Adenosine Deaminase Deficiency

Clinical characteristics.

Adenosine deaminase (ADA) deficiency is a systemic purine metabolic disorder that primarily affects lymphocyte development, viability, and function. The clinical phenotypic spectrum includes:

• Severe combined immunodeficiency disease (SCID), often diagnosed by age six months and usually by age 12 months;
• Less severe "delayed" onset combined immune deficiency (CID), usually diagnosed between age one and ten years;
• "Late/adult onset" CID, diagnosed in the second to fourth decades;
• Benign "partial ADA deficiency" (very low or absent ADA activity in erythrocytes but greater ADA activity in nucleated cells), which is compatible with normal immune function.

Infants with typical early-onset ADA-deficient SCID have failure to thrive and opportunistic infections associated with marked depletion of T, B, and NK lymphocytes, and an absence of both humoral and cellular immune function. If immune function is not restored, children with ADA-deficient SCID rarely survive beyond age one to two years. Infections in delayed- and late-onset types (commonly, recurrent otitis, sinusitis, and upper respiratory) may initially be less severe than those in individuals with ADA-deficient SCID; however, by the time of diagnosis these individuals often have chronic pulmonary insufficiency and may have autoimmune phenomena (cytopenias, anti-thyroid antibodies), allergies, and elevated serum concentration of IgE. The longer the disorder goes unrecognized, the more immune function deteriorates and the more likely are chronic sequelae of recurrent infection.

Diagnosis/testing.

The diagnosis of ADA deficiency is established in a proband:

• With <1% of normal ADA catalytic activity in hemolysates (in untransfused individuals) or in extracts of other cells (e.g., blood mononuclear cells, fibroblasts); AND/OR
• By the identification of biallelic pathogenic variants in ADA by molecular genetic testing.

Management.

Treatment of manifestations: Infections are treated with specific antibiotic, antifungal, and antiviral agents and administration of intravenous immunoglobulin (IVIg); prophylaxis is provided for Pneumocystis jiroveci infection.

Prevention of primary manifestations: Restoration of a functional immune system is essential. The preferred treatment is bone marrow/stem cell transplantation (BMT/SCT) from an HLA-identical healthy sib or close relative. However, most individuals with ADA-deficient SCID lack an HLA-identical related donor. For these individuals, alternative therapies can be considered:

• BMT/SCT from a "non-ideal" donor, which may be an HLA-matched unrelated donor, an HLA-haploidentical donor (usually a parent), or umbilical cord-derived stem cells
• Enzyme replacement therapy (ERT) with polyethylene glycol-modified bovine adenosine deaminase (PEG-ADA)
• Gene therapy, which is currently experimental

Surveillance: Annual or more frequent evaluation of lymphocyte counts, serum immunoglobulin levels, and various in vitro tests of cellular and humoral immune function following BMT/SCT and during ERT (more frequent monitoring and other specialized testing would be required for participants in gene therapy trials). Individuals on ERT also require periodic monitoring of PEG-ADA levels in plasma and metabolite levels in erythrocytes, and under some circumstances testing for anti-ADA antibodies.

Agents/circumstances to avoid: The use of adenine arabinoside (a substrate for ADA) as an antiviral agent or for chemotherapy of malignancies should be avoided; pentostatin, a potent ADA inhibitor used to treat some lymphoid malignancies, would be ineffective in persons who lack ADA, and would interfere with PEG-ADA.

Evaluation of relatives at risk: In the newborn sibs of a proband, it is appropriate to either assay ADA catalytic activity or perform molecular genetic testing (if the family-specific pathogenic variants are known), so that morbidity and mortality can be reduced by early diagnosis and treatment.

Genetic counseling.

ADA deficiency 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 an asymptomatic carrier, and a 25% chance of being unaffected and not a carrier. Carrier testing for at-risk family members and prenatal testing for pregnancies at increased risk are possible once the pathogenic variants have been identified in the family.

Suggestive Findings

Adenosine deaminase (ADA) deficiency should be suspected in individuals with the following newborn screening results, clinical findings (by age), and supportive laboratory findings.

Newborn screening

• In newborns found to have reduced T cell receptor excision circles (TRECs), particularly if follow-up lymphocyte subset analysis reveals a deficiency of T, B, and NK lymphocytes (T-B- NK- phenotype), biochemical testing for ADA deficiency should be performed.
• Historically, about 15% of all cases of SCID result from ADA deficiency, but the frequency among newborns identified as having SCID by screening for a reduced number of TRECs is not yet known.

