Biomarkers and sustainable innovation in cardiovascular drug development: lessons from near and far afield.

Future innovative therapies targeting cardiovascular disease (CVD) have the potential to improve health outcomes and to contain rising healthcare costs. Unsustainable increases in the size, cost and duration of clinical trial programs necessary for regulatory approval, however, threaten the entire innovation enterprise. Rising costs for clinical trials are due in large part to increasing demands for hard cardiovascular clinical endpoints as measures of therapeutic efficacy.

Kaposi's sarcoma associated herpesvirus G-protein coupled receptor activation of cyclooxygenase-2 in vascular endothelial cells.

Background: Kaposi's sarcoma associated herpesvirus (KSHV) is the etiologic agent of Kaposi's sarcoma (KS), a highly vascularized neoplasm characterized by endothelial-derived spindle-shaped tumor cells. KSHV-infected microvascular endothelial cells demonstrate increased cyclooxygenase-2 (COX-2) expression and KS lesions have high levels of prostaglandin E2 (PGE2), a short-lived eicosanoid dependent on cyclooxygenase activity that has been linked to pathogenesis of other neoplasias. To determine whether increased COX-2 expression and PGE2 production is mediated by the angiogenic and tumorigenic KSHV-encoded G-protein coupled receptor (vGPCR), we developed a recombinant retrovirus to express vGPCR in Human Umbilical Vascular Endothelial Cells (HUVEC).

Human herpesvirus 8 presence and viral load are associated with the progression of AIDS-associated Kaposi's sarcoma.

Objective: We present the largest longitudinal study to date that examines the association between Kaposi's Sarcoma (KS) disease progression and the presence and viral load of human herpesvirus 8 (HHV-8).

Methods: Ninety-six men were enrolled at HIV clinics in Atlanta, Georgia, who had KS (n = 47) or were without KS but seropositive for HHV-8. Visits occurred at 6-month intervals for 2 years at which the patient's KS status was evaluated and oral fluid and blood were collected for quantification of HHV-8 DNA and antibodies.

Targeting human herpesvirus-8 for treatment of Kaposi's sarcoma and primary effusion lymphoma.

Purpose Of Review: Human herpesvirus-8, also called the Kaposi's sarcoma herpesvirus, is present in all cases of Kaposi's sarcoma and primary effusion lymphoma and in some cases of multicentric Castleman's disease. This review discusses mechanisms by which human herpesvirus-8 contributes to tumorigenesis and how this knowledge can be used to target the virus for the treatment of these tumors.

Recent Findings: Most primary effusion lymphomas and Kaposi's sarcoma tumor cells are latently infected with human herpesvirus-8 and hence resistant to antiherpesvirus drugs that are dependent on lytic replication.

Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus.

As sentinels of host defense, intestinal epithelial cells respond to the viral pathogen rotavirus by activating a gene expression that promotes immune cell recruitment and activation. We hypothesized that epithelial sensing of rotavirus might target dsRNA, which can be detected by TLR3 or protein kinase R (PKR). Accordingly, we observed that synthetic dsRNA, polyinosinic acid:cytidylic acid (poly(I:C)), potently induced gene remodeling in model intestinal epithelia with the specific pattern of expressed genes, including both classic proinflammatory genes (e.

Apoptosis induced by the toll-like receptor adaptor TRIF is dependent on its receptor interacting protein homotypic interaction motif.

TLRs detect specific molecular features of microorganisms and subsequently engage distinct signaling networks through the differential use of Toll/IL-1R (TIR)-domain-containing adapter proteins. In this study, we investigated the control of apoptosis by the TIR domain-containing adapter proteins MyD88, TIR-domain containing adapter protein (TIRAP), TIR-domain-containing adapter-inducing IFN-beta (TRIF), TRIF-related adapter molecule (TRAM), and sterile alpha motifs and beta-catenin/armadillo repeats (SARM). Upon overexpression, TRIF was the sole TIR-adapter to potently engage mammalian cell death signaling pathways.

The targeting of primary effusion lymphoma cells for apoptosis by inducing lytic replication of human herpesvirus 8 while blocking virus production.

