Unexpected combination: DiGeorge syndrome and myeloperoxidase deficiency

Abstract

We report a case of a 3-year-old boy who presented with recurrent bacterial and fungal infections and a known diagnosis of partial DiGeorge (22q11.2 deletion) syndrome. The nature and severity of his infections were more than normally expected in partial DiGeorge syndrome with normal T-cell counts and T-cell proliferative response to phytohaemagglutinin. This prompted further investigation of the immune system. An abnormal neutrophil respiratory oxidative burst, but normal protein expression of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system, led to the identification of myeloperoxidase deficiency. DiGeorge syndrome has a heterogeneous clinical phenotype and may not be an isolated diagnosis. It raises awareness of the possibility of two rare diseases occurring in a single patient and emphasises that even when a rare diagnosis is confirmed, if the clinical features remain atypical or unresponsive, then further investigation for additional cofactors is warranted.

Background

DiGeorge syndrome was first named after Dr Angelo DiGeorge who described infants with thymic aplasia and hypoparathyroidism in 1965. The condition is now known to be caused by a microdeletion on chromosome 22q11.2. Ninety per cent of 22q11.2 microdeletions are de novo. It is a heterogeneous condition characterised by cardiovascular abnormalities (predominantly conotruncal heart defects), palatal abnormalities, immunodeficiency and autoimmune disease, endocrinopathies (hypoparathyroidism with resultant hypocalcemia, thyroid dysfunction, growth hormone deficiency), genitourinary and gastrointestinal abnormalities, and neurodevelopmental abnormalities.¹ This report focuses on the immunological features.

Myeloperoxidase (MPO) deficiency is a rare autosomal recessive disorder caused by mutations in the MPO gene, located in chromosome 17. MPO deficiency is usually asymptomatic and constitutes an incidental finding.² Studies state that only 50% of the complete MPO-deficient patients have infectious complications. The rest are asymptomatic.³

In this case, we describe a patient with partial DiGeorge syndrome characterised by a mild clinical phenotype (mild dysmorphic facial features, a submucous cleft palate, global developmental delay, normal T-cell numbers and normal T-cell proliferative response to phytohaemagglutinin (PHA) stimulus). His frequent and severe bacterial and fungal infections were more than what would be normally seen with partial DiGeorge syndrome with normal T-cell counts, hence he was investigated for coexisting disease and other factors that could influence his infection burden. Further immunological testing led to the identification of coexisting MPO deficiency.

Case presentation

A 3-year-old boy with a known diagnosis of DiGeorge syndrome presented with a history of recurrent bacterial and fungal infections. He was born prematurely at 29 weeks to healthy non-consanguineous parents. At birth, he was admitted to neonatal intensive care unit and required respiratory support until day 48 of life. The neonatal period was complicated by necrotising enterocolitis which was managed conservatively, an episode of sepsis and a patent ductus arteriosus (PDA). He was discharged from the neonatal intensive care unit on day 77 of life with ongoing paediatric and cardiology follow-up. He subsequently had two admissions to paediatric intensive care unit (PICU), requiring ventilatory support for a bronchiolitic type illness. During the first admission, Pneumocystis jirovecii was identified on respiratory secretions at a low copy number in keeping with that normally seen in immune-competent children at this age, and was not thought to be significant. Candida albicans was identified in pulmonary secretions during the second PICU admission.

He underwent ligation of his PDA with a transluminal coil at 11 months of age. Following the recognition of mild facial dysmorphic features and global developmental delay, the diagnosis of DiGeorge syndrome was confirmed at 20 months of age by fluorescence in situ hybridisation. The patient’s parents did not carry the 22q deletion, indicating that our patient’s mutation was de novo.

In light of the diagnosis of DiGeorge syndrome, further assessment was undertaken looking for other manifestations of this condition.

