PEDIATRICS Vol. 107 No. 4 April 2001, p. e48

ELECTRONIC ARTICLE:
High Levels of Interferon Gamma in the Plasma of Children With Complete Interferon Gamma Receptor Deficiency

Claire Fieschi, MD, MSc^*, Stéphanie Dupuis, MSc^*, Capucine Picard, MD, MSc^*, C. I. Edvard Smith, MD, PhD, Steven M. Holland, MD^§, and Jean-Laurent Casanova, MD, PhD^*,

From the ^* Laboratory of Human Genetics of Infectious Diseases, Necker Medical School, Paris, France, European Union (EU);  Karolinska Institute, Huddinge, Sweden, EU; ^§ Laboratory of Host Defense, National Institutes of Health, Bethesda, Maryland; and  Pediatric Immunology Unit, Necker-Enfants Malades Hospital, Paris, France, EU.

	ABSTRACT

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We have found that children with complete interferon gamma (IFN) receptor deficiency, unlike patients with other genetic defects predisposing them to mycobacterial diseases, have very high levels of IFN in their plasma. This unexpected observation provides a simple and accurate diagnostic method for complete IFN receptor deficiency in children with clinical disease caused by bacille Calmette-Guérin vaccines or environmental nontuberculous mycobacteria.

	ARTICLE

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Mendelian susceptibility to mycobacterial infection (MIM 209950) is a rare and heterogeneous syndrome.^1-3 Affected individuals develop severe clinical disease caused by weakly virulent mycobacterial species, such as bacille Calmette-Guérin (BCG) vaccines and environmental nontuberculous mycobacteria (NTM). The clinical phenotype ranges from fatal disseminated infection in early childhood to focal recurrent infection in adults. In the last 5 years, considerable genetic heterogeneity has been documented. Mutations have been found in 4 genes: IFNGR1, encoding the interferon gamma (IFN) receptor ligand-binding chain; IFNGR2, encoding the IFN receptor signal-transducing chain; IL12B, encoding the p40 subunit of interleukin (IL)-12; and IL12RB1 encoding the IL-12 receptor 1 chain. Different types of mutations define 8 inherited disorders: complete recessive IFNR1 deficiency with^4-6 or without⁷ receptor surface expression; partial, as opposed to complete, IFNR1 deficiency with recessive⁸ or dominant inheritance⁹; recessive complete¹⁰ or partial¹¹ IFNR2 deficiency; complete recessive IL-12p40¹²; and IL-12R1 deficiency.^13,14 However, a molecular etiology is still lacking for a majority of the patients. Complete IFNR1^1-7 and IFNR2⁸ deficiency are responsible for early-onset overwhelming mycobacterial disease. Partial IFNR1 defects^8,9 and partial IFNR2 deficiency,¹¹ like complete IL-12p40¹² and IL-12R1 deficiency,^13,14 are responsible for milder clinical forms.^1-3

The diversity of the genes and pathogenic mutations involved renders molecular diagnosis challenging. For example, complete IFNR1 deficiency may be caused by mutations preventing expression of the receptor^4-6 or binding of the surface receptor to IFN.⁷ Moreover, cells with complete IFNR deficiency do not respond to IFN,^4-7 whereas cells with partial IFNR deficiency respond to IFN at high concentration.^8,9,11 Finally, most patients display low levels of IFN production by peripheral blood cells.^1,2 Cumbersome diagnostic investigations combining highly specialized functional, biochemical, and genetic assays are, therefore, required in most patients with the syndrome. An accurate and rapid molecular diagnosis is, however, essential for the rational and efficient treatment of the patient. Indeed, children with complete IFNR deficiency do not achieve sustained remission with antibiotics alone and do not respond to exogenous IFN, resulting from a lack of functional receptors. The outcome seems to be often fatal and bone marrow transplantation should be considered.^1-3,15 In contrast, the administration of subcutaneous IFN together with antibiotics is often beneficial in patients with other genetic defects, and full remission of mycobacterial disease has been achieved.^1-3 The lack of a simple method for rapidly discriminating between patients with complete IFNR deficiency and patients with other genetic etiologies greatly compromises the management of these patients.

