Nevertheless, the key finding from these experiments is that LRBA-deficient mice mount a remarkably normal immune response to a chronic, systemic viral infection. Open in a separate window Figure 5 Response of LRBA-deficient mice to chronic systemic viral infection. LRBA-deficient mice, pointing to compensatory mechanisms for controlling CD86 in the face of low CTLA-4. These results add to the experimental rationale for treating LRBA deficiency with the CTLA4-Ig fusion protein, Abatacept, and pose questions about the limitations of laboratory experiments in mice to reproduce human disease mutations were discovered in 2012 as the cause of a new human immunodeficiency disorder characterised by recurrent infections and defects in B-lymphocyte activation, low numbers of isotype-switched memory B cells and diminished IgG and IgA antibody formation4 and by chronic diarrhoea.5 Subsequently, homozygous or compound CDC42 heterozygous mutations without clinical disease, although this may PI3k-delta inhibitor 1 relate to difficulty in detecting asymptomatic individuals.6, 12 The pathogenesis of immunodeficiency and autoimmunity caused by LRBA deficiency is not understood. Laboratory findings from children with LRBA deficiency are also variable in presentation and raise many questions about pathogenesis.6, 18 Hypogammaglobulinemia is found in 57C58% of patients.6, 18 Total B-lymphocyte counts are often normal or sometimes reduced, but isotype-switched memory B cells are decreased in >80% of patients6, 18 and plasmablasts are reduced in 92% of patients.18 Natural killer (NK) cells are normal or decreased in LRBA deficient patients.6, 18 Counts of CD4+ and CD8+ T cells are generally normal; however, individual patients have presented with either increases or decreases in their numbers,6, 18 and the percentage of CD45RO+ RA?-activated/memory T cells and CXCR5+ PD-1+ follicular helper T cells is increased.8 FOXP3+ CD4+ T-regulatory (Treg) cells are decreased as a percentage of CD4+ cells in the majority of LRBA-deficient patients6, 8, 18 and the Tregs that are present have decreased levels per cell of FOXP3, HELIOS, CD25 and CTLA-4.8, 11 These pleiotropic lymphocyte abnormalities, together with the broad expression of mRNA across leucocyte subsets and other tissues, make it unclear if LRBA deficiency causes intrinsic deficits in B-cell isotype switching and memory formation,4 a primary, generalised deficit in FOXP3 Treg cells,8 or a problem in nonlymphoid organs such as the gut. An important insight into the pathogenesis of LRBA-deficiency syndrome came from the finding in 2015 that the immune dysregulation responds exceptionally well to treatment with soluble CTLA4-Ig fusion protein, Abatacept.11 Experimental analysis of cells in culture revealed that CTLA-4 and LRBA interact through specific sequences in the CTLA-4 cytoplasmic tail, colocalise at recycling endosomes and the trans-Golgi network, and that LRBA protects CTLA-4 from being sorted to and degraded in lysosomes.11 Hence, an attractive hypothesis is that low CTLA-4 expression on activated T cells or FOXP3+ Treg cells is responsible for some or all of the immune dysregulation in LRBA deficiency. CTLA-4 on T cells removes CD86 from antigen-presenting cells,19 and exaggerated expression of CD86 on anergic self-reactive B cells switches the outcome of their interaction with T cells from FAS-mediated deletion to plasma cell differentiation and autoantibody secretion,20 providing a plausible mechanism for the pathogenesis of autoimmune haemolytic anaemia and thrombocytopenia and its correction with Abatacept therapy. However, it is unclear how this mechanism would explain the humoral immunodeficiency and low numbers of switched memory B cells, which appear less responsive to Abatacept.11 To resolve the many questions summarised above, we generated and analysed an LRBA-deficient mouse strain. The results reveal no evidence for an intrinsic requirement for LRBA in B-cell activation, germinal centre (GC) formation, isotype switching and affinity maturation. LRBA deficiency greatly decreased CTLA-4 on activated CD4+ T cells and FOXP3+ Tregs in a cell-autonomous manner, but other Treg markers and Treg frequency were unaffected in young mice. We conclude that partial CTLA-4 deficiency is a primary component of the immune dysregulation that occurs in LRBA deficiency, but is compensated to prevent progression to autoimmunity and immunodeficiency under standard mouse housing conditions. PI3k-delta inhibitor 1 Results CTLA-4 deficiency in T cells of LRBA-deficient mice LRBA-deficient mice were generated on the C57BL/6 background using CRISPR/Cas9-mediated gene targeting to produce an 8?bp deletion in exon 37 of and WT mice. This revealed the serum of unimmunised mice to contain significantly higher levels of IgG2b than age- and sex-matched WT mice (Figure 2g). There was also a trend towards an increase in IgM levels, although this was not significant (and WT bone marrow distinguished PI3k-delta inhibitor 1 by CD45.2 and CD45.1.