Zap70-Related Combined Immunodeficiency

Clinical characteristics.

ZAP70-related combined immunodeficiency (ZAP70-related CID) is a cell-mediated immunodeficiency caused by abnormal T-cell receptor (TCR) signaling. Affected children usually present in the first year of life with recurrent bacterial, viral, and opportunistic infections, diarrhea, and failure to thrive. Severe lower-respiratory infections and oral candidiasis are common. Affected children usually do not survive past their second year without hematopoietic stem cell transplantation (HSCT).

Diagnosis/testing.

The diagnosis of ZAP70-related CID is suggested by low to absent CD8⁺ T cells in an individual with normal CD3+ and CD4+ T-cell counts. Additional supportive laboratory features include absent proliferation of CD4⁺ T cells in response to mitogens and antigens, and absent ZAP-70 protein expression. The diagnosis is established in a proband by identification of biallelic pathogenic variants in ZAP70 on molecular genetic testing.

Management.

Treatment of manifestations: Supportive care includes immediate intravenous immunoglobulin (IVIG) and antibacterial, antifungal, antiviral, and Pneumocystis jiroveci prophylaxis to control and reduce the occurrence of infections.

Prevention of primary manifestations: Allogeneic HSCT to reconstitute the immune system, preferably prior to the onset of infections.

Prevention of secondary complications: Use of irradiated, leukoreduced, cytomegalovirus (CMV)-safe blood products; deferment of immunizations until immune reconstitution; consideration for formula feeds in place of breast feeding until CMV status of mother is known.

Surveillance: Individuals with milder findings or those who have not undergone HSCT need to be monitored for worsening of immune function with periodic assessment of clinical status and functional lymphocyte responsiveness. Following a successful HSCT, the following should be routinely monitored: growth, psychomotor development, complete blood counts, liver and renal function, immune status, donor and recipient chimerism, development of post-transplant complications.

Agents/circumstances to avoid: Non-irradiated blood products; live viral, live mycobacterial, and live bacterial vaccinations; contaminated water sources; exposure to fungus-enriched environments (e.g., construction sites, agricultural areas with active soil disruption, mulch, hay).

Evaluation of relatives at risk: Because the outcome of HSCT in children with ZAP70-related CID is significantly improved by performing HSCT prior to the onset of severe infections, early testing of at-risk sibs should be considered. In addition, any sibs considered as bone marrow donors must be evaluated for ZAP70-related CID prior to donation.

Genetic counseling.

ZAP70-related CID is inherited in an autosomal recessive manner. At conception, each sib of an affected individual has a 25% chance of being affected, a 50% chance of being a carrier, and a 25% chance of being unaffected and not a carrier. Carrier testing for at-risk family members and prenatal diagnosis for pregnancies at increased risk are possible if the pathogenic variants in the family are known.

Suggestive Findings

ZAP70-related combined immunodeficiency (ZAP70-related CID) should be suspected in individuals who present with the following findings within the first two years of life:

• Recurrent viral, bacterial, and opportunistic infections
• Chronic diarrhea and failure to thrive
• Characteristic results of lymphocyte subset analysis of CD3, CD4, and CD8 T cells, lymphocyte functional testing, and ZAP-70 protein expression (see Lymphocyte development and numbers)

Note: Individuals with non-classic ZAP70-related CID may present at an older age with symptoms of autoimmunity, lymphoproliferation, and/or immune dysregulation with or without evidence of immunodeficiency.

Lymphocyte development and numbers. In ZAP70-related CID, total lymphocyte counts can range from normal to high.

