The right “Job” for STAT3 mutant mice!

In this issue of Blood, Steward-Tharp et al report the generation of a murine model of the human primary immunodeficiency autosomal dominant hyper-immunoglobulin E syndrome (AD-HIES) and reveal novel insights and therapeutic outcomes for this fascinating human monogenic disorder.¹ 

“So Satan went forth from the presence of the Lord, and smote Job with sore boils from the sole of his foot unto his crown.” This quote from the book of Job prefaced the subject of a clinical report by Davis et al that, in 1966, provided the first description of a new primary immunodeficiency. It was named Job's syndrome after the biblical figure Job who was covered in boils, not unlike the patients detailed in that report.²  Job's syndrome is characterized by recurrent staphylococcal infections of the lung and skin, as well as chronic mucocutaneous candidiasis.^2-5  Strikingly, viral susceptibility is not a common clinical complication of Job's syndrome patients, although recent reports suggest problems controlling reactivation of some herpes viruses.⁶  Following the discovery of immunoglobulin E (IgE) in the early 1970s, Buckley noted that these patients had extremely high levels of IgE,³  which led to renaming Job's syndrome as autosomal dominant hyper-immunoglobulin E syndrome (AD-HIES). Buckley also reported defective humoral immune responses in patients with Job's syndrome, despite normal levels of serum IgM, IgG, and IgA.³  Additional features of AD-HIES include eosinophilia, susceptibility to B-cell lymphoma, and nonimmunologic defects affecting musculoskeletal, connective tissue, dental, and circulatory systems.^4,5 

Despite increased understanding and awareness of the clinical features of AD-HIES over the ensuing decades, it was not until 2007—41 years after the initial description² —that the genetic defect underlying this condition was revealed. Elegant studies by Minegishi et al⁷  and Holland et al⁸  discovered that heterozygous mutations in STAT3 caused AD-HIES. Subsequent studies of lymphocytes from patients with AD-HIES identified critical roles for STAT3 in generating Th17 cells, Tfh cells, and memory B and CD8⁺ T cells and the ability of naïve B cells to differentiate into plasmablasts in response to STAT3-activating cytokines.⁵  These findings provided a cellular basis for several key clinical features of AD-HIES, namely susceptibility to recurrent fungal and possibly staphylococcal infections (Th17 deficit), impaired humoral immunity (Tfh and B-cell defects), and susceptibility to B-cell lymphoma and latent herpes viruses (reduced memory CD8⁺ T cells).^5,6  However, studies in humans are limited by ethical and logistical constraints; thus, an animal model of STAT3 deficiency would be a valuable resource for further exploring molecular and cellular mechanisms underlying disease pathogenesis in AD-HIES.

Although Stat3-gene targeted mice were generated in 1997, their utility as an experimental model was curtailed by embryonic lethality following constitutive Stat3 deletion.⁹  This finding had at least two important outcomes. First, Stat3-null mice provided little indication that STAT3 mutations could underlie a primary immune-deficient condition in humans. Second, there were substantial differences between Stat3-null mutations in mice and heterozygous mutations in humans, with the former being fatal and the latter, although compatible with life, were deleterious for human health. This reflected residual activity of the wild-type STAT3 allele that permitted placental implantation and embryogenesis but was clearly insufficient for immune cell function. However, the generation of mice lacking Stat3 in specific lineages did reveal defects in Th17 cells, Tfh cells, and antigen-specific antibody (Ab) responses when Stat3 was selectively deleted from CD4⁺ T cells or B cells.⁵  Even with the knowledge that STAT3 mutations caused AD-HIES, Stat3 conditionally deficient mice had limited application as an in vivo model because loss of Stat3 in individual cell lineages failed to recapitulate the full phenotype of disease.

Thus, a new mouse model was required that would be more faithful to the human condition. To achieve this, Steward-Tharp et al¹  generated BAC-transgenic mice expressing a deletion mutation (V463del) in Stat3 that is relatively common in AD-HIES patients.^7,8  Strategically, the authors selected 1 mouse strain expressing 2 copies of the transgene, thus recreating the heterozygous and dominant negative nature of the human STAT3 mutation. Detailed analysis revealed significant similarities between these mice and AD-HIES patients, including normal lymphocyte development but impaired generation of Th17 cells, which resulted in susceptibility to infection with the gut-tropic bacteria Citrobacter rodentium, poor Ab responses and, importantly, hyper-IgE.¹  Although it could be argued that susceptibility to C rodentium is a feature of mice rather than the human condition, it is consistent with poor immunity in AD-HIES to infections at mucocutaneous sites which, like protection against C rodentium, require interleukin-17 (IL-17) and IL-22 production by CD4⁺ T cells and IL-22–induced STAT3-dependent expression of antimicrobial proteins in epithelial cells. Stat3-mutant mice also developed severe C rodentium–induced inflammatory bowel disease and significant mortality following lipopolysaccharide challenge¹ ; both of these were attributed to increased production of proinflammatory cytokines (tumor necrosis factor α [TNF-α], IL-12, interferon gamma [IFN-γ]) and reduced IL-10, which signals through STAT3. A curious feature of AD-HIES is a marked lack of inflammation,^2,4  which may reflect a predominant role for IL-6/STAT3 signaling in human inflammation that is not phenocopied in mice or may be unique to C rodentium infection in rodents.

Although AD-HIES is not a fatal immunodeficiency, it is nonetheless life shortening.⁴  Thus, insights into therapeutic options would be valuable. Although stem cell transplantation (SCT) of AD-HIES patients has been attempted, results have been inconsistent.¹⁰  Given the ubiquitous expression of STAT3 and the nonimmunologic features of AD-HIES, it is perhaps not surprising that SCT has had mixed outcomes. Reconstitution of Stat3-mutant mice with wild-type bone marrow restored Th17 responses, but this provided only partial protection against C rodentium, demonstrating that, although transplanted mice could generate appropriate cytokines for protection against this pathogen, these effector cytokines—probably IL-22—need to exert their function via STAT3 in nonhematopoietic cells (ie, epithelia) to provide complete protection against infection. These findings suggest that SCT of AD-HIES patients may have only a partial impact on infectious susceptibility, with additional prophylactic treatments required. However, SCT will probably cure defects in Ab responses in AD-HIES.

Overall, this is a very exciting model that, unlike germline Stat3 deletion, mimics the genetic lesion in AD-HIES, thereby allowing detailed dissection of cellular defects and disease in affected individuals. And although this model may not tell us what causes retention of primary teeth in AD-HIES patients, it may provide answers to the age-old question of what causes hyper-IgE and may identify mechanisms causing nonimmunologic defects. Furthermore, it could represent a preclinical model to test drugs considered for treating AD-HIES. So, there are plenty of “Job's” in store for these mice!