Anthony L. Cunningham

Genomic Structure of an Attenuated Quasi Species of HIV-1 from a Blood Transfusion Donor and Recipients

A blood donor infected with human immunodeficiency virus-type 1 (HIV-1) and a cohort of six blood or blood product recipients infected from this donor remain free of HIV-1-related disease with stable and normal CD4 lymphocyte counts 10 to 14 years after infection. HIV-1 sequences from either virus isolates or patient peripheral blood mononuclear cells had similar deletions in the nef gene and in the region of overlap of nef and the U3 region of the long terminal repeat (LTR). Full-length sequencing of one isolate genome and amplification of selected HIV-1 genome regions from other cohort members revealed no other abnormalities of obvious functional significance. These data show that survival after HIV infection can be determined by the HIV genome and support the importance of nef or the U3 region of the LTR in determining the pathogenicity of HIV-1.

Diversity of receptors binding HIV on dendritic cell subsets.

The ability of HIV-1 to use dendritic cells (DCs) for transport and to transfer virus to activated T cells in the lymph node may be crucial in early HIV-1 pathogenesis. We have characterized primary DCs for the receptors involved in viral envelope attachment and observed that C-type lectin receptor (CLR) binding was predominant in skin DCs, whereas binding to emigrating and tonsil DCs was CD4-dependent. No one CLR was solely responsible for envelope binding on all skin DC subsets. DC-SIGN (DC-specific ICAM-3–grabbing nonintegrin) was only expressed by CD14+CDlalo dermal DCs. The mannose receptor was expressed by CD1ahi and CD14+CDlalo dermal DCs, and langerin was expressed by Langerhans cells. The diversity of CLRs able to bind HIV-1 in skin DCs may reflect their ability to bind a range of microbial glycoproteins.

Famciclovir for the Treatment of Acute Herpes Zoster: Effects on Acute Disease and Postherpetic Neuralgia: A Randomized, Double-Blind, Placebo-Controlled Trial

