VZV Infection Research Articles

Bacterial artificial chromosome derived simian varicella virus is pathogenic in vivo

BackgroundVaricella zoster virus (VZV) is a neurotropic alphaherpesvirus that infects humans and results in chickenpox and herpes zoster. A number of VZV genes remain functionally uncharacterized and since VZV is an obligate human pathogen, rigorous evaluation of VZV mutants in vivo remains challenging. Simian varicella virus (SVV) is homologous to VZV and SVV infection of rhesus macaques (RM) closely mimics VZV infection of humans. Recently the SVV genome was cloned as a bacterial artificial chromosome (BAC) and BAC-derived SVV displayed similar replication kinetics as wild-type (WT) SVV in vitro.MethodsRMs were infected with BAC-derived SVV or WT SVV at 4x105 PFU intrabronchially (N=8, 4 per group, sex and age matched). We collected whole blood (PBMC) and bronchoalveolar lavage (BAL) at various days post-infection (dpi) and sensory ganglia during latent infection (>84 dpi) at necropsy and compared disease progression, viral replication, immune response and the establishment of latency.ResultsViral replication kinetics and magnitude in bronchoalveolar lavage cells and whole blood as well as rash severity and duration were similar in RMs infected with SVV BAC or WT SVV. Moreover, SVV-specific B and T cell responses were comparable between BAC and WT-infected animals. Lastly, we measured viral DNA in sensory ganglia from both cohorts of infected RMs during latent infection.ConclusionsSVV BAC is as pathogenic and immunogenic as WT SVV in vivo. Thus, the SVV BAC genetic system combined with the rhesus macaque animal model can further our understanding of viral ORFs important for VZV pathogenesis and the development of second-generation vaccines.

Association of asthma and herpes zoster in children

The incidence of asthma and herpes zoster (HZ) rose concurrently at the end of the 20th century. Investigators took advantage of the setting of Olmstead County, Minnesota, where medical care is “virtually self-contained,” to perform a population-based epidemiologic study to explore whether HZ was associated with asthma. The study was performed before substantial use of varicella vaccine, and no case had received systemic corticosteroid in the three months prior to HZ. Twenty-three percent of cases of HZ had asthma compared with 12.6% of matched controls (aOR: 2.09, 95% CI 1.24-3.52; P = .006). The authors provide arguments about why detection bias is likely not responsible for their finding. They conclude that asthma may be an unrecognized risk factor for reactivation of non—airway-related latent VZV infection and speculate that altered cell-mediated innate immune response could be the biologic link.Article page 816▶ The incidence of asthma and herpes zoster (HZ) rose concurrently at the end of the 20th century. Investigators took advantage of the setting of Olmstead County, Minnesota, where medical care is “virtually self-contained,” to perform a population-based epidemiologic study to explore whether HZ was associated with asthma. The study was performed before substantial use of varicella vaccine, and no case had received systemic corticosteroid in the three months prior to HZ. Twenty-three percent of cases of HZ had asthma compared with 12.6% of matched controls (aOR: 2.09, 95% CI 1.24-3.52; P = .006). The authors provide arguments about why detection bias is likely not responsible for their finding. They conclude that asthma may be an unrecognized risk factor for reactivation of non—airway-related latent VZV infection and speculate that altered cell-mediated innate immune response could be the biologic link. Article page 816▶ Increased Risk of Herpes Zoster in Children with Asthma: A Population-Based Case-Control StudyThe Journal of PediatricsVol. 163Issue 3PreviewTo determine the risk of herpes zoster (HZ) in children with and without asthma. Full-Text PDF

Genome-Wide Analysis of T Cell Responses during Acute and Latent Simian Varicella Virus Infections in Rhesus Macaques

Varicella zoster virus (VZV) is the etiological agent of varicella (chickenpox) and herpes zoster (HZ [shingles]). Clinical observations suggest that VZV-specific T cell immunity plays a more critical role than humoral immunity in the prevention of VZV reactivation and development of herpes zoster. Although numerous studies have characterized T cell responses directed against select VZV open reading frames (ORFs), a comprehensive analysis of the T cell response to the entire VZV genome has not yet been conducted. We have recently shown that intrabronchial inoculation of young rhesus macaques with simian varicella virus (SVV), a homolog of VZV, recapitulates the hallmarks of acute and latent VZV infection in humans. In this study, we characterized the specificity of T cell responses during acute and latent SVV infection. Animals generated a robust and broad T cell response directed against both structural and nonstructural viral proteins during acute infection in bronchoalveolar lavage (BAL) fluid and peripheral blood. During latency, T cell responses were detected only in the BAL fluid and were lower and more restricted than those observed during acute infection. Interestingly, we identified a small set of ORFs that were immunogenic during both acute and latent infection in the BAL fluid. Given the close genome relatedness of SVV and VZV, our studies highlight immunogenic ORFs that may be further investigated as potential components of novel VZV vaccines that specifically boost T cell immunity.

