Background
The recent SARS‐CoV‐2 pandemic, which has recently affected Italy since February 21, constitutes a threat to normal subjects, as the coronavirus disease‐19 (COVID‐19) can manifest with a ...broad spectrum of clinical phenotypes ranging from asymptomatic cases to pneumonia or even death. There is evidence that older age and several comorbidities can affect the risk to develop severe pneumonia and possibly the need of mechanic ventilation in subjects infected with SARS‐CoV‐2. Therefore, we evaluated the outcome of SARS‐CoV‐2 infection in patients with inborn errors of immunity (IEI) such as X‐linked agammaglobulinemia (XLA).
Methods
When the SARS‐CoV‐2 epidemic has reached Italy, we have activated a surveillance protocol of patients with IEI, to perform SARS‐CoV‐2 search by nasopharyngeal swab in patients presenting with symptoms that could be a manifestation of COVID‐19, such as fever, cough, diarrhea, or vomiting.
Results
We describe two patients with X‐linked agammaglobulinemia (XLA) aged 34 and 26 years with complete absence of B cells from peripheral blood who developed COVID‐19, as diagnosed by SARS‐CoV‐2 detection by nasopharyngeal swab, while receiving immunoglobulin infusions. Both patients developed interstitial pneumonia characterized by fever, cough, and anorexia and associated with elevation of CRP and ferritin, but have never required oxygen ventilation or intensive care.
Conclusion
Our report suggests that XLA patients might present with high risk to develop pneumonia after SARS‐CoV‐2 infection, but can recover from infection, suggesting that B‐cell response might be important, but is not strictly required to overcome the disease. However, there is a need for larger observational studies to extend these conclusions to other patients with similar genetic immune defects.
The underlying genetic mechanisms have been elucidated in the last few years in less than 10% to 15% of the cases and involve mutations in CD19, MS4A1 (CD20), CR2 (CD21), ICOS, TNFRSF13C, TNFRSF13B, ...PLCG2 (phospholipase Cg2), CD81, LRBA, and PRKCD (protein kinase Cd).1-3 Recently, germline heterozygous mutations in NFKB2 were identified in 10 patients to be associated with early-onset CVID with autoimmunity in most cases,4,5 profound B-cell deficiency,6 or a CVID-like phenotype.7 All affected patients had hypogammaglobulinemia with variable association of the following clinical and immunologic features: central adrenal insufficiency (ACTH insufficiency), alopecia totalis or areata, trachyonychia, variable natural killer (NK) cell numbers, and defects in peripheral T and B cells. Appendix Lymphocyte subsets Index patient (%) Normal range for age (%) Proliferation (cpm) Index patient Healthy control T cells (CD3+) 89.8 60.5-79.8 CD3+CD4+ 63.8 30.3-48.3 Naive (CD45RA+CCR7+) 85.8 34.3-74.6 RTE (CD45RA+CCR7+CD31+) 66.5 21.1-63.5 Central memory (CD45RA-CCR7+) 8.5 13.0-43.5 Effector memory (CD45RA-CCR7-) 4.4 8.5-28.1 Terminally differentiated (CD45RA+CCR7-) 1.2 0.7-6.6 CD3+CD8+ 22.3 13.8-37.5 Naive (CD45RA+CCR7+) 92.0 26.7-72.9 Central memory (CD45RA-CCR7+) 1.8 1.2-11.6 Effector memory (CD45RA-CCR7-) 4.0 6.0-53.6 Terminally differentiated (CD45RA+CCR7-) 2.3 3.9-72.0 TCR γ/δ 1.7 0.5-21.5 B cells (CD19+) 4.8 5.7-19.7 RBE (CD38hiCD21dim/loCD27-) 31.4 15.0-35.3 Naive (CD38dim/loCD21hiCD27-) 60.5 33.8-79.6 CD19hiCD21lo 1.9 1.1-10 Switched memory (IgD-CD27+) 0.4 2.7-20.6 IgM memory (IgD+CD27+) 2.0 3.5-24.1 Terminally differentiated (CD38hiCD27hiCD21lo) 0.17 0.16-8.70 Plasma cells (CD38hiCD20-CD138+) 0.00 0.04-3.20 NK cells (CD3-CD16+CD56+) 4.5 4.6-27.8 CD3 227,000 77,000 CD3 + IL2 200,000 152,000 PHA 116,000 120,000 PMA + Ionomycin 418,000 285,000 Background 4,000 7,000 Table I Immunologic profile from the index patient mutated in NFKB2 PMA, Phorbol 12-myristate 13-acetate; RBE, recent bone marrow emigrants; RTE, recent thymic emigrants; TCR, T-cell receptor.
