Common variable immunodeficiency: autoimmune cytopenias and advances in molecular diagnosis Free

A 33-year-old man had onset of acute vomiting and diarrhea in 2022 that was ascribed to an undercooked meat dish. He had, however, a low white cell count of 1200 and a low neutrophil count of 400 at this time. His hemoglobin was 8.4  g/dl, and he had a platelet count of 12 000 units. A bone marrow examination suggested a degree of multilineage dysplasia but revealed no clearly abnormal findings. A CT scan showed an enlarged spleen and some lung nodules. Other laboratory data showed low immune globulins, IgG  =  249, IgA  =  22, IgM <25  mg/dl, so he was referred to immunology and was found to have a poor pneumococcal vaccine response (only 4 of 23 serotypes became positive). However, his past medical history included 2 episodes of immune thrombocytopenia (ITP), at age 16 and age 22, each time successfully treated with steroids. He had not had pneumonias, cough, bronchitis, or other significant infectious history suggestive of an immune deficiency. However, with the low immune globulin levels and lack of antibody response to vaccination, our patient was diagnosed with common variable immunodeficiency (CVID) and started on immune globulin replacement. He has since remained stable with normal platelet counts.

CVID is one of the most frequently encountered primary immunodeficiencies in clinical practice, with an estimated incidence of about 1 in 20 000 to 50 000, depending on the population studied.¹ The diagnosis is made in male or female patients with reduced serum levels of IgG, IgA, and/or IgM, with documented defects of antibody production to both protein and carbohydrate antigens, and the exclusion of other causes of hypogammaglobulinemia, such as physiologic immaturity, medications, malignancy, or protein losses.^1-5 The standard treatment of CVID, when antibody production is established, is immune globulin replacement, given by intravenous or subcutaneous route.⁶ While CVID is considered a primary immune defect, most newly diagnosed patients are between the ages of 20 and 40 years.^1,7,8 More than half of all patients have infections as the primary manifestation; however, a significant proportion have autoimmune cytopenias,^8-13 such as sudden-onset immune thrombocytopenia, hemolytic anemia, or both (Evans syndrome), or less commonly, neutropenia, as the only manifestation of the immune defect. In some, the autoimmune conditions are concurrently or later associated with additional inflammatory features, such as interstitial lung disease or liver or gastrointestinal complications.¹⁴ These conditions in aggregate with other inflammatory features lead to increased morbidity and mortality in CVID.^8,15,16 

With increased use of whole-exome sequencing (WES) and whole-genome sequencing (WGS), monogenic causes of the CVID phenotype now account for about 25% to 30% of patients.^17-21 In fact, dissecting the genes that underlie CVID syndrome has elucidated many of the complex requirements of B-cell development in normal human immunity, starting with B-cell generation in the bone marrow, proliferation, antigen signaling, cell activation, class-switch recombination, B-cell migration, long-term survival, and finally, the retention and maturation of antibody-secreting memory B cells and ultimately, the development of long-lived plasma cells. Here we outline the demographic, immunologic, and genetic analyses of a cohort of CVID patients with immune thrombocytopenia, hemolytic anemia, or neutropenia drawn from a large patient population in the United States.

Individuals with CVID were seen in the Immune Deficiency Clinic at the Icahn School of Medicine at Mount Sinai over a 25-year period. Patients were diagnosed with CVID using established criteria, including serum IgG and IgA and/or IgM deficiency with proven loss-of-antibody production.^2,4,22 Clinical, demographic, and immunologic information was collected on each patient. Genetic evaluation was done by WES.^19,22,23 In a number of cases, patient exomes were also examined for mutations in a panel of 429 genes associated with a primary immune deficiency disease (Invitae Diagnostics). Ethical permission for these studies was obtained from the Mount Sinai Institutional Review Board, and informed consent was obtained from all individuals and/or their legal guardians.

In this study, 408 CVID patients were analyzed. The median age for the group was 44 years (range 5 to 77 years). Table 1 outlines the baseline immune globulin levels of these participants. Of these, 122 (29%) patients (60 females and 62 males) had had 1 or more episodes of immune thrombocytopenia, hemolytic anemia, and/or neutropenia, or more rarely, a few other hematologic complications, while the other 286 CVID participants in this cohort did not have these conditions (Figure 1). ITP was the most common complaint and was reported in 68 individuals (16.7%); ITP had also occurred in 24 others (5.8%) as part of Evans syndrome, having had 1 or more concomitant or usually separate episodes of warm autoimmune hemolytic anemia (AIHA). The third most common autoimmune condition was warm AIHA in 10 (2.5%) patients. Fewer had neutropenia (N  =  3) or had combinations of these or other hematologic autoimmune conditions (N  =  8) (Table 3). As noted in the case described above, ITP and/or AIHA had recurred in the same patient over time, and the immune defect was not identified on the first or sometimes even the second of these occasions. As for the patient presented, the diagnosis of CVID was not made until further medical care was required, often for another reason. Thirty-three of these 122 individuals (27%) had had a splenectomy to treat the autoimmune disease and/or massive splenic enlargement, which was considered to lead to hypersplenism and low counts. Of the group of 122 individuals who had experienced hematologic autoimmune disorders, 19 (15.5%) are no longer alive.