Varicella zoster virus decreases transcription and translation of STAT1 and blocks translocation of pSTAT1 to the nucleus in cerebrovascular adventitial fibroblasts. (P6149)

Abstract Varicella zoster virus (VZV) is the only human virus known to replicate in cerebral arteries and produce stroke. After reactivation, virus travels transaxonally from ganglia and initially infects the arterial adventitia. To analyze the virus-host relationship, we infected primary human cerebrovascular adventitial fibroblasts with VZV and studied the interferon-α initiated STAT pathway, which is commonly activated after virus infection. Quantitative PCR showed that expression of STAT1, STAT2 and Mx1 mRNA were significantly decreased in VZV-infected, but not uninfected adventitial fibroblasts (p<0.05). Western blotting confirmed decreased STAT1 and Mx1 expression in VZV-infected fibroblasts compared to uninfected cells. Although Western blotting did not reveal phosphorylated STAT1 (pSTAT1) in VZV-infected cells, interferon-α is known to trigger STAT1 phosphorylation; thus, we treated uninfected adventitial fibroblasts with interferon-α, after which pSTAT1 expression was readily seen on Western blots. Furthermore, immunocytochemical staining detected pSTAT1 exclusively in the cytoplasm of VZV-infected cells. In interferon-α -treated uninfected cells, pSTAT1 was found exclusively in the nucleus, suggesting that VZV infection blocks egress of pSTAT1 from the cytoplasm to the nucleus in cerebral adventitial fibroblasts.

T cell responses against structural proteins develop rapidly during acute simian varicella virus infection and are maintained during viral latency (P6133)

Abstract Varicella zoster virus (VZV) is the etiological agent of varicella and herpes zoster. Clinical observations suggest that successful control of VZV during both primary infection and reactivation is mediated by cellular rather than humoral immunity. Numerous studies have characterized T cell responses directed against select VZV open reading frames (ORF), but a comprehensive analysis of the T cell response to the entire VZV genome has not yet been conducted. We have shown that intrabronchial inoculation of young rhesus macaques with simian varicella virus (SVV), a homolog of VZV, recapitulates the hallmarks of acute and latent VZV infection in humans. In this study, we characterized the specificity of T cell responses during acute and latent SVV infection. We identified several SVV ORFs targeted by T cell responses in majority of animals examined during both acute and latent infection. We show that structural proteins are targeted by both CD4 and CD8 T cells, and immediate early/early viral proteins are primarily targeted by CD8 T cells. Also, while both CD4 and CD8 T cell responses are detected during acute SVV infection, latent infection elicits primarily CD8 T cell responses. These results suggest that CD4 T cell immunity may wane faster than CD8 T cell immunity. Given the close genome relatedness of SVV and VZV, our studies highlight immunogenic ORFs to be further investigated as potential components of second-generation VZV vaccines designed to boost T cell immunity.

Results of a Phase 2 Clinical Trial Testing the Efficacy of Plerixafor in Combination with Chemotherapy in the Mobilization of Autologous Blood Hematopoietic Progenitor Cells

s / Biol Blood Marrow Transplant 19 (2013) S178eS193 S191 herpetic neuralgia (n1⁄45), severe pain (n1⁄43), scarring (n1⁄41), and motor weakness (n1⁄41); 2 patients required hospitalization and 3 patients developed disseminated zoster. Conclusions: Our limited retrospective analysis suggests a significant reduction in rates of post auto-HCT rates of VZV infection with extended 12month antiviral prophylaxis. VZV infection is a significant complication post auto-HCT, and extended prophylaxis appears to be safe and effective in this setting.

Attenuation of the Adaptive Immune Response in Rhesus Macaques Infected with Simian Varicella Virus Lacking Open Reading Frame 61

Varicella zoster virus (VZV) is a neurotropic alphaherpesvirus that causes chickenpox during primary infection and establishes latency in sensory ganglia. Infection of rhesus macaques (RM) with the homologous simian varicella virus (SVV) recapitulates hallmarks of VZV infection. We have shown that an antisense transcript of SVV open reading frame 61 (ORF61), a viral transactivator, was detected most frequently in latently infected RM sensory ganglia. In this study, we compared disease progression, viral replication, immune response, and the establishment of latency following intrabronchial infection with a recombinant SVV lacking ORF61 (SVVΔORF61) to those following infection with wild-type (WT) SVV. Varicella severity and viral latency within sensory ganglia were comparable in RMs infected with SVVΔORF61 and WT SVV. In contrast, viral loads, B and T cell responses, and plasma inflammatory cytokine levels were decreased in RMs infected with SVVΔORF61. To investigate the mechanisms underlying the reduced adaptive immune response, we compared acute SVV gene expression, frequency and proliferation of dendritic cell (DC) subsets, and the expression of innate antiviral genes in bronchoalveolar lavage (BAL) samples. The abundance of SVV transcripts in all kinetic classes was significantly decreased in RMs infected with SVVΔORF61. In addition, we detected a higher frequency and proliferation of plasmacytoid dendritic cells in BAL fluid at 3 days postinfection in RMs infected with SVVΔORF61, which was accompanied by a slight increase in type I interferon gene expression. Taken together, our data suggest that ORF61 plays an important role in orchestrating viral gene expression in vivo and interferes with the host antiviral interferon response.