Herpes zoster (shingles) develops in as many as 20% of all persons [1]. The characteristic zoster rash is often accompanied by substantial pain, dysesthesias, and skin hypersensitivity. The unmet challenge in the management of patients with acute zoster is the alleviation of chronic pain. In many patients, pain resolves once the affected area of skin returns to normal. However, some patients continue to experience pain long after the lesions have healed; this pain is commonly called postherpetic neuralgia. Postherpetic neuralgia is by far the most common complication of herpes zoster and is one of the most intractable pain disorders [2, 3]. The incidence of postherpetic neuralgia increases sharply with increasing age [4, 5]; nearly half of patients older than 60 years have this complication [2, 5]. Postherpetic neuralgia is also more severe and persists longer in older than in younger patients [3]. The disorder is clearly the most distressing component of the disease process for both the patient and the physician. Although for many years acyclovir has been the only oral antiviral agent approved for the treatment of patients with acute herpes zoster, its effect on postherpetic neuralgia remains controversial [6-10]. Famciclovir, a new antiviral agent, was approved for marketing by the Food and Drug Administration in June 1994 for the management of acute herpes zoster. Famciclovir is the well-absorbed (77% bioavailable) [11] oral form of penciclovir, with activity against varicella-zoster virus, herpes simplex virus types 1 and 2, and Epstein-Barr virus [12-15]. Penciclovir is selectively activated in virus-infected cells through phosphorylation to the antiviral compound penciclovir triphosphate. Viral thymidine kinase converts penciclovir to penciclovir monophosphate, which is converted by cellular enzymes to penciclovir triphosphate [12, 16, 17]. Penciclovir triphosphate inhibits viral DNA polymerase, thereby halting DNA synthesis and viral replication [16-19]. The in vitro potencies of penciclovir and acyclovir depend on the tissue culture cell line and assay method used but are generally similar [14, 15, 20].The 50% plaque inhibitory concentrations in human lung fibroblasts infected with varicella-zoster virus were 4.0 1.5 g/mL for penciclovir and 4.0 1.1 g/mL for acyclovir [14]. A potentially clinically important characteristic of penciclovir triphosphate is that it persists in virus-infected host cells longer than acyclovir triphosphate [19]. For cells infected with varicella-zoster virus, the intracellular half-life of penciclovir triphosphate is 9.1 hours; in contrast, the half-life for acyclovir triphosphate is 0.8 hours [19, 21]. Thus, penciclovir triphosphate has the potential to continue to inhibit viral replication even if serum concentrations are below the inhibitory level. Consistent oral bioavailability linked to a favorable intracellular half-life of penciclovir triphosphate in cells infected with varicella-zoster virus suggested that famciclovir could offer clinically important advantages in the management of herpes zoster over currently available therapy. These advantages include a reduced total daily dose and a reduced dosing frequency. Because persistent antiviral activity throughout each day of dosing was expected to lead to improved clinical efficacy, especially with regard to postherpetic neuralgia, we examined the effects of famciclovir on acute herpes zoster and postherpetic neuralgia. Methods Study Design In our randomized, double-blind, placebo-controlled, multicenter trial, we compared the efficacy and safety of famciclovir, 500 mg or 750 mg, given three times daily for 7 days, with that of placebo in the treatment of patients with uncomplicated acute herpes zoster. Eligible participants were immunocompetent patients 18 years or older with clinically diagnosed uncomplicated herpes zoster who gave written informed consent. Exclusion criteria included zoster rash that had been present for more than 72 hours; complications of herpes zoster (for example, ocular or visceral involvement, disseminated herpes zoster); presence of crusts at enrollment; other serious underlying disease (such as immune disorders or human immunodeficiency virus [HIV] infection); or pregnancy or lactation. Patients were prohibited from receiving any concomitant antiviral or immunomodifying therapy and any topical medication that would be applied to zoster lesions during the study. Patient Assessments Patients were instructed to return to the clinic for evaluation of lesions and pain on each of the 7 therapy days and every day for the 7 days after therapy. Patients with lesions that were fully crusted by day 7 were examined every other day during the week after therapy. After day 14, weekly visits were required of all patients until all lesions had lost their crusts. After the lesions had healed, patients were assessed for the presence of postherpetic neuralgia (defined as pain after healing) at monthly intervals for an additional 5 months. These monthly pain assessments continued for all patients, even if postherpetic neuralgia had resolved before completion of the study period. Patients attending the clinic for fewer than 80% of required visits were deemed noncompliant. The number of papules, vesicles, ulcers, and crusts within the primary dermatome was classified as none, mild (<25 lesions), moderate (25 to 50 lesions), or severe (>50 lesions). A specimen for viral culture was obtained at baseline and daily thereafter while vesicles were present. Patients were asked to rate the intensity of their pain as none, mild, moderate, or severe. Definitions for mild, moderate, and severe pain were not prospectively provided. Adverse events were assessed at each visit according to the time of onset, duration, severity, and investigator-defined relation to the study medication. Blood samples were obtained for chemical and hematologic assessment, and urine samples were obtained for dipstick analysis before initiation of therapy and at the last therapy visit. Statistical Analysis Efficacy end points were analyzed by standard survival methods. We used the Cox proportional-hazards regression model in our analyses [22]. Time-dependent covariates were modeled to evaluate the proportional hazards assumption, and a model that included the main effects of treatment was used to examine efficacy. Statistical conclusions were based on the significance of estimated hazard ratios. Hazard ratios greater than 1 indicated a faster rate of event occurrence in patients receiving famciclovir than in those receiving placebo. An estimated hazard ratio of 2 indicated that the event of interest occurred twice as rapidly in patients receiving famciclovir as it did in placebo recipients. Two comparisons were made for each end point: 500-mg famciclovir compared with placebo and 750-mg famciclovir compared with placebo. We also used Kaplan-Meier estimates of the cumulative proportion of patients achieving an event to graphically illustrate the trial results. We used the Fisher exact test (two-tailed) to analyze proportion data. The primary efficacy variable was the time to full crusting of the lesions. Secondary variables included duration of viral shedding; time to resolution of vesicles, ulcers, crusts, and acute pain; and duration of postherpetic neuralgia (that is, the time to resolution of pain after the lesions had healed). Healing was defined as the first time in which a patient had no papules, vesicles, ulcers, or crusts and after which did not develop them at any later visit. Similarly, the time to the resolution of a clinical variable was the time to the complete cessation of that variable with no further occurrence at any later date. Duration of viral shedding was measured as the number of days from the first dose of the study medication to the last positive culture. The enrollment objective before the study was 400 patients; this number was selected to ensure a target of 300 evaluable patients (100 per group). The sample provided an 80% power to detect a significant difference from placebo, assuming a true hazard ratio of 1.5 and exponential time to full crusting (or time to event). We analyzed data from both the intention-to-treat group (patients receiving at least one dose of study medication) and the efficacy-evaluable group (patients complying with the protocol). We prospectively defined the efficacy-evaluable group before unblinding the randomization code. Because the results of both analyses were generally similar, we present data for the intention-to-treat group, unless otherwise specified. We also examined prospectively defined subgroups with respect to age, duration of rash at enrollment, and severity of rash at enrollment. The safety analysis included all patients who had received at least one dose of the study medication. Each analysis included all patients who provided information for the respective end point. For example, the analysis for the time to resolution of crusts included all patients who presented with crusts during the study. Therefore, the number of patients in each analysis of the time to resolution of a condition may vary because only patients experiencing the specific condition were included in the analysis. Results Demographic Characteristics Characteristics of patients in the intention-to-treat group (n = 419), including sex, age, duration of rash, location of rash, severity of rash, and severity of pain, are shown in Table 1. Of the 419 patients enrolled, 138 received 500 mg of famciclovir, 135 received 750 mg of famciclovir, and 146 received placebo. Approximately half of the patients were female, and the mean age was 50 years. More than half of the patients had severe rash (>50 lesions) at enrollment, and more than 60% had moderate or severe zoster pain at enrollment. Table 1. Patient Characteristics at Study Enrollment The intention-to-treat group and the efficacy-evaluable group (n = 323) had similar ba