• Thymic architecture is largely preserved in individuals with ZAP70-related CID; however, compared to controls, a smaller medullary zone, decreased numbers of AIRE+ medullary thymic epithelial cells, and decreased numbers of dendritic cell numbers may be seen [Poliani et al 2013].
• T-cell counts:
  • CD3⁺ cell counts are typically normal.
  • CD4⁺ cell counts are normal or elevated and may account for 60%-80% of the lymphocytes in individuals with ZAP70-related CID. Numbers of CD4+ recent thymic emigrants may be decreased [Hauck et al 2015].
  • CD8⁺ cells are absent or extremely low, often comprising only 0%-2% of the child's total T-cell count in individuals with ZAP70-related CID [Arpaia et al 1994, Noraz et al 2000].
  • Numbers of regulatory T cells may be normal or decreased [Poliani et al 2013, Hauck et al 2015].
• B cell counts and NK cell counts are normal.

Lymphocyte function. T-cell responses to stimuli that act through the T-cell receptor (TCR) are absent or severely diminished:

• Absent or decreased proliferation to CD3 antibody [Roifman et al 2010]
• Absent or decreased proliferation of CD4⁺ cells in response to mitogens (e.g., PHA, ConA) [Roifman et al 2012]
• Intact proliferative response to mitogenic stimuli that bypass the TCR (e.g., PMA/Ionomycin) [Elder et al 1994, Elder 1997]
• Normal TCR Vβ repertoire in both CD4+ and CD8+ T cells [Roifman et al 2010]
• In CD4 cells: limited differentiation into Th2 cells and resistance to Fas-induced cell death [Roifman et al 2010]
• In regulatory T cells: potentially decreased expression of CTLA4 and TGFB leading to increased risk of autoimmunity [Roifman et al 2010]

ZAP-70 protein expression. Testing of CD4⁺ T cells reveals absence or near absence of ZAP-70 protein in most affected individuals. Recent reports suggest that the amount of residual ZAP-70 protein expression influences the clinical phenotype [Picard et al 2009, Chan et al 2016, Gavino et al 2017]. Expression of Syk, a related tyrosine kinase important in T-cell signaling, may partially compensate for ZAP-70 signaling in some individuals [Toyabe et al 2001, Hauck et al 2015].

Immunoglobulin concentrations and function

• Immunoglobulin levels vary by individual. Many affected individuals have severe hypogammaglobulinemia, but normal immunoglobulins or elevated IgA, IgM, and/or IgE can also be seen [Turul et al 2009, Cuvelier et al 2016].
• Although functional antibody responses to immunization are present in a few persons [Turul et al 2009, Hauck et al 2015], this finding does not indicate that all specific antigenic responses are intact.

Newborn screening. The utility of TREC screening for individuals with ZAP70-related CID continues to be controversial. Although some have shown that TREC levels in individuals with confirmed ZAP70-related CID are very low compared to age-matched controls [Roifman et al 2010, Roifman et al 2012, Aluri et al 2017], others have shown that a substantial portion of individuals with ZAP70-related CID have TREC levels above the cutoffs used for newborn screening and could be missed [Grazioli et al 2014, Jilkina et al 2014, Hauck et al 2015, Schroeder et al 2016]. Therefore, newborn screening results must be interpreted with caution in individuals with clinical findings consistent with ZAP70-related CID or in populations with a high prevalence of ZAP70-related CID.

Establishing the Diagnosis

The diagnosis of ZAP70-related CID is established in a proband by identification of biallelic pathogenic variants in ZAP70 on molecular genetic testing (see Table 1).

Molecular genetic testing approaches can include single-gene testing or use of a multigene panel:

• Single-gene testing. Sequence analysis of ZAP70 is performed.
  Note: (1) No deletions or duplications of ZAP70 have been reported. (2) Targeted analysis for the c.1624-11G>A variant can be performed first in individuals of Mennonite ancestry.
• A multigene panel that includes ZAP70 and other genes of interest (see Differential Diagnosis) may also be considered.
  Note: (1) The genes included in the panel and the diagnostic sensitivity of the testing used for each gene varies by laboratory and are likely to change over time. (2) Some multigene panels may include genes not associated with the condition discussed in this GeneReview; thus, clinicians need to determine which multigene panel is most likely to identify the genetic cause of the condition at the most reasonable cost while limiting identification of variants of uncertain significance and pathogenic variants in genes that do not explain the underlying phenotype. (3) In some laboratories, panel options may include a custom laboratory-designed panel and/or custom phenotype-focused exome analysis that includes genes specified by the clinician. (4) Methods used in a panel may include sequence analysis, deletion/duplication analysis, and/or other non-sequencing-based tests.
  For an introduction to multigene panels click here. More detailed information for clinicians ordering genetic tests can be found here.