RAC2D57N also disrupts the polymerization of filamentous actin, thereby reducing myeloid cell and lymphocyte chemotaxis and adhesion.2,3 Although conditional deletion of either RAC1 or RAC2 in ...lymphocytes is sufficient for intact T-cell and B-cell development in mice, reduced numbers of CD4+ T cells and B cells have been found in one patient with RAC2D57N and in another kindred lacking RAC2 protein expression due to a homozygous RAC2W56X mutation.4,5 Somatic activating variants in RAC2 have been associated with melanoma,1 but there are no published reports of germline gain-of-function mutations in any RAC family members. The mutation was confirmed by Sanger sequencing (Fig 1, B) and had a deleterious Combined Annotation Dependent Depletion Phred-like score of 32.7 The mutated P34 residue resides within the highly conserved Switch 1 domain important for interactions with guanine exchange factors and downstream effectors (Fig 1, C).1 Immunoblotting revealed similar levels of RAC2 in patients and controls (Fig E1, B). Because the patients' primary cells express both wild-type (WT) and RAC2P34H, we expressed Flag-tagged WT or mutant RAC2 in HEK293T cells and confirmed that WT and mutant forms of RAC2 had comparable expression (Fig 1, D). Filamentous actin accumulation is known to reduce mitochondrial membrane potential, increase reactive oxidant species, and trigger proapoptotic pathways, all of which increased susceptibility to apoptosis.8 In addition, the conserved Switch 1 domain in the Rac guanosine triphosphatases binds to the p100β catalytic subunit and activates phosphoinositide-3-kinase (PI3K).9 PI3K overactivation, due to germline gain-of-function mutations in PIK3CD or PIK3R1, results in a combined immunodeficiency with certain features shared by our patients, including reduced percentages of naive T cells, the accumulation of senescent CD8+CD57+ lymphocytes, and increased T- and B-cell apoptosis.10 The lymphopenia in patients with PI3K overactivation has been attributed to TCR-mediated apoptotic signaling as well as metabolic shifts that favor the expansion of memory and senescent T cells over naive cells.10 However, because patients with activated phosphoinositide 3-kinase δ syndrome have additional features, including increased IgM levels, lymphoproliferative disease, and autoimmunity, additional patients and studies are needed to investigate the PI3K-AKT axis in patients with gain-of-function mutations in RAC2. Variable Patient 1 Patient 2 Patient 3 Age at the time of testing (y) 31 7 2 Hemogram (normal range) Hemoglobin (g/dL) 15.6 (10.9-15) 12.6 (10.5-13.8) 11.8 (11-12.8) WBCs (103 cells/μL) 3.670 (5.8-9.3) 1.640 (5.4-9.7) 2.960 (5.9-10.4) Neutrophils (103 cells/μL) 2.822 (3.3-6.3) 0.730 (2.5-5.9) 1.557 (2.5-6) Lymphocytes (103 cells/μL) 0.499 (1.14-2.28) 0.350 (1.28-2.76) 0.400 (1.33-4.77) Monocytes (103 cells/μL) 0.184 (0.29-0.7) 0.172 (0.19-0.81) 0.266 (0.19-0.94) Platelets (103 cells/μL) 153 (174-333) 214 (187-367) 216 (208-413) Lymphocyte subsets CD3+ (103 cells/μL) 359 (1000-2600) 248 (1400-3700) 252 (2100-6200) CD3+CD4+ (103 cells/μL) 72 (530-1500) 140 (700-2200) 98 (1300-3400) CD45RA+CD31+ recent thymic emigrants, % CD4+ 0.0 (9.8-43) 32 (45-63) 29 (57-76) CD45RA+CCR7+CD31– naive, % CD4+ 0.7 (21-61) 36.4 (57.1-84.8) 32.7 (65-84) CD45RA–CCR7– effector memory, % CD4+ 95.8 (7.6-25) 44.1 (3.3-15.2) 35.7 (2.9-9.8) CD45RA–CCR7+ central memory, % CD4+ 3.3 (26-62) 16.9 (11.2-26.7) 29.8 (10-22.3) CD3+CD8+ (103 cells/μL) 224 (330-1100) 55 (490-1300) 65 (620-2000) CD45RA+CCR7+CD31– naive, % CD8+ 0.4 (11-66) 4.6 (28-80) 14.7 (39-89) CD45RA–CCR7– effector memory, % CD8+ 60.9 (16-54) 56.2 (6.2-29.3) 36.4 (3.4-28) CD45RA–CCR7+ central memory, % CD8+ 0.8 (3.7-32) 1.2 (1.2-4.5) 6.3 (1-5.7) CD45RA+CCR7− TEMRA, % CD8+ 38.1 (5.6-43) 38 (9.1-49.1) 42.7 (4.8-30) CD57+, % CD8+∗ 67.9 (2-38) 50 (2-20) 42.8 (2-16) CD19+ (103 cells/μL) 12 (110-570) 40 (390-1400) 19 (720-2600) CD27−IgD+IgM+ naive, % CD19+ 60.8 (48-97) 30.2 (47-77) 25.0 (54-88) CD27+IgD+ unswitched memory, % CD19+ 27.4 (7-23) 8.8 (5.2-20.4) 11.9 (2.7-19) CD27+IgD– switched memory, % CD19+ 9.4 (8.8-27) 21.7 (10.9-30.4) 19.6 (3.3-7.4) CD38hiIgMhitransitional, % CD19+ 0.8 (2.2-13) 35.6 (7.2-23) 37.8 (10-30) CD3–CD56+ (103 cells/μL) 101 (70-480) 56 (130-0.720) 116 (160-920) Immunoglobulins IgG (mg/dL) 804† (639-1349) 353 (463-1236) 239 (345-1213) IgM (mg/dL) 83 (56-352) 14 (43-196) 12 (14-106) IgA (mg/dL) 226 (70-312) 17 (25-154) 13 (43-173) Tetanus vaccine titer n.d. before IVIG started UD UD Hepatitis vaccine titer n.d. before IVIG started UD UD Proliferation‡ Anti-CD3 (% CD4+ divided) 65.3 (84.2-93.8) 89.05 (84.2-93.8) 24.5 (84.2-93.8) Phytohemmaglutinin (% CD4+ divided) 6.04 (62.9-85) 60 (62.9-85) 63.85 (62.9-85) Anti-CD3 (% CD8+ divided) 54.5 (87-92.7) 64.45 (87-92.7) 34.3 (87-92.7) Phytohemmaglutinin (% CD8+ divided) 3.81 (83.1-93.5) 21.6 (83.1-93.5) 20.75 (83.1-93.5) Table I Immune phenotyping of the patients' lymphocytes Chromosome Position Reference Patient Consequence Gene name CADD score 1 3328670 A C Non-synonymous PRDM16 19.7 1 22310701 C A Non-synonymous CELA3B 19.63 1 27023871 G T Non-synonymous ARID1A 21.1 1 27679943 C T Non-synonymous SYTL1 34 1 27695833 C T Non-synonymous FCN3 23.3 1 27695833 C T Upstream MAP3K6 23.3 2 220420994 G A Non-synonymous OBSL1 32 3 100373932 G A Non-synonymous GPR128 18.75 5 40681232 G T Non-synonymous PTGER4 33 5 79354543 G A Non-synonymous THBS4 23.9 5 102887994 A G Non-synonymous NUDT12 24.2 5 112884695 G A Non-synonymous YTHDC2 22.9 5 137713429 