DiagnosisClinical Diagnosis ZAP70-related severe combined immunodeficiency (ZAP70-related SCID) is a cell-mediated immunodeficiency caused by abnormal T-cell receptor (TCR) signaling leading to a selective absence of CD8⁺ T cells and normal or elevated numbers of non-functional CD4⁺ T cells [Arpaia et al 1994, Chan et al 1994, Elder et al 1994].Children usually present in the first year of life with failure to thrive and recurrent viral, bacterial, and opportunistic infections.TestingThe diagnosis of ZAP70-related SCID relies on lymphocyte subset analysis of CD3, CD4, and CD8 T cells, lymphocyte function testing, ZAP-70 protein expression, and ZAP70 molecular genetic testing. Lymphocyte counts and lymphocyte cell surface expression. In ZAP70-related SCID, total lymphocyte counts can range from normal to high. T cell countsCD8⁺ cells are absent or cell counts are very low. Note: T cells expressing CD8⁺ make up 0%-2% of the child’s total T-cell count [Monafo et al 1992, Arpaia et al 1994, Elder et al 1995, Gelfand et al 1995, Matsuda et al 1999, Noraz et al 2000]. CD4⁺ cell counts are normal or elevated. Note: CD4⁺ cells account for 60%-80% of mononuclear cells in the lymphocyte count of individuals with ZAP70-related SCID.CD3⁺ cell counts are normal. Note: Most CD3⁺ cells are composed of CD4⁺ cells in ZAP70-related SCID.B cell counts and NK cell counts. NormalLymphocyte function. T-cell responses to stimuli that act through the T-cell receptor (TCR) are absent or severely diminished:Absence of proliferation of CD4⁺ cells in response to mitogens (e.g., PHA)Absence of proliferation of CD4⁺ cells in response to antigens (e.g., ConA) Defective CD4⁺ cell-cell activation manifest as impaired Ca²⁺ flux in response to CD3 (OKT3) stimulation [Arpaia et al 1994, Chan et al 1994, Gelfand et al 1995, Elder et al 2001, Turul et al 2009]Note: T-cell responses to phorbol myristic acetate and ionomycin stimulation (which bypasses the TCR) are normal [Elder et al 1994, Elder 1997].ZAP-70 protein expression. Immunocytochemistry testing of CD4⁺ T cells reveals absence of ZAP-70 protein in most cases. Note: (1) Two reports have described defects leading to protein expression with either rapid protein degradation [Matsuda et al 1999] or no catalytic function [Elder et al 2001]. (2) One report describes a hypomorphic ZAP70 mutation leading to decreased protein expression and function with late-onset combined immunodeficiency [Picard et al 2009]. Immunoglobulin concentrations and functionImmunoglobulin levels vary by individual. A majority of affected individuals have severe hypogammaglobulinemia, but hypergammaglobulinemia and normal immunoglobulin levels have been seen [Turul et al 2009].Although functional antibody responses to immunization are present in a few persons [Turul et al 2009], this finding does not indicate that all specific antigenic responses are intact. Molecular Genetic Testing Gene. ZAP70 is the only gene in which mutations cause ZAP70-related severe combined immunodeficiency.Clinical testing Table 1. Summary of Molecular Genetic Testing Used in ZAP70-Related Severe Combined ImmunodeficiencyView in own windowGene SymbolTest MethodMutations DetectedMutation Detection Frequency by Test Method ^{1Test AvailabilityZAP70Sequence analysisSequence variants 215/15 3 Clinical1. The ability of the test method used to detect a mutation that is present in the indicated gene2. Examples of mutations detected by sequence analysis may include small intragenic deletions/insertions and missense, nonsense, and splice site mutations; typically, exonic or whole-gene deletions/duplications are not detected.3. At least 15 individuals meeting diagnostic criteria have had identifiable ZAP70 mutations [Arpaia et al 1994, Chan et al 1994, Elder et al 1994, Matsuda et al 1999, Noraz et al 2000, Elder et al 2001, Meinl et al 2001, Toyabe et al 2001, Picard et al 2009, Turul et al 2009]. Interpretation of test results. For issues to consider in interpretation of sequence analysis results, click here.Information on specific allelic variants may be available in Molecular Genetics (see Table A. Genes and Databases and/or Pathologic allelic variants).Testing Strategy To confirm/establish the diagnosis in a proband, the following evaluations should be considered: Lymphocyte cell surface expression of CD3, CD4, and CD8Lymphocyte functional response to mitogens, antigens, and biochemical agents like phorbol myrsitate acetate and ionomycineImmunohistochemistry for presence of ZAP-70 proteinImmunoglobulin level and specific antibody responsesMolecular genetic testing for mutations in ZAP70 may be helpful to confirm a diagnosis due to the clinical heterogeneity of ZAP70-related SCID [Turul et al 2009] (see Testing).Newborn screening. The use of routine newborn screening for T– SCID by measuring T-cell receptor excision circle (TREC) levels continues to increase. Despite normal quantitative levels of CD4 cells in individuals with ZAP70-related SCID, TREC levels in already known cases of ZAP70-related SCID were found to be very low compared to age-matched controls [Roifman et al 2010], suggesting that detection of ZAP70-related SCID may be possible through newborn screening methods utilizing TREC quantitation. Carrier testing for at-risk relatives requires prior identification of the disease-causing mutations in the family. Note: Carriers are heterozygotes for this autosomal recessive disorder and are not at risk of developing the disorder.Prenatal diagnosis and preimplantation genetic diagnosis (PGD) for at-risk pregnancies require prior identification of the disease-causing mutations in the family.Genetically Related (Allelic) DisordersNo other phenotypes are known to be associated with mutations in ZAP70.}

