Respuesta inmune y terapia gó©nica con vectores virales adenoasociados / Immune response and gene therapy with adenoassociated viral vectors

En los ó;ºltimos aó;±os la terapia gó;©nica se ha posicionado como una opció;n real y segura en el desarrollo de alternativas terapó;©uticas para la cura y la prevenció;n de diferentes enfermedades. Consiste en la inserció;n de material genó;©tico en un tejido o có;©lula defectuosa, mediante el uso de un vector. Existen varias consideraciones para seleccionar el vector más apropiado, incluyendo el potencial de unió;n y entrada a la có;©lula diana, la capacidad de transferencia del material genó;©tico al nó;ºcleo, la habilidad de expresió;n del inserto y la ausencia de toxicidad. En el panorama actual, los vectores virales más utilizados son los derivados de los virus adenoasociados (AAV). Características como su bioseguridad, baja toxicidad y tropismo selectivo, han posibilitado su evaluació;n como opció;n terapó;©utica en un amplio nó;ºmero de enfermedades monogó;©nicas o complejas. A pesar de sus ventajas, los vectores AAV presentan inconvenientes, siendo el más importante la respuesta inmune del paciente al vector, especialmente la respuesta mediada por anticuerpos neutralizantes (NAb). Los NAb disminuyen la transducció;n del vector e impiden la expresió;n del gen que transporta, limitando su aplicació;n clínica. Por lo tanto, identificar y cuantificar la presencia y actividad de los NAbs, es el primer paso en cualquier protocolo de terapia gó;©nica con vectores AAV. La presencia de NAb depende principalmente de la exposició;n al virus en la naturaleza y varía drásticamente segó;ºn edad, localizació;n geográfica y estado de salud de la persona evaluada.

In recent years, gene therapy has been positioned as a real and safe option in the development of therapeutic alternatives for the cure and prevention of different diseases. It consists in the insertion of genetic material in a defective tissue or cell, through the use of a vector. There are several considerations for selecting the most appropriate vector, including the potential for binding and entry to the target cell, the ability of the genetic material to transfer to the nucleus, the ability to express the insert, and the absence of toxicity. In the current scenario, the most commonly used viral vectors are those derived from adeno-associated viruses (AAV). Characteristics such as biosafety, low toxicity and selective tropism have enabled its evaluation as a therapeutic option in many monogenic or complex diseases. Despite their advantages, AAV vectors have drawbacks, the most important being the patient’s immune response to the vector, especially the response mediated by neutralizing antibodies (NAb). NAbs decrease the transduction of the vector and prevent the expression of the gene it transports, limiting its clinical application. Therefore, identifying and quantifying the presence and activity of NAbs is the first step in any gene therapy protocol with AAV vectors. The presence of NAbs depends mainly on exposure to the virus in nature and varies drastically according to age, geographic location and health status of the person evaluated.

Estudio de marcadores de infección e inmunidad frente a parvovirus B19 en Córdoba, Argentina

RESUMEN: El parvovirus B19 (B19V), descubierto en 1974, es un patógeno humano de distribución global. Si bien su infección puede transcurrir de manera asintomática, es conocido por ser capaz de causar un amplio espectro de manifestaciones clínicas como son el eritema infeccioso, artropatías, crisis aplásica en individuos con recambio acelerado de glóbulos rojos por una enfermedad de base, y anemia crónica en inmunocomprometidos, entre otras patologías en las que aún se investiga la causalidad del B19V. La infección durante el embarazo es un riesgo potencial para el feto...

ABSTRACT: Parvovirus B19 (B19V), discovered in 1974, is a pathogen of global distribution. While the infection can be asymptomatic, it is known for being able to cause a wide range of clinical manifestations such as erythema infectiosum (or fitth disease), arthropathy, aplastic crisis in persons with accelerated red blood cell renewal due to a base disease, and chronic anemia in immunocompromised individuals, among other pathologies inwhich B19V causality is still under research. In addition, infection during pregnancy is a potential risk for the fetus...