Note: (1) Individuals with delayed or late/adult ADA deficiency may have TRECs in the normal range. (2) Reduced levels of a B lymphocyte marker, kappa-deleting recombination excision circles (KRECs), have also been found in DNA from dried blood spots (DBS) of individuals with delayed- or late-onset ADA deficiency [Speckmann et al 2012].

Clinical findings

• Infancy (severe combined immunodeficiency disease or SCID phenotype):
  • Failure to thrive
  • Absence of lymphoid tissues (tonsils, lymph nodes)
  • Opportunistic infections
  • Persistent diarrhea
  • Extensive dermatitis
  • Recurrent pneumonia
• Childhood ("delayed" onset combined immune deficiency [CID]) and adulthood ("late/adult onset" CID):
  • Recurrent otitis media
  • Sinusitis
  • Upper respiratory infections
  • Chronic pulmonary insufficiency
  • Allergies or autoimmunity

Supportive laboratory findings

• Immune function
  • Lymphopenia is present at birth. The total blood lymphocyte count is usually <500/µL (normal for neonates: 2,000 to >5,000).
  • All major lymphoid lineages (T-, B-, and NK-cells) are depleted as demonstrated by flow cytometry.
  • In vitro lymphocyte function, as measured by proliferative response to mitogens and antigens, is low or absent.
  • Serum immunoglobulins are low and no specific antibody response to infections and immunizations is observed. However, those with a delayed or late-onset phenotype may have elevated serum IgE levels.
• Elevated deoxyadenosine (dAdo) triphosphate (dATP) or total dAdo nucleotides (dAXP, measured as the sum of dAMP+dADP+dATP) in erythrocytes
  • Marked increase in dATP and total dAXP, a finding pathognomonic for ADA deficiency.
  • In untreated affected individuals, the level of dAXP in hemolysates or dried blood spot (DBS) extracts correlates with clinical phenotype (SCID > "delayed/late" onset > "partial ADA deficiency").
  • Even after transfusion, some elevation in erythrocyte dAXP persists and strongly indicates underlying ADA deficiency.
• Reduced S-adenosylhomocysteine hydrolase (AdoHcyase, SAHase) activity in erythrocytes, typically <5% of normal
• Elevated urinary dAdo in untreated affected individuals
• Elevated levels of adenosine (Ado) and dAdo in extracts of dried blood spots (DBS), quantified using tandem mass spectrometry [Azzari et al 2011, Speckmann et al 2012, la Marca et al 2013]

Establishing the Diagnosis

The diagnosis of ADA deficiency is established in a proband:

• With <1% of normal ADA catalytic activity in hemolysates or in extracts of dried blood spots (DBS) prepared with EDTA or heparin-anticoagulated blood; AND/OR
• By the identification of biallelic pathogenic variants in ADA on molecular genetic testing (see Table 1).

Note: (1) In recently transfused individuals, a deficiency of ADA catalytic activity can be demonstrated in extracts of non-erythroid cells (e.g., blood mononuclear cells, fibroblasts). (2) Analysis of ADA enzymatic activity in plasma is not useful for diagnosis because ADA activity is much lower in plasma than in cells, even in normal (non ADA-deficient) individuals. Additionally, plasma contains adenosine deaminase activity as a result of another enzyme known as ADA2 (see Differential Diagnosis).

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

• Single-gene testing. Sequence analysis of ADA is performed first and followed by gene-targeted deletion/duplication analysis if only one or no pathogenic variant is found.
• Multigene panel. A panel that includes ADA 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 vary 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

Adenosine deaminase (ADA) deficiency is a systemic purine metabolic disorder that primarily affects lymphocyte development, viability, and function; various effects on several non-lymphoid organs are also observed in some affected individuals [Hirschhorn 1999, Hershfield & Mitchell 2001, Hershfield 2004, Whitmore & Gaspar 2016]. The phenotypic spectrum associated with ADA deficiency includes early-onset SCID diagnosed in infancy; less severe "delayed/late" -onset combined immune deficiency (CID) diagnosed in older children and adults; and benign "partial ADA deficiency," which is discovered by screening populations or relatives of individuals with SCID for deficiency of erythrocyte ADA activity.

Genotype-Phenotype Correlations

Most known ADA pathogenic variants have been discovered through research into the relationship of genotype to phenotype [Hirschhorn et al 1990, Santisteban et al 1993, Arredondo-Vega et al 1994, Ozsahin et al 1997].