Efficacy of an Adjuvanted Herpes Zoster Subunit Vaccine in Older Adults

BACKGROUND In previous phase 1-2 clinical trials involving older adults, a subunit vaccine containing varicella-zoster virus glycoprotein E and the AS01B adjuvant system (called HZ/su) had a clinically acceptable safety profile and elicited a robust immune response. METHODS We conducted a randomized, placebo-controlled, phase 3 study in 18 countries to evaluate the efficacy and safety of HZ/su in older adults (≥50 years of age), stratified according to age group (50 to 59, 60 to 69, and ≥70 years). Participants received two intramuscular doses of the vaccine or placebo 2 months apart. The primary objective was to assess the efficacy of the vaccine, as compared with placebo, in reducing the risk of herpes zoster in older adults. RESULTS A total of 15,411 participants who could be evaluated received either the vaccine (7698 participants) or placebo (7713 participants). During a mean follow-up of 3.2 years, herpes zoster was confirmed in 6 participants in the vaccine group and in 210 participants in the placebo group (incidence rate, 0.3 vs. 9.1 per 1000 person-years) in the modified vaccinated cohort. Overall vaccine efficacy against herpes zoster was 97.2% (95% confidence interval [CI], 93.7 to 99.0; P<0.001). Vaccine efficacy was between 96.6% and 97.9% for all age groups. Solicited reports of injection-site and systemic reactions within 7 days after vaccination were more frequent in the vaccine group. There were solicited or unsolicited reports of grade 3 symptoms in 17.0% of vaccine recipients and 3.2% of placebo recipients. The proportions of participants who had serious adverse events or potential immune-mediated diseases or who died were similar in the two groups. CONCLUSIONS The HZ/su vaccine significantly reduced the risk of herpes zoster in adults who were 50 years of age or older. Vaccine efficacy in adults who were 70 years of age or older was similar to that in the other two age groups. (Funded by GlaxoSmithKline Biologicals; ZOE-50 ClinicalTrials.gov number, NCT01165177.).