Cynomolgus monkeys (Macaca fascicularis) experimentally infected with B19V and hepatitis A virus: no evidence of the co-infection as a cause of acute liver failure

This study was conducted to analyse the course and the outcome of the liver disease in the co-infected animals in order to evaluate a possible synergic effect of human parvovirus B19 (B19V) and hepatitis A virus (HAV) co-infection. Nine adult cynomolgus monkeys were inoculated with serum obtained from a fatal case of B19V infection and/or a faecal suspension of acute HAV. The presence of specific antibodies to HAV and B19V, liver enzyme levels, viraemia, haematological changes, and necroinflammatory liver lesions were used for monitoring the infections. Seroconversion was confirmed in all infected groups. A similar pattern of B19V infection to human disease was observed, which was characterised by high and persistent viraemia in association with reticulocytopenia and mild to moderate anaemia during the period of investigation (59 days). Additionally, the intranuclear inclusion bodies were observed in pro-erythroblast cell from an infected cynomolgus and B19V Ag in hepatocytes. The erythroid hypoplasia and decrease in lymphocyte counts were more evident in the co-infected group. The present results demonstrated, for the first time, the susceptibility of cynomolgus to B19V infection, but it did not show a worsening of liver histopathology in the co-infected group.

Clinical features and laboratory findings of human parvovirus B19 in human immunodeficiency virus-infected patients

Immunocompromised patients may develop severe chronic anaemia when infected by human parvovirus B19 (B19V). However, this is not the case in human immunodeficiency virus (HIV)-infected patients with good adherence to highly active antiretroviral treatment (HAART). In this study, we investigated the clinical evolution of five HIV-infected patients receiving HAART who had B19V infections confirmed by serum polymerase chain reaction. Four of the patients were infected with genotype 1a strains and the remaining patient was infected with a genotype 3b strain. Anaemia was detected in three of the patients, but all patients recovered without requiring immunoglobulin and/or blood transfusions. In all cases, the attending physicians did not suspect the B19V infections. There was no apparent relationship between the infecting genotype and the clinical course. In the HAART era, B19V infections in HIV-positive patients may be limited, subtle or unapparent.

Parvovirus B19 seroconversion in a cohort of human immunodeficiency virus-infected patients

Erythrovirus B19 (B19V) infection may cause red cell aplasia in patients infected with human immunodeficiency virus (HIV). The introduction of highly active antiretroviral therapy (HAART) has improved the immune function of these patients by modifying the course of B19V infection. The purpose of this study was to estimate the frequency of B19 seroconversion in a cohort of HIV-infected patients and evaluate the occurrence of B19V-related anaemia during the seroconversion period. Adult HIV-infected patients were studied at a public hospital in Niterói, state of Rio de Janeiro, Brazil. IgG and IgM antibodies against B19V were detected by an enzyme-linked immunosorbent assay and B19 viraemia was assayed by polymerase chain reaction. Medical records were reviewed for any clinical evaluation of anaemia. Seroconversion was detected in 31.8% of the 88 individuals who began the study as anti-B19V IgG-negative. No clinical manifestations of B19V infection were detected during the period of seroconversion. Patients who seroconverted were 5.40 times more likely to have anaemia than those who did not [odds ratio 5.40 (95% confidence interval: 1.33-22.93)]. Anaemia was detected in eight patients. All patients recovered from anaemia by either beginning or continuing HAART, without requiring blood transfusions. In the HAART era, B19V infection may only be associated with a course of disease characterised by less severe chronic anaemia. This milder course of B19V-associated disease is likely due to the increased immune function of HAART-treated patients.

Profile of circulating levels of IL-1Ra, CXCL10/IP-10, CCL4/MIP-1β and CCL2/MCP-1 in dengue fever and parvovirosis