• A good correlation has been established between the effect of pathogenic variants on ADA activity and both clinical and metabolic phenotype [Arredondo-Vega et al 1998].
• In several individuals, the relationship of genotype to phenotype was modulated by mosaicism in lymphoid cells for reversion of deleterious pathogenic variants [Hirschhorn et al 1994, Hirschhorn et al 1996, Ariga et al 2001a, Arredondo-Vega et al 2002, Liu et al 2009, Moncada-Vélez et al 2011].

Systematic expression in E coli of more than 30 cDNAs with single missense variants identified in ADA-deficient individuals has shown that the total ADA activity expressed by both of an individual's mutated alleles correlates with age at diagnosis and the level of erythrocyte dAXP measured prior to treatment [Arredondo-Vega et al 1998]. A system for ranking the severity of genotypes has been proposed based on these data and the potential of other alleles to provide ADA activity. For this purpose, individual pathogenic ADA alleles are clustered in groups, as follows:

• Group 0. "Null" alleles (deletion, frameshifting, or nonsense variants)
• Groups I-IV. Missense variants ranked in order of increasing activity expressed in the E coli system
• Splice site variants. A separate group, as a low level of normal splicing may result in variable levels of ADA activity

Phenotype correlation with pathogenic variant type was assessed for 52 clinically diverse individuals with 43 genotypes composed of 42 different pathogenic alleles [Arredondo-Vega et al 1998]:

• ADA-deficiency SCID. Both alleles scored as Group 0 or I.
• Delayed-/late-onset ADA deficiency. At least one allele in Group II or III was detected.
• Partial ADA deficiency. At least one Group IV allele was detected.

Discordance in phenotype among first-degree ADA-deficient relatives in several families has been attributed to the following:

• Individual differences in "leakiness" of a splice site variant [Arredondo-Vega et al 1994]
• Mosaicism for reversion of a pathogenic allele in lymphoid cells [Arredondo-Vega et al 2002]
• The segregation of both alleles that cause a severe phenotype and a mild phenotype in a family [Santisteban et al 1995, Ozsahin et al 1997, Ariga et al 2001b]

Prevalence

ADA deficiency has been estimated to occur in from 1:200,000 to 1:1,000,000 births. As information from newborn screening becomes available, estimates of the incidence of ADA deficiency may change.

All racial and ethnic groups are affected. Prevalence is higher in some geographic areas where a high degree of consanguinity exists in certain population groups.

Evaluations Following Initial Diagnosis

To establish the extent of disease and needs in an individual diagnosed with adenosine deaminase (ADA) deficiency, the following evaluations are recommended, some of which may be performed as part of the diagnostic evaluation:

• Identification of specific disease-causing viral, fungal, or bacterial organisms (both normal pathogens and opportunistic agents)
• Complete blood count (CBC)
• Flow cytometry to quantify lymphocyte subsets (T-, B-, NK-cells)
• Assessment of humoral immune function by measuring serum immunoglobulins and the titer of specific antibodies related to infections and immunizations
• Evaluation of cellular immune function by in vitro response of blood mononuclear cells to mitogens and antigens
• Measurement of erythrocyte dAXP concentration to evaluate metabolic severity
• Liver function testing to assess for metabolic hepatitis
• Auditory testing
• Consultation with a clinical geneticist and/or genetic counselor

Treatment of Manifestations

The following are appropriate:

• Treatment of infections with specific antibiotic, antifungal, and antiviral agents
• Prophylaxis for Pneumocystis jiroveci
• Intravenous immunoglobulin (IVIg)

Prevention of Primary Manifestations

Restoring a functional immune system is essential and can be achieved in several ways. The choice of therapy is complex and depends on a number of factors, including the affected person’s age and clinical status, the expectations and desires of the parents, and the specific experience and expertise of physicians in treating ADA-deficient SCID. A workshop held in 2008 developed consensus guidelines for therapy [Gaspar et al 2009] (full text).

Bone marrow/stem cell transplantation (BMT/SCT) from an HLA-identical healthy sib is the method of choice for treating all forms of SCID.

• This can be performed without cytoreductive conditioning of the affected individual, and without depletion of donor T-cells.
• Results vary among transplant centers, but the procedure is curative in approximately 70% or more of affected individuals.
• The main risks are graft-versus-host disease and delayed or incomplete recovery of humoral immune function, requiring continued immunoglobulin replacement.

For the majority of individuals with ADA-deficient SCID who lack an HLA-identical related donor, the following two forms of treatment can be considered [Gaspar et al 2009, Gaspar 2010, Candotti et al 2012, Baffelli et al 2015].