Immunological investigations at that stage showed a normal lymphocyte subset analysis (results shown in table 1). T-cells proliferated to PHA. Immunoglobulins showed a normal IgG (8.89 g/L (reference range 3.1–13.8)), slight elevation in IgA (1.32 g/L (reference range 0.3–1.2)) and slight reduction in IgM (0.44 g/L (reference range 0.5–2.2)).

Table 1.

Lymphocyte subset analysis at the time DiGeorge was diagnosed

Tests	Results	Units	Reference ranges
Lymphocyte count	4.86	×109/L	3.46–11.62
CD3%	51	%	50–78
CD4%	29	%	26–53
CD8%	19	%	16–39
CD19%	35	%	16–33
CD16/56%	13	%	5–19
CD3 abs	2.49	×109/L	2.20–8.19
CD4 abs	1.43	×109/L	1.09–4.55
CD8 abs	0.91	×109/L	0.75–3.75
CD19 abs	1.71	×109/L	0.70–2.71
CD16/56 abs	0.62	×109/L	0.18–1.58

A further cardiology review confirmed closure of the patent PDA, with an otherwise structurally normal heart. An oral medicine assessment identified a submucus cleft palate. Conservative observation rather than surgical intervention was recommended. There was no evidence of aspiration on speech and language assessment and video fluoroscopy. He had mild eczema, which did not require specialist input. Non-IgE-mediated cow’s milk protein allergy was diagnosed based on reflux, loose stools and perianal skin breakdown. This responded to exclusion of cow’s milk protein in the diet. Imaging did not identify any skeletal or renal abnormality. He had no evidence of autoimmune disease or endocrinopathy.

He received continued input from the paediatric team, speech and language therapy, occupational therapy and physiotherapy in view of the global developmental delay, and to monitor for complications associated with DiGeorge syndrome.

Despite normal T-cell numbers and functional response to PHA, he continued to have frequent and significant upper and lower respiratory tract infections, with frequent episodes of oral candidiasis. Due to recurrent tonsillitis with associated cervical adenitis, he was listed for tonsillectomy following ear nose and throat review.

Respiratory consultation was sought in view of ongoing respiratory tract infections. Chest CT identified a non-specific ground glass pattern, which was most likely secondary to infection.

In the absence of other cofactors identified for his increased infection burden, further immunological testing was then undertaken.

Further immunological investigations

These confirmed normal classical and alternative complement pathway function, and mannan binding lectin levels. Neutrophil respiratory oxidative burst assay (ROB) showed reduced dihydrorhodamine oxidation signal on several samples, raising the possibility of chronic granulomatous disease (CGD). Subsequent protein analysis failed to identify any abnormality of the NADPH oxidase system, therefore a deficiency of MPO was suspected. A homozygous mutation in the MPO gene (c.2031-2A>C) was identified in the patient. Both parents are heterozygous for the same mutation. Testing of the patient’s three male siblings showed that one sibling is homozygous, another is heterozygous, and one has wild-type sequence. All siblings, including the homozygous MPO-deficient sibling, are clinically well and do not have a history of frequent or severe infections.

Treatment

He commenced itraconazole and co-trimoxazole prophylaxis with input from respiratory, infectious disease and ENT teams. The oral medicine and speech and language team continued to monitor the effects of the submucus cleft palate and to ensure he was not at risk of aspiration. Surgical intervention for the cleft palate was not recommended.

This significantly reduced his infection burden; however, he continued to develop recurrent episodes of tonsillitis and associated cervical adenitis with systemic upset. He has subsequently underwent tonsillectomy, which was uncomplicated.

Outcome and follow up

Our patient is now 6 years and 8 months of age. He continues on antimicrobial prophylaxis, with the need for this kept under periodic review. His infection burden has significantly improved, with no recent episodes of severe bacterial or fungal infections. He has had no recent mucocutaneous candidiasis. He has developmentally progressed; however, some behavioural issues have become apparent. He attends a school with a dedicated developmental team who continues to monitor his progress and provide educational support.

