Research

Guidelines for genetic studies in single patients: lessons from primary immunodeficiencies.

February 20, 2015 By Manish Butte

Guidelines for genetic studies in single patients: lessons from primary immunodeficiencies.

J Exp Med. 2014 Oct 20;211(11):2137-49

Authors: Casanova JL, Conley ME, Seligman SJ, Abel L, Notarangelo LD

Abstract
Can genetic and clinical findings made in a single patient be considered sufficient to establish a causal relationship between genotype and phenotype? We report that up to 49 of the 232 monogenic etiologies (21%) of human primary immunodeficiencies (PIDs) were initially reported in single patients. The ability to incriminate single-gene inborn errors in immunodeficient patients results from the relative ease in validating the disease-causing role of the genotype by in-depth mechanistic studies demonstrating the structural and functional consequences of the mutations using blood samples. The candidate genotype can be causally connected to a clinical phenotype using cellular (leukocytes) or molecular (plasma) substrates. The recent advent of next generation sequencing (NGS), with whole exome and whole genome sequencing, induced pluripotent stem cell (iPSC) technology, and gene editing technologies-including in particular the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology-offer new and exciting possibilities for the genetic exploration of single patients not only in hematology and immunology but also in other fields. We propose three criteria for deciding if the clinical and experimental data suffice to establish a causal relationship based on only one case. The patient’s candidate genotype must not occur in individuals without the clinical phenotype. Experimental studies must indicate that the genetic variant impairs, destroys, or alters the expression or function of the gene product (or two genetic variants for compound heterozygosity). The causal relationship between the candidate genotype and the clinical phenotype must be confirmed via a relevant cellular phenotype, or by default via a relevant animal phenotype. When supported by satisfaction of rigorous criteria, the report of single patient-based discovery of Mendelian disorders should be encouraged, as it can provide the first step in the understanding of a group of human diseases, thereby revealing crucial pathways underlying physiological and pathological processes.

PMID: 25311508 [PubMed – indexed for MEDLINE]

Human T-lymphotropic virus type 1-infected cells secrete exosomes that contain Tax protein.

February 20, 2015 By Manish Butte

Human T-lymphotropic virus type 1-infected cells secrete exosomes that contain Tax protein.

J Biol Chem. 2014 Aug 8;289(32):22284-305

Authors: Jaworski E, Narayanan A, Van Duyne R, Shabbeer-Meyering S, Iordanskiy S, Saifuddin M, Das R, Afonso PV, Sampey GC, Chung M, Popratiloff A, Shrestha B, Sehgal M, Jain P, Vertes A, Mahieux R, Kashanchi F

Abstract
Human T-lymphotropic virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis. The HTLV-1 transactivator protein Tax controls many critical cellular pathways, including host cell DNA damage response mechanisms, cell cycle progression, and apoptosis. Extracellular vesicles called exosomes play critical roles during pathogenic viral infections as delivery vehicles for host and viral components, including proteins, mRNA, and microRNA. We hypothesized that exosomes derived from HTLV-1-infected cells contain unique host and viral proteins that may contribute to HTLV-1-induced pathogenesis. We found exosomes derived from infected cells to contain Tax protein and proinflammatory mediators as well as viral mRNA transcripts, including Tax, HBZ, and Env. Furthermore, we observed that exosomes released from HTLV-1-infected Tax-expressing cells contributed to enhanced survival of exosome-recipient cells when treated with Fas antibody. This survival was cFLIP-dependent, with Tax showing induction of NF-κB in exosome-recipient cells. Finally, IL-2-dependent CTLL-2 cells that received Tax-containing exosomes were protected from apoptosis through activation of AKT. Similar experiments with primary cultures showed protection and survival of peripheral blood mononuclear cells even in the absence of phytohemagglutinin/IL-2. Surviving cells contained more phosphorylated Rb, consistent with the role of Tax in regulation of the cell cycle. Collectively, these results suggest that exosomes may play an important role in extracellular delivery of functional HTLV-1 proteins and mRNA to recipient cells.

PMID: 24939845 [PubMed – indexed for MEDLINE]

Specific retrograde transduction of spinal motor neurons using lentiviral vectors targeted to presynaptic NMJ receptors.