C A Non-synonymous KDM3B 17.27 5 147796767 G A Non-synonymous FBXO38 27.8 5 150227995 T A Non-synonymous IRGM 22.7 9 1052014 G T Non-synonymous DMRT2 33 9 121976315 T A Non-synonymous BRINP1 16.83 9 130826079 C G Upstream SLC25A25 22.1 9 130826079 C G Non-synonymous NAIF1 22.1 9 140057403 G C Non-synonymous GRIN1 24 10 14870199 A G Non-synonymous CDNF 24.3 10 71966072 G A Non-synonymous PPA1 22.9 12 10962022 C T Non-synonymous TAS2R9 15.52 12 10962022 C T Upstream TAS2R8 15.52 12 29786258 T C Non-synonymous TMTC1 29.3 12 56030724 T G Non-synonymous OR10P1 22.1 12 57578143 C T Non-synonymous LRP1 22.4 14 21780617 A G Non-synonymous RPGRIP1 27.3 14 47600951 A T Non-synonymous MDGA2 22.5 14 72196852 C G Non-synonymous SIPA1L1 22.6 17 26684394 T G Upstream TMEM199 15.59 19 1592525 A C Non-synonymous MBD3 28.3 19 1592543 G C Non-synonymous MBD3 23.3 19 6452423 C T Non-synonymous SLC25A23 19.64 19 44662108 A G Non-synonymous ZNF234 25 19 50832152 T C Upstream NR1H2 16.25 19 50832152 T C Non-synonymous KCNC3 16.25 22 24953707 C T Upstream GUCD1 22.8 22 24953707 C T Non-synonymous SNRPD3 22.8 22 37637633 G T Non-synonymous RAC2 32 Table E1 Variants shared by all 3 patients, which are rare (<30 alleles in gnomAD) and deleterious (CADD PHRED-like score >15)
SARS-CoV-2 occurs in the majority of children as COVID-19, without symptoms or with a paucisymptomatic respiratory syndrome, but a small proportion of children develop the systemic Multi Inflammatory ...Syndrome (MIS-C), characterized by persistent fever and systemic hyperinflammation, with some clinical features resembling Kawasaki Disease (KD).
With this study we aimed to shed new light on the pathogenesis of these two SARS-CoV-2-related clinical manifestations.
We investigated lymphocyte and dendritic cells subsets, chemokine/cytokine profiles and evaluated the neutrophil activity mediators, myeloperoxidase (MPO), and reactive oxygen species (ROS), in 10 children with COVID-19 and 9 with MIS-C at the time of hospital admission.
Patients with MIS-C showed higher plasma levels of C reactive protein (CRP), MPO, IL-6, and of the pro-inflammatory chemokines CXCL8 and CCL2 than COVID-19 children. In addition, they displayed higher levels of the chemokines CXCL9 and CXCL10, mainly induced by IFN-γ. By contrast, we detected IFN-α in plasma of children with COVID-19, but not in patients with MIS-C. This observation was consistent with the increase of ISG15 and IFIT1 mRNAs in cells of COVID-19 patients, while ISG15 and IFIT1 mRNA were detected in MIS-C at levels comparable to healthy controls. Moreover, quantification of the number of plasmacytoid dendritic cells (pDCs), which constitute the main source of IFN-α, showed profound depletion of this subset in MIS-C, but not in COVID-19.