Natural History Individuals with ZAP70-related SCID usually present in the first year of life with recurrent bacterial, viral, and opportunistic infections, diarrhea, and failure to thrive. Severe lower respiratory infections are typically seen, most notably Pneumocystis jiroveci infections and viral infections. Oral moniliasis is common. Whereas the above presentation is characteristic of ZAP70-related SCID, there are exceptions:Reports of milder phenotypes in sibs of children who had died from ZAP70-related SCID 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]. Picard et al [2009] described a child age nine years with a ZAP70 hypomorphic intronic mutation and an attenuated clinical and immunologic phenotype (see Genotype-Phenotype Correlations). Newell et al [2011] reported a child age 11 months with ZAP70-related SCID who presented with lymphoma. Santos et al [2010] reported ZAP70 defects in cousins (ages 5 and 6 months) presenting as axillary lymphadenitis following BCG vaccine. Other presenting findings [Elder et al 1995, Parry et al 1996, Turul et al 2009]: Subcutaneous nodulesLymphadenopathyExfoliative dermatitisThrombocytopeniaChronic gastroenteritisUlcerative colitis The long-term prognosis of untreated ZAP70-related SCID is death from infection. Affected children have a declining quality of life and usually do not survive past their second year without HSCT. A long-term study found that following HSCT, three-year survival rates were 77% and 54% for HLA-identical and HLA-mismatched transplants respectively [Antoine et al 2003, Müller & Friedrich 2005]. Note: Survival rates given are for cohorts that comprise various forms of SCID, including ZAP70-related SCID; statistical outcomes specifically for ZAP70-related SCID are unknown. Children with preexisting viral infections are at increased risk of developing graft-versus-host disease (GVHD) following HSCT, leading to a poor prognosis [Dvorak & Cowan 2008].

Differential DiagnosisInfants positive for human immunodeficiency virus (HIV+) may present with recurring infections and failure to thrive similar to SCID. Individuals with HIV have CD4⁺ lymphopenia, in contrast to the CD8⁺ lymphopenia in ZAP70-related SCID. In a neonate the definitive diagnosis of HIV should be made by detection of cell-associated human immunodeficiency proviral DNA by polymerase chain reaction (PCR) amplification. See Table 2.Table 2. Combined Immunodeficiencies in the Differential Diagnosis of ZAP70-Related SCIDView in own windowDisease NameGene InvolvedMode of InheritanceLymphocyte PhenotypeTBNKOtherFamilial CD8 deficiencyCD8AAR+++CD4⁺/CD8^– CD25 deficiency IL2RAAR+++CD4^–/CD8^{+MHC II deficiency (BLS)See Major histocompatibility complex (below) AR+++CD4–/CD8+MHC II = major histocompatibility complex class II BLS = bare lymphocyte syndromeFamilial CD8 deficiency (OMIM 608957) may have a presentation similar to ZAP70-related SCID, but the diagnosis can be confirmed with CD8A molecular genetic testing. The two individuals reported with this disease had recurring infections from early childhood and lived past their twenties [de la Calle-Martin et al 2001, Mancebo et al 2008]. CD25 deficiency (OMIM 606367) also presents with recurring infections early in life with low to normal T-cell counts. However, the T cells are CD4–/CD8+. The diagnosis can be confirmed with molecular genetic testing of IL2RA (CD25), which encodes the interleukin-2 receptor alpha chain. Major histocompatibility complex (MHC) class II deficiency (also known as bare lymphocyte syndrome) (OMIM 209920) may have normal or elevated T-cell counts; however, the T cells are CD4–/CD8+. As in other forms of SCID, pathologic findings manifest within the first year of life. Major histocompatibility complex II expression is decreased. Molecular genetic testing may reveal mutations in RFX5, RFXAP, MHC2TA, or RFXANK, the four genes in which mutation is known to cause this disorder. Table 3 differentiates several forms of severe combined immunodeficiency. Since SCID presents as a phenotypically heterogeneous class of diseases, it is useful to recognize forms of SCID that present with low to normal T-cell counts. Lymphocyte subset testing and molecular genetic testing can implicate or eliminate these other forms of SCID. Table 3. T-Cell-Negative Forms of SCID in the Differential Diagnosis of ZAP70-Related SCIDView in own windowDisease NameGene InvolvedDefectMode of InheritanceLymphocyte PhenotypeTBNKZAP70-related SCIDZAP70Decreased protein expressionAR+++JAK3-related SCIDJAK3AR–+–IL7RA-related SCIDIL7RAAR–++CD45 deficiencyCD45AR–+–ADA deficiencyADADecreased protein productionAR–––RAG1/2 deficiency RAG1, RAG2AR––+SCID AthabascanARTEMISAR––+X-linked SCIDIL2RGDysfunctional receptorXLR–+–Omenn syndrome (OMIM 603554). Two children with ZAP70-related SCID presented with an Omenn syndrome-like phenotype that included lymphadenopathy, hepatosplenomegaly, and eosinophilia. Lymphocyte subset tests consistent with ZAP70-related SCID in both cases eliminated Omenn syndrome as a possible diagnosis. Note to clinicians: For a patient-specific ‘simultaneous consult’ related to this disorder, go to , an interactive diagnostic decision support software tool that provides differential diagnoses based on patient findings (registration or institutional access required).}

