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FRIMED2-FRIPRO forskerprosjekt, medisin og helse

Intracellular immunity

Alternative title: Intracellulær immunitet

Awarded: NOK 4.7 mill.

Antibodies are key molecules in the immune response. Although previously thought to carry out their protective role solely outside of cells, in the blood, tissues and on mucous membranes, recent research has revealed that antibodies also function to block infections inside infected cells. Specifically, antibodies bind TRIM21, a receptor molecule found in the cytosol of all cells. Using adenovirus infection as model, we have studied how IgG binding to both virus and TRIM21 triggers virus degradation inside cells and signals inflammation before the virus has had time to multiply and spread. We have produced panels of engineered antibody variants and tested them for their ability to inhibit both virus production and the signaling that initiates inflammation. We find that IgG coated viral particles are neutralized even under suboptimal conditions, whereas induction of immune signaling is balanced according to the strength of the incoming stimuli. Furthermore, the complement system is vital for the antibody mediated defense. In the classical pathway, microbe-bound antibodies recruit complement components. We have unraveled how antibody mediated complement activation synergizes with TRIM21 to block adenovirus infection. We have also elucidated the intracellular anti-virus activity of the IgG subclasses, as humans have four, namely IgG1, IgG2, IgG3 and IgG4. The subclasses differ in structure, but bind equally well to TRIM21 via their Fc part. However, we found that IgG3 induces a much more potent TRIM21 dependent antiviral response than IgG1, IgG2 and IgG4. Our data indicate that flexibility of the Fabs and Fc accessibility determines the TRIM21 activity. Furthermore, the complement dependent neutralization activity was IgG subclass dependent, with IgG3 again being the most efficient mediator. Interestingly, adenoviral vectors hold great promise for gene therapy, the induction of antitumor immunity, and vaccination against viral infections. We have studied how preexisting antibodies elicited by natural infection with particular types of adenovirus limit the efficacy of gene therapy in a TRIM21 dependent manner. TRIM21 induced virus degradation prevents the desired expression of the transgene carried by the vector. Efficient gene therapy can be partially restored by Fab-mediated virus shielding from TRIM21 attachment. In summary, the data generated in the project broadens our understanding of intracellular antiviral immunity. Furthermore, the new knowledge will pave the way for design of engineered IgG variants with improved antiviral activity, and the results will guide improved Ad5-based gene delivery therapy.

Adenoviral vectors hold great promise for gene therapy and the induction of antitumor immunity. However, preexisting antibodies elicited by natural infection with adenovirus may prevent th expression of the transgene carried by such vectors. We have shown how a cytoplasmic receptor, TRIM21, participates in neutralizing adenovirus and related viruses. Once the antibodies on the gene-delivery vehicle are in the cytoplasm, they interact with TRIM21 and initiate destruction of the virus. In addition, the antibody-TRIM21 interaction sets off cascades of inflammatory signaling, which can be deleterious. Importantly, efficient gene therapy can be improved by Fab- mediated virus shielding. We have produced panels of engineered antibody variants that we have tested for their ability to neutralize virus in a TRIM21 and complement dependent manner. The results will guide the design of engineered IgGs with improved antiviral activity.

Antibodies (Abs) are key molecules in the defence against pathogens such as viruses and bacteria. Although previously thought to give protection solely in the extracellular environment and inside intracellular vesicles, recent research has revealed that Abs also function in the cytosol of infected cells. Most virus infections occur at mucosal surfaces, where IgG molecules are present as they are carried across polarized cells by the neonatal Fc receptor (FcRn). This project aims to elucidate the intracellular anti-viral activity of IgG, and how intracellular receptors for the Fc region of IgG expressed by epithelial cells contribute. Humans have four subclasses of IgG; IgG1, 2, 3 and 4. All four bind FcRn, an endosomal Fc receptor, as well as TRIM21, a recently characterized cytosolic Fc receptor. While FcRn captures IgG-virus complexes in vesicular transit across cell layers, virus-IgG complexes that enter the cytosol, are recognized by TRIM21. Upon binding, TRIM21 activates proinflammatory signalling pathways and targets the complexes for proteasomal degradation. My research group has new and exciting results showing that the intracellular anti-viral activity of IgG3 is far better than that of the other subclasses. While signalling was found to be TRIM21 dependent, inhibition of virus production was not, and strong in cells that did not express TRIM21. Thus, IgG3 uses a TRIM21 independent, as yet unknown mechanism. This project aims to characterize the molecular mechanism at play and will surely provide groundbreaking new insights into anti-viral immune responses, and pave the way for design of new therapeutics. We will also design IgG variants to study how and why IgG3 differs from the other subclasses. The IgG variants will have specificity for human adenovirus (AdV5) or human immunodeficiency virus (HIV) and be tested for antiviral activity in established cellular assays by us at the University of Oslo (AdV5) or The Scripps Research Institute (HIV).

Funding scheme:

FRIMED2-FRIPRO forskerprosjekt, medisin og helse

Funding Sources