Inborn errors of immunity (IEI) are a large heterogeneous group of diseases characterized by immunodeficiency, immune dysregulation, allergy, auto-inflammation, and predisposition to malignancies. Most are inherited as an autosomal recessive trait. We studied a patient with severe combined immunodeficiency (SCID) and immune dysregulation who harbored two distinct biallelic IEI-associated genetic mutations. Clinical, immunological, and genetic data were collected. Genetic investigation included whole-exome sequencing on DNA extracted from skin fibroblasts. Family segregation was performed by Sanger sequencing. Immunological evaluation included absolute and functional evaluations of lymphocytes and chimerism analysis post-hematopoietic stem cell transplantation (HSCT). Treg subsets, lipopolysaccharide-responsive and beige-like anchor (LRBA), and Cytotoxic T-Lymphocyte Associated protein 4 (CTLA4) expression levels were measured by flow cytometric analysis. A 19-year-old female patient from a consanguineous background underwent unconditioned matched sibling-related HSCT during infancy due to the clinical presentation of SCID with an Omenn phenotype. At that time, her underlying genetic defect was not defined. Years after HSCT, severe autoimmune phenomena were noted, including a systemic lupus erythematosus-like syndrome and ophthalmic manifestations. Genetic evaluation revealed biallelic homozygous mutations in recombination activating gene-2 (c.685C>T, p.Arg229Trp) and a previously undescribed mutation in LRBA (c.3325G>T, p.Asp1109Tyr). LRBA and CTLA4 expression levels were normal, suggesting that the LRBA variant identified in these kindred is unlikely to be pathogenic. Multiple genetic defects causing complex IEIs may be identified in the same individual in highly consanguineous populations. Functional immunological testing is essential for the evaluation of novel genetic variants.

Dedicator of cytokinesis protein 8 (DOCK8) is a crucial regulator for the formation of immune synapses, allowing for a proper function of innate and adaptive immune systems. DOCK8 deficiency is a primary immunodeficiency, currently known as Inborn Errors of Immunity (IEI) affecting both cellular and humoral immunity, thus leading to various clinical presentations ranging from infections, autoimmunity, inflammations and malignancies. We aimed to identify unique molecular signatures in the adaptive immune repertoire of DOCK8-deficient patients through comprehensive sequencing and analysis of the T-cell receptor (TCR) and B-cell receptor (BCR) repertoires, correlating these signatures with clinical features of the patients, ultimately providing a “fingerprint” of both individual’s immunological status and disease conditions. We characterized the TCR repertoires of αβ T- and γδ T-cells, together with BCR repertoires of B-cells, in peripheral blood of 13 patients and age matched healthy donor controls using next generation sequencing. TCR repertoire in patients with DOCK8 deficiency demonstrated restricted diversity and clonal expansions, favorable use of specific gene in the V-D-J recombination process, and a unique finding of longer complementarity-determining-region-3 (CDR3) length, revealing an impaired T-cell development and activation. Furthermore, BCR repertoire demonstrated high diversity without clonal expansions, portraying an abnormal B-cell development leading to a possible mechanism for atopy and autoimmunity, alongside inadequate activation in DOCK8 deficiency. Lastly, various DOCK8 mutations demonstrated a more profound defects in immune repertoire. These findings expand our knowledge on the role DOCK8 plays in shaping the TCR and BCR repertoires in DOCK8 deficiency.

IntroductionMitochondrial metabolism is essential for T-cell function, but the roles of individual electron transport chain (ETC) components are unclear. Here, we aimed to explore the role of mitochondrial complex I (CI) subunit NADH:ubiquinone oxidoreductase iron-sulfur protein 4 (NDUFS4) in T-cell metabolic fitness and immunity.MethodsWe used a T cell-specific Ndufs4 knockout mouse model to find that NDUFS4 deficiency disrupts CI function, leading to metabolic and redox imbalances. Additionally, T cells from a patient with Leigh syndrome induced by NDUFS4 loss-of-function were analyzed.ResultsNdufs4-deficient T cells exhibit impaired OXPHOS, reduced respiratory capacity, and increased glycolysis, accompanied by reactive oxygen species (ROS) accumulation and defective TCR-driven activation, including reduced proliferation and cytokine production. In vivo, Ndufs4(-/-) mice show T-cell lymphopenia and impaired humoral and cytotoxic immunity. Importantly, T cells from a single Leigh syndrome patient with an NDUFS4 loss-of-function variant showed similar defects, including impaired activation and proliferation.DiscussionThese findings highlight the importance of NDUFS4 for human immunity and establish a mechanistic link between complex I dysfunction and T-cell immunodeficiency. Our results identify NDUFS4 as a key regulator connecting mitochondrial integrity to adaptive immune function.

