FOXP3 TSDR Measurement Could Assist Variant Classification and Diagnosis of IPEX Syndrome

Abstract

Pathogenic FOXP3 variants cause immune dysregulation polyendocrinopathy enteropathy X-linked (IPEX) syndrome, a progressive autoimmune disease resulting from disruption of the regulatory T cell (Treg) compartment. Assigning pathogenicity to novel variants in FOXP3 is challenging due to the heterogeneous phenotype and variable immunological abnormalities. The number of cells with demethylation at the Treg cell-specific demethylated region (TSDR) is an independent biomarker of IPEX. We aimed to investigate if diagnosing IPEX at presentation with isolated diabetes could allow for effective monitoring of disease progression and assess whether TSDR analysis can aid FOXP3 variant classification and predict disease course. We describe a large genetically diagnosed IPEX cohort (n = 65) and 13 individuals with other monogenic autoimmunity subtypes in whom we quantified the proportion of cells with FOXP3 TSDR demethylation, normalized to the number with CD4 demethylation (%TSDR/CD4) and compare them to 29 unaffected controls. IPEX patients presenting with isolated diabetes (50/65, 77%) often later developed enteropathy (20/50, 40%) with a median interval of 23.5 weeks. %TSDR/CD4 was a good discriminator of IPEX vs. unaffected controls (ROC-AUC 0.81, median 13.6% vs. 8.5%, p < 0.0001) with higher levels of demethylation associated with more severe disease. Patients with other monogenic autoimmunity had a similar %TSDR/CD4 to controls (median 8.7%, p = 1.0). Identifying increased %TSDR/CD4 in patients with novel FOXP3 mutations presenting with isolated diabetes facilitates diagnosis and could offer an opportunity to monitor patients and begin immune modulatory treatment before onset of severe enteropathy.

Epigenetic immune monitoring for COVID-19 disease course prognosis

“…Furthermore, given the feasibility to perform EICC from nasopharyngeal swabs and DBS, this approach lends itself to unsupervised home testing and possibly facilitate medical surveillance. The concept shown here may be transferred to other clinical applications, such as newborn screening for primary immunodeficiencies using DBS ( 31 ) or determination of immune cell infiltration in solid tumor tissues ( 32 ), where flow cytometry in particular is not suitable or reaches its limits, as well as to situations where patients suffer from chronic inflammatory pathologies like rheumatoid arthritis or systemic lupus erythematosus ( 33 , 34 ).…”

FOXP3 deficiency, from the mechanisms of the disease to curative strategies

“…We and others have consistently found an increased frequency of cells containing the epigenetic marker of Treg cells in IPEX patients 22,43,44 . The prototypical Treg epigenetic marker is the demethylation of the Treg‐specific demethylated region (TSDR), located within a noncoding region of the FOXP3 gene 45 .…”

“…The increased frequency of TSDR demethylated cells in IPEX patients is even more prominent when normalized to CD3+ or CD4+ T cell numbers 22,43,44 . This phenotype is not present at birth but may precede the first clinical manifestations, suggesting that the increase in TSDR demethylated cells likely reflects a dynamic process of Treg cell expansion in the periphery rather than an increased thymic output of Treg cells 22 .…”

“…The increased frequency of TSDR demethylated cells in IPEX patients is even more prominent when normalized to CD3+ or CD4+ T cell numbers. 22,43,44 This phenotype is not present at birth but may precede the first clinical manifestations, suggesting that the increase in TSDR demethylated cells likely reflects a dynamic process of Treg cell expansion in the periphery rather than an increased thymic output of Treg cells. 22 Collectively, the data suggest that Treg cells are expanded in IPEX, the expansion is associated with autoimmunity, and that TSDR demethylation analyses could assist and/or complement genetic diagnoses, particularly in patients carrying a new mutation of unknown significance.…”

“…The exact mechanisms that lead to the competitive advantage of FOXP3 wild‐type Treg cells remain unclear. Given that healthy donors' FOXP3 knock‐out Treg cells expand better in vitro compared to controls 27 and IPEX patients have expanded Treg cells, 22,43,44 the competitive advantage of wild‐type Treg cells in vivo is somehow surprising. Several mechanisms may play a role in this phenomenon.…”