Case Report: Reactive Lymphohistiocytic Proliferation in Infant With a Novel Nonsense Variant of IL2RG Who Received BCG Vaccine

We present here a male young infant with X-linked severe combined immunodeficiency (MIM#300400) due to the novel nonsense variant of IL2RG (interleukin 2 receptor, gamma; MIM#308380), NM_000206.2(IL2RG):c.820_823dup p.Ser275Asnfs*29. He developed aggressive reactive lymphohistiocytic proliferation after receiving the live-attenuated Bacillus Calmette-Guérin (BCG) vaccine at birth. This report advocates for modifying the current practice of early use of BCG. The natural history of his disease also suggests considering IL2RG variants as a potential cause of “X-linked recessive Mendelian susceptibility to mycobacterial disease” (MSMD). His reactive lymphohistiocytic proliferation and massive hepatosplenomegaly simulated hemophagocytic lymphohistiocytosis (HLH, likely triggered by the BCG disease). This entity was masked by the absence of fever and markedly elevated inflammatory biomarkers. Thus, his findings stimulate discussion on the need to modify the diagnostic criteria of HLH, in order to accommodate conditions, such IL2RG variants that block systemic inflammation.


YOUNG INFANT
This 5-month-old male infant was born at term to asymptomatic, non-consanguineous emirati parents who belonged to the same tribal descent. The pregnancy and delivery were uneventful. His birth weight was 2.75 kg. He had received the BCG vaccine in the upper left arm at birth (0.05 mL, intradermal injection; manufactured by Serum Institute of India Pvt. Ltd., India). He had a healthy 16-month-old sibling, who had an oozing discharge from the BCG vaccine site for 2 months; he was currently asymptomatic. Otherwise, the family history is negative for any type of IEI.
His growth and development were normal. He was never genuinely febrile during the entire course of this illness; his temperatures were always ≤37.2 • C. At 3½ months of age, he developed a tender swelling in the right lower forearm (Figure 1). Abdominal examination revealed massive hepatosplenomegaly. The skin examination was remarkable for a prominent BCG scar (Figure 1), and skin-colored soft nodules such as the one in the lateral chest cage (Figure 1). He had mild cough; but on examination, he had no shortnessof-breath and his breath sounds and oxygen saturation were normal.
Radiograph and magnetic resonance imaging (MRI) of the right forearm showed areas of bone destruction (resorption) in the distal ulnar, surrounded by a soft tissue mass (Figure 2).
Chest radiograph and chest CT showed nodular lesions in the lungs (Figure 2). Ultrasound and computed tomography (CT) scan of the abdomen showed organomegaly with two focal hypodense splenic lesions (Figure 2).
Biopsy of a skin nodule revealed dermal histiocytic infiltrate forming granuloma ( Figure 3F).
Diagnostic exome sequencing revealed the hemizygous, pathogenic, novel non-sense variant of exon 6 of IL2RG (interleukin 2 receptor, gamma; MIM#308380): NM_ 000206.2(IL2RG):c.820_823dup, p.Ser275Asnfs * 29. This novel variant causes "severe combined immunodeficiency, X-linked" (SCIDX1 or XSCID; MIM#300400). This IL2RG variant was not detected in the mother's blood sample. Thus, the variant was assumed de novo in the affected infant. Germline mosaicism, however, could not be excluded. It is also important to note that the only significant variant reported on his diagnostic exome sequencing test was that involving the IL2RG gene. In addition, there was no clinically relevant copy number variant by NGS-CNV (copy number variation detection by next generation sequencing read counts) analysis.
His management included intravenous immunoglobulin, sulfamethoxazole-trimethoprim, isoniazid, rifampin, vancomycin, piperacillin/tazobactam, azithromycin, and amphotericin B liposomal. He continued breastfeeding and occasionally required oxygen supplementation by nasal cannula. He had no human leukocyte antigen (HLA) identical match. Thus, he was transferred to a transplant center for parental donor haploidentical hematopoietic stem cell transplantation (HSCT).