Clinical Description

Individuals with ZAP70-related CID characteristically present in the first two years of life with recurrent bacterial, viral (including live-virus vaccine strains), and opportunistic infections, diarrhea, and failure to thrive. Severe lower-respiratory infections are seen, most notably Pneumocystis jiroveci infections and viral infections. Oral candidiasis is also common [Cuvelier et al 2016, Schroeder et al 2016].

Other presentations have also been reported:

• Reports of milder phenotypes in sibs of children who had died from ZAP70-related CID include a child age five months with recurrent lower-respiratory disease but no severe infections [Turul et al 2009] and a child with persistent dermatitis resistant to therapy [Katamura et al 1999].
• A child age nine years with a ZAP70 hypomorphic intronic pathogenic variant had an attenuated clinical and immunologic phenotype [Picard et al 2009] (see Genotype-Phenotype Correlations).
• A child age 11 months with ZAP70-related CID presented with lymphoma [Newell et al 2011].
• BCG-related complications including axillary lymphadenitis or disseminated mycobacterial disease following BCG vaccination may be presenting features [Santos et al 2010, Esenboga et al 2016].
• Two individuals presented with recurrent infections and silent brain infarcts; one also had congenital nephrotic syndrome and autoimmune hemolytic anemia [Akar et al 2015].
• An individual with isolated treatment-refractory immune thrombocytopenia (ITP) has been described [Cuvelier et al 2016].
• Two sibs had refractory autoimmune features including nephrotic syndrome/proteinuria, bullous pemphigoid, and colitis; one also developed autoantibodies to factor VIII caused by a hypomorphic and weakly activating ZAP70 pathogenic variant [Chan et al 2016].
• An adult with a history of infantile-onset colitis, recurrent respiratory-tract infections, mucocutaneous candidiasis, HSV stomatitis, VZV infection, EBV lymphoproliferative disorder, recurrent CMV viremia, polyomaviremia, and epidermodysplasia verruciformis-like lesions due to HHV-23 was recently reported. A novel pathogenic variant (c.1272C>T) that leads to an altered splice site and low levels of wild-type ZAP-70 protein was identified in this individual [Gavino et al 2017].

The long-term prognosis of untreated ZAP70-related CID is death from infection. Affected children have a declining quality of life and usually do not survive past their second year without hematopoietic stem cell transplantation (HSCT).

Genotype-Phenotype Correlations

There is very little genotype-phenotype correlation reported in ZAP70-related CID; however, the amount of residual ZAP-70 protein expressed may modulate the clinical phenotype, as suggested by the following:

• A child with a hypomorphic intronic pathogenic variant (c.837+121G>A in intron 7) who had had chronic eczema from age two months and recurrent infections from age two years. The frequency of infections declined at age six following introduction of co-trimoxazole and IVIG prophylaxis. Of note, an older sib with a history of multiple infections had died at age one year [Picard et al 2009].
• Two sibs with refractory autoimmune features including nephrotic syndrome/proteinuria, bullous pemphigoid, and colitis. One sib also developed autoantibodies to factor VIII. These sibs had compound heterozygous ZAP70 pathogenic variants including a hypomorphic variant (c.574C>T) and a weakly activating variant (c.1079G>C) [Chan et al 2016].

Nomenclature

ZAP70-related combined immunodeficiency may also be referred to as zeta-associated protein 70 deficiency or immunodeficiency 48.