ManagementEvaluations Following Initial Diagnosis To establish the extent of disease in an individual diagnosed with ZAP70-related severe combined immunodeficiency (SCID), the following evaluations are recommended:Evaluation for common and opportunistic viral, bacterial, and fungal disease-causing agentsAssessment of growthComplete metabolic panel (liver and renal function) and CBC with differential and platelet countMedical genetics consultationImmunology consultation, if not performed alreadyTreatment of ManifestationsTreatment relies on prompt reconstitution of the individual’s immune system (see Prevention of Primary Manifestations). Short-term treatment includes immediate intravenous immunoglobulin (IVIG) and antibacterial, antifungal, and anti-protozoal prophylaxis to control and reduce the occurrence of infections. Prevention of Primary ManifestationsThe standard of therapy to cure SCID is allogeneic hematopoietic stem cell transplantation (HSCT). The outcome of HSCT in children with SCID is significantly improved by performing HSCT in the first three months of life [Buckley 2004]. Several children with ZAP70-related SCID have been transplanted [Arpaia et al 1994, Elder et al 1994, Elder et al 2001, Noraz et al 2000]. Outcomes are the best with HLA-matched, related donors.If a related, HLA-matched donor is not available, alternatives include:Matched unrelated donorUmbilical cord blood donorHaploidentical parental bone marrow or mobilized peripheral blood stem cells that have been T cell depletedIn contrast to individuals with other forms of SCID, individuals with ZAP70-related SCID are typically treated with a chemotherapeutic conditioning regimen prior to HSCT. Hönig et al [2012] described the successful use of lymphocyte transfusion from a previously transplanted HLA identical sib without the use of conditioning for reconstituting the immune system in an individual with ZAP70-SCID. Cellular reconstitution following HSCT takes three to four months and restoration of humoral immunity can take one to two years or more.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].Affected individuals with poor humoral reconstitution are maintained on long-term immunoglobulin replacement. Individuals with mild initial findings are maintained on immunoglobulin replacement and prophylactic antimicrobial therapy. They need to be monitored for worsening of immune function manifest by increased susceptibility to severe or opportunistic infections (see also Surveillance). If clinical status worsens, curative HSCT should be considered. Prevention of Secondary ComplicationsThe following are appropriate:Use of irradiated, CMV-negative blood products
Note: While not routinely screened for, the use of blood products from a known Epstein-Barr virus (EBV)-negative source should be considered as EBV-related lymphoma has been described in ZAP70-SCID [Newell et al 2011].Delay of immunizations until immune reconstitution SurveillanceFollowing a successful HSCT, the following should be monitored every six to 12 months: Immune status Liver and renal function Complete blood count Growth Psychomotor development Individuals with milder findings need to be monitored for worsening of immune function with at least semiannual assessment of clinical status and functional lymphocyte responsiveness.Agents/Circumstances to AvoidIndividuals with ZAP70-related SCID should never receive 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 endemicEvaluation of Relatives at RiskBecause the outcome of HSCT in children with SCID is significantly improved by performing HSCT in the first three months of life, early testing of at-risk sibs should be considered. See Genetic Counseling for issues related to testing of at-risk relatives for genetic counseling purposes.Pregnancy ManagementAn affected pregnant mother should receive prenatal counseling. Appropriately screened blood products should be available, if needed, during the course of the pregnancy or delivery.Therapies Under InvestigationGene therapy. While gene therapy has been used for other forms of SCID (notably ADA deficiency and X-linked SCID), it has not been performed in ZAP70-related SCID. Experimental studies utilizing gene therapy for this disease 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]. Adverse oncogenic reactions have been documented in some individuals with X-linked SCID transduced with retroviral vector [Buckley 2003, Fischer et al 2005]. The role of nonviral transfer methods (e.g., electro-gene transfer) have been used to correct ZAP-70 deficiency in a murine model [Irla et al 2008]. Search ClinicalTrials.gov for access to information on clinical studies for a wide range of diseases and conditions.