Our results show a pattern of immune response which is suggestive of type I interferon activation in COVID-19 children, probably related to a recent interaction with the virus, while in MIS-C the immune response is characterized by elevation of the inflammatory cytokines/chemokines IL-6, CCL2, and CXCL8 and of the chemokines CXCL9 and CXL10, which are markers of an active Th1 type immune response. We believe that these immunological events, together with neutrophil activation, might be crucial in inducing the multisystem and cardiovascular damage observed in MIS-C.
To the Editor: The nuclear factor kappa B (NF-kB) signaling pathway plays an important role in immune cell biology.1 Both the classical (nuclear factor kappa B, subunit 1 NF-kB1; p105/p50) and the ...alternative (nuclear factor kappa B, subunit 2 NF-kB2; p100/p52) NF-kB pathways have been largely studied mainly in animal models.1 Regarding B cells, the role of NF-kB1 was underlined in a murine nf-kb1 knockout model in which peripheral B cells showed defective maturation, defective isotype switching, and impaired humoral immune responses.1-3 A similar, although more pronounced, immunologic phenotype was observed in the nf-kb2 knockout mice, with defective secondary lymphoid organ development and impaired B-cell development both in early (bone marrow) and in late (periphery) stages with defective humoral responses both to T-dependent and to T-independent antigens.1 The role of NF-kB2 in human B-cell development was recently defined in patients carrying monoallelic mutations in NF-kB2, leading to common variable immunodeficiency (CVID)-like disease with autoimmunity and defects in late stages of peripheral B-cell maturation.4,5 Monoallelic mutations in NF-kB1 leading to p50 haploinsufficiency were recently described in a limited number of patients with CVID6; however, data regarding the effect of monoallelic NF-kB1 mutations on B-cell development are scarce. Appendix Features Patient 1 Patient 2low * Sex Male Female Age at diagnosis (y) 7 30 Current age (y) 40 55 IgG (mg/dL)dagger 160 181 IgA (mg/dL)dagger 14 6 IgM (mg/dL)dagger 14 48 Recall response to vaccinations Tetanus toxoid Absent NA Hepatitis B Absent NA Lymphocyte subsetsdagger CD3 (%) 83.984.3 Â CD4 (%) 39.758 Â CD8 (%) 53.523.2 Â CD19 (%) 3.210.5 Â CD56 (%) 10.64.7 Â Proliferative response to mitogens Anti-CD3, anti-CD3 + IL-2, PMA + I, PHA Normal NA Respiratory infections Pneumonia, sinusitis Pneumonia, recurrent sinusitis Bronchiectasis -- -- Autoimmunity Autoimmune thyroiditis, autoimmune enteropathy --
Agammaglobulinemia is the most profound primary antibody deficiency that can occur due to an early termination of B-cell development. We here investigated 3 novel patients, including the first known ...adult, from unrelated families with agammaglobulinemia, recurrent infections, and hypertrophic cardiomyopathy (HCM). Two of them also presented with intermittent or severe chronic neutropenia. We identified homozygous or compound-heterozygous variants in the gene for folliculin interacting protein 1 (FNIP1), leading to loss of the FNIP1 protein. B-cell metabolism, including mitochondrial numbers and activity and phosphatidylinositol 3-kinase/AKT pathway, was impaired. These defects recapitulated the Fnip1−/− animal model. Moreover, we identified either uniparental disomy or copy-number variants (CNVs) in 2 patients, expanding the variant spectrum of this novel inborn error of immunity. The results indicate that FNIP1 deficiency can be caused by complex genetic mechanisms and support the clinical utility of exome sequencing and CNV analysis in patients with broad phenotypes, including agammaglobulinemia and HCM. FNIP1 deficiency is a novel inborn error of immunity characterized by early and severe B-cell development defect, agammaglobulinemia, variable neutropenia, and HCM. Our findings elucidate a functional and relevant role of FNIP1 in B-cell development and metabolism and potentially neutrophil activity.