February 20, 2015 By Manish Butte

Specific retrograde transduction of spinal motor neurons using lentiviral vectors targeted to presynaptic NMJ receptors.

Mol Ther. 2014 Jul;22(7):1285-98

Authors: Eleftheriadou I, Trabalza A, Ellison S, Gharun K, Mazarakis N

Abstract
To understand how receptors are involved in neuronal trafficking and to be able to utilize them for specific targeting via the peripheral route would be of great benefit. Here, we describe the generation of novel lentiviral vectors with tropism to motor neurons that were made by coexpressing onto the lentiviral surface a fusogenic glycoprotein (mutated sindbis G) and an antibody against a cell-surface receptor (Thy1.1, p75(NTR), or coxsackievirus and adenovirus receptor) on the presynaptic terminal of the neuromuscular junction. These vectors exhibit binding specificity and efficient transduction of receptor positive cell lines and primary motor neurons in vitro. Targeting of each of these receptors conferred to these vectors the capability of being transported retrogradely from the axonal tip, leading to transduction of motor neurons in vitro in compartmented microfluidic cultures. In vivo delivery of coxsackievirus and adenovirus receptor-targeted vectors in leg muscles of mice resulted in predicted patterns of motor neuron labeling in lumbar spinal cord. This opens up the clinical potential of these vectors for minimally invasive administration of central nervous system-targeted therapeutics in motor neuron diseases.

PMID: 24670531 [PubMed – indexed for MEDLINE]

Epstein-Barr virus: dermatologic associations and implications: part II. Associated lymphoproliferative disorders and solid tumors.

February 19, 2015 By Manish Butte

Epstein-Barr virus: dermatologic associations and implications: part II. Associated lymphoproliferative disorders and solid tumors.

J Am Acad Dermatol. 2015 Jan;72(1):21-34; quiz 35-6

Authors: Eminger LA, Hall LD, Hesterman KS, Heymann WR

Abstract
Epstein-Barr virus (EBV) was the first human virus to be associated with oncogenesis. Over the past few decades, cumulative research has revealed that latent EBV infection may be implicated in the pathogenesis of a heterogeneous group of lymphoproliferative disorders and malignancies occurring in both immunocompetent and immunocompromised hosts. Many of these diseases have either primary or secondary cutaneous manifestations. Serologic studies and EBV-encoded RNA in situ hybridization stains have been used to show the association of EBV with disease; while these findings may imply a role, they do not equate with causation. In part II of this continuing medical education review, the salient features of EBV-associated lymphoproliferative disorders and solid tumors are detailed.

PMID: 25497918 [PubMed – indexed for MEDLINE]

CEACAM1 induces B-cell survival and is essential for protective antiviral antibody production.

February 19, 2015 By Manish Butte

CEACAM1 induces B-cell survival and is essential for protective antiviral antibody production.

Nat Commun. 2015;6:6217

Authors: Khairnar V, Duhan V, Maney SK, Honke N, Shaabani N, Pandyra AA, Seifert M, Pozdeev V, Xu HC, Sharma P, Baldin F, Marquardsen F, Merches K, Lang E, Kirschning C, Westendorf AM, Häussinger D, Lang F, Dittmer U, Küppers R, Recher M, Hardt C, Scheffrahn I, Beauchemin N, Göthert JR, Singer BB, Lang PA, Lang KS

Abstract
B cells are essential for antiviral immune defence because they produce neutralizing antibodies, present antigen and maintain the lymphoid architecture. Here we show that intrinsic signalling of CEACAM1 is essential for generating efficient B-cell responses. Although CEACAM1 exerts limited influence on the proliferation of B cells, expression of CEACAM1 induces survival of proliferating B cells via the BTK/Syk/NF-κB-axis. The absence of this signalling cascade in naive Ceacam1(-/-) mice limits the survival of B cells. During systemic infection with cytopathic vesicular stomatitis virus, Ceacam1(-/-) mice can barely induce neutralizing antibody responses and die early after infection. We find, therefore, that CEACAM1 is a crucial regulator of B-cell survival, influencing B-cell numbers and protective antiviral antibody responses.