ZBTB24 is a member of a protein family containing a Broad-Complex, Tramtrack, and Bric a Brac (BTB) domain, which functions in protein-protein interactions. ZBTB24, a transcription factor, binds its DNA targets through its C-terminal zinc finger (ZF) domain. Biallelic ZBTB24 pathogenic variants lead to the rare autosomal recessive Immunodeficiency, Centromeric instability and Facial anomalies type 2 (ICF2) syndrome. The majority of ICF2 patients carry biallelic loss-of-function variants in ZBTB24. The remaining patients harbor missense variants in the ZF domain that compromise the ability of ZBTB24 to transcriptionally activate CDCA7, the gene responsible for ICF subtype 3 syndrome. Although an ICF2 patient with compound heterozygous pathogenic variants, including a missense variant (p.Ser59Gly) in the BTB domain, has been reported, no ICF2 patients with biallelic missense variants in any ZBTB24 domains other than the zinc finger domain have been described. Similar to all subtypes of ICF syndrome, ZBTB24 pathogenic variants lead to significant DNA hypomethylation throughout the genome. Here we describe a patient with severe infections initiating during her first year of life, significant developmental delay and an abnormal facial shape, who carries a homozygous p.Val43Leu substitution in the BTB domain of ZBTB24. The patient's peripheral blood cells demonstrate whole genome DNA hypomethylation with patterns identical to those found in verified ICF2 patients. Both the p.Val43Leu and p.Ser59Gly variants cause significant ZBTB24 protein instability. Thus, we demonstrate that pathogenic missense variants in the BTB domain of ZBTB24 can functionally act as loss-of-function variants that result in ICF2 syndrome.

Telomere biology disorders (TBDs) are characterized by bone marrow failure (BMF) and dysfunctional telomeres. So far, inherited mutations in 18 genes have been identified in TBDs. Here, we describe a child presenting with early BMF, immunodeficiency, and severely short and defective telomeres, carrying a homozygous splicing mutation (c.409-2 A > G; p.Q136_K138del) in RPA2 - a known replication factor and telomerase accessory factor. The Q136_K138del mutation is predicted to compromise the binding of RPA2 to the single-stranded telomeric DNA. Sequencing of single telomeres revealed that telomere variant repeats (TVRs), present also in healthy individuals, became more prominent in the short telomeres of the patient. Such TVRs may aggravate the telomere dysfunction due to lower affinity to the shelterin proteins. The severe telomere shortening and the activation of DNA damage response at telomeres indicate that this RPA2 mutation causes TBD in a similar manner to other known mutations and should be considered in patients displaying clinical TBD features.

BACKGROUND:Bone marrow failure syndromes (BMFs) comprise a heterogeneous group of genetic disorders characterized by impaired hematopoiesis and multisystem involvement. Dyskeratosis congenita (DC) is a telomere biology disorder caused by defects in telomerase or telomere maintenance, leading to progressive BMF and variable extra-hematopoietic manifestations. OBJECTIVES:To describe the clinical, immunologic, genetic, and telomere biology findings in a patient with DC caused by a novel biallelic telomerase reverse transcriptase (TERT) mutation, and to highlight diagnostic and therapeutic considerations in telomere-associated BMFs. METHODS:We assessed cellular and humoral immune functions. Genetic analysis was conducted using whole-exome sequencing (WES) with segregation analysis. Telomere length was assessed by flow-FISH. Functional and radiologic evaluations were performed to define disease extent. RESULTS:A 2-year old male born to consanguineous parents presented with multisystemic clinical features, and hypocellular bone marrow. WES identified a novel homozygous TERT missense variant (c.3052G>A; p.Ala109Thr) supported by markedly shortened telomeres. Neuroimaging revealed cerebellar hypoplasia consistent with Hoyeraal-Hreidarsson syndrome. Immunologic evaluation demonstrated skewed CD4:CD8 ratios. An incidental heterozygous MEN1 variant was also detected. CONCLUSIONS:We expand the clinical and genetic spectrum of TERT-associated DC and illustrate the critical role of genomic diagnostics and telomere assessment in BMFs. Early molecular diagnosis enables targeted evaluation, informs prognosis, and guides personalized management in telomere biology disorders. In addition, the identification of actionable secondary variants further highlights both the power and complexity of comprehensive genomic testing.