Varicella-Zoster Virus ORF12 Protein Activates the Phosphatidylinositol 3-Kinase/Akt Pathway To Regulate Cell Cycle Progression

Varicella-zoster virus (VZV) activates the phosphatidylinositol 3-kinase (PI3K)/Akt pathway and alters cell cycle progression, but the viral protein(s) responsible for these activities is unknown. We previously reported that the VZV open reading frame 12 (ORF12) protein triggers phosphorylation of ERK. Here, we demonstrate that the VZV ORF12 protein also activates the PI3K/Akt pathway to regulate cell cycle progression. Transfection of cells with a plasmid expressing the ORF12 protein induced phosphorylation of Akt, which was dependent on PI3K. Infection of cells with wild-type VZV triggered phosphorylation of Akt, while infection with an ORF12 deletion mutant induced less phosphorylated Akt. The activation of Akt by ORF12 protein was associated with its binding to the p85 subunit of PI3K. Infection of cells with wild-type VZV resulted in increased levels of cyclin B1, cyclin D3, and phosphorylated glycogen synthase kinase 3β (GSK-3β), while infection with the ORF12 deletion mutant induced lower levels of these proteins. Wild-type VZV infection reduced the G(1) phase cell population and increased the M phase cell population, while infection with the ORF12 deletion mutant had a reduced effect on the G(1) and M phase populations. Inhibition of Akt activity with LY294002 reduced the G(1) and M phase differences observed in cells infected with wild-type and ORF12 mutant viruses. In conclusion, we have found that the VZV ORF12 protein activates the PI3K/Akt pathway to regulate cell cycle progression. Since VZV replicates in both dividing (e.g., keratinocytes) and nondividing (neurons) cells, the ability of the VZV ORF12 protein to regulate the cell cycle is likely important for VZV replication in various cell types in the body.

Reduced-Dosage of Three Drugs Combination of Cyclophosphamide, Bortezomib and Dexamethasone (reduced-CyBorD) Retains High Response Rate with Less Toxicities in Elderly Patients with Multiple Myeloma

AbstractAbstract 1865 Background:The three drugs combination of cyclophosphamide, bortezomib and dexamethasone (CyBorD) has been proven to be a powerful regimen for the treatment of the patients with multiple myeloma (MM). However, twice-weekly schedule of bortezomib and high dose dexamethasone has been shown to cause significant toxicities, including severe peripheral neuropathy (PN). Recently, once-weekly bortezomib induction therapy with CyBorD (modified-CyBorD) has shown less toxicity with high response rate equal to original regimen (Reeder, CB et al. Leukemia 2009). However, more than half of the patients developed bortezomib induced PN (BiPN) in modified-CyBorD. Thus it was still concerned adverse event for the transplant ineligible patients, especially frail elderly patients because it often caused interruption or withdrawal of the treatments, resulting in fewer efficacies. Under these circumstances, we introduced the reduced-dosage of CyBorD regimen (reduced-CyBorD) in the transplant ineligible patients with multiple myeloma for continuation of the treatment. Methods:The protocol consisted of bortezomib given intravenously at a dose of 1.3 mg/m2once a week on days 1, 8, 15, cyclophosphamide orally at a dose of 50 mg daily on days 1–21, and dexamethasone orally or intravenously at a dose of 20 mg daily on days 1,2,8,9,15,16 in 4-week cycles. Total of 20 patients, including 12 newly diagnosed and 8 refractory, were treated with reduced-CyBorD and evaluated its efficacy and safety. Results:The median age was 72 years (range from 62 to 81). 14 patients were more than 70 year-old (70%). A half of the patients were female. According ISS, 3 patients were classified in stage I, 6 were in II, and 11 were in III. The overall response was 86.6 % with 26.7 % CR/nCR (1 CR, 3 nCR, 5 VGPR and 4 PR). Hematological adverse events were neutropenia (35%; G1/2 n=7), lymphocytopenia (35%; G1/2 n=1, G3/4, n=1), thrombocytopenia (10%; G1/2 n=2). Non-hematological adverse events were pneumonia (20%; G2 n=2, G3 n=2), VZV infection (15%; G2 n=3), cerebral infarction (5%; G2 n=1). Importantly, only three patients (15%) developed grade 1 PN, and no patient reduced or discontinued bortezomib due to PN. Conclusions:Reduced-CyBorD with three times once-weekly bortezomib retained high efficacy seen in standard and modified CyBorD (4 times bortezomib administration). Furthermore, it was found that this regimen obviously revealed less toxicity, especially BiPN compared with those previous regimens (See Table). Our results suggested that reduced-CyBorD might be safe and effective approach to the transplant ineligible patients, especially elderly frail patients with MM. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Early Consolidation with High Dose Therapy and Autologous Stem Cell Transplantation in HIV-Associated Non Hodgkin Lymphoma At High Risk (Aaipi 2–3). an Interim Report of a Multicenter Phase II Trial.

