Mukesh Desai

Autoimmune lymphoproliferative syndrome caused by a homozygous null FAS ligand (FASLG) mutation

BACKGROUND Autoimmune lymphoproliferative syndrome (ALPS) is characterized by chronic nonmalignant lymphoproliferation, accumulation of double-negative T cells, hypergammaglobulinemia G and A, and autoimmune cytopenia. OBJECTIVES Although mostly associated with FAS mutations, different genetic defects leading to impaired apoptosis have been described in patients with ALPS, including the FAS ligand gene (FASLG) in rare cases. Here we report on the first case of complete FAS ligand deficiency caused by a homozygous null mutant. METHODS Double-negative T-cell counts and plasma IL-10 and FAS ligand concentrations were determined as ALPS markers. The FASLG gene was sequenced, and its expression was analyzed by means of Western blotting. FAS ligand function was assessed based on reactivation-induced cell death. RESULTS We describe a patient born to consanguineous parents who presented with a severe form of ALPS caused by FASLG deficiency. Although the clinical presentation was compatible with a homozygous FAS mutation, FAS-induced apoptosis was normal, and plasma FAS ligand levels were not detectable. This patient carries a homozygous, germline, single-base-pair deletion in FASLG exon 1, leading to a premature stop codon (F87fs x95) and a complete defect in FASLG expression. The healthy parents were each heterozygous for the mutation, confirming its recessive trait. CONCLUSION FAS ligand deficiency should be screened in patients presenting with ALPS features but lacking the usual markers, including plasma soluble FAS ligand and an in vitro apoptotic defect. An activation-induced cell death test could help in discrimination.

Hemolytic anemia and distal renal tubular acidosis in two Indian patients homozygous for SLC4A1/AE1 mutation A858D

Familial distal renal tubular acidosis (dRTA) can be caused by mutations in the Cl2/HCO32 exchanger of the renal Type A intercalated cell, kidney AE1/SLC4A1. dRTA-associated AE1 mutations have been reported in families from North America, Europe, Thailand, Malaysia, Papua-New Guinea, Taiwan, and the Philippines, but not India. The dRTA mutation AE1 A858D has been detected only in the context of compound heterozygosity. We report here two unrelated Indian patients with combined hemolytic anemia and dRTA who share homozygous A858D mutations of the AE1/SLC4A1 gene. The mutation creates a novel restriction site that is validated for diagnostic screening.

Current Updates on Classification, Diagnosis and Treatment of Hemophagocytic Lymphohistiocytosis (HLH)

Hemophagocytic lymphohistiocytosis (HLH) is a life threatening hyperinflammatory syndrome characterized by excessive activation of macrophages and T cells resulting from defective cytotoxicity. Severe hyperinflammation caused by uncontrolled proliferation of activated lymphocytes and histiocytes (macrophages) secreting high amounts of inflammatory cytokines threatens the life of the patient and may lead to death unless arrested by appropriate treatment. HLH can be caused either by certain underlying genetic diseases (familial HLH), or may also occur due to particular triggers in patients with no known inherited disorder (acquired HLH). Due to life threatening nature of the disease, early diagnosis and initiation of immunosuppressive therapy is extremely important. HLH diagnosis is based on constellation of clinical manifestations and laboratory parameters which often overlap with those of severe infection or sepsis. Identification of patients with familial HLH and their underlying genetic defects requires specialized laboratory tests and is important for predicting relapses and planning early therapeutic hematopoietic stem cell transplantation (HSCT). A high suspicion and thorough clinical, immunological and genetic work-up is required for diagnosis of HLH. Prompt initiation of adequate treatment is essential for the survival. Substantial progress has been made in exploring the complex cause and pathophysiology of HLH and also in management of HLH patients.

Spectrum of perforin gene mutations in familial hemophagocytic lymphohistiocytosis (FHL) patients in India

BACKGROUND Inherited perforin deficiency is a rare autosomal recessive disorder that causes severe form of hemophagocytic lymphohistiocytosis (FHL2). The main aim of this study was to analyze the nature of gene mutations in a cohort of Indian patients with FHL2 and to utilize this knowledge for genetic counseling and prenatal diagnosis. METHODS 13 HLH patients with abnormal perforin expression on NK cells by flow cytometry were included in the study. The entire coding region and intronic splice sites of the PRF1 gene were sequenced from the genomic DNA of these patients. RESULTS 10 patients from the present series had an early presentation with severe clinical manifestations, while 3 had a delayed onset with unusual presenting features viz Hodgkins lymphoma, tuberculosis and acute lymphoblastic leukemia. Sequence analysis revealed 11 different mutations (8 novel and 3 previously reported) spread over the entire coding region of PRF1 gene. Missense mutation Trp129Ser in heterozygous state was present in all the 3 patients with a delayed onset of the disease. CONCLUSION A wide heterogeneity was observed in the nature of mutations in Indian FHL2 patients. Molecular characterization of PRF1 gene was not only used in the confirmation of diagnosis but also in genetic counseling and pre-natal diagnosis in affected families.

Clinical profile of leukocyte adhesion deficiency type I

Leukocyte adhesion deficiency type I (LAD-I) is a rare, inherited immunodeficiency with defect in the recruitment of leukocyte to the site of inflammation. Patients with severe LAD-I have absent or markedly reduced expression of CD18 and CD11. Here we report clinical profile of 7 cases of LAD-I diagnosed at our center over a period of 3 years. Recurrent skin and mucous membrane infections were the major presenting manifestations. All children had a history of delayed cord separation.