Molecular GeneticsInformation in the Molecular Genetics and OMIM tables may differ from that elsewhere in the GeneReview: tables may contain more recent information. —ED.Table A. ZAP70-Related Severe Combined Immunodeficiency: Genes and DatabasesView in own windowGene SymbolChromosomal LocusProtein NameLocus SpecificHGMDZAP702q11​.2Tyrosine-protein kinase ZAP-70Resource of Asian Primary Immunodeficiency Diseases (RAPID)
ZAP70 homepage - Mendelian genesZAP70Data are compiled from the following standard references: gene symbol from HGNC; chromosomal locus, locus name, critical region, complementation group from OMIM; protein name from UniProt. For a description of databases (Locus Specific, HGMD) to which links are provided, click here.Table B. OMIM Entries for ZAP70-Related Severe Combined Immunodeficiency (View All in OMIM) View in own window 176947ZETA-CHAIN-ASSOCIATED PROTEIN KINASE; ZAP70 269840SELECTIVE T-CELL DEFECT; STCDNormal allelic variants. ZAP70 spans 26.3 kb of genomic DNA. The gene consists of 14 exons comprising 2450 bp. Pathologic allelic variants. ZAP70 pathologic allelic variants reside mostly in the kinase domain, although mutations that result in loss of transcription or are located in the N-terminal SH2 domain and result in rapid degradation of ZAP-70 protein have been reported [Matsuda et al 1999, Au-Yeung et al 2009]. More than a dozen mutations consisting of single point mutations, splice defects, and intragenic deletions have been reported. A mutation in the arginine residue (p.Arg465Cys) of the DLAARN motif of the kinase domain has been described (see Abnormal gene product). Selected mutations can be viewed in Table 4 and Table A. Details on other mutations may also be found in the review article Wang et al [2010] and in Fischer et al [2010].Table 4. Selected ZAP70 Pathologic Allelic VariantsView in own windowDNA Nucleotide Change
(Alias ¹) Protein Amino Acid ChangeReference Sequencesc.239C>A
(448C>A)p.Pro80GlnNM_001079​.3
NP_001070​.2c.837+121G>A
(836+121G>A)See Abnormal gene productc.1393C>Tp.Arg465Cysc.1714A>T
(1923A>T)p.Met572LeuSee Quick Reference for an explanation of nomenclature. GeneReviews follows the standard naming conventions of the Human Genome Variation Society (www​.hgvs.org).1. Variant designation that does not conform to current naming conventionsNormal gene product. ZAP70 codes for an enzyme in the Syk-protein tyrosine kinase family that plays a role in T-cell development and activation. This enzyme is phosphorylated at tyrosine residues upon T-cell receptor (TCR) stimulation and functions in the initial step of TCR-mediated signal transduction with Src family kinases. ZAP-70 comprises 619 amino acids and contains two SH2 domains and one kinase domain. The mutations that lead to ZAP70-related SCID occur in or close to the region coding the kinase domain. Abnormal gene product. Most noted mutations affect the kinase domain of ZAP70 and cause a lack of protein expression. The p.Arg465Cys alteration in the highly conserved DLAARN motif of the kinase domain compromises ZAP-70 protein stability and eliminates the protein’s catalytic function [Elder et al 2001]. Both the p.Pro80Gln and p.Met572Leu altered proteins undergo temperature-sensitive degradation [Matsuda et al 1999]. Picard et al [2009] described a hypomorphic mutation in ZAP70 intron 7 (c.837+121G>A) that creates a new in-frame splice product with a new stop codon within intron 7. In addition to the truncated product, the mutation allowed residual expression of the wild type protein (20% expression level in the patient’s T cells) and an attenuated clinical and immunologic phenotype (see Genotype-Phenotype Correlations) [Picard et al 2009].