Discussion

DiGeorge, immune deficiency and infection

The immune deficiency associated with DiGeorge syndrome varies in severity. Some infants have normal T-cell numbers. Many have low T-cell counts which tend to normalise with time. Longitudinal studies have shown resolution of CD3, CD4 and CD8 cell numbers with a median age of 1.9, 2.5 and 2.2 years, respectively.⁴ Patients with complete DiGeorge syndrome have no detectable T cells, constituting a form of severe combined immune deficiency that requires immune reconstitution with a thymic or haematopoietic stem cell transplant.^{5 6}

Humoral defects are often seen in older children. Specific IgA deficiency, reduced differentiation of class switched memory B cells and a decline in immunoglobulin production and function are reported.⁷

A range of infections are seen in patients with DiGeorge syndrome. A longitudinal follow-up of a cohort of patients found respiratory tract infection was the most common infection occurring in patients with DiGeorge syndrome. Septicaemia with fungal infections, particularly Candida is described, but this was in patients with a reduction in T-cell count at the time of infection.⁴

Due to his serious infections, which appeared atypical for partial DiGeorge syndrome with normal T-cell counts and function, further investigations were undertaken to look for any other possible contributing immunological defect. The abnormal ROB test was a ‘key’ to look for MPO deficiency.

MPO, immune deficiency and infection

MPO deficiency is an autosomal recessive condition encoded by a single gene located on band 17q22-23. It can be complete or partial. It occurs in 1/1000–1/4000 people.^{8 9}

MPO is an enzyme in the granules of neutrophils and lysosomes of monocytes acting against invading microorganisms. During the respiratory oxidative burst, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in neutrophils generates reactive oxygen species. MPO catalyses the formation of reactive oxygen intermediates, forming part of the innate host defence against invading microorganisms. Patients with CGD have a deficiency in the NADPH oxidase activity and are unable to produce O₂ ⁻ resulting in a susceptibility to fungal infections and catalase positive bacteria.¹⁰ A low neutrophil dihydrorhodamine oxidation signal can be seen in both CGD and complete MPO deficiency.¹¹ Many patients with MPO deficiency are asymptomatic; however, there is a recognised susceptibility to fungal infections such as C . albicans.¹² Severe infections are reported in up to 5% of patients.¹³

MPO deficiency may become apparent in patients with relevant comorbidities that also increase susceptibility to Candida infections, such as diabetes.¹³ As discussed above, fungal infections in DiGeorge syndrome are more commonly seen in patients with low T-cell numbers. However, the detrimental effect of both immunological defects in defence against fungal infections in our patient may have had a synergistic effect, resulting in his increased susceptibility to fungal infections.

DiGeorge and inflammatory/autoimmune conditions

Our patient to date does not have autoimmune disease; however, there is a recognised increased risk of autoimmune manifestations in DiGeorge syndrome. Impaired central and peripheral tolerance is implicated as the underlying mechanism for increased autoimmune disease, but this remains poorly defined.^{14 15} A variety of autoimmune conditions are described, including thrombocytopaenia, hypothyroidism and juvenile idiopathic arthritis.¹⁶

MPO and inflammatory/autoimmune conditions

MPO is also believed to play a role in suppression of the adaptive immune response by inhibition of dendritic cell function, resulting in attenuation of some T-cell-mediated inflammatory disorders.¹⁷

Previous human observation studies have shown an increased incidence of lupus nephritis in patients with a polymorphism causing reduced MPO expression.¹⁸ Murine studies have shown that MPO-deficient mice with antigen-induced arthritis developed more severe joint inflammation and damage.^{19 20} An increased incidence in chronic inflammatory conditions is seen in MPO-deficient patients.²¹

Conversely, a pathogenic role for MPO in driving autoimmune inflammation was demonstrated using MPO-deficient mice in the K/BxN arthritis and collagen-induced arthritis models exhibiting reduced disease severity.²² Also, increased MPO levels and activity have been observed in many inflammatory conditions and autoimmune diseases including and rheumatoid arthritis.²³

As discussed by Aratani,¹⁰ both over and under expression of MPO have been linked to adverse disease outcome in inflammatory disease. More research into this enzyme is required to ascertain its role in these inflammatory conditions.