Long-term symptomless HIV-1 infection in recipients of blood products from a single donor

There have been reported cases of long-term symptomless human immunodeficiency virus type 1 (HIV-1) infection, but it is not clear whether the benign course of infection was due to host, viral, or other unknown factors. During follow-up of subjects with transfusion-acquired HIV-1 infection in New South Wales, Australia, we identified a group of 6 subjects who had been infected through a single common donor. We were therefore able to study the contributions of various factors to the course of infection. Throughout follow-up (range 6.8-10.1 years after infection), 5 of the recipients and the donor (last follow-up 10.2 years after infection of the first recipient) remained clinically free of symptoms, with normal CD4 cell counts and no p24 antigenaemia. HIV-1 was isolated from only 1 recipient; the isolate did not induce syncytia in a SUPT1 co-culture assay and had a limited in-vitro host range. 1 infected recipient (who had received extensive immunosuppressive treatment for systemic lupus erythematosus) developed Pneumocystis carinii pneumonia and died 4.3 years after infection. The frequency of progression to AIDS or a CD4 cell count below 0.50 x 10(9)/l was significantly lower among the 6 subjects with a common donor (1/6) than among 101 other HIV-infected transfusion recipients for whom data from 7 years of follow-up were available (94/101; p less than 0.0001). These findings suggest that the subjects were infected by a less virulent strain of HIV-1. The identification of this group of subjects should stimulate a search for other similar groups, which will provide important information on the immunopathogenesis of HIV-1 disease.

Prospects for Control of Herpes Simplex Virus Disease through Immunization

Herpes simplex viruses (HSVs) can cause a variety of infections, including genital herpes. Despite effective antiviral therapy, HSV infections remain a significant worldwide public health problem. Vaccines offer the best hope for controlling spread and limiting HSV disease. This article discusses the pathogenesis and immunobiology of mucocutaneous HSV infections, summarizes the spectrum of diseases caused by HSV, and provides a review of the field of HSV vaccine research. This article also discusses what might be realistically expected of a vaccine intended for control of genital herpes and explores the question of whether a vaccine that is effective in controlling genital HSV disease might also be effective in controlling nongenital HSV disease. The efficacy of such vaccines for the full spectrum of HSV disease will eventually determine the timing and targeting of immunization, ranging from selective immunization in preadolescence to universal childhood immunization as part of the routine childhood regimen.

Determination of Interactions between Tegument Proteins of Herpes Simplex Virus Type 1

ABSTRACT The aim of this study was to elucidate protein-protein interactions between tegument proteins of herpes simplex virus type 1 (HSV-1). To do so, we have cloned and expressed in the LexA yeast (Saccharomyces cerevisiae) two-hybrid system, 13 of the 21 currently known tegument proteins of HSV-1. These included the tegument proteins essential for replication in cell lines, UL17, UL36, UL37, UL48, and UL49, and the nonessential tegument proteins US11, UL11, UL14, UL16, UL21, UL41, UL46, and UL47. A total of 104 combinations were screened in the yeast two-hybrid assay, with 9 interactions identified. These included: UL11-UL16, UL36-UL37, UL36-UL48, UL46-UL48, UL47-UL48, and UL48-UL49. The remaining interactions consisted of self-associations that were observed for US11, UL37, and UL49. The interactions UL36-UL37, UL36-UL48, UL37-UL37, UL46-UL48, and UL47-UL48 have not been previously reported for HSV-1. The interaction of UL46-UL48 was verified using an in vitro pull-down assay. The interactions of UL36-UL37 and UL37-UL37 were verified with a coimmunoprecipitation assay. Knowledge of HSV-1 tegument protein-protein interactions will provide insights into the pathways of tegument assembly, and the identified interactions are potential targets for new antiviral drugs.

