Type 3 IFNs do share a lot of activities.
The Lambdas are most interesting because of the distribution of their receptors. While many cells can make Lambdas, the Lambda receptors are distributed only throughout the epithelia and lymphoid tissues.
When one considers that infection - the actual act of infection - takes place at the interface between the inside and outside of the body, then one realises that the lambda interferons are dedicated to protecting us at this point of infection; because their receptors are only found on epithelial and immune cells they only act at this interface.
There are clear antiviral activities associated with all the Lambdas, but IFNL3 has the greatest activity in this regard. At HUMIGEN, our lab focussed on the immunoregulatory activity of IFNL, particularly IFNL1, and this was how we came to understand the specific action on the Th2 response.
What are T helper 2 (Th2) cells and what role are they thought to they play in asthma and severe allergies?
Th2 cells are the immune cells that secrete IL-4, IL-5 and IL-13, the cytokines associated with allergy, atopy and asthma. IL-5 has a key role in attracting a major asthma cell type, the eosinophil.
Many asthma patients, particularly those with severe asthma, have pathology derived from eosinophils and often, high levels of IgE (which activates eosinophils in the airway).
IL-4, also secreted by Th2 cells, is important in promoting the specific production of IgE from B-cells and IL-13 promotes the secretion of eosinophil chemotactic factors - the “eotaxins” - from airway epithelial cells. So, it’s clear that these three Th2-derived cytokines play an important role in asthma pathology.
In addition to their effect on eosinophils, these cytokines also activate mast cells, resulting in the release of histamines and other asthma-promoting substances.
Therefore, by coordinating and activating asthma-driving responses from these three key cell-types - the B-cell, the eosinophil and the mast cell - Th2 cells are really at the centre of asthmatic and severe allergic responses.
When was the ability of Type 3 IFNs to restrict Th2 cells development via suppression of GATA3 expression first described?
We made our first observations on this when Joyce Eskdale and Bill Jordan in my lab showed that IFNL1 could prevent the secretion of chemotactic factors such as IP-10 and I-TAC, and inhibit IL-13 secretion, without affecting IFN-gamma production.
These experiments were the first indication that Th2 cells were being targeted specifically by IFN lambda, and that the diminished Th2 response was not the consequence of an activation of the Th1 response.
This work was continued and expanded, and we showed that IL-13 in particular was affected by IFNL1. This was the point that led us to examine the role of IFNL1 in modifying the development of newly-stimulated Th2 cells; it became clear that the effect was at the very core of Th2 development, because IFNL1 was preventing the expression of GATA3.
Since GATA3 is the signature transcription factor that drives all aspects of Th2 development, then when GATA3 does not get expressed, downstream events typical of Th2 responses - IL-4 receptor expression, IL-4, 5 and 13 cytokine secretion for example - do not happen.
This really modified our understanding how fundamental IFNL1’s role was in regulating Th2. The work was published in four papers from our lab, between 2007 and 2009, and the key investigators in this process were: Jihong Dai, Joyce Eskdale, Grant E Gallagher, Bill Jordan, Nick Megjugorac, Vera Pekarek, Shekar Srinivas and Raymond Yu.
A supporting paper from an independent laboratory showing that Type-1 IFNs also had some aspects of Th2 inhibitory activity was published almost exactly a year later in 2010, and it followed the experimental model we presented in 2009.
Why was this an important finding?
This was an important finding for three reasons, in my view. Firstly, it defined clearly a functional niche for IFNL out of the anti-viral space where the vast majority of investigators were pursuing it.
Secondly, it indicated a role for Lambda in a key human disease (asthma) that was completely consistent with its known locations of action - the epithelial and immune cell populations - in a way that tied these two cell compartments together.
Finally, it provided a mechanism that explained the observation in 2006 where a UK study of asthma patients and healthy individuals showed that asthma patients are deficient in Lambda production; the absence of Lambda would remove a key regulator of IL-13 (as well as diminish the response against respiratory viruses so often associated with asthma exacerbations in asthma patients), allowing the Th2 response to proceed unchecked and so accelerate asthma pathology. This in turn provided HUMIGEN with a stimulus to develop therapeutic strategies that would promote Lambda expression in the airway.
Please can you outline the work your lab has been doing since this observation?
Since our discovery of IFN regulation of GATA3 expression and Th2 development, we have explored two pathways.