Fifty-three of the 122 (43%) individuals with hematologic autoimmunity had 1 or more genetic mutations considered to be related to the immune defect. As noted in other cohorts of CVID patients, heterozygous mutations were the most commonly observed in this cohort.^17,18 Of the 408 participants, only 26 (6%) had a family history of an immune defect (Table 3). As shown previously, a significant proportion of CVID patients have variants in the TNFRSF13B/TACI gene (transmembrane activator and CAML interactor) and here, 13% of those with autoimmunity had deleterious variants in this gene. These included a monoallelic variant in most patients but a homozygous mutation in one patient. A mutation was also noted in our index case as this man had one of the common mutations in this gene, A181E, which is in the transmembrane region. Seven other participants on this autoimmune cohort had autosomal dominant NFKB1 deficiency (nuclear factor kappa B subunit 1), and 3 other patients each had heterozygous mutations in CTL4, IKZF1, or STAT3. Two sisters and the son of one of them, with no warts and moderate neutropenia but recurrent severe autoimmune thrombocytopenia and autoimmune hemolytic anemia, had a heterozygous deleterious mutation in CXCR4 (C-X-C motif chemokine receptor 4), a gene associated with WHIM syndrome (warts, hypogammaglobulinemia, infections, and myelokathexis).^17,19,24 Two patients with Kabuki syndrome and a CVID phenotype had mutations in KMT2D (lysine methyltransferase 2), and 3 others had heterozygous chromosomal deletion on chromosome 22 or a TBX1 gene mutation, or DiGeorge syndrome. Other gene variants were also noted in these patients, as outlined in Table 3. For 3 of these patients, 2 different deleterious mutations were noted, a TACI mutation along with pathogenetic mutations in NFKB2, TBK1, or TCF3. Of the 33 patients who had a splenectomy, 7 had a TACI variant. Massive lymphoid hyperplasia with splenomegaly has been noted commonly in patients with TACI mutations.^25,26 Eight of these 122 patients (6.6%) had a history of lymphoma; 1 had had a thymoma. (The mutations in these patients included 1 with an NFKB2 mutation and a TACI mutation, 1 with a pathogenic mutation in TET2, and 2 different pathogenic TACI mutations.)

In addition to the CVID patients with hematologic autoimmune disease, the genetic associations noted in the 286 other individuals (139 females and 147 males) were also examined. Of these, 68 (24%) had 1 or more genetic variants associated with CVID (Table 3). For these as well, genetic variants were also noted in the TACI (23%) and NFKB1 (13%) genes, with other genetic alterations also noted. Fourteen (4.9%) of these patients had had either lymphoma or Hodgkin's disease. (One had a mutation in PIK3CD.) Of the group of 286, 18 (6.3%) are no longer alive.

Hematologic manifestations occur in a substantial proportion of patients with CVID.^27-29 Most commonly, this is ITP, or less commonly, AIHA and/or neutropenia. Other participants in this cohort had other autoimmune hematologic manifestations, with aplastic anemia, antinuclear antibodies, pancytopenia, or combinations of these issues. These features of immune deficiency were in some cases the presenting manifestation of the immune defect, as it was for our patient, and the reason that the diagnosis of CVID was finally made. As for participants with CVID in general, most were adults at the time of presentation, but the patient may have had a history of an earlier episode of autoimmunity, a significant previous infection, or another medical event in the past that suggested an immune defect might be present. In most cases, steroids were used as the first line of treatment of the hematologic condition, often coupled with high doses of intravenous immune globulin; however, in the past decade many patients have been treated with rituximab,^30,31 which in most cases is successful (coupled with immune replacement immune globulin). As the added immune globulin protects against most infections, the risks of using rituximab appear quite limited. In addition, the therapy also avoids long-term use of steroids and splenectomy. Using steroids in CVID is generally for limited periods due to the risk of opportunistic infections, which can develop with long-term use. While splenectomy is also discouraged, in a review of 45 CVID patients from 16 centers, this procedure did successfully treat the cytopenia. However, it was associated with more episodes of overwhelming post-splenectomy infections, especially if there was absent or inadequate immunoglobulin replacement therapy.³² 

One of the other hematologic complications that may be diagnosed in CVID is lymphoid malignancy.¹¹ In a group of 647 CVID patients followed over 4 decades, 45 patients (15 males and 30 females, 7%) developed 49 lymphoid malignancies, almost all of B-cell linage. Some of these patients had a history of only recurrent infections (36.3%); others had autoimmunity (33%), enteropathy (20%), and/or granulomatous disease (11%). Another 6 patients in this group were also diagnosed with Non-Hodgkin lymphoma, but on review of pathology this diagnosis was reversed, as the findings were more consistent with benign lymphoproliferation, demonstrating that there can be diagnostic challenges due to immune dysregulation characteristic of this immune defect.¹¹ 