•FNIP1 deficiency causes agammaglobulinemia, variable neutropenia, and hypertrophic cardiomyopathy.•FNIP1 deficiency alters B-cell development and metabolism.
Display omitted
A female offspring of consanguineous parents, showed features of Wiskott-Aldrich syndrome (WAS), including recurrent infections, eczema, thrombocytopenia, defective T cell proliferation and ...chemotaxis, and impaired natural killer cell function. Cells from this patient had undetectable WAS protein (WASP), but normal WAS sequence and messenger RNA levels. WASP interacting protein (WIP), which stabilizes WASP, was also undetectable. A homozygous c.1301C>G stop codon mutation was found in the WIPF1 gene, which encodes WIP. Introduction of WIP into the patient's T cells restored WASP expression. These findings indicate that WIP deficiency should be suspected in patients with features of WAS in whom WAS sequence and mRNA levels are normal.
Background Combined immunodeficiency with multiple intestinal atresias (CID-MIA) is a rare hereditary disease characterized by intestinal obstructions and profound immune defects. Objective We sought ...to determine the underlying genetic causes of CID-MIA by analyzing the exomic sequences of 5 patients and their healthy direct relatives from 5 unrelated families. Methods We performed whole-exome sequencing on 5 patients with CID-MIA and 10 healthy direct family members belonging to 5 unrelated families with CID-MIA. We also performed targeted Sanger sequencing for the candidate gene tetratricopeptide repeat domain 7A ( TTC7A ) on 3 additional patients with CID-MIA. Results Through analysis and comparison of the exomic sequence of the subjects from these 5 families, we identified biallelic damaging mutations in the TTC7A gene, for a total of 7 distinct mutations. Targeted TTC7A gene sequencing in 3 additional unrelated patients with CID-MIA revealed biallelic deleterious mutations in 2 of them, as well as an aberrant splice product in the third patient. Staining of normal thymus showed that the TTC7A protein is expressed in thymic epithelial cells, as well as in thymocytes. Moreover, severe lymphoid depletion was observed in the thymus and peripheral lymphoid tissues from 2 patients with CID-MIA. Conclusions We identified deleterious mutations of the TTC7A gene in 8 unrelated patients with CID-MIA and demonstrated that the TTC7A protein is expressed in the thymus. Our results strongly suggest that TTC7A gene defects cause CID-MIA.
Purpose
Jacobsen syndrome (JS) is a rare form of genetic disorder that was recently classified as a syndromic immunodeficiency. Available detailed immunological data from JS patients are limited.
...Methods
Clinical and immunological presentation of twelve pediatric patients with JS by means of revision of clinical records, flow cytometry, real-time PCR, and lymphocyte functional testing were collected.
Results
Recurrent infections were registered in 6/12 patients (50%), while bleeding episodes in 2/12 (16.7%). White blood cell and absolute lymphocyte counts were reduced in 8/12 (66.7%) and 7/12 (58.3%) patients, respectively. Absolute numbers of CD3
+
and CD4
+
T cells were reduced in 8/12 (66.7%) and 7/12 (58.3%), respectively. Of note, recent thymic emigrants (RTE) were reduced in all tested patients (9/9), with T-cell receptor excision circle analysis (TRECs) showing a similar trend in 8/9 patients; naïve CD4
+
T cells were low only in 5/11 patients (45.4%). Interestingly, B-cell counts, IgM memory B cells, and IgM serum levels were reduced in 10/12 (83.3%) patients. Natural killer (NK) cell counts were mostly normal but the percentages of CD16
+
CD56
low/−
cells were expanded in 7/7 patients tested. The observed immunological alterations did not correlate with patients’ age. Finally, responses to proliferative stimuli were normal at presentation for all patients, although they may deteriorate over time.
Conclusions
Our data suggest that patients affected with JS may display important numeric and maturational alterations in the T-, B-, and NK-cell compartments. These findings suggest that JS patients should be regularly monitored from an immunological point of view.
PDF