The first was to understand whether Lambda expression was itself responsive to the presence of IL-4 and/or IL-13 in the immunological environment, creating a negative feedback loop, and this turned out to be the case.
We defined a negative feedback loop where IL-13 or IL-4 act on macrophages to up-regulate the expression of IL-1 receptor antagonist, and this in turn acted on plasmacytoid dendritic cells (pDCs), which up-regulate Lambda and consequently diminished IL-4/IL-13 expression.
We were also able to show that Lambda pre-sensitises pDCs to Lambda, changing their cell surface and up-regulating the Lambda receptor, and the consequence of these changes was that they modified their interaction with T-cells, resulting in lower IL-13 production, amongst other things. These studies were published in 2009 and 2010. The key investigators in this arm of the work were Grant E Gallagher and Nick Megjugorac, and again were duplicated in part by an independent pDC laboratory in 2012.
In parallel we decided to understand the regulation of IFNL1 expression in the human airway. Understanding this was our first step to developing novel therapeutic strategies that could elevate IFNL in the airway while still retaining a physiological regulatory context.
This work uncovered showed that Lambda is under extremely tight repression in the human airway. This regulation has three aspects: following viral stimulation, a change in NFKappaB homo and hetero-dimers allows transcription with the aid of IRF1 but this does not progress until the repressive transcription factor ZEB1 leaves the promoter. After a very short period, ZEB1 returns and begins rapid re-repression of Lambda transcription and this is assisted by an additional repressor, BLIMP-1, which appears to displace IRF1.
We designed an anti-sense therapautic approach and showed that siRNA knockdown of ZEB1 or BLIMP-1 extended the expression of IFNL1 and led to increased secretion of IFNL1 protein.
This work was published in 2010 and the key investigators were Joyce Eskdale and Rachael Siegel. It stands today as the only report of IFNL regulation in an airway epithelial model.
What have been your main findings and what impact do you hope they will have?
Our main findings then have been to define a role for Lambda in regulating Th2 responses in the airway and understand how it works, and how it is regulated.
We have used these data to develop therapeutic strategies based around relief of Lambda repression in the airway to extend its presence without having to deliver the protein itself, and to develop cell-based therapies also.
We’ve protected these with a set of layered Patents that are available to protect the interests of partners who might wish to move our observations towards a clinical space.
We hope that these approaches will make a difference to the accelerating numbers of children and adults suffering from asthma and severe atopy, such as food or contact allergies.
What further research is needed to increase our understanding of the relationship between Type 3 IFNs and Th2 cells?
The next step in this work is to determine whether the approaches we have developed can be made suitable for clinical administration in humans. Before this, there needs to be a thorough investigation in a range of pre-clinical models.
Interestingly, the in vivo role for Lambda as a protective agent in asthma is predicted by studies in Lambda Receptor knock-out models, where clinical parameters were worsened when the receptor was absent.
What are HUMIGEN’s plans for the future?
HUMIGEN’s plans for the future are to aggressively pursue partnering with interested parties to address the pre-clinical validation of our approaches with Lambda in allergy and asthma. We are also bringing other pipelines forward that address the IL-23 pathway.
Where can readers find more information?
Readers can get more information directly from HUMIGEN by contacting Grant Gallagher, at: [email protected]
Information about Genesis Biotechnology Group can be obtained from Stephanie Thompson, at: [email protected]
Information about partnering HUMIGEN or GBG technologies can be obtained from Siu Lo, at: [email protected]
The publications referred to here can be obtained via their PMID numbers: 12483210; 17082759; 17252004; 18547367; 19346497; 19759281; 20233967; 22058416
The patents surrounding our work with IFN-lambda can be obtained from USPTO: 8,282,964; 8,349,808; 8,354,125; 8,771,666; 8,802,648; 20140057960
About Grant Gallagher
Grant Gallagher obtained his Bachelor’s degree in Biochemistry in 1980 and his PhD in T-cell immunology in 1984. Following a successful career in academia in the UK and the USA, he joined what is now the Genesis Biotechnology Group (GBG) in July of 2006; HUMIGEN was formed in 2007.
Since then through HUMIGEN, Dr Gallagher has led the GBG effort in immunology with regard to autoimmunity and more recently, immune-oncology.
Dr Gallagher is an author on 85 peer-reviewed publications and an inventor on 21 published patents, 12 of which are issued or allowed. He has been married for 33 years and has two children.
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