Projects / Programmes
Intercellular transfer of the signaling complex by the extracellular vesicles secreted from the WM-associated MyD88 mutation carrying cells
| Code |
Science |
Field |
Subfield |
| 3.01.00 |
Medical sciences |
Microbiology and immunology |
|
| Code |
Science |
Field |
| B500 |
Biomedical sciences |
Immunology, serology, transplantation |
| Code |
Science |
Field |
| 3.01 |
Medical and Health Sciences |
Basic medicine |
extracellular vesicles, MyD88, chronic inflammation, cancer
Researchers (15)
Organisations (2)
Abstract
Abstract
Innate immune system is the first line of defense against pathogens and endogenous damage. This is however a double-edged sword as unregulated inflammation may lead to pathology, including cancer. Toll-like receptors, interleukin-1 receptor and inflammasomes are pivotal in these processes. Cytosolic protein adaptor MyD88 plays a key role in innate immune response. After activation of TLRs and IL-1R MyD88 is recruited to the membrane, where it forms large myddosomal complex with IRAK4 and IRAK2 thus leading to activation of the inflammatory signaling pathway. Mutation within TIR domain of MyD88 (L265P), which has been observed in )90% of patients with Waldenström’s macroglobulinaemia (WM) and in other types of B-cell lymphomas leads to a spontaneous myddosome formation and activation of inflammatory pathways, which represents a survival signal in WM cells. Inflammatory environment is crucial for the establishment and survival of cancer as inhibitors of MyD88 pathway kill cancer cells.
In this project we propose the existence of a new, never before reported mechanism of intercellular communication via transfer of the myddosomal complex from cancer cells via extracellular vesicles (EVs). EVs are submicron membrane-delimited vesicles that can transfer cytosolic components. Recent studies highlighted the role of miRNA transfer via EVs in establishment of the microenvironment in cancer, however transfer of proteins may also affect the phenotype of the recipient cells.
Osnova naše hipoteze je dejstvo, da je midosomski kompleks povezan z membrano in sproži oligomerizacijo, kateri sta značilnosti proteinov, ki se vnesejo v EV. Naša skupina je nedavno pokazala, da lahko MyD88L265P veže endogeni MyD88wt in aktivira celice neodvisno od receptorja (Avbelj et al., Blood, 2015), torej bi EV iz rakastih celic lahko sprožili signalizacijo v tarčnih celicah v mikro okolju raka.
The main objectives of the project are:
Detect MyD88 and other myddosomal components (IRAK4, IRAK2) in the EVs isolated from cells overexpressing MyD88 and from B cell lymphoma cell lines.
Investigate activation of the MyD88-mediated signaling in different recipient cell types by the uptake of MyD88L265P containing EVs in wt and MyD88 deficient cells, test the inhibition of MyD88 signaling in the recipient cells by a BTK inhibitor drug ibrutinib, recently approved for the therapy of WM and inhibitory peptide.
Investigate the mechanism of the EV uptake and detect the intracellular localization of the MyD88L265P, delivered by the EVs and interaction with MyD88wt in the recipient cells using confocal fluorescence microscopy.
Investigate the effect of EVs on the recipient cell response for shaping the tumor microenviroment.
Investigate the effect of EVs comprising MyD88L265P in an animal model by introduction of EVs into the bone marrow of mice.
Investigate the presence of MyD88L265P in EVs from bone marrow samples of patients with WM and their ability to trigger signaling in the recipient cells.
Preliminary results confirm that the MyD88 is indeed released via EVs and, most importantly, that the EVs carrying myddosomal complex can activate TLR independent signaling in the recipient cells. Physiological relevance of these findings will be tested in the animal model and, importantly, the presence of myddosomal components and the ability to activate nontransformed cells will be investigated in EVs from bone marrow aspirates from patients with WM, provided through a collaboration with a leading researcher and clinician in WM prof. Steven Treon from Dana Farber Cancer Institute and Harvard Medical School, who has one of the largest collection of clinical samples of patients with WM in the world. Results of the project promise to introduce a new important paradigm on cell communication and cancer immunology with potentials for new therapeutic approaches.
Significance for science
Although the mechanisms by which inflammation promotes neoplastic transformation are not fully understood, it seems that tumor development is linked to chronic inflammation. Persistent inflammation establishes a microenvironment, which influences tumor formation and growth. Why inflammation develops and how it is spread in the tissue or a body is still detrimental, although knowledge on molecular mechanisms is prerequisite for development of efficient therapies and diagnostics. Role of MyD88 in chronic inflammation is evident. The transmission by EVs is new and as such it has a strong breakthrough potential that is limited not only to DLBCL, WM and MyD88 as it is likely that other signaling pathway complexes might be transferred and exert the effect in the recipient cells in a similar way.
The methods that we will use are known, but for example the injection of EVs into the bone marrow has not yet been described in the literature, so we are expanding the use of methods for analyzing the function of EVs. Additionally, there are currently no publications on the detection of proteins inside isolated EVs using TEM, so this will be a new contribution to the EV field as well. Implementing new methods in addition to the novelty of the topic itself has the potential to be published in high ranking journal and thus contribute to the development of the science in Slovenia and broadly.
Using innovative strategies and new ideas we will also educate students (undergraduate and PhD students) for research and will pass our knowledge to scientific community through workshops and conferences.
Significance for the country
The five year survival of patients with B cell lymphoma increased in the last decade due to the introduction of new therapeutic approaches, however survival is still low and new therapeutic approaches that can only originate from better understanding of the cancer development contributing to the development and progression of lymphoma are still needed. Identification of the unexpected role of MyD88 identified NFkB signaling as the cancer survival signal five years ago strongly contributed to the recent approval of ibrutinib for the therapy of WM. This project will investigate a completely new mechanism of cellular communication through the transfer of signaling mediators. As such it has a strong breakthrough potential that is limited not only to DLBCL, WM and MyD88 as it is likely that other signaling pathway complexes might be transferred and exert the effect in the recipient cells in a similar way.
In addition to this fundamental discovery, that is likely to have a significant impact for the basic understanding of physiological processes, its impact may also have a strong translational potential. Several mechanisms of interference with the formation and uptake of EVs have been described (e.g. targeting sphingomyelinase, tetraspanins or endocytosis), therefore it is likely that the therapeutic efficiency of targeting EV formation and delivery could be investigated in different types of cancer. Additionally, EVs have already been used as a prognostic marker for the cancer progression and the presence of myddosome and its components in EVs in different body fluids may turn out to provide a diagnostically relevant information that could be used to select the most appropriate therapy. While EVs from bone marrow biopsies of WM patients and comparison to EVs from other types of lymphoma will be used in this project it is likely that the EVs from cancer cells are present in the peripheral blood where they could provide a much easier accessible marker.
Therefore the impact of this project includes both the fundamental advancement of science as well as the translation for human health for one of the most devastating diseases. Project has strong interdisciplinary components and strong connections between molecular and cellular immunology and clinicians. A link to the world’s top clinicians who have been involved in the testing and registration of ibrutinib for the therapy of WM will provide an excellent clinically relevant feedback and possibility for translation of the new therapeutic approaches.
Most important scientific results
Interim report,
final report
Most important socioeconomically and culturally relevant results
Final report