Dengue virus (DENV) and parvovirus B19 (B19V) infections are acute exanthematic febrile illnesses that are not easily differentiated on clinical grounds and affect the paediatric population. Patients with these acute exanthematic diseases were studied. Fever was more frequent in DENV than in B19V-infected patients. Arthritis/arthralgias with DENV infection were shown to be significantly more frequent in adults than in children. The circulating levels of interleukin (IL)-1 receptor antagonist (Ra), CXCL10/inducible protein-10 (IP-10), CCL4/macrophage inflammatory protein-1 beta and CCL2/monocyte chemotactic protein-1 (MCP-1) were determined by multiplex immunoassay in serum samples obtained from B19V (37) and DENV-infected (36) patients and from healthy individuals (7). Forward stepwise logistic regression analysis revealed that circulating CXCL10/IP-10 tends to be associated with DENV infection and that IL-1Ra was significantly associated with DENV infection. Similar analysis showed that circulating CCL2/MCP-1 tends to be associated with B19V infection. In dengue fever, increased circulating IL-1Ra may exert antipyretic actions in an effort to counteract the already increased concentrations of IL-1β, while CXCL10/IP-10 was confirmed as a strong pro-inflammatory marker. Recruitment of monocytes/macrophages and upregulation of the humoral immune response by CCL2/MCP-1 by B19V may be involved in the persistence of the infection. Children with B19V or DENV infections had levels of these cytokines similar to those of adult patients.

A pilot study on parvovirus B19 infection in paediatric haematological malignancies.

Background & objectives: Leukaemia and lymphoma are common paediatric haematological malignancies acquiring human parvovirus B19 (B19) infection. In some studies anaemia has been found in children with acute lymphoblastic leukaemia (ALL) during maintenance therapy and rarely in lymphoma. We studied frequency of B19 infection and its implications in new onset acute leukaemia (mostly ALL) and lymphoma in children. Methods: Seventy serum samples from 35 children (age <12 yr, 29 males) newly diagnosed with haematological malignancies (on induction therapy) were collected together with 34 controls (solid tumours). Children were examined clinically and for anti-B19 IgM antibodies by quantitative ELISA and B19 DNA by PCR (VP1-VP2) and nested-PCR (VP1 unique). Bone marrow aspirates were examined histopathologically, whenever possible. Results: Of the 35 children, 22 had acute leukaemia while 13 had lymphoma. B19 infection was seen in six (17.1%) of 35 children (5 ALL, 1 NHL), two at diagnosis and four during follow up compared to none in the control. Among five B19 IgM positive ALL (n=18) children, two had B19 genome and two had giant pronormoblasts (lantern cells; but one lacked B19 DNA). Of the 70 serum samples tested, eight (11.4%) had anti-B19 IgM as two children had persistent B19 infection and one showed atypical maculopapular rashes (lower limbs) while 12 (34.3%) had anti-B19 IgG antibodies. B19 infected children had unexplained anaemia (80%), required more blood transfusions (6.6 ± 4.8 Units vs 3.0 ± 2.6 Units) besides induction chemotherapy was delayed (60%) and required longer duration of therapy (29.2 ± 20 vs 6.3 ± 7.8 days) (P<0.02). Five children (2 ALL, 2 AML, 1 NHL) died but none were infected with B19. Interpretation & conclusions: B19 infection should be considered in children with ALL as it frequently caused unexplained anaemia and delay in induction chemotherapy.

Parvovirus B19 antibodies and correlates of infection in pregnant women attending an antenatal clinic in central Nigeria

Human parvovirus B19 infection is associated with spontaneous abortion, hydrops foetalis, intrauterine foetal death, erythema infectiosum (5th disease), aplastic crisis and acute symmetric polyarthropathy. However, data concerning Nigerian patients with B19 infection have not been published yet. The purpose of this study was to establish the prevalence of B19 IgG and IgM antibodies, including correlates of infection, among pregnant women attending an antenatal clinic in Nigeria. Subsequent to clearance from an ethical committee, blood samples were collected between August-November 2008 from 273 pregnant women between the ages of 15-40 years who have given their informed consent and completed self-administered questionnaires. Recombinant IgG and IgM enzyme linked immunosorbent assay kits (Demeditec Diagnostics, Germany) were used for the assays. Out of the 273 participants, 111 (40.7 percent) had either IgG or IgM antibodies. Out of these, 75 (27.5 percent) had IgG antibodies whereas 36 (13.2 percent) had IgM antibodies, and those aged 36-40 years had the highest prevalence of IgG antibodies. Significant determinants of infection (p < 0.05) included the receipt of a blood transfusion, occupation and the presence of a large number of children in the household. Our findings have important implications for transfusion and foeto-maternal health policy in Nigeria. Routine screening for B19 IgM antibodies and accompanying clinical management of positive cases should be made mandatory for all Nigerian blood donors and women of childbearing age.