While most patients diagnosed with CVID do not have a clarified molecular cause, the genetic discoveries in antibody defects continue to reveal the complex immunologic pathways needed to initiate and sustain normal B-cell development and the long-term maintenance of B-cell memory. In the current cohort of 408 CVID patients, 119 (29%) had an identifiable causative or associated genetic variant, similar to other investigated cohorts.^17,18 These recognized mutations are in selected genes that are important in B-cell biology, including the TACI,^25,33,34CTLA4,³⁵NFKB1,³⁶NFKB2,³⁷STAT3,³⁸PI3KCD,³⁹ and LRBA genes,⁴⁰ as well as gain and loss of function in IKZF1.⁴¹ However, the largest number of patients had variants in the TACI gene. (This includes our patient, as he had the common transmembrane TACI mutation, A181E.) While TACI gene variants are not thought to be disease causing, they are still commonly associated with autoimmunity when found in CVID, potentially functioning as a genetic modifier^25,26,34 and possibly explaining this enrichment. In contrast to those CVID patients who had not experienced hematologic conditions, a genetic connection was also clearly increased in those patients with hematologic conditions, as 43% of these individuals had an identified genetic association, in contrast to those with no hematologic conditions at 24%. Analysis of patients from the genetic perspective is also important in some cases, as the gene lesion may lead to a more precise method of therapy. For example, the inflammatory complications in patients with mutations in CTLA4 have been shown to be ameliorated by the anti-inflammatory compound abatacept (a CTLA4-Fc fusion protein),³⁵ patients with STAT3 gain-of-function mutations may benefit from JAK inhibitors,⁴² and patients with activating mutations in PI3K-delta may benefit from treatment with the specific inhibitor leniolisib.⁴³ 

Why autoimmunity occurs in patients with impaired immunity is of course a complex topic and opens an interesting discussion about the mechanisms of central and peripheral tolerance and how these develop to prevent autoimmunity in healthy humans.⁴⁴ Some of these mechanisms involve the same genetic pathways that are defective in CVID. In this regard, it is notable that the majority of genetic defects noted in CVID with autoimmunity are mutations that affect both T- and B-cell interactions, and not those aligning with a purely B-cell intrinsic defect. While many of the mutations are autosomal dominant in inheritance, a family history of immune deficiency is infrequently obtained in CVID, as noted here. In some cases, we found that the mutation was de novo, while in others, there was an asymptomatic parent or sibling with the same mutation, demonstrating the variable penetrance of these alleles. With further exploration, more digenic or even polygenic causes of CVID are likely to be dissected. Considering the intersecting immunologic pathways, it is possible that aside from genetics, these different clinical manifestations may result from metabolic factors, environmental factors, or epigenetic causes.^45-50 Analysis of the patient from the genetic perspective is also important in some cases, as the gene lesion may lead to a more precise method of therapy. For example, the inflammatory complications in patients with mutations in CTLA4 have been shown to be ameliorated by the anti-inflammatory compound abatacept (a CTLA4-Fc fusion protein), and patients with STAT2 gain-of-function mutations may benefit from a JAK inhibitor.

Limitations of this study include the fairly narrow demographic background of the patients referred, important when considering the genetics of different CVID patient cohorts,^1,17,18 and the possibility of incomplete clinical information, as these data were collected over varying periods of time. A further limitation is that confirmation of the pathogenicity of the identified variants depends on previous publications, in vitro assays where available, and the genetic methods used to assign the likelihood of a variant of dexterous variant exerting a deleterious change.

In summary, for 408 patients with CVID, 122 patients (29%) had had 1 or more episodes of immune thrombocytopenia, hemolytic anemia, and/or neutropenia, or more rarely, another benign hematologic condition. Of this group, 53 of the 122 (43%) with hematologic autoimmunity had 1 or more genetic mutations considered to be related to the immune defect. For the 286 other patients, 68 (24%) had 1 or more genetic variants associated with CVID. For the group, 22 (5%) had a history of lymphoma or Hodgkin's disease, and 1 had had a thymoma.

This work was partially supported by the National Institutes of Health's National Institute of Allergy and Infectious Diseases, AI-061093, AI-08603, and the David S. Gottesman Chair.

Charlotte Cunningham-Rundles: no competing financial interests to declare.

Jean-Laurent Casanova: no competing financial interests to declare.

Bertrand Boisson: no competing financial interests to declare.

Charlotte Cunningham-Rundles: None to declare.

Jean-Laurent Casanova: None to declare.

Bertrand Boisson: None to declare.