Targeted immunotherapy against autoreactive B cells in autoimmune diseases

‍

Perico et al. Bi-specific autoantigen-T cell engagers as targeted immunotherapy for autoreactive B cell depletion in autoimmune diseases, Frontiers in Immunology, 2024

‍

The availability of treatments for autoimmune diseases is limited, and they are often associated with undesirable side effects. In this paper the authors explore the use of a new T-cell engager (TCE) for treating an autoimmune disease of the kidney and validate their strategy using a humanized genO-panhCD3 mouse model provided by genOway.

Membranous nephropathy (MN), is an autoimmune disease of the kidney, characterized by the attack of antibodies to proteins expressed in the glomerulus, responsible for blood filtration. These autoantibodies¹ are produced by autoreactive B cells, which comprise 0.1% to 0.5% of the total circulating B cells and target PLA₂R, the phospholipase A₂ receptor, the primary autoantigen in MN². 

‍

Development and in vitro testing of a Bi-specific AutoAntigen-T cell Engager (BiAATE)

Building on the identification of this biomarker² and their previous work on targeting B cells for the treatment of MN³, Perico and colleagues developed a new class of immunotherapeutic molecules, the Bi-specific AutoAntigen-T cell Engagers (BiAATEs) to specifically target these autoreactive B cells⁴. These BiAATEs are engineered to express an autoantigen specific to MN, such as PLA₂R, and the single-chain variable fragment (scFv) of an antibody that targets CD3, as shown in the figure below (Figure 1).

‍

‍Figure 1: A) Schematic representation of the BiAATE, a bi-specific molecule that expresses the autoantigen involved in a specific autoimmune disease linked to the single-chain variable fragment (scFv) of an anti-CD3 by a small flexible linker. B) The BiAATE redirects the lytic activity of T cells against the surface immunoglobulin (sIg)⁺ included in the B-cell receptor (BCR) of autoreactive B cells. Activation of T cells leads to the production of perforin (PFN), granzyme B (GzmB), interferon gamma (IFNγ), and tumor necrosis factor alpha (TNFα), which induce the selective removal of autoreactive B cells. Adapted from Perico et al., 2024⁴.

‍

The ability of the BiAATE to simultaneously bind T cells’ CD3 and these autoantibodies was evaluated, and binding was only observed when T-cell extracts were incubated with the BiAATE and sera from MN patients. This finding indicates that the construct provides the link between T cells and anti-PLA₂R antibodies. The authors then tested whether BiAATE would specifically target autoreactive B cells without affecting the remaining population of B cells, and limit the secretion of anti-PLA₂R antibodies. Treatment with both BiAATE and blinatumomab, an anti-CD3 and anti-CD19 bispecific antibody which helps T cells deplete malignant B cells⁵ resulted in a decrease in anti-PLA₂R antibodies. Blinatumomab induced a strong decrease in B-cell viability in the blood while BiAATE did not, demonstrating not only that BiAATE specifically targets the small subpopulation of autoreactive B cells but also that, contrary to blinatumomab, it does not deplete healthy B cells.

‍

In vivo validation and advantages of this construct

To validate the function of the construct in vivo, Perico and colleagues employed a mouse model expressing the humanized CD3εγδ subunits (genO-panhCD3), thanks to a collaborative effort with genOway. This model displays unaltered thymocyte maturation and cellularity when compared with WT mice, suggesting that humanization has not altered the functionality of CD3. Indeed, T cells from the genO-panhCD3 model respond to anti-human CD3, as evidenced by IFNγ production and T-cell proliferation. Since the TCR/CD3 complex is functional, this model can be used for the assessment of new TCEs, such as BiAATE. Therefore, the authors proceeded with the immunization of genO-panhCD3 mice, to analyze whether BiAATE treatment decreased the levels of anti-PLA₂R antibodies in vivo. BiAATE treatment led to the reduction of anti-PLA₂R antibodies, when compared to the vehicle control (Figure 2). These results confirm that BiAATE can efficiently deplete autoantibodies, by stimulating T cells to eliminate autoreactive B cells. Nonetheless, it remains to be confirmed whether, in the context of the MN disease, this treatment would lead to improvements of the pathology and its symptoms.

‍

Figure 2: Quantification over time of IgG anti-PLA₂R expressed as % changes over D28 in the sera of immunized genO-panhCD3 mice (n=9 per group) treated at D28 and D35 (black arrows) with vehicle or 1 mg/kg BiAATE. *p-value<0.05 vs BiAATE at the corresponding time; °°p-value<0.01, and °°°p-value<0.001 vs BiAATE at D28; and #p-value<0.05 vs BiAATE at D42. Adapted from Figure 3 of Perico et al., 2024⁴.

‍

While the use of TCEs has been assessed for the treatment of autoimmune diseases^6–8, this study provides the first proof-of-concept that TCEs can be used to selectively deplete autoreactive B cells, a finding that was confirmed in vivo using a humanized CD3 mouse model (genO-panhCD3). The use of BiAATEs provides additional specificity that has often not been achieved with B-cell targeted therapies, as it targets B cells producing a specific type of autoantibody, as well as plasma cells. By limiting unwanted side effects, BiAATEs have the potential to serve as a promising treatment for autoimmune diseases. 

As shown in this study, the genO-panhCD3 mouse model made available by genOway enables the study of T-cell engagers and different types of immunotherapies (see posters), but it is also a useful model for the testing and validation of different compounds/biologics and safety studies.

‍

The genO-panhCD3 mouse model used in this publication is available off the shelf at genOway, designer and provider of multiple preclinical models in several research areas, including immuno-oncology, metabolism, cardiovascular diseases and neuroscience.

‍

References

1. Pisetsky, D. S. Pathogenesis of autoimmune disease. Nat. Rev. Nephrol. 19, 509–524 (2023).
2. Beck, L. H. et al. M-Type Phospholipase A ₂ Receptor as Target Antigen in Idiopathic Membranous Nephropathy. N. Engl. J. Med. 361, 11–21 (2009).
3. Remuzzi, G. et al. Rituximab for idiopathic membranous nephropathy. The Lancet 360, 923–924 (2002).
4. Perico, L. et al. Bi-specific autoantigen-T cell engagers as targeted immunotherapy for autoreactive B cell depletion in autoimmune diseases. Front. Immunol. 15, 1335998 (2024).
5. Löffler, A. et al. Efficient elimination of chronic lymphocytic leukaemia B cells by autologous T cells with a bispecific anti-CD19/anti-CD3 single-chain antibody construct. Leukemia 17, 900–909 (2003).
6. Remuzzi, G. et al. Rituximab for idiopathic membranous nephropathy. doi:https://doi.org/10.1016/S0140-6736(02)11042-7.
7. Bucci, L. et al. Bispecific T cell engager therapy for refractory rheumatoid arthritis. Nat. Med. 30, 1593–1601 (2024).
8. Subklewe, M. et al. Application of blinatumomab, a bispecific anti-CD3/CD19 T-cell engager, in treating severe systemic sclerosis: A case study. Eur. J. Cancer 204, 114071 (2024).