X-linked hyper IgM syndrome: Clinical, immunological and molecular features in patients from India☆☆☆

BACKGROUND X-linked hyper-IgM (XHIM) is a primary immunodeficiency disorder characterized by recurrent infections, low serum IgG and IgA and normal or elevated IgM. It results from mutations in the CD40 ligand (CD40L) gene. Confirmation of diagnosis with identification of underlying molecular defect is important for the initiation of appropriate therapeutic interventions, including immunoglobulin replacement, antibiotics and bone marrow transplantation. METHODS To investigate the molecular basis of XHIM, we evaluated 7 patients with suspected XHIM and abnormal CD40L expression on activated CD4(+) T lymphocytes. The entire coding region and intronic splice sites of the CD40L gene were sequenced from the genomic DNA of the patients. RESULTS 7 mutations; 3 nonsense (c.172delA, c.A229T, c.C478T), 1 missense (c.A506G) and 3 splice sites [c.346+2(T→C), c.289-1(G→C), c.346+1(G→T)] were identified, out of which 5 were novel. CONCLUSION A wide heterogeneity in the nature of mutations has been observed in Indian XHIM patients in the present study. Identification of mutations in this rare disorder will help in genetic diagnosis in affected families which could be further useful in prenatal diagnosis.

Molecular characterization of leukocyte adhesion deficiency-I in Indian patients: Identification of 9 novel mutations

PURPOSE Leukocyte adhesion deficiency type-I (LAD-I) is caused by mutations in the ITGB2 gene, encoding the β2-subunit of β2-integrin (CD18) which leads to markedly reduced expression of CD18 on leukocytes resulting into recurrent life threatening infections. Here we aim to identify the molecular defects underlying LAD-I in Indian patients and correlate with the clinical presentation. METHODS Blood was collected from 30 patients and their parents for absolute neutrophil count, expression of CD18 and CD11 by flow cytometry and DNA extraction. PCR and DNA sequencing of the ITGB2 gene was done for mutation characterization. RESULTS Phenotypically, 22 patients were LAD-I(0), 1 was LAD-I(-) and 7 were LAD-I(+) showing no expression and reduced expression of CD18 respectively. Nine novel mutations in 15 patients and 11 known mutations in 16 patients were detected. Prenatal diagnosis was performed for 5 families. CONCLUSION In this study 30 patients were phenotypically and genotypically evaluated for a less known disease LAD-I. Unavailability of curative options to majority of the patients and high cost of supportive care emphasize the need to increase awareness about a suspicious case so that timely management can be given to the patient and prenatal diagnosis can be offered to their families.

Purine nucleoside phosphorylase deficiency with a novel PNP gene mutation: a first case report from India

The authors report a case of purine nucleoside phosphorylase (PNP) deficiency for the first time from India. The case presented with recurrent severe infections, developmental delays, seizures and progressive neurological deterioration. The diagnosis of primary immunodeficiency disorder was delayed in spite of recurrent infection due to predominant neurological symptoms. Sequencing of the PNP gene revealed a novel mutation resulting in a premature stop codon.

Clinical, Immunological, and Molecular Findings in Five Patients with Major Histocompatibility Complex Class II Deficiency from India

Major histocompatibility complex (MHC) class II deficiency is a rare autosomal recessive form of primary immunodeficiency disorder (PID) characterized by the deficiency of MHC class II molecules. This deficiency affects the cellular and humoral immune response by impairing the development of CD4+ T helper (Th) cells and Th cell-dependent antibody production by B cells. Affected children typically present with severe respiratory and gastrointestinal tract infections. Hematopoietic stem cell transplantation (HSCT) is the only curative therapy available for treating these patients. This is the first report from India wherein we describe the clinical, immunological, and molecular findings in five patients with MHC class II deficiency. Our patients presented with recurrent lower respiratory tract infection as the most common clinical presentation within their first year of life and had a complete absence of human leukocyte antigen-antigen D-related (HLA-DR) expression on B cells and monocytes. Molecular characterization revealed novel mutations in RFAXP, RFX5, and CIITA genes. Despite genetic heterogeneity, these patients were clinically indistinguishable. Two patients underwent HSCT but had a poor survival outcome. Detectable level of T cell receptor excision circles (TRECs) were measured in our patients, highlighting that this form of PID may be missed by TREC-based newborn screening program for severe combined immunodeficiency.

Clinical, Immunological, and Molecular Findings in four cases of B cell Expansion with NF-κB and T cell Anergy (BENTA) Disease for the first time from India.

B cell expansion with NF-κB and T cell anergy (BENTA) is a rare primary immunodeficiency disorder caused by mutations in the CARD11 gene and results in constitutive NF-κB activation in B and T cells. Affected patients present with polyclonal expansion of B cells at an early age with splenomegaly, lymphadenopathy, and mild autoimmunity. Here, we discuss four BENTA cases with unusual clinical manifestations not previously reported. All patients showed previously reported gain-of-function mutations (G123S, G123D, and C49Y) in the CARD11 gene. Severe autoimmune manifestations were noted for the first time in all our patients.