It is not known if the combination of DiGeorge syndrome and MPO deficiency will result in an even further increased susceptibility to autoimmune disease and/or increased inflammatory response.

Coexisting genetic diseases

22q deletion syndrome and MPO deficiency are genetically distinct conditions located on separate chromosomes. Our case demonstrates the occurrence of two genetic disorders, one occurring de novo and the other that was inherited from carrier parents.

DiGeorge syndrome is the most frequent copy number variant (CNV) affecting infants, with a variable clinical phenotype both in relation to the immune deficiency and the other associated complications. Coexisting diagnosis due to additional genome-wide mutations, CNVs or mutations/CNVs on the other allele are increasingly described in patients with DiGeorge syndrome.²⁴ Examples of dual diagnoses of genetic disorders related to DiGeorge syndrome are Bernard-Soulier syndrome type B²⁵ and CEDNIK syndrome (cerebral dysgenesis, neuropathy, ichthyosis and keratoderma).²⁶ In these autosomal recessive disorders, in addition to the deletion in 22q11.2, the patient has a deletion/mutation in the relevant gene on the other allele. Likewise, there are reports of patients with DiGeorge syndrome and another disorder that is coincidental. Such conditions are cystic fibrosis, glycose 6 phosphate dehydrogenase deficiency and severe combined deficiency.²⁴ These coexisting conditions are relevant to the medical management and long-term outcome in these patients.

To the best of our knowledge, this is the first case described of 22q deletion with MPO deficiency. It is possible that the two immunological defects are working in consort to produce a significant immune-deficient clinical phenotype. Long-term follow-up of our patient will determine if the two conditions will have any effect on the risk of autoimmune/inflammatory disease.

In conclusion, this case highlights the need to consider the possibility of a second diagnosis, should the clinical presentation be more severe, or atypical than expected for the known immune deficiency. The association of immunodeficiencies with other diseases and the molecular mechanisms behind their interactions is an area for future research.

Patient’s perspective.

When we discovered our son had 22qDS, we understood it came with the prospect of immunological problems, so when we started landing regularly at the hospital and in PICU we were not surprised. However, we kept being told by various doctors that they could not figure out why he was so bad, bacterial swabs were coming back clear but he was obviously fighting something. He always fought his way back but it was never quite ‘normal’. His constant illnesses and infections had his medical team baffled so his blood was sent for further genetic testing, and it was discovered he had a further genetic disorder, which could also cause immunological issues. In hindsight, since finding out about his MPO deficiency, it seems obvious he was being floored by C andida infections, which I am almost certain they found in swabs but which would not normally cause serious issues for most people, and so was overlooked. Constant infections and hospitalisation have been difficult but since the discovery of his second genetic disorder our son has improved constantly. His medical team have been exceptional. Going forward we are slightly anxious because we do not know what the interaction of the two conditions will mean for our son, or what his future may hold. We recently had our other sons tested to see if they have the condition too. One of his brothers also has MPO deficiency, but thus far has displayed no real symptoms. For us this clearly points to the fact that our eldest son’s combination of conditions is what leads him to be unwell so often, and so abhorrently. Lots of people say their child is one in a million, we know that medically speaking ours definitely is.

Learning points.

• Severe or recurrent infections indicate a disruption in host defence that should warn us for immune deficiencies.

• A possible combination of immunodeficiencies should be considered despite the low frequency of these diseases.

• DiGeorge syndrome increases the susceptibility to bacterial and fungal infections as a result of thymus hypoplasia.

• Myeloperoxidase deficiency can cause severe infections in already immunosuppressed patients.

• If the clinical presentation is more than normally expected for a known diagnosis, then investigation should focus on an additional defect. DiGeorge syndrome is heterogeneous disorder, and there should be a low threshold for considering the possibility of coexisting disease.