AIMS:Paediatric cancer survivors often experience treatment-induced immunosuppression, requiring post-treatment revaccination. However, immune recovery timelines vary, and current revaccination guidelines, largely based on data of varied quality derived from studies on acute-lymphoblastic-leukaemia (ALL), may not be applicable across all paediatric malignancies. This study evaluated a personalised revaccination approach based on immune recovery. METHODS:Immune recovery was evaluated using antibody titers to common childhood vaccines, immunoglobulin levels, B- and T-cell subpopulations, and T-cell receptor excision circles (TREC) and kappa-deleting recombination excision circles (KREC) markers. The recommended revaccination window was defined as 3-6 months post-treatment. Patients completing immune and serology testing within this window were categorised as the per-protocol-group (PPG), while others were categorised as the protocol-deviated-group (PDG). RESULTS:Among 19 PPG patients, 57.9% required major and 36.8% minor modifications to recommended revaccination guidelines, with major modifications more common among hematologic malignancies than solid tumours (83.3% vs. 46.2%, p < 0.0001). In the intention-to-treat (ITT) cohort (n = 52), 40.4% required major and 57.7% minor modifications; hematologic survivors again had higher rates of major modifications (52.9% vs. 34.3%, p = 0.008). CONCLUSIONS:Immune recovery following paediatric cancer therapy is highly variable, particularly among hematologic malignancies. Personalised revaccination strategies incorporating comprehensive immunological workups may optimise protection against vaccine-preventable diseases.

Background: Hematopoietic stem cell transplantation (HSCT) is a curative treatment for infants with severe combined immunodeficiency (SCID). Different factors determine HSCT success and overall survival (OS). Specifically, prompt diagnosis of SCID, preferably through newborn screening (NBS), is critical. Objective: We explored variable factors including the impact of NBS that are associated with HSCT outcomes and OS in a multiethnic SCID cohort. Methods: One hundred patients with SCID diagnosed and treated with HSCT at Sheba Medical Center in Israel between 1996 and 2024 were studied. Results: We distinguished 3 groups: Palestinians (62%), Israeli Jews (16%), and Israeli non-Jews (22%). The OS rate was 68%, increasing to 84% when excluding mortality in the first month after transplantation. Better outcomes were significantly associated with the use of bone marrow (BM) as a stem cell source (P = .003), the availability of matched related donors (P = .045), and the use of conditioning (P = .0006). As a result of delayed diagnosis, Palestinian patients had more infections, more events of early post-HSCT death, and inferior OS rates compared to other patients. SCID cases identified by NBS demonstrated superior OS (93%) compared to cases identified by clinical presentation (P = .04). Improvement in OS was most significant after the implementation of the NBS program for SCID in Israel (P = .03). Conclusion: Our study delineates and reinforces specific factors that influence OS after undergoing HSCT for SCID. Importantly, it raises the value of early diagnosis and treatment of affected infants, highlighting the benefit of NBS for SCID in determining the clinical outcome. (J Allergy Clin Immunol 2025;155:1800-12.)

PURPOSE:Baraitser-Winter syndrome type 1 (BRWS1) is a rare disorder characterized by intellectual disability, short stature, facial dysmorphism, cortical malformations, macrothrombocytopenia, and recurrent infections. BRWS1 is caused by loss-of-function variants in ACTB, leading to β-actin deficiency. Given the essential role of the actin cytoskeleton in T-cell activation, the immunological consequences of ACTB mutations remain unexplored. Here, we characterize immune dysfunction associated with a novel ACTB variant in a patient with BRWS1. METHODS:Whole-exome sequencing identified a heterozygous ACTB p.Gln360ProfsTer4 variant in a patient with BRWS1 and combined immunodeficiency. Functional studies were performed in HEK293T cells transfected with wild-type and mutant ACTB constructs. Patient-derived T cells were analyzed for immunological synapse formation, cytokine production, activation, and proliferation. The therapeutic effects of exogenous IL-2 and dupilumab were evaluated. RESULTS:The mutant β-actin protein was rapidly degraded and exerted a dominant-negative effect on wild-type β-actin, thereby disrupting cytoskeletal integrity. Patient-derived T cells demonstrated defective immunological synapse formation, reduced intra-synaptic IL-2 levels, and impaired activation and proliferation. Supplementation with exogenous IL-2 partially restored T-cell function in vitro. Notably, dupilumab treatment led to significant clinical and immunological improvement, suggesting a role in restoring immune regulation. CONCLUSION:BRWS1 represents a novel primary immune regulatory disorder. Our findings highlight actinopathy-driven immunodeficiency as a target for therapeutic intervention, with broader implications for cytoskeletal disorders.