BMT/SCT from a "non-ideal" donor

• Donor-derived T-cells are depleted to minimize the risk of graft-versus-host disease.
• Pre-transplant cytoreductive "conditioning" of the recipient (individual with SCID) is often performed to prevent graft loss, which occurs with relative frequency in those with ADA-deficient SCID who are not conditioned.
  Note: Some transplant centers do not perform conditioning of the recipient prior to a haploidentical transplant because of the risk of peri-transplant morbidity [Buckley et al 1999]. However, this latter approach has frequently been associated with a failure to achieve stable engraftment [Gaspar et al 2009, Gaspar 2010, Hassan et al 2012].

Following a T-cell-depleted transplant, return of functional T-cells requires three to four months. B-cell reconstitution is delayed longer, or may not be adequately achieved, requiring long-term immunoglobulin replacement therapy.

Note: Universal agreement regarding the best methods for performing partially mismatched BMT/SCT does not exist [Cancrini et al 2010, Gaspar 2010, Hassan et al 2012]. When considering therapeutic options, it is therefore important for parents to obtain specific information about prior experience and long-term results of transplants for ADA-deficient SCID at the center where their child will be treated.

Enzyme replacement therapy (ERT) has been used as a primary therapy in individuals who lack an HLA-identical marrow/stem cell donor when the risks associated with a partially mismatched transplant are deemed too great or when the risk of graft failure is high, as in older individuals with a delayed- or late-onset phenotype.

• Polyethylene glycol-modified bovine adenosine deaminase (PEG-ADA) is composed of purified bovine ADA covalently linked to multiple strands of PEG (average mass: 5 kd) in order to prolong circulating life and reduce immunogenicity. It is administered by intramuscular injection once or twice a week (~15-60 U/kg per week).
• By maintaining a high level of ADA activity in plasma, PEG-ADA eliminates extracellular Ado and dAdo, preventing the toxic metabolic effects that interfere with lymphocyte viability and function and that may injure other organs (liver, lung, brain) [Hershfield et al 1987, Hershfield & Mitchell 2001, Hershfield 2004, Gaspar et al 2009].
• ERT is not curative; PEG-ADA must be given regularly and at a sufficient dose to maintain a nontoxic metabolic environment.
• PEG-ADA has also been used as a secondary therapy in affected individuals who have failed to engraft following an unconditioned BMT/SCT, or in whom an acceptable recovery of immune function has not been achieved following experimental gene therapy.

Most individuals treated with PEG-ADA recover partial immune function that is sufficient to prevent opportunistic infections and other clinical manifestations of SCID. A lag of approximately two to four months occurs before T-cell function appears, but B-cells often increase earlier than after BMT/SCT. Lymphocyte counts and in vitro lymphocyte function usually increase during the first year of ERT, but beyond the first year or two most PEG-ADA-treated individuals remain lymphopenic and in vitro lymphocyte function fluctuates widely. Most individuals remain clinically well, but over time both T and B lymphocytes gradually decline in number and display various functional abnormalities [Chan et al 2005, Malacarne et al 2005, Serana et al 2010, Brigida et al 2014]. Approximately half of those maintained on ERT were continuing to receive immunoglobulin replacement.

More than 300 affected individuals have received PEG-ADA. Survival of PEG-ADA-treated individuals beyond five years and through approximately ten years is approximately 75%-80%, comparable or superior to that achieved with BMT/SCT (i.e. in individuals who lack an HLA-identical donor). Most deaths occurred during the first six months of treatment, with the majority in the first month due to life-threatening infections present at diagnosis.

Lymphoproliferative disorders have developed in eight individuals who received PEG-ADA for eight to 22 years [Hershfield 2004; Chan et al 2005; Kaufman et al 2005; Husain et al 2007; Author, unpublished data]. Hepatocellular carcinoma developed in one affected individual after 13 years of ERT, and was present in another at the time ERT was initiated following an unsuccessful stem cell transplant. A third affected individual died of hepatoblastoma after 2.5 years of ERT; the tumor was thought to have been present but undetected prior to ERT. Several other affected individuals have developed persistent hemolytic anemia, which in some cases began in association with a viral infection or with central catheter sepsis [Hershfield 2004, Lainka et al 2005].

The limitations of PEG-ADA therapy include primary failure to recover protective immune function, the development of neutralizing antibodies that reduce or eliminate efficacy, immune dysregulation (particularly in the first few months of therapy), and a risk that immune function will eventually (i.e., beyond 10-15 years) decline to an inadequate level. Approximately 20% of affected individuals have discontinued ERT in order to undergo BMT/SCT. In most of these cases, the transplant had been intended at the time of diagnosis but not performed because a suitable donor was not available or the affected individual had been too ill to undergo the procedure. In a minority of individuals, the transplant was performed because of declining immune function while receiving PEG-ADA. Overall, approximately half of these secondary transplants have been successful [Hershfield 2004, Gaspar et al 2009].