AbstractAbstract 3124▪▪This icon denotes a clinically relevant abstract Background.Outcome of aggressive HIV-associated NHL (HIV-NHL) with adverse prognostic features is not satisfactory with standard therapy. High dose therapy (HDT) and autologous stem cell transplantation (ASCT) has been demonstrated safe and active in HIV-NHL in the salvage setting. Aims.To define the role of HDT and ASCT in the upfront treatment of HIV-NHL at high risk, in terms of efficacy and toxicity. We report the interim results of a multicenter study including HDT and ASCT as consolidation after first-line treatment of HIV-NHL at high-risk. Patients and methods.Inclusion criteria: untreated aggressive HIV-NHL (including DLBCL, plasmablastic, anaplastic lymphoma), aa-IPI 2–3, age 18–60, CD4+ count >50/mcl and availability of effective HAART. Burkitt and CNS lymphoma are excluded. Patients (pts) receive R-CHOP-21 (CHOP-21 or CHOP-14 for CD20 negative lymphoma) for 6 cycles and, if responsive, undergo stem cells mobilization and collection after Cyclophosphamide 4gr/ms + G-CSF followed by HDT with BEAM and ASCT. Involved-field radiotherapy on previous bulky or residual disease is recommended. Pts receive HAART during the entire treatment program. Overall survival (OS) at 2 years is the primary endpoint. Results.Since January 2007 to July 2012, 23 pts were registered and 20 entered the study. Median age was 47.5 years (range, 27–62). Fifteen pts (75%) had DLBCL, 4 (20%) plasmablastic and 1 (5%) anaplastic lymphoma. ECOG PS was >1 in 11 pts (55%); Ann-Arbor stage III/IV, 5(25%)/15(75%); B symptoms, 12 (60%); LDH > n.v., 18 (90%); aaIPI 2/3, 10 (50%)/10(50%). Fourteen pts (70%) had a prior history of HIV-positivity and 13 (65%) were on HAART at NHL diagnosis; thirteen (65%) had detectable HIV-viremia. Median CD4+ count was 283/mcl (58–571). Nine pts (45%) had HCV infection. Seventeen pts received (R)-CHOP-21 and 3 pts with plasmablastic histology received CHOP-14. One pt is still on treatment and 19/20 are evaluable for (R)-CHOP response. One had toxic death (due to hepatic failure), 2 had prolonged cytopenia (1 with severe hepatic toxicity) that lead to withdrawal of therapy and 16 completed (R)-CHOP therapy: 11 pts had complete remission (CR), 4 partial remission (PR) and 1 disease progression (PD) (ORR 79%,CR 58%, PR 21%, according to intention to treat). One pt is ongoing and 14 collected CD34+ cells after Cyclophosphamide + G-CSF (median CD34+ cells 7.35 × 10e6/Kg, range 2.65–10.0). Two pts had early disease progression after collection, 1 did not receive HDT because of cardiac ejection fraction < 50% at evaluation before BEAM and 1 is ongoing. So far, 10 pts received ASCT according to the protocol. Moreover, three pts received radiotherapy (2 on previous bulky and 1 on testicular disease).HDT-related toxicities included 2 grade II and 5 grade III GI and 1 grade III hepatic toxicity. Prior to engraftment 1 VZV infection, 1 Clostridium difficile colitis, 1 sigmoiditis and 5 episodes of FUO were registered. There were no transplant-related deaths. One case of CMV reactivation and no opportunistic infections were registered during observation. All transplanted pts are alive and relapse-free after a median of 43 ms (range, 4–58) after transplant (Figure 1). In the entire series, with a median f-up of 29.5 ms (range, 4–65), the 2-years PFS and OS of the entire series from study entry were 73% (+10.3%) and 76% (+10.6%), respectively (Figure 2). Conclusions.This is the first prospective trial addressing the role of HDT and ASCT in first line treatment of HIV-LNH. The procedure was well tolerated and the clinical results highly encouraging. Interim evaluation of OS in this very high risk series of pts is satisfactory and accrual is ongoing. [Display omitted] [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Prolonged Hypogammaglobulinemia After Rituximab Therapy for Post-Transplant EBV-Lymphoproliferative Disorder Following Hematopoietic Stem Cell Transplantation in Pediatric Patients