Efficacy of the Herpes Zoster Subunit Vaccine in Adults 70 Years of Age or Older

BACKGROUND A trial involving adults 50 years of age or older (ZOE-50) showed that the herpes zoster subunit vaccine (HZ/su) containing recombinant varicella-zoster virus glycoprotein E and the AS01B adjuvant system was associated with a risk of herpes zoster that was 97.2% lower than that associated with placebo. A second trial was performed concurrently at the same sites and examined the safety and efficacy of HZ/su in adults 70 years of age or older (ZOE-70). METHODS This randomized, placebo-controlled, phase 3 trial was conducted in 18 countries and involved adults 70 years of age or older. Participants received two doses of HZ/su or placebo (assigned in a 1:1 ratio) administered intramuscularly 2 months apart. Vaccine efficacy against herpes zoster and postherpetic neuralgia was assessed in participants from ZOE-70 and in participants pooled from ZOE-70 and ZOE-50. RESULTS In ZOE-70, 13,900 participants who could be evaluated (mean age, 75.6 years) received either HZ/su (6950 participants) or placebo (6950 participants). During a mean follow-up period of 3.7 years, herpes zoster occurred in 23 HZ/su recipients and in 223 placebo recipients (0.9 vs. 9.2 per 1000 person-years). Vaccine efficacy against herpes zoster was 89.8% (95% confidence interval [CI], 84.2 to 93.7; P<0.001) and was similar in participants 70 to 79 years of age (90.0%) and participants 80 years of age or older (89.1%). In pooled analyses of data from participants 70 years of age or older in ZOE-50 and ZOE-70 (16,596 participants), vaccine efficacy against herpes zoster was 91.3% (95% CI, 86.8 to 94.5; P<0.001), and vaccine efficacy against postherpetic neuralgia was 88.8% (95% CI, 68.7 to 97.1; P<0.001). Solicited reports of injection-site and systemic reactions within 7 days after injection were more frequent among HZ/su recipients than among placebo recipients (79.0% vs. 29.5%). Serious adverse events, potential immune-mediated diseases, and deaths occurred with similar frequencies in the two study groups. CONCLUSIONS In our trial, HZ/su was found to reduce the risks of herpes zoster and postherpetic neuralgia among adults 70 years of age or older. (Funded by GlaxoSmithKline Biologicals; ZOE-50 and ZOE-70 ClinicalTrials.gov numbers, NCT01165177 and NCT01165229 .).

Immature Monocyte-Derived Dendritic Cells Are Productively Infected with Herpes Simplex Virus Type 1

ABSTRACT Herpes simplex viruses (HSV) have developed several immunoevasive strategies. Here we demonstrate a novel mechanism by which HSV type 1 may interfere with the immune response through infection of immature dendritic cells (DC) and selective downmodulation of costimulatory molecules. In our study we show productive infection of immature monocyte-derived DC, which closely resemble sessile Langerhans cells, by sequential expression of immediate-early, early, and late viral proteins and of glycoprotein D mRNA, as well as production of infectious virus of moderate titers. Infection was cytopathic, with the progressive loss of 20 to 45% of cells from 24 to 48 h after infection, with no more than 80% of DC found to be infected. These results are in contrast to those of previous findings of nonpermissive or abortive infection of monocytes and mature monocyte-derived DC. Infection of immature DC also led to selective and asynchronous downregulation of CD1a, CD40, CD54 (ICAM-1) (12 h postinfection), CD80 (24 h postinfection), and CD86 (48 h postinfection) but not of CD11c or major histocompatibility complex class I and II molecules when compared to DC exposed to UV-inactivated virus. Thus, we propose that productive infection of epidermal Langerhans cells in vivo may lead to delayed activation of T cells, allowing more time for replication of HSV type 1 in epidermal cells.

Herpes Simplex Virus Infection of Human Dendritic Cells Induces Apoptosis and Allows Cross-Presentation via Uninfected Dendritic Cells

HSV efficiently infects dendritic cells (DCs) in their immature state and induces down-regulation of costimulatory and adhesion molecules. As in mice, HSV infection of human DCs also leads to their rapid and progressive apoptosis, and we show that both early and late viral proteins contribute to its induction. Because topical HSV infection is confined to the epidermis, Langerhans cells are expected to be the major APCs in draining lymph nodes. However, recent observations in murine models show T cell activation to be mediated by nonepidermal DC subsets, suggesting cross-presentation of viral Ag. In this study we provide an explanation for this phenomenon, demonstrating that HSV-infected apoptotic DCs are readily phagocytosed by uninfected bystander DCs, which, in turn, stimulate virus-specific CD8+ T cell clones.