Most individuals treated with PEG-ADA for longer than a year develop antibodies that bind specifically to bovine ADA and are detectable by an enzyme-linked immunosorbent assay (ELISA); these are of no clinical significance. Neutralizing antibodies that inhibit catalytic activity and enhance clearance of PEG-ADA (and which do compromise efficacy) have developed in fewer than 10% of treated individuals [Chaffee et al 1992, Hershfield 1997]. No allergic or hypersensitivity reactions to PEG-ADA have occurred, and the treatment has generally been well tolerated.

Gene therapy, while still technically experimental, is still another treatment option for those who have failed all other options (see Therapies Under Investigation).

Prevention of Secondary Complications

As noted under Treatment of Manifestations, affected individuals receive antibiotic prophylaxis for Pneumocystis, and also IVIG, prior to immune reconstitution. The use of such prophylaxis following transplantation and while receiving ERT varies and depends on the level of immune reconstitution achieved.

Surveillance

Annual or more frequent evaluation of lymphocyte counts, serum immunoglobulin levels, and various in vitro tests of cellular and humoral immune function (i.e., as listed above for the evaluation of individuals suspected of having SCID) should be performed following BMT/SCT and during ERT [Gaspar et al 2009].

Individuals on ERT also require periodic monitoring as follows:

• Plasma levels of PEG-ADA activity
• Erythrocyte dAXP concentration
• Development of neutralizing antibodies, particularly if plasma ADA activity or clinical or immunologic status declines unexpectedly
• Monitoring for the appearance or recurrence of dermatofibrosarcoma protuberans

Agents/Circumstances to Avoid

The use of adenine arabinoside (a substrate for ADA) as an antiviral agent or for chemotherapy of malignancies should be avoided.

Pentostatin, a potent ADA inhibitor used to treat some lymphoid malignancies, would be ineffective in persons who lack ADA, and would interfere with PEG-ADA.

Evaluation of Relatives at Risk

It is appropriate to evaluate newborn sibs of a proband so that morbidity and mortality can be reduced by early diagnosis and treatment. Evaluations can include:

• Molecular genetic testing if the pathogenic variants in the family are known;
• Assay of ADA catalytic activity if the pathogenic variants in the family are not known.

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

Therapies Under Investigation

Experimental gene therapy for ADA-deficient SCID employing gamma retroviral vectors has been under clinical investigation for more than 20 years [Engel et al 2003, Cavazzana-Calvo et al 2005]. Clinical trials since about 2000 have employed an approach that was first reported for two affected individuals treated in Milan, Italy and later updated for ten others [Aiuti et al 2002, Aiuti et al 2009]. This strategy involved discontinuing PEG-ADA (in those receiving ERT) and administering non-myeloablative conditioning prior to the infusion of vector carrying human ADA cDNA in order to transduce autologous CD34+ stem cells. Variations on this protocol have also been under investigation in the UK, US, and Japan. More than 40 affected individuals (most of whom had been receiving PEG-ADA) have been treated at these centers [Aiuti et al 2002, Gaspar et al 2006, Engel et al 2007, Aiuti et al 2009, Cappelli & Aiuti 2010, Selleri et al 2011, Candotti et al 2012, Sauer et al 2012, Gaspar et al 2013]. In most treated individuals, stable ADA expression in lymphoid cells has been achieved, along with correction of metabolic abnormalities in erythrocytes, which has been accompanied by reconstitution of both T and B cell immune function, although this reconstitution may take a year or more. Good health has been maintained without the need for ERT.

In contrast to the experience with gene therapy for X-linked SCID, no individuals with ADA deficiency have developed leukemia as a result of vector-associated insertional mutagenesis, and at this time no deaths have been reported [Cicalese et al 2016]. In May of 2016, GlaxoSmithKline announced that the European Commission had approved the gamma retroviral vector developed in Milan for use in treatment of SCID due to ADA deficiency. However, because of remaining concerns about potential insertional mutagenesis, and in order to achieve more effective ADA expression, clinical investigation of gene therapy using lentiviral vectors has now begun [Farinelli et al 2014, Candotti 2016].

Search ClinicalTrials.gov in the US and EU Clinical Trials Register in Europe for access to information on clinical studies for a wide range of diseases and conditions.