Genetic and Small Molecule Disruption of the AID/RAD51 Axis Similarly Protects Nonobese Diabetic Mice from Type 1 Diabetes through Expansion of Regulatory B Lymphocytes

Abstract

B-lymphocytes play a key role in type 1 diabetes (T1D) development by serving as a subset of APC preferentially supporting expansion of autoreactive pathogenic T-cells. As a result of their pathogenic importance, B-lymphocyte-targeted therapies have received considerable interest as potential T1D interventions. Unfortunately, B-lymphocyte-directed T1D interventions tested to date failed to halt β-cell demise. IgG autoantibodies marking humans at future T1D risk indicate B-lymphocytes producing them have undergone the affinity maturation processes of class switch recombination (CSR) and possibly somatic hypermutation (SHM). This study found that CRISPR/Cas9-mediated ablation of the Aicda gene required for CSR/SHM induction, inhibits T1D development in the NOD mouse model. The Aicda encoded AID molecule induces genome-wide DNA breaks that, if not repaired through RAD51-mediated homologous recombination (HR), result in B-lymphocyte death. Treatment with the RAD51 inhibitor 4,4′-diisothiocyanatostilbene-2, 2′-disulfonic acid (DIDS) also strongly inhibited T1D development in NOD mice. Both the genetic and small molecule-targeting approaches expanded CD73+ B-lymphocytes exerting regulatory activity suppressing diabetogenic T-cell responses. Hence, an initial CRISPR/Cas9 mediated genetic modification approach has identified the AID/RAD51 axis as a target for a potentially clinically translatable pharmacological approach that can block T1D development by converting B-lymphocytes to a disease inhibitory CD73+ regulatory state.

Activation-induced cytidine deaminase deficiency accelerates autoimmune diabetes in NOD mice

“…Furthermore, AID deficiency may also enhance the escape of these pathogenic B cell clones from GCs of peripheral lymphoid tissues by evading the selection pressures and circulating to target the systemic host organs (39). AID (23), in which the results obtained could be due to the different approach used for genetic targeting and a different environment. Foxp3 + Tregs were also not affected here by AID deficiency in our model system.…”

A hypermorphic Nfkbid allele represents an Idd7 locus gene contributing to impaired thymic deletion of autoreactive diabetogenic CD8+ T-cells in NOD mice

“…6G). We recently found that partially overlapping populations of CD73 expressing and IL-10 producing Bregs can inhibit autoimmune T1D development in NOD mice (44). Ablation of Nfkbid also reduced levels of such IL-10 producing and CD73 expressing Bregs in NOD mice (Fig.…”

Transient BAFF Blockade Inhibits Type 1 Diabetes Development in Nonobese Diabetic Mice by Enriching Immunoregulatory B Lymphocytes Sensitive to Deletion by Anti-CD20 Cotherapy

“…The administration of an IL-10 neutralizing antibody (JES5-2A5) in NOD mice following transient BAFFR-Fc treatment abrogated the T1D protective effects of this reagent (Figure 4H). We recently identified a subset of CD73 + B-lymphocytes capable of suppressing the activity of NOD diabetogenic T-cells (29). Thus, we characterized the extent B-lymphocytes expressed CD73 following the various depletion regimens.…”

“…T1D resistance elicited in NOD mice by transient BAFFR-Fc treatment also coincided with an initial expansion of a CD73 + B-lymphocytes. We recently found B-lymphocytes in NOD- Aicda null mice convert to a CD73 + phenotype capable of suppressing diabetogenic T cell responses through production of immunosuppressive adenosine (29). However, following cessation of treatment, proportions of CD73 + B-lymphocytes do not remain expanded in NOD mice in which long term T1D resistance was established by transient BAFFR-Fc mono-therapy.…”

“…However, the capacity of B-lymphocyte depletion to modulate levels of B 10 cells in the context of T1D is currently unknown. We recently found a sizeable proportion of B-lymphocytes in NOD- Aicda null mice which are unable to undergo class switch recombination (CSR) and somatic hyper-mutation (SHM) not only fail to support T1D development but also convert to a CD73 + phenotype capable of actively suppressing pathogenic T-cells (29). CD39 and CD73 are ecto-enzymes that respectively convert pro-inflammatory ATP to AMP and then to anti-inflammatory adenosine (30).…”