Immunodeficiency and bone marrow failure with mosaic and germline TLR8 gain of function

Abstract

Inborn errors of immunity (IEI) are a genetically heterogeneous group of disorders with a broad clinical spectrum. Identification of molecular and functional bases of these disorders is important for diagnosis, treatment and an understanding of the human immune response. We identified six unrelated males with neutropenia, infections, lymphoproliferation, humoral immune defects, and in some cases bone marrow failure associated with three different variants in the X-linked gene TLR8, encoding the endosomal Toll-like receptor 8 (TLR8). Interestingly, five patients had somatic variants in TLR8 with less than 30% mosaicism, suggesting a dominant mechanism responsible for the clinical phenotype. Mosaicism was also detected in skin-derived fibroblasts in three patients, demonstrating that mutations were not limited to the hematopoietic compartment. All patients had refractory chronic neutropenia, and three patients underwent allogeneic hematopoietic cell transplantation. All variants conferred gain-of-function to TLR8 protein, and immune phenotyping demonstrated a pro-inflammatory phenotype with activated T cells and elevated serum cytokines associated with impaired B cell maturation. Differentiation of myeloid cells from patient-derived induce pluripotent stem cells demonstrated increased responsiveness to TLR8. Together these findings demonstrate that gain-of-function variants in TLR8 lead to a novel childhood-onset IEI with lymphoproliferation, neutropenia, infectious susceptibility, B and T cell defects, and in some cases bone marrow failure. Somatic mosaicism is a prominent molecular mechanism of this new disease.

Genetic Mosaicism as a Cause of Inborn Errors of Immunity

“…Since mosaicism of <30% was sufficient to cause the disease, we anticipate that full donor chimerism may be important for HCT. This was evidenced by one patient who had recurrence of neutropenia with low donor chimerism, which resolved when full donor engraftment was once again achieved following treatment with donor lymphocytes [41]. TLR8 has not been previously associated with IEI, and our discovery again suggests that mosaicism should be considered in the "firstline" approach of genetic analysis.…”

“…Cultured skin fibroblasts derived from patients with mosaic TLR8 variants were reprogrammed into induced pluripotent stem cells (iPSCs), which were single cell cloned to generate clonal iPSC lines with wild type (WT) or TLR8 variant. These clones were differentiated into neutrophils or macrophages and tested for their response to high or low doses of TLR8 stimulation [41]. There was no difference in response to stimulation with a high-dose of TLR8 ligand.…”

“…We recently described somatic mutations in the TLR8 gene as a primary mechanism of monogenic IEI [41]. We identified six unrelated boys with neutropenia, B cell defects, and lymphoproliferation, all with novel genetic variants in TLR8, the gene encoding toll-like receptor 8 (TLR8), an endosomal TLR that recognizes single-stranded RNA.…”

Genetic screening of children with marrow failure. The role of primary Immunodeficiencies

“…In fact, although it is well known that impairment of the immune system represents the major pathogenic mechanism of acquired SAA, 31 we firstly showed that a considerable number of these patients, usually considered to have an idiopathic disease, are actually affected with monogenic disorders of immunity which are known to have heterogeneous phenotypes and unpredictable presentations in children. An immune-mediated attack of marrow precursors generating MF has been sporadically reported in the setting of PIDs 17,18 such as CTLA4 deficiency 32 or, more recently, in a cohort of patients carrying mutations on the TLR8 gene 33 which have been shown to present MF in addition to immune-deficiency features.…”

Toll-Like Receptor Signaling in the Establishment and Function of the Immune System

“…Recently, we described germline and somatic variants in TLR8 as an underlying monogenic cause of IEI in patients with recurrent infections, neutropenia, lymphoproliferation, hypo-gammaglobulinemia, and bone marrow failure [105]. In our cohort of six unrelated male patients, five patients harbored somatic mosaic variants in TLR8, with four patients having the same mosaic variant (P432L) and the fifth patient with a different mosaic variant (F494L).…”