Abstract 4480Post-transplant lymphoproliferative disorder (PTLD) caused by Epstein Barr Virus (EBV) is a severe complication in allogeneic hematopoietic stem cell transplantation (HSCT). Rituximab is an effective treatment, now commonly used as early therapy against B-cell PTLD and preemptive treatment of EBV reactivation. In first trials, rituximab was considered to have little adverse events. As exposure to this therapy is increasing worldwide, special toxicities are recognized including prolonged hypogammaglobulinemia (PH). Few details are known about duration and severity of rituximab induced low serum immunoglobulin status, especially in pediatric population treated with this drug for PTLPD after HSCT.This retrospective study was conducted in one pediatric HSCT center (Robert Debré Hospital, Paris, France). Between December 2008 and June 2011, among 138 transplanted children, 39 children received rituximab (Mabthera®, Roche, Paris, France) for either EBV reactivation or B-cell PTLD. 28 children were then followed for immune status more than 12 months after rituximab based therapy. Median age at HSCT was 7 years (range 1 to 18). There were 17 males and 11 females. Indications for HSCT were hematological malignancy (n=17), sickle cell disease (n=3), B thalassemia (n=1), severe aplastic anemia (n=4) and inherited bone marrow failure (n=3). 23 children underwent 1 HSCT, 4 children received 2 while 1 received 3 HSCT. They received HSCT from either MSD (n=10), 9 to 10/10 HLA MUD (n=16), haploidentical familial donor (n=1) or unrelated 5/6 cord blood (n=1). PH was defined as serum Ig G level < 3.3g/l (1 year old), < 5g/l (1 to 5 year old), < 5.5 g/l (5 to 15 years old) and <6.5 g/l after 15 years old or serum IgM level < 0.5g/l, more than 12 months after HSCT. IV-Ig replacement therapy was used when serum immunoglobulin level was under the median range for age.Median duration of hypogammaglobulinemia was 13 months (range 0 to 38): 60% (17/28) of patients had PH that last in 3 cases more than 24 months (24, 27 and 38 months) after HSCT. 6 children still have a low immunoglobulin status. Among patients with PH, 6 underwent HSCT from 9/10 MUD, 1 from haploidentical family donor and 1 from unrelated cord blood. 3 patients received 2 HSCT and 1 patient 3 HSCT. 10 children with PH had abnormal levels of immunoglobulin before rituximab therapy. Regarding B-cell reconstitution, normalization of B cell count for age was obtained in every child, by a median time of 7.5 months (range 5 to 19) after HSCT in PH cases and 7 months (range 4 to 9) in non PH cases. Among patients with PH, 5 presented repeated lower or upper respiratory track infections (vs. 1 without PH). 2 had pancytopenia secondary to a Parvovirus B 19 infection, 1 presented an extensive VZV infection 17 month after HSCT, and 1 had Pneumocystis jirovecii pneumonia 9 month after HSCT. 1 child presented invasive aspergillosis 8 months after HSCT and 2 months after IV-Ig supplementation interruption.Although rituximab induces almost complete depletion of normal B lymphocytes in peripheral blood for an average of 6–9 months, prolonged serum immunoglobulin suppression is unusual. However, PH has been reported in patients treated with rituximab in association with chemotherapy, or after either autologous or allogeneic HSCT. In some of these studies, phenotypical analysis of B cell recovery help to find abnormalities in naïve B cell differentiation into memory B cells and plasma cells. It may help to explain that normalization of B-cell count is not systematically followed by a rapid increase in immunoglobulin. It seems that during the period of post HSCT immune reconstitution, rituximab could affect, in association with other immunosuppressive factors, not only B cell quantity but also B cell quality. PH may be then a frequent event as we report in our study. Consequences of PH on infection severity and frequency are controversial. Infectious complications secondary to rituximab-associated PH are rare but some are reported as a fatal invasive aspergillosis, frequent pulmonary infections, parvovirus or VZV infections. We also report such severe infections in our patients with PH. Thus, every EBV-PTLD patient treated with rituximab must be monitored closely with regard to serum Ig levels and probably have to receive IV-Ig supplementation, even if its interest remains unproved. B-cell sub-population monitoring may probably help to understand mechanism of PH and identify high-risk patients. Disclosures:No relevant conflicts of interest to declare.

Adult Patients with Hemophagocytic Lymphohistiocytosis (HLH) in Germany: A Retrospective Multicenter Case Study of 29 Patients

