Dominant NFKB1 Mutations Cause Antibody Deficiency and Autoinflammatory Episodes

The NFkB signaling pathway is a master regulator of the immune response. Recently, dominant mutations in NFKB2 have shown to cause antibody deficiency with adrenal insufficiency.

We investigated two families with antibody deficiency and autoinflammatory features, with onset in teenage years and early adulthood (Fig. 1 and Table 1).  In family 1, all patients available to study presented with respiratory tract infections and B cell dysfunction marked by hypogammaglobulinemia, poor antibody response to vaccines, or low switched memory B cell counts. Several experienced recurrent episodes of aphthous mucositis in upper gastrointestinal and genital area, sometimes accompanied by abdominal pain, monoarthritis, or fever with elevated inflammatory markers (peripheral blood leukocytes >10x10⁶ cells/ml, C reactive protein >100 mg/L). A lesional biopsy revealed lymphocytic small cell vasculitis reminiscent of Behcet’s disease. In family 2, all patients presented with respiratory tract infections, hypogammaglobulinemia, and poor vaccine responses.

Based on the pedigrees, we assumed autosomal dominant inheritance. For family 1, we performed genotyping and linkage analysis coupled with whole genome sequencing to pinpoint the causative variant. Linkage analysis showed a 15.4 Mb haplotype on chr4 segregating with the phenotype in all affected individuals. When focusing our search on novel, heterozygous variants negatively affecting conserved residues, we identified only one variant in this genomic location, localizing to NFKB1 gene (p.H67R).

In family 2, we exome sequenced one affected individual and filtered the data for novel heterozygous variants negatively affecting conserved residues. This resulted in 17 variants, out of which the I553M NFKB1 variant was considered the best candidate.

Several functional analyses showed that the mutations had a negative impact on NFKB1 function. The H67R variant increased NFKB1 affinity to NEMO and delayed protein entry to nucleus, when assessed by affinity purification mass spectrometry and immunofluorescence microscopy. The mutation also decreased NFKB1 transcriptional activity, when expressed together with an NFkB-responsive luciferase reporter, and showed a neomorphic DNA binding pattern on chromatin immunoprecipitation-sequencing. Furthermore, the cell growth was hampered when the mutant NFKB1 was overexpressed in cultured HEK293 cells. The I553M showed a similar negative effect on cultured cell proliferation. Furthermore, the I553M variant negatively affected serine S907 and S893 phosphorylation, leading to defective posttranslational processing of the full-length NFKB1 upon TNF stimulation.

To conclude, we show that hypogammaglobulinemia and Behcet’s disease-like autoinflammatory syndrome can be caused by mutations in NFKB1. This suggests that a subset of Behcet’s disease cases may be of monogenic origin, and highlights the role of NFkB complex in B cell maturation and innate immune regulation.

Figure 1. Family pedigrees.

 

Table 1. Patient characteristics

Patient

1

2

3

4

5

6

7

8

9

10

11

12

Age

55

43#

25

29

40

30

37

7

10

60#

61

32

Sex

F

M

F

F

F

F

M

F

M

M

F

M

Mutation status

H67R

ND

H67R

H67R

H67R

H67R

H67R

H67R

H67R

ND

I553M

I553M

Immunodeficiency

Infections

URTI

RTI

URTI

-

URTI

URTI

URTI

URTI

-

RTI

RTI

RTI

Antibody deficiency

Hypogammaglobulinemia, SAD

ND

Hypogammaglobulinemia, SAD

-

IgG subclass deficiency, SAD

Hypogammaglobulinemia, SAD

Hypogammaglobulinemia, SAD

Hypogammaglobulinemia

Hypogammaglobulinemia

Hypogammaglobulinemia

Hypogammaglobulinemia, SAD

SAD

Immunoglobulin replacement

+

-

+

-

-

+

+

-

-

-

+

-

Immune dysregulation

Febrile attacks

-

+

+

+

-

-

+

+

-

-

-

-

Complex aphtae

Mouth, genitalia

ND

Mouth, genitalia

Mouth, genitalia

-

Mouth

Esophagus

Mouth, esophagus

-

-

-

-

Arthritis

Monoarthritis

-

Monoarthritis

Monoarthritis

-

-

-

-

-

-

Oligoarthritis

-

Gut disease

Periodic abdominal pain, chronic idiopathic diarrhea

ND

Periodic abdominal pain, microscopic colitis

Periodic abdominal pain

-

-

-

Periodic abdominal pain

-

ND

Chronic idiopathic diarrhea

Coeliac disease

Other

-

Hyperinflammatory response and death after gallbladder surgery

-

Iritis, Hyperinflammatory response to tooth excision

Kidney tumor

-

Rudimentary left kidney

-

-

-

Astma, hypothyreosis

-

URTI, upper respiratory tract infections; RTI, respiratory tract infections including recurrent pneumonia; SAD, Specific antibody deficiency; ND, No data; ^#Death age