Prevalence

ZAP70-related CID was first described in 1994 in three individuals of Mennonite descent [Arpaia et al 1994]. Since that time, more than 50 affected individuals have been described in the literature [Arpaia et al 1994, Chan et al 1994, Elder et al 1994, Elder et al 1995, Gelfand et al 1995, Katamura et al 1999, Matsuda et al 1999, Meinl et al 2000, Noraz et al 2000, Barata et al 2001, Elder et al 2001, Toyabe et al 2001, Fagioli et al 2003, Picard et al 2009, Turul et al 2009, Santos et al 2010, Newell et al 2011, Hönig et al 2012, Roifman et al 2012, Karaca et al 2013, Kim et al 2013, Grazioli et al 2014, Akar et al 2015, Hauck et al 2015, Cuvelier et al 2016, Esenboga et al 2016, Schroeder et al 2016, Aluri et al 2017, Gavino et al 2017, Shirkani et al 2017].

The prevalence of ZAP70-related CID is unknown but much lower than that of all forms of SCID, which is estimated at 1:50,000. Prevalence is higher in the Canadian Mennonite population, where a c.1624-11G>A pathogenic variant resulting in destabilization of the ZAP-70 protein is frequently seen [Arpaia et al 1994, Jilkina et al 2014, Cuvelier et al 2016, Schroeder et al 2016].

Evaluations Following Initial Diagnosis

The care of individuals diagnosed with ZAP70-related CID is best managed with a multidisciplinary team of providers including hematology/oncology/bone marrow transplantation, immunology, genetics, and infectious disease specialists. To establish the extent of disease and needs in an individual diagnosed with ZAP70-related combined immunodeficiency (CID), the following evaluations are recommended:

• Assessment of growth
• Evaluation for common and opportunistic viral, bacterial, and fungal disease-causing agents
• Complete metabolic panel (liver and renal function), complete blood count (CBC) with differential and platelet count, lymphocyte subsets and mitogen proliferation, and quantitative immunoglobulins
• Consultation with a clinical geneticist and/or genetic counselor
• Consultation with a clinical immunologist
• Consultation for hematopoietic stem cell transplantation

Treatment of Manifestations

Treatment relies on prompt reconstitution of the individual's immune system (see Prevention of Primary Manifestations).

Supportive treatment includes IVIG and antibacterial, antifungal, antiviral, and Pneumocystis jiroveci prophylaxis to control and reduce the occurrence of infections.

Prevention of Primary Manifestations

The only curative therapy for ZAP70-related CID is allogeneic hematopoietic stem cell transplantation (HSCT). Extrapolated data show that the outcome of HSCT in children with SCID is significantly improved by performing HSCT prior to the onset of infections [Pai et al 2014]. Children with ZAP70-related CID have been successfully transplanted using a variety of donors including haploidentical donors and unrelated umbilical cord blood [Noraz et al 2000, Elder et al 2001, Hönig et al 2012, Cuvelier et al 2016].