AbstractAbstract 3271 Introduction:Hemophagocytic Lymphohistiocytosis (HLH) is a rare and potentially life-threatening disorder in adults. Patients (pts) with fever of unknown origin, lymphadenopathy, deteriorating performance status, encephalopathy and elevated systemic inflammatory parameters frequently show no clear cut diagnostic criteria specific for HLH. Among adults, late onset of inherited HLH is possible, but acquired HLH triggered by infection, neoplasia or autoimmune disease is more frequent than in pediatrics. Due to the variability of clinical presentation, large-scale unreporting can be assumed. We therefore commenced a German registry for HLH pts, aiming to sensitize clinicians and to establish an HLH network as starting point for research and clinical guideline activities. Methods:We performed a retrospective multicenter case study in Germany. Diagnostic criteria were derived from the pediatric revised HLH-04 protocol. Treatment and outcome parameters were evaluated. Results:In 29 adult pts registered so far, median age was 46 years (18–81), 13 pts were female. HLH in 10 pts (34%) was malignancy-associated (aggressive B-cell lymphoma, n=6; B-CLL n=1; AML n=2; melanoma n=1), infection-associated (IHLH) in 13 pts (45%), 2 pts (7%) had connective tissue disease (CTD), in 1 pt HLH developed after therapeutic plasma exchange for TTP and in 3 pts (10%) the cause could not be determined. The main pathogen in IHLH pts was EBV (9/29, 31%) followed by other viruses (CMV n=1; HSV n=1; VZV n=1; Parvovirus B19 n=1). Of the 13 IHLH pts, 4 pts showed an EBV-reactivation post-allogeneic stem cell transplantation (SCT) for acute leukemia (n=3) and severe aplastic anemia (n=1) and 1 pt a systemic VZV infection after autologous peripheral blood SCT for aggressive B-cell NHL. 2 pts showed an EBV- and a lymphoma-triggered HLH respectively during late pregnancy. In 2 pts with EBV-associated HLH sequencing revealed mutations in XIAP and XLP-1 genes. The median time from onset of symptoms to diagnosis was 14 days (range 1–60 days). Most pts received polyvalent immunoglobulin (20/29, 69%) and HLH-04 based therapy, including steroids (14/29, 48%), cyclosporine (6/29, 21%), and etoposide (10/29, 34%). 5 pts (17%) received rituximab and 3 pts (10%) R-CHOP for lymphoma. 2 pts each (7%) received alemtuzumab and cyclophosphamide. One patient underwent allogeneic SCT but died. With a median follow up of 413 days (range, 54–1204 days), 12 pts (41.3%) survived. 2 pts were lost to follow-up. Median survival after diagnosis was 151 days (range, 14–1204+ days). Conclusions:Adult pts with HLH in our cohort showed a heterogeneous clinical syndrome most frequently triggered by EBV. HLH represents a final common pathway of an aberrantly activated immune system responding to a variety of triggers. If not recognized and treated early, HLH in adults has a very poor prognosis. Clinical registries need to include adult HLH pts in order to facilitate basic and clinical research to improve outcome of this frequently fatal immune response syndrome.Table 1Clinical and laboratory findings in adult HLH pts (n = 29)N(%)MedianRangeClinical findingsFever29 (100.0)––Splenomegaly23/27 (85.2)––Neurological abnormalities4 (15.3)––Laboratory findingsHemoglobin (g/dL; RR 12.0–15.7)29 (100)7.86.6–13.8Platelet count (/nl; RR 150–350)29 (100)329–245Neutrophil count (/nl; RR 1.8–7.7)29 (100)1.30.08–11.6Triglycerides (mmol/L; RR 0.83–2.3)25 (86.2)5.312.24–14.4Fibrinogen (g/L; RR 1.8–3.5)29 (100)1.60.13–3.8Ferritin (μg/L; RR 13–150)29 (100)45,553479–523,673Ferritin >10,000 μg/L23 (79.3)––sIL2-R (U/ml; RR <1000)20 (68.9)6,7911,194–24,032Low NK-cell count (FACS)11/18 (61%)––Morphologically proven hemophagocytosis19 (65.5)––RR, reference range; sIL2-R, soluble interleukin-2 receptor. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

COMMENTARY

COMMENTARY

Human Anti-Varicella-Zoster Virus (VZV) Recombinant Monoclonal Antibody Produced after Zostavax Immunization Recognizes the gH/gL Complex and Neutralizes VZV Infection

Varicella-zoster virus (VZV) is a ubiquitous, highly cell-associated, and exclusively human neurotropic alphaherpesvirus. VZV infection is initiated by membrane fusion, an event dependent in part on VZV glycoproteins gH and gL. Consistent with its location on the virus envelope, the gH/gL complex is a target of neutralizing antibodies produced after virus infection. One week after immunizing a 59-year-old VZV-seropositive man with Zostavax, we sorted his circulating blood plasma blasts and amplified expressed immunoglobulin variable domain sequences by single-cell PCR. Sequence analysis identified two plasma blast clones, one of which was used to construct a recombinant monoclonal antibody (rec-RC IgG). The rec-RC IgG colocalized with VZV gE on the membranes of VZV-infected cells and neutralized VZV infection in tissue culture. Mass spectrometric analysis of proteins immunoprecipitated by rec-RC IgG identified both VZV gH and gL. Transfection experiments showed that rec-RC IgG recognized a VZV gH/gL protein complex but not individual gH or gL proteins. Overall, our recombinant monoclonal anti-VZV antibody effectively neutralizes VZV and recognizes a conformational epitope within the VZV gH/L protein complex. An unlimited supply of this antibody provides the opportunity to analyze membrane fusion events that follow virus attachment and to identify multiple epitopes on VZV-specific proteins.