Falla hepática fulminante por parvovirus B 19 y trasplante hepático, caso clínico / Acute liver failure by Parvovirus B19 and liver transplant: case report

Introducción: La infección por parvovirus humano B19 (PHB 19) produce un amplio rango de enfermedades que van desde eritema infeccioso en niños hasta artritis aguda en adultos. Algunos estudios sugieren un rol patogénico del PHB 19 en el desarrollo de la hepatitis aguda (HA) y falla hepática fulminante (FHF) en niños y adultos. La Anemia aplástica (AA) es una complicación reconocida de la HA y FHF por PHB 19. Objetivo: Reportar un caso de FHF por infección por PHB 19 y revisar la literatura. Caso clínico: Niña de 7 años de edad con HA que en una semana desarrolló FHF con serología IgM anti-PHB 19 positiva. Otras causas virales, autoinmunes, metabólicas o toxicas fueron descartadas. Fue sometida a trasplante hepático ortotópico (THO) y un año después no ha presentado complicaciones. Conclusiones: El PHB 19 puede causar HA y FHF, su oportuno diagnóstico y tratamiento, que en el caso de la FHF incluye el THO puede resultar en un pronóstico favorable.

Anti-VP1 and anti-VP2 antibodies detected by immunofluorescence assays in patients with acute human parvovirus B19 infection

Acute human parvovirus B19 infection is followed by an antibody response to the structural proteins of the viral capsid (VP1 and VP2). We used 80 sera collected from 58 erythema infectiosum and 6 transient aplastic crisis patients to test IgM and IgG antibodies against these two proteins in an immunofluorescence assay (IFA) using Sf9 cells infected with recombinant baculovirus expressing either VP1 or VP2 antigen. Although less sensitive than IgM capture enzyme immunoassay using native antigen (MACEIA), we could detect anti-VP1 or anti-VP2 IgM antibodies by IFA in 49 patients with acute infection (76.6 percent). Detection of IgG anti-VP1 and anti-VP2 by IFA, however, was as sensitive as IgG detection by indirect enzyme immunoassay. By applying IgG avidity IFA to sera of the 15 IgM IFA negative patients we were able to confirm acute infection in further 12 cases by IFA. Overall, acute infection was confirmed by IFA in 61 (95.3 percent) of the 64 patients

Avaliaçäo da resposta sorológica de suínos imunizados contra parvovirose suína com uma vacina inativada experimental / Serological evaluation of immunized pigs with an inactivated experimental vaccine against swine parvovirus

Foi estudada a resposta sorológica de leitoas e porcas vacinadas e revacinadas com intervalo de 15 e 21 dias com uma vacina inativada contra parvovirose suína. Os animais vacinados assim como o rebanho de origem eram livres de anticorpos inibidores de hemoaglutinaçäo (HI). Aos 15 dias após a revacinaçäo, 87,5 por cento dos animais vacinados apresentaram títulos HI variando de 1:40 a 1:1280. O intervalo de revacinaçäo e a diferença de títulos HI säo discutidos

Human B19 parvovirus infetion: an example of multiple pathogenic effects determined by differences in host susceptibility

B19 infection offers some general lessons about human viruses and their possible effects on the human host, as follows: (1) Ubiquitous apparently benign viruses may have severe effects on a compromissed host. The virus may be invariable but the host can have diverse susceptibilities. (2) B19 and some other human viruses (through for none is the evidence so clear as for B19) have narrowly targetted effects. The host cell of B19 is a specialised progenitor of mature red cells: impairment of the function of this cell by B19 may cause profound anaemia. (3) The 'normal'host response to B19 may also cause disease, though this is slef limiting. (4) The effects of malfunction of the virus'target cell are exacerbated when the immune response is impaired by congenital or acquired immunodeficiency, immunosupressive therapy or, in the case of the fetus, developmental immaturity that allows the virus to persist

Eritema infeccioso: una parvovirosis / Infectious erythema: a parvovirosis

Se realizó una actualización de eritema infeccioso, una enfermedad exantemática de la infancia que recientemente se ha asociado a parvovirus humanos y se presentan 2 casos en que se comprobó el diagnóstico por alza de IgM específica, mediante la técnica de Radioinmunoanálisis