PMID: 25692415 [PubMed – as supplied by publisher]

Melanoma in patients with chronic lymphocytic leukemia and non-Hodgkin lymphoma.

February 19, 2015 By Manish Butte

Melanoma in patients with chronic lymphocytic leukemia and non-Hodgkin lymphoma.

J Am Acad Dermatol. 2015 Jan;72(1):78-84

Authors: Famenini S, Martires KJ, Zhou H, Xavier MF, Wu JJ

Abstract
BACKGROUND: The relationship between melanoma and chronic lymphocytic leukemia (CLL) or non-Hodgkin lymphoma (NHL) has been minimally investigated.
OBJECTIVE: The objective of this study was to examine the incidence of melanoma in patients with a history of CLL or NHL, and their associated mortality.
METHODS: Cohorts of Kaiser Permanente Southern California members with a history of CLL and NHL were identified. Age-adjusted incidence density rates of melanoma among patients with CLL or NHL were compared with rates of melanoma among the general population of Kaiser Permanente Southern California patients. The mortality of patients with melanoma was examined using Cox proportional hazards modeling.
RESULTS: The age-adjusted incidence rate per 100,000 person-years for melanoma among patients with either CLL or NHL was 107 (95% confidence interval 84.4-129.6) versus 25.9 among the general population (95% confidence interval 84.4-129.6, P < .001). Patients with melanoma and a history of CLL or NHL had 2.46 greater odds of death compared with those without CLL or NHL (95% confidence interval 1.77-3.41).
LIMITATIONS: This study was retrospective in nature; the International Classification of Diseases, Ninth Revision codes used may contain diagnostic errors; and only overall survival was used in our analysis.
CONCLUSIONS: Patients with a history of CLL or NHL have a higher incidence of melanoma. Patients with CLL or NHL who are subsequently given the diagnosis of melanoma have a higher mortality than patients with melanoma without a preceding diagnosis of CLL.

PMID: 25440434 [PubMed – indexed for MEDLINE]

Novel TTC37 Mutations in a Patient with Immunodeficiency without Diarrhea: Extending the Phenotype of Trichohepatoenteric Syndrome.

February 18, 2015 By Manish Butte

Novel TTC37 Mutations in a Patient with Immunodeficiency without Diarrhea: Extending the Phenotype of Trichohepatoenteric Syndrome.

Front Pediatr. 2015;3:2

Authors: Rider NL, Boisson B, Jyonouchi S, Hanson EP, Rosenzweig SD, Cassanova JL, Orange JS

Abstract
Unbiased genetic diagnosis has increasingly associated seemingly unrelated somatic and immunological phenotypes. We report a male infant who presented within the first year of life with physical growth impairment, feeding difficulties, hyperemesis without diarrhea, and abnormal hair findings suggestive of trichorrhexis nodosa. With advancing age, moderate global developmental delay, susceptibility to frequent viral illnesses, otitis media, and purulent conjunctivitis were identified. Because of the repeated infections, an immunological evaluation was pursued and identified impaired antibody memory responses following pneumococcal vaccine administration. Immunoglobulin replacement therapy and nutritional support were employed as mainstays of therapy. The child is now aged 12 years and still without diarrhea. Whole exome sequencing identified compound heterozygous mutations in the TTC37 gene, a known cause of the trichohepatoenteric syndrome (THES). This case extends the known phenotype of THES and defines a potential subset for inclusion as an immune overlap syndrome.

PMID: 25688341 [PubMed]

MyD88 in Macrophages Is Critical for Abscess Resolution in Staphylococcal Skin Infection.

February 15, 2015 By Manish Butte

MyD88 in Macrophages Is Critical for Abscess Resolution in Staphylococcal Skin Infection.