Acquired deficiency of coagulation factor XIII—possible evidence for a new link between coagulation and infection from a case

Dear Editor,We would like to present a case with a parainfectious dropof factor XIII (FXIII) pointing to a so far not described linkbetween coagulation and infection. A 3-year-old girl wasadmitted to our clinic due to acute joint swelling and hae-matomas during an uncomplicated infection of the upperrespiratory tract lasting since 3 weeks. The child has so farhad a normal history with respect to alimentation and de-velopment and has had the usual vaccinations. On admis-sion, the patient had haematomas at her right cheek, rightelbow andabove both heels anda mild hepatosplenomegaly.By clinical evaluation and laboratory results (Table 1), aninfection of unknown origin was suspected, presenting witha mild acute phase reaction (elevated CRP, immunoglobu-lins and fibrinogen) and signs of a mild coagulation activa-tion (indicated by slightly elevated D-dimers). A screeningshowed that the patient has already had HHV-6, HSV 1,VZV and RSV infection (IgG positive, IgM negative) andprotection from vaccination against mumps, measles andrubella. An acute infection by CMV, adenovirus, RSV, my-coplasma, cocksackie, ECHO, enterovirus, Parvo B19,EBV, Yersinia, Borrelia, Salmonella and Streptococcus pyo-genes was excluded. FXIII was very low and was the onlysignificant deviation in the coagulation system (Table 1).After a few days, the patient’s condition improved undersymptomatic therapy and could be discharged. Two weekslater, a control check-up in the outpatients’ clinic showed agood clinical recovery and normalisation of the laboratoryresults. The usual causes for FXIII deficiencies like geneticdefects[1],inhibitors, andunderlying diseases like Henoch–Schoenlein purpura, Crohn’s disease or ulcerative colitis[2-4] can be excluded by the clinical and laboratory find-ings. No interventions had taken place, which would poten-tially explain the transient deficiency. Also, it seems lessprobable that a cut coagulation activation lead to consump-tion of FXIII, as D-dimers were only slightly elevated and

Vaccination in solid-organ transplantation candidates: time for a benefit/risk assessment.

Vaccination in solid-organ transplantation candidates: time for a benefit/risk assessment.

Seroprevalence of Varicella Zoster Virus Infection among Primary school Children In Northern Nigeria

Background: Varicella Zoster infection occurs exclusively in man and commonly affects children with predilection for school age children. Following infection the course of the disease in immunocompetent children is often mild and self limited but in older age groups infection may be severe with poor outcomes ranging from complicating bacterial morbidities to death. In Kaduna State, Northern Nigeria the seroprevalence ofVaricella - zoster virus (VZV) infection is unknown The current cross - sectional study was conducted to assess the seroprevalence of VZV infection among nursery and primary school pupils in Kaduna State. Mathods: Nursery and primary school pupils between the ages 4 and 15 years were randomly selected from the three geopolitical zones of Kaduna State . Demographic data on each subject&nbsp; were obtained by administration of a questionnaire and blood samples were collected for serum analysis of Varicella - zoster virus immunoglobulin G (IgG) using the Automat ion ELISA IgG Kit manufactured by Automation INC. USA. Data obtained were summarized using percentages and frequency tables. Results were analyzed using Epi- info version 3.0. Results: Three hundred and fifty three pupils were recruited for the study. The overall prevalence rate for VZV infection was 66.3% with the value in males being 68.5% and in females 63.9%. The prevalence of VZV infection increased with age in both sexes. A high percentage of children (60%) were seropositive at 4-6 years. Conclusion: The study shows a 66.3% prevalence of VZV infection among children in Kaduna State. Thi s high pr eva l enc e r a t e necessitates further studies to establish the burden of the problem in Nigeria and rationalize the institution of preventive measures against the infection, particularly immunocompromi sed young children.

Retrospective Analysis of Infections In An Unselected Cohort of Patients with Multiple Myeloma Treated with Lenalidomide Combinations Therapies