• Outcomes are the best with HLA-matched, related donors.
• If an HLA-matched, related donor is not available, alternatives include:
  • Matched unrelated donor;
  • Umbilical cord blood donor;
  • Haploidentical parental bone marrow or mobilized peripheral blood stem cells that have been T-cell depleted.
• In contrast to individuals with SCID, individuals with ZAP70-related CID are typically treated with a chemotherapeutic conditioning regimen prior to HSCT, although some individuals have received unconditioned transplants with variable success, suggesting that conditioning may not be essential in some circumstances [Hönig et al 2012, Kim et al 2013, Cuvelier et al 2016].
• The largest series of eight individuals with ZAP70-related CID who received HSCT using a variety of stem cell sources showed the following:
  • All individuals were alive at a median of 13.5 years of follow up.
  • Two-thirds of the individuals who did not receive conditioning failed to have myeloid engraftment but have maintained stable mixed chimerism. In addition, three individuals who received stem cells from a matched sib did not receive conditioning prior to transplant and achieved engraftment.
  • 75% of individuals developed acute graft-versus-host disease (GVHD) and 50% developed chronic GVHD.
  • Seven of eight individuals achieved freedom from IVIG and show evidence of class switching with resolution of dysregulated immunoglobulin production and six of the eight show evidence of antibody production to both protein and polysaccharide vaccines.
  • Two individuals receiving myeloablative conditioning have developed premature ovarian failure.
• Cellular reconstitution following HSCT takes up to one year, while restoration of humoral immunity can take significantly longer, and may not occur in some individuals.
• Complications from HSCT include graft-versus-host disease, failure to reconstitute the humoral immune compartment, graft failure over time, and post-transplant lymphoproliferative disease [Skoda-Smith et al 2001, Dvorak & Cowan 2008, Pai et al 2014].
• Affected individuals with poor humoral reconstitution are maintained on long-term immunoglobulin replacement.

Individuals who do not undergo HSCT require close monitoring for worsening of immune function manifested by increased susceptibility to severe or opportunistic infections (see also Surveillance). If clinical status worsens, rapid transition to HSCT should be considered.

Prevention of Secondary Complications

The following are appropriate:

• Use of irradiated, leukoreduced, cytomegalovirus (CMV)-safe blood products
• Delay of immunizations until immune reconstitution
• Consideration for formula feeds in place of breast feeding until CMV status of mother is known. Caution should be taken to assess the quality of the water source for the infant formula.

Surveillance

Following a successful HSCT, the following should be routinely monitored:

• Growth
• Psychomotor development
• Complete blood counts
• Liver and renal function
• Immune status
• Donor and recipient chimerism
• Development of post-transplant complications, particularly chronic graft-versus-host disease, decreased bone density, pulmonary and cardiac function, and gonadal function

Individuals with milder findings or those who have not undergone HSCT also need to be monitored for worsening of immune function with periodic assessment of clinical status and functional lymphocyte responsiveness.

Agents/Circumstances to Avoid

Individuals with ZAP70-related CID should avoid the following:

• Non-irradiated blood products
• Live virus vaccinations
• Mycobacterium bovis (BCG) vaccine against tuberculosis, Salmonella typhi (Ty21a) vaccine against typhoid fever, and Vibrio cholerae (CVD 103-HgR) vaccine against cholera, which may be part of the routine vaccination schedule in countries where these diseases are endemic
• Contaminated water sources
• Exposure to fungus-enriched environments (e.g., construction sites, agricultural areas with active soil disruption, mulch, hay)

Evaluation of Relatives at Risk

Because the outcome of HSCT in children with ZAP70-related CID is significantly improved by performing HSCT prior to the onset of severe infections, early testing of at-risk sibs should be considered. In addition, any sibs considered as bone marrow donors must be evaluated for ZAP70-related CID prior to donation.

• If the ZAP70 pathogenic variants in the family are known, molecular genetic testing can be used to clarify the genetic status of at-risk sibs.
• If the pathogenic variants in the family are not known, CBC, quantitative immunoglobulins, and lymphocyte subsets and proliferation can be used to clarify the genetic status (immunologic status) of at-risk sibs.

See Genetic Counseling for issues related to testing of at-risk relatives for genetic counseling purposes.

Pregnancy Management

Appropriately screened blood products should be available, if needed, during the course of the pregnancy and delivery.

Therapies Under Investigation

Gene therapy. Gene therapy has not been performed in ZAP70-related CID. Experimental studies utilizing gene therapy have been conducted on murine models [Adjali et al 2005, Irla et al 2008] as well as human cells in vitro [Steinberg et al 2000, Kofler et al 2004, Gavino et al 2017]. Nonviral transfer methods (e.g., electro-gene transfer) have also been used to correct ZAP-70 deficiency in a murine model [Irla et