J Immunol. 2015 Feb 13;

Authors: Feuerstein R, Seidl M, Prinz M, Henneke P

Abstract
When Staphylococcus aureus penetrates the epidermis and reaches the dermis, polymorphonuclear leukocytes (PMLs) accumulate and an abscess is formed. However, the molecular mechanisms that orchestrate initiation and termination of inflammation in skin infection are incompletely understood. In human myeloid differentiation primary response gene 88 (MyD88) deficiency, staphylococcal skin and soft tissue infections are a leading and potentially life-threatening problem. In this study, we found that MyD88-dependent sensing of S. aureus by dermal macrophages (Mϕ) contributes to both timely escalation and termination of PML-mediated inflammation in a mouse model of staphylococcal skin infection. Mϕs were key to recruit PML within hours in response to staphylococci, irrespective of bacterial viability. In contrast with bone marrow-derived Mϕs, dermal Mϕs did not require UNC-93B or TLR2 for activation. Moreover, PMLs, once recruited, were highly activated in an MyD88-independent fashion, yet failed to clear the infection if Mϕs were missing or functionally impaired. In normal mice, clearance of the infection and contraction of the PML infiltrate were accompanied by expansion of resident Mϕs in a CCR2-dependent fashion. Thus, whereas monocytes were dispensable for the early immune response to staphylococci, they contributed to Mϕ renewal after the infection was overcome. Taken together, MyD88-dependent sensing of staphylococci by resident dermal Mϕs is key for a rapid and balanced immune response, and PMLs are dependent on intact Mϕ for full function. Renewal of resident Mϕs requires both local control of bacteria and inflammatory monocytes entering the skin.

PMID: 25681348 [PubMed – as supplied by publisher]

A novel BTK gene mutation creates a de-novo splice site in an X-linked agammaglobulinemia patient.

February 15, 2015 By Manish Butte

A novel BTK gene mutation creates a de-novo splice site in an X-linked agammaglobulinemia patient.

Gene. 2015 Feb 10;

Authors: Chear CT, Ripen AM, Mohamed SA, Dhaliwal JS

Abstract
Bruton’s tyrosine kinase (BTK), encoded by the BTK gene, is a cytoplasmic protein critical in B cell development. Mutations in the BTK gene cause X-linked agammaglobulinemia (XLA), a primary immunodeficiency with characteristically low or absent B cells and antibodies. This report describes a five year-old boy who presented with otitis externa, arthritis, reduced immunoglobulins and no B cells. Flow cytometry showed undetectable monocyte BTK expression. Sequencing revealed a novel mutation at exon 13 of the BTK gene which created a de novo splice site with a proximal 5 nucleotide loss resulting in a truncated BTK protein. The patient still suffered from ear infection despite intravenous immunoglobulin replacement therapy. In this study, mosaicism was seen only in the mother’s genomic DNA. These results suggest that a combination of flow cytometry and BTK gene analysis is important for XLA diagnosis and carrier screening.

PMID: 25680287 [PubMed – as supplied by publisher]

What has happened in the last 50 years in immunology.

February 14, 2015 By Manish Butte

What has happened in the last 50 years in immunology.

J Paediatr Child Health. 2015 Feb;51(2):135-139

Authors: Wong M

Abstract
Fifty years ago, in 1964, our understanding of the immune system was very rudimentary. Gell and Coombs had just described classes of hypersensitivity reactions, and Bruton had described and commenced immunoglobulin replacement in agammaglobulinaemia. The distinction between T and B cells was not identified and characterised until the 1960s and 1970s. This was followed by increasing recognition of T and B cell collaboration in immune responses and identification of significant immunodeficiencies. CD4 and CD8 T cells were only recognised in the 1970s and 1980s. We now know of five CD4 subsets; dysfunction of each is associated with different disorders. By 2014, advances in technology have enabled identification of the genetic basis of over 240 primary immunodeficiencies. Research into the gut microbiome and vitamin D holds promise for the understanding, treatment and prevention of autoimmune and allergic diseases. Immunoglobulin preparations for the treatment of antibody deficiencies improved with the development of preparations for intravenous then subcutaneous administration, giving patients choice and the ability for home-based treatment, especially if experiencing infusion associated adverse effects. Newborn screening for severe combined immunodeficiency is a reality. Improvements in haemopoietic stem cell transplantation and now gene therapy, albeit still only available in the research setting, are improving long-term survival in primary immunodeficiencies. Biologic therapeutic agents are improving the control of autoimmune disease but potentially leading to secondary immunodeficiency, increasing the risk of opportunistic infection and malignancy. It is an exciting time.

PMID: 25677480 [PubMed – as supplied by publisher]