DISCUSSION
This male infant had SCID (MIM#300400) due to the novel non-sense variant, NM_000206.2(IL2RG):c.820_823dup p.Ser275Asnfs * 29 (11). He developed an aggressive reactive lymphohistiocytic proliferation after the BCG vaccine (4). Evidenced by the skin nodule histology, this illness is most likely a BCG disease as a result of his inborn error of immunity (IEI). It is well to know that sensitivities of the special stains acid-fast bacillus (AFB) and Grocott-Gomori's methenamine silver (GMS) are low. Therefore, false negative stains in situations of infection are relatively common. In a recent review, the sensitivity of "AFB microscopy" ranges from 20 to 70%, with a specificity of 95% or higher (12). In one study, the sensitivity of GMS stain for invasive pulmonary aspergillosis was 92% and specificity 82%; the negative predictive value was 75%, and the positive predictive value was 95% (13).
The clinical and pathologic data in this infant confer a predisposition to the weakly virulent mycobacteria, such as that of the BCG vaccine (4). Thus, this monogenic inherited condition provides an additional insight into the known list of MSMD (Mendelian susceptibility to mycobacterial diseases) (1)(2)(3). This report also advocates for modifying the current practice of early use of BCG (8)(9)(10).
The natural history of his disease suggests considering IL2RG variants as a potential cause of "X-linked recessive  Mendelian susceptibility to mycobacterial disease." His reactive lymphohistiocytic proliferation and hepatosplenomegaly simulated hemophagocytic lymphohistiocytosis (HLH), likely triggered by the BCG disease. The lack of fever and the just mildly elevated inflammatory biomarkers (serum ferritin, triglyceride, CRP, and ESR) are likely explained by the defective "interleukin 2 receptor, gamma" (14).
There are no in silico pathogenicity predictions for this novel variant (p.Ser275Asnfs * 29) from Ensembl Variant Effect Predictor (VEP) (15), but Varsome (16) indicates that it is pathogenic. The frameshift variation results in the alternation of the polypeptide sequence starting from position 275. This position is located in the middle of the single transmembrane helix (amino acids 263-283) of the IL2RG cytokine receptor. The 4-base duplication (c.820_823dup) results in a frameshift and the truncation of the polypeptide 29 amino acids downstream. Transmembrane helix prediction using TOPCONS (https:// topcons.cbr.su.se) indicates that the variant peptide could still harbor a transmembrane helix in this region. Critically, the region 286-294, termed the "Box 1 motif, " is required for the activation of Janus kinase (JAK), a tyrosine kinase that links cytokine signaling to transcription factors, by ILR2G (14). The consensus sequence of the "Box 1 motif " includes several hydrophobic residues and conserved prolines. The duplication significantly alters the protein sequence between 275 and 302; including the intracellular "Box 1 motif, " and truncates the polypeptide at position 303 (Figure 4). The "Box 1 motif " region lacks the necessary hydrophobic residues and the conserved prolines and, therefore, the variant protein is unlikely to activate JAK.
Interleukin 2 receptor gamma (IL2RG) is present in the receptors for IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 (17)(18)(19)(20). Stimulation of these receptors causes specific tyrosine phosphorylation and activation of JAK1 and JAK3. JAK3 is selectively associated with the c-terminal region of IL2RG, which is vital for the thymic development of T cells (14). Thus, the profound T cell lymphopenia in this infant with IL2RG:p.Ser275Asnfs * 29 is a result of the failure of IL2RG-JAK3 complex to promote thymic T cell selection and maturation (21,22).

DATA AVAILABILITY STATEMENT
The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.

ETHICS STATEMENT
The studies involving human participants were reviewed and approved by Tawam Human Research Ethics Committee -Signed informed consent was obtained from the parents. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin. Written informed consent was obtained from the individual(s) for the publication of any potentially identifiable images or data included in this article.

AUTHOR CONTRIBUTIONS
A-KS, AY, and SA-H: conceived, designed, structured the report, edited, and reviewed the paper. NA, SA, AY, AA, MA, and RK: analyzed and interpreted the clinical data. AE: analyzed and interpreted the pathology data. RV: analyzed and interpreted the variant. A-KS, AY, AE, and RV: wrote the first draft. All authors contributed to the article and approved the submitted version.