Background Hematopoietic stem cell transplantation (HSCT) is a curative treatment for infants with severe combined immunodeficiency (SCID). Different factors determine HSCT success and overall survival (OS). Specifically, prompt diagnosis of SCID, preferably through newborn screening (NBS), is critical. Objective To explore variable factors including the impact of NBS that are associated with HSCT outcomes and OS, in a multi-ethnic SCID cohort. Methods 100 SCID patients diagnosed and treated with HSCT at Sheba Medical Center in Israel between the years 1996 to 2024 were studied. Results We distinguished three groups: Palestinians (62%), Israeli Jews (16%), and Israeli non-Jews (22%). The OS rate was 68%, increasing to 84% when excluding mortality in the first month post-transplantation. Better outcomes were significantly associated with the use of bone marrow (BM) as a stem cell source (P =0.003), the availability of matched related donors (MRD, P =0.045) and use of conditioning (P =0.0006). Due to delayed diagnosis, Palestinian patients had more infections, more events of early post-HSCT death, and inferior OS rates compared to other patients. SCID cases identified by NBS demonstrated superior OS (93%) compared to cases identified by clinical presentation (P =0.04). Improvement in OS was most significant following the implementation of the NBS program for SCID in Israel (P = 0.03). Conclusion Our study delineates and reinforces specific factors that influence OS after undergoing HSCT for SCID. Importantly, it raises the value of early diagnosis and treatment of affected infants, highlighting the benefit of NBS for SCID in determining the clinical outcome.

BACKGROUND:Mutations in RELA, a key component of NF-κB signaling, are associated with dysregulated immune responses and inflammatory disorders. While immunodeficiency phenotypes associated with RELA haploinsufficiency have been reported, gastrointestinal manifestations remain poorly described. This study aimed to characterize the clinical, genomic, and immunological features of a patient presenting with an atypical Crohn's-like phenotype driven by RELA haploinsufficiency. METHODS:Whole-exome sequencing was performed, and results were confirmed by Sanger sequencing. Protein modeling, Western blotting, immunofluorescence, and nuclear extract-based NF-κB activation assays were conducted to assess the functional impact of the identified variant. Immune profiling was performed using mass cytometry time of flight (CyTOF) and single-cell RNA sequencing (scRNA-seq) and compared to controls. RESULTS:We studied a 17-year-old male diagnosed with pan-enteric Crohn's disease (CD), perianal fistulas, chronic mucocutaneous candidiasis, and chronic lymphopenia. Sequencing identified a heterozygous missense variant in RELA (c.587T>C, p.V196A) that potentially impairs RelA (p65) protein stability, confirmed by reduced activity and diminished protein expression. CyTOF analysis revealed decreased circulating T regulatory cells (Tregs), absence of mucosal Tregs, high apoptotic rates, and elevated IFN-γ induced levels, while scRNA-seq demonstrated a robust type I/II interferon signature in multiple immune subsets. Dysregulated mucosal-associated invariant T (MAIT) and cytotoxic CD4+ T cells exhibited upregulation of IL23R and ADAM12, further linking RELA dysfunction to enhanced pro-inflammatory T cell response and tissue inflammation. CONCLUSION:This study links RELA haploinsufficiency with CD-like features, Th1/Th17 polarization, and interferon-driven inflammation, emphasizing the importance of genetic evaluation in patients with atypical or refractory IBD.

BACKGROUND:Chronic recalcitrant dermatophyte infection (CRDI) remains poorly understood, with limited research on its genetic, immunological and treatment aspects. OBJECTIVES:To investigate the genetic and immunological profiles, clinical characteristics and treatment outcomes in otherwise healthy patients with CRDI. METHODS:This was a prospective study of patients with isolated superficial dermatophyte infections persisting for > 5 years and refractory to at least three oral antifungal agents. Whole-exome sequencing, interleukin (IL)-17 quantification and lymphocyte subset analysis were performed. RESULTS:Of 5910 patients with superficial cutaneous fungal infections, 11 (10 men; 91%) with a mean (SD) disease duration of 26.4 (8.7) years were included in this study. All patients presented with extensive dermatophytosis, involving a mean (SD) of 5.6 (1.4) skin and nail sites. Immune alterations were identified in 55% (n = 6/11) of the patients, including three with monogenic immunodeficiency mutations (CARD9 and FOXN1), one with acquired immunodeficiency (Good syndrome) and two with isolated IL-17 deficiencies. Most patients (n = 10/11) showed a partial clinical response to continuous oral antifungal therapy, while only one patient remained a complete responder after discontinuing treatment. CONCLUSIONS:CRDI is a rare condition in patients with superficial cutaneous fungal infections. Genetic and immunological evaluation for immunodeficiency should be considered in patients with CRDI. Management may require prolonged or lifelong oral antifungal therapy.