Abstract 5135IMiDs exert various effects on the immune system altering cytokine production, regulating T cell costimulation and enhancing NK cell cytotoxicity. Particularly, lenalidomide is 50000 times more potent than thalidomide in inhibiting tumour necrosis factor a (TNFa), playing an important role in immune response against bacterial and virus infection. Moreover, lenalidomide causes myelosuppression, mainly neutropenia, more often than thalidomide.We assessed the incidence, type and major factors affecting infections in 127 patients with multiple myeloma (MM) receiving lenalidomide-based regimens. Median age was 67 years (range 40–88) and 53.5% were older than 65 years. Fifty-four patients (42.5%) had newly diagnosed MM whereas the remaining 73 (57.5%) had relapsed/refractory disease. Among this group, 67% of patients had previous received more than one line of therapy and 26% had undergone APSCT. ISS stage 2–3 and renal failure were recognized in 54.5% and 8.5% of patients, respectively, and 15% of them had a ECOG PS >= 2. Eighty patients (63%) received lenalidomide plus dexamethasone and 47 (37%) lenalidomide combined with steroids and chemotherapy. Median courses of lenalidomide administered was 6 (range 1–28). Nearly all patients (95%) received trimethoprim-sulfamethoxazole (TMP-SMZ) as prophylaxis for infections and granulocyte-colony stimulating factor was used according to guidelines.Twenty-six patients (20.5%) developed infections resulting of grade 1–2 in 8 patients (6%) and 3–5 in the remaining 18 (14%). There were two deaths (1.5%) due to infections. Type of infection were: pneumonia in 15 (58%), upper respiratory tract infections in 3 (11.5%), FUO in 3 (11.5%), septic shock by gram-negative microorganisms in 2 (8%), cholecystitis in 2 (8%) and VZV infection in one (4%). Risk of grade 3–5 infection was 16% at 12 months; 62.5%, 69% and 94% of infections occurred at 3, 4 and 6 months, respectively. Univariate analysis showed that ISS 2–3 (16.5% vs 7%; p=0.017), creatinine > 2 mg/dl (23.5 vs 10.5%; p=0.087) and WBC < 4000/mcl (33% vs 12.7%: p=0.087) prior therapy were associated with development of severe infection, whereas all other variables as sex, age, PS, disease status, monoclonal component level, thrombocytopenia prior therapy, renal failure, type of therapy and number of prior lines of therapy did not affect infections. Multivariate analysis selected only ISS as factor affecting severe infection development. Particularly, the risk of grade 3–5 infections at 6 months was 18% in patients with ISS 2–3 if compared with 6% in those with ISS 1 (p=0.034). As regard outcome parameters, there was a trend for a longer PFS in patients without infection (median PFS = 8 vs 16 months in patients without infections; p=0.064); however, OS of patients developing infection was significantly shorter compared to that of patients who did not develop infections (median OS=26 vs 33 months; p=0.001). Multivariate analysis showed that infections significantly affected OS (HR=3.2; 95%CI=1.5–6.7; p=0.002) adjusted for age, PS, ISS, renal function and therapy with 2 or 3 drugs.In clinical practice, infections represent a frequent complication in patients with MM receiving lenalidomide-based regimens, as reported in phase II-III trials. Respiratory infections accounted for a large majority (nearly 70%), although severe gram-negative septic shock should be taken into account in neutropenic patients. Nearly all patients who developed infections during the first 6 months of therapy and those with higher tumour burden have been found to be at higher risk.In conclusion, a broader antibiotic prophylaxis (ie TMP-SFZ plus fluoroquinolones) should be taken into consideration to prevent severe infections and ameliorate final outcome of patients treated with lenalidomide. Disclosures:Caravita:Celgene: Honoraria, Research Funding; Janssen: Honoraria. Offidani:Celgene: Honoraria. Gentili:celgene: Honoraria.

Rothmund-Thomson syndrome: Immuno-osseous challenges

Results All patients had characteristic poikiloderma as well as thumb anomalies. They were born dysmaturely and presented with failure to thrive. Age at genetic diagnosis was 5y, 4y and 3y for P1, P2, P3. Osteopenia and abnormal metaphyseal trabeculation of bones were striking on the initial skeletal survey in all patients. Z-scores on DXA scan were -0.1, -1.1 and -1.2 for P1, P2, P3 respectively at presentation. The presentation in P2 was dramatic with 6 fractures in upper and lower extremities and subluxation of both radii. All patients were suffering from recurrent chest infections. P1 had granulomatous skin inflammation following primo VZV infection. All patients have low switched memory B cells for age, P1 has IgG2 deficiency. P1 and P3 have IgM deficiency. P1 and P3 have specific polysaccharide antibody deficiency. Results are pending for P2. All receive prophylactic antibiotics. P1 is treated with subcutaneous immunoglobulin substitution.

Reporter cell line-based screening for anti-varicella-zoster virus compounds

Objective To establish a novel method to screen for anti-varicella-zoster virus (VZV) compounds with our previously generated reporter cell line for VZV, MV9G. Methods MV9G cells were directly infected with cell-free virus of Oka vaccine strain (vOka) for 2 hours( CFV direct-infection) or cocultured with vOka-infected MeWo cells containing cell-associated virus for 48 hours (CAV co-culture) to promote expression of the reporter gene firefly luciferase. Antiviral compounds including heparin, mannose-6-phosphate( M-6-P), acyclovir( ACV ), resveratrol and roscovitine were added in the medium before or after the virus infection. Inhibitory effects( IC50 ) of the antiviral compounds were analyzed by comparing firefly luciferase activities of MV9G cells in the presence of antiviral compounds with those in the absence. Results Antiviral compounds inhibited luciferase activities of MV9G cells activated by CFV direct-infection and/or CAV co-culture in different levels. The reductions of luciferase activities statistically correlated with those of viral foci shown by immunostaining with a monoclonal antibody against VZV immediate early 62 antigens (IE62) in controls. Among these compounds, heparin, M-6-P, and 2.5 μmol/L of roscovitine inhibited CFV-activated more strongly than CAV-activated luciferase activities, whereas ACV and resveratrol inhibited CAV-activated more strongly than CFV-activated luciferase activities. Cell-associated ACV-resistant strains,Kanno and rOka YSR, activated luciferase activities of MV9G cells, too. However, the inhibitory concentrations (IC50) of ACV to the ACV-resistant strains were much higher than those to the ACV-sensitive strains,pOka and CaGu. Conclusion The CFV direct-infection and CAV co-culture assays were useful to screen for antiviral compounds targeting the early and late phases of VZV infection, respectively. The VZV reporter cell-based assays may provide a simple, rapid, sensitive, and high-throughput method to screen for anti-VZV compounds. Key words: Varicella-zoster virus; Reporter cell line; Mannose-6-phosphate; Resveratrol; Roscovitine