We measured IFN by enzyme-linked immunosorbent assay (ELISA) in the plasma of healthy individuals and patients with various forms of Mendelian susceptibility to mycobacterial infection. IFN is undetectable (<5 pg/mL) in the serum and plasma of 6 healthy individuals tested (not shown). All patients had suffered from BCG and/or NTM clinical disease when the blood sample was taken. Patients with IL-12p40 (n = 3) and with IL-12-receptor 1 chain deficiency (n = 5)like patients with partial dominant IFNR1 deficiency (n = 7) and partial recessive IFNR2 deficiency (n = 1)had no detectable IFN in the plasma (Fig 1). We found low levels of IFN (median = 57 ± 9.9 pg/mL) in the plasma of patients with partial recessive IFNR1 deficiency (n = 2). Remarkably, we found very high levels of IFN in the plasma of patients with complete IFNR1 deficiency (n = 5; median = 252 ± 113 pg/mL) and complete IFNR2 deficiency (n = 2; median = 433 ± 306 pg/mL). To validate these results, we measured IFN in the serum of 40 other children with unexplained BCG and/or NTM clinical disease. High levels of IFN were found in 1 child, who was subsequently diagnosed with complete IFNR1 deficiency (n = 6; median 249 ± 101 pg/mL). Complete IFNR deficiency was functionally and genetically excluded in the remaining 39 patients.

View larger version (13K): [in this window] [in a new window]   Fig. 1.   Serum levels of IFN in patients with known genetic etiologies for Mendelian susceptibility to mycobacterial infection. Known genetic etiologies include complete recessive IFNR1 deficiency (CR1), complete recessive IFNR2 deficiency (CR2), and partial recessive IFNR1 deficiency (PR1). Other cases (others) include partial dominant IFNR1 deficiency (n = 7), partial recessive IFNR2 deficiency (n = 1), complete IL-12p40 deficiency (n = 3), and complete IL-12R1 deficiency (n = 5). The experiment was performed in a single laboratory using the IFN ELISA kit: PeliKine Compact, Human IFN ELISA kit (CLB, The Netherlands). Its sensitivity is 5 pg/mL and the linear range is 5 pg/mL to 500 pg/mL. Our study was performed in compliance with institutional requirements and an informed consent was obtained from each patient's family.

These results may reflect the more severe course of mycobacterial disease in patients with complete IFNR deficiency, resulting in more intense and sustained IFN secretion. However, high plasma levels of IFN (35 pg/mL) in 1 asymptomatic child with a family history, diagnosed at birth suggests that it is not the case (not shown). Paradoxically, patients with complete IFNR deficiency have previously been shown to have impaired secretion of IFN, attributable to a secondary defect in IL-12 production.⁶ As IFNR is ubiquitously expressed in the organism, our results suggest that patients with complete IFNR deficiency cannot eliminate blood IFN, resulting from a lack of binding (IFNR1 deficiency) or a lack of internalization (IFNR2 deficiency) of the cytokine. This would also account for the detectable levels of IFN in the plasma of patients with partial recessive IFNR1 deficiency, in whom the receptor mutation probably reduces but does not abolish the affinity of the receptor for IFN.⁸ Receptors from patients with dominant IFNR1 deficiency probably bind and/or recycle sufficient amounts of IFN to keep serum levels undetectable. Profound defects of IFN production (IL-12p40 and IL-12R1 deficiencies) in patients with functional IFN receptors are not associated with high levels of IFN in the plasma.

In any event, plasma IFN determination by ELISA is a simple, cheap, rapid, and efficient way to guide molecular diagnosis and to provide a rational basis for the treatment of patients with Mendelian susceptibility to mycobacterial infection. High levels (our threshold of 80 pg/mL is >2 standard deviations above the mean level in patients with partial recessive IFNR1 defects) of IFN in the serum of a patient with BCG and/or NTM clinical disease should lead to the consideration of bone marrow transplantation options while searching for and validating null mutations of IFNGR1 or IFNGR2. Undetectable or low levels of IFN should lead to the child being treated with subcutaneous IFN while searching for mild mutations of IFNGR1 and IFNGR2, or null mutations of IL12B and IL12RB1.

	FOOTNOTES

Received for publication Oct 25, 2000; accepted Jan 8, 2001.

Address correspondence to Jean-Laurent Casanova, MD, PhD, Laboratory of Human Genetics of Infectious Diseases, Necker-Enfants Malades Medical School, 156 rue de Vaugirard, 75015 Paris, France. E-mail: casanova{at}necker.fr

	ABBREVIATIONS

BCG, bacille Calmette-Guérin; NTM, nontuberculous mycobacteria; IFN, interferon; IL, interleukin; ELISA, enzyme-linked immunosorbent assay.

	REFERENCES

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