Aim Pediatricians are trained to identify recurrent or unusual infections in children, prompting evaluation for inborn errors of immunity (IEI). Some monogenic IEI, however, may present atypically. This study describes our experience with children diagnosed with activated phosphoinositide 3-kinase delta syndrome (APDS2) including unusual presentations. Methods A retrospective review was conducted on two children diagnosed with APDS2 at Shaare Zedek and Sheba Tel-Hashomer Medical Centers in Israel. Both patients underwent immune assessments, genetic testing, and treatment between 2019 and 2024. Results Two patients, a 17-year-old female (P1) and a 7-year-old male (P2), were diagnosed with APDS2 after presenting with recurrent juvenile parotitis (P1) and severe lymphadenopathy (P2). Immunologic evaluation revealed hypogammaglobulinemia and combined immune deficiency. Genetic testing identified PIK3R1 variants (c.1425 + 1G > T in P1 and c.1425 + 1G > C in P2). Both received intravenous immunoglobulins and prophylactic antibiotics. P2 was treated with rapamycin, leading to resolution of lymphadenopathy. Conclusion This report highlights the clinical presentation of APDS2, a rare monogenic IEI in children, including the atypical manifestation of RJP and the common feature of lymphadenopathy. Pediatricians should stay vigilant for red flags of IEI during clinical evaluations, as early diagnosis and multidisciplinary care are crucial for effective management.

Mutations in proteasome β-subunits or their chaperone and regulatory proteins are associated with proteasome-associated autoinflammatory disorders (PRAAS). We studied six unrelated infants with three de novo heterozygous missense variants in PSMB10, encoding the proteasome β2i-subunit. Individuals presented with T-B-NK± severe combined immunodeficiency (SCID) and clinical features suggestive of Omenn syndrome, including diarrhea, alopecia, and desquamating erythematous rash. Remaining T cells had limited T cell receptor repertoires, a skewed memory phenotype, and an elevated CD4/CD8 ratio. Bone marrow examination indicated severely impaired B cell maturation with limited V(D)J recombination. All infants received an allogeneic stem cell transplant and exhibited a variety of severe inflammatory complications thereafter, with 2 peri-transplant and 2 delayed deaths. The single long-term transplant survivor showed evidence for genetic rescue through revertant mosaicism overlapping the affected PSMB10 locus. The identified variants (c.166G>C [p.Asp56His] and c.601G>A/c.601G>C [p.Gly201Arg]) were predicted in silico to profoundly disrupt 20S immunoproteasome structure through impaired β-ring/β-ring interaction. Our identification of PSMB10 mutations as a cause of SCID-Omenn syndrome reinforces the connection between PRAAS-related diseases and SCID.

Genetic variants in Folliculin interacting protein 1 (FNIP1) were recently discovered as monogenic causes for immunodeficiency and cardiomyopathy, with only a few patients diagnosed thus far. In this study, we describe a patient harboring a novel genetic variant in FNIP1 causing immunodeficiency with cardiac involvement. Clinical and immunological workups were performed. Genetic evaluation utilizing whole-exome sequencing (WES) and Sanger sequencing was conducted. The index patient (subject II-4) presented with hypertrophic cardiomyopathy, recurrent infections, and chronic diarrhea during infancy. Immune workup revealed agammaglobulinemia and a lack of B lymphocytes. Genetic evaluation identified a homozygous 13-bp duplication variant in FNIP1 (c.52_64dupGCGCCCGGCCGCG, p. Asp22GlyfsTer21) resulting in a frameshift in exon 1/18. She was treated with supplemental intravenous immunoglobulins (IVIg) with good control of sinopulmonary and gastrointestinal manifestations. Her sibling (subject II-1) had similar clinical features, along with dysmorphic facial features and hypotony, and succumbed to cardiogenic shock at the age of 2 months, prior to genetic evaluation. Diagnosis of novel immunodeficiencies promotes our understanding of the immune system, enabling genetic counseling as herein, and may assist in the development of novel medical therapies in the future. FNIP1 loss-of-function should be considered in patients presenting in infancy with cardiac manifestations along with agammaglobulinemia (and B-cell lymphopenia).

Severe combined immunodeficiency (SCID) is a group of disorders caused by mutations in genes involved in the process of lymphocyte maturation and function. CRISPR-Cas9 gene editing of the patient's own hematopoietic stem and progenitor cells (HSPCs) ex vivo could provide a therapeutic alternative to allogeneic hematopoietic stem cell transplantation, the cur-rent gold standard for treatment of SCID. To eliminate the need for scarce patient samples, we engineered genotypes in healthy donor (HD)-derived CD34+ HSPCs using CRISPR-Cas9/rAAV6 gene-editing, to model both SCID and the therapeutic outcomes of gene-editing therapies for SCID via multiplexed homology-directed repair (HDR). First, we developed a SCID disease model via biallelic knockout of genes critical to the development of lymphocytes; and second, we established a knockin/knockout strategy to develop a proof-of-concept single-allelic gene correction. Based on these results, we performed gene correction of RAG2-SCID patient-derived CD34+ HSPCs that successfully developed into CD3+ T cells with diverse TCR repertoires in an in vitro T cell differentiation platform. In summary, we present a strategy to determine the optimal configuration for CRISPR-Cas9 gene correction of SCID using HD-derived CD34+ HSPCs, and the feasibility of translating this gene correction approach in patient-derived CD34+ HSPCs.

Mutations affecting T-cell receptor (TCR) signaling typically cause combined immunodeficiency (CID) due to varying degrees of disturbed T-cell homeostasis and differentiation. Here, we describe two cousins with CID due to a novel nonsense mutation in LCK and investigate the effect of this novel nonsense mutation on TCR signaling, T-cell function, and differentiation. Patients underwent clinical, genetic, and immunological investigations. The effect was addressed in primary cells and LCK-deficient T-cell lines after expression of mutated LCK. RESULTS: Both patients primarily presented with infections in early infancy. The LCK mutation led to reduced expression of a truncated LCK protein lacking a substantial part of the kinase domain and two critical regulatory tyrosine residues. T cells were oligoclonal, and especially naïve CD4 and CD8 T-cell counts were reduced, but regulatory and memory including circulating follicular helper T cells were less severely affected. A diagnostic hallmark of this immunodeficiency is the reduced surface expression of CD4. Despite severely impaired TCR signaling mTOR activation was partially preserved in patients' T cells. LCK-deficient T-cell lines reconstituted with mutant LCK corroborated partially preserved signaling. Despite detectable differentiation of memory and effector T cells, their function was severely disturbed. NK cell cytotoxicity was unaffected. Residual TCR signaling in LCK deficiency allows for reduced, but detectable T-cell differentiation, while T-cell function is severely disturbed. Our findings expand the previous report on one single patient on the central role of LCK in human T-cell development and function.

Introduction: Cartilage-hair hypoplasia (CHH) syndrome is a rare autosomal recessive syndrome associated with skeletal dysplasia, varying degrees of combined immunodeficiency (CID), short stature, hair hypoplasia, macrocytic anemia, increased risk of malignancies, and Hirschsprung disease.Purpose: To provide clinical and immunological insights obtained from 2 unrelated patients who displayed clinical characteristics of CHH.Methods: Two patients with suspected CHH syndrome due to skeletal dysplasia and immunodeficiency underwent an immunological and genetic work-up using flow cytometry, next-generation sequencing (NGS) of the immune repertoire, and Sanger sequencing to identify the underlying defects.Results: Patient 1 presented with low birth weight and skeletal dysplasia. Newborn screening was suggestive of T-cell immunodeficiency, as T-cell receptor excision circle levels were undetectable. Both the T-cell receptor (TCR) VB and TCR-g (TRG) repertoires were restricted, with evidence of clonal expansion. Genetic analysis identified compound heterozygous RMRP variants inherited from both parents. Patient 2 presented with recurrent lung and gastrointestinal infections, skeletal dysplasia, failure to thrive, and hepatomegaly. The polyclonal pattern of the TCRB repertoire was normal, with only slight overexpression of TCR-BV20 and restricted expression of VBs. TRG expressed a normal diverse repertoire, similar to that of the healthy control sample. Genetic analysis identified biallelic novel regulatory variants in RMRP. Both parents are carriers of this mutation.Conclusion: Our findings demonstrate how the immunological work-up, supported by genetic findings, can dramatically change treatment and future outcome in patients with the same clinical syndrome.