Primary effusion lymphoma (PEL) is a B-cell lymphoma in which human herpesvirus-8 (HHV-8) is found within all tumor cells and represents a target for selectively destroying tumor cells. HHV-8 is latent in most PEL cells and, hence, resistant to antiviral agents that inhibit lytic replication. We demonstrate that PEL cell lines containing HHV-8 without and with coinfection with Epstein-Barr virus responded to the antiseizure medication valproate with entry into the lytic cascade and production of infectious virus.

Inhibition of infectious human herpesvirus 8 production by gamma interferon and alpha interferon in BCBL-1 cells.

Human herpesvirus-8 (HHV-8) is aetiologically linked to Kaposi's sarcoma and primary effusion lymphoma. Although interferon-alpha (IFN-alpha) and interferon-gamma (IFN-gamma) are both antiviral cytokines, IFN-alpha blocks entry of HHV-8 into the lytic phase, whereas IFN-gamma induces an increase in the percentage of cells undergoing lytic replication. Multiple events in the lytic cascade must be completed to produce infectious virus.

Repeated measures study of human herpesvirus 8 (HHV-8) DNA and antibodies in men seropositive for both HHV-8 and HIV.

Objective: To study the natural history and pathogenesis of human herpesvirus 8 (HHV-8) infection in HHV-8-seropositive, immunosuppressed men.

Design: Longitudinal study of 87 HHV-8- and HIV-seropositive men [42 with Kaposi's sarcoma (KS)] during four visits over a 2 month period.

Methods: : Patients provided oral fluid and blood.

Inhibition of infection and replication of human herpesvirus 8 in microvascular endothelial cells by alpha interferon and phosphonoformic acid.

Infection of endothelial cells with human herpesvirus 8 (HHV-8) is an essential event in the development of Kaposi's sarcoma. When primary microvascular endothelial cells (MECs) were infected with HHV-8 at a low multiplicity of infection, considerable latent replication of HHV-8 occurred, leading to a time-dependent increase in the percentage of virus-infected cells that was accompanied by cellular spindling and growth to a high density with loss of contact inhibition. Only a low percentage of MECs supported lytic replication of HHV-8 and produced infectious virus.

Short duration of elevated vIRF-1 expression during lytic replication of human herpesvirus 8 limits its ability to block antiviral responses induced by alpha interferon in BCBL-1 cells.

Human herpesvirus 8 (HHV-8) encodes multiple proteins that disrupt the host antiviral response, including viral interferon (IFN) regulatory factor 1 (vIRF-1). The product of the vIRF-1 gene blocks responses to IFN when overexpressed by transfection, but the functional consequence of vIRF-1 that is expressed during infection with HHV-8 is not known. These studies demonstrate that BCBL-1 cells that were latently infected with HHV-8 expressed low levels of vIRF-1 that were associated with PML bodies, whereas much higher levels of vIRF-1 were transiently expressed during the lytic phase of HHV-8 replication.

IFN-alpha sensitizes human umbilical vein endothelial cells to apoptosis induced by double-stranded RNA.

The ability of endothelial cells to mount an efficient antiviral response is important in restricting viral dissemination and eliminating viral infection from the endothelium and surrounding tissues. We demonstrate that dsRNA, a molecular signature of viral infection, induced apoptosis in HUVEC, and priming with IFN-alpha shortened the time between when dsRNA was encountered and when apoptosis was initiated. IFN-alpha priming induced higher levels of mRNA for dsRNA-activated protein kinase, 2'5'-oligoadenylate synthetase, and Toll-like receptor 3, transcripts that encode dsRNA-responsive proteins.

Characterization of entry mechanisms of human herpesvirus 8 by using an Rta-dependent reporter cell line.

To analyze the mechanisms of entry of human herpesvirus 8 (HHV-8), we established a reporter cell line T1H6 that contains the lacZ gene under the control of the polyadenylated nuclear RNA promoter, known to be strongly activated by a viral transactivator, Rta. We found that infection with cell-free virus, as well as cocultivation with HHV-8-positive primary effusion lymphoma cell lines, activated the lacZ gene of T1H6 in a sensitive and dose-dependent manner. Addition of Polybrene and centrifugation enhanced, but polysulfonate compounds inhibited, the HHV-8 infectivity.

Risk factors for Kaposi's sarcoma in men seropositive for both human herpesvirus 8 and human immunodeficiency virus.

Objective: To identify risk factors for Kaposi's sarcoma (KS) among men seropositive for both human herpesvirus 8 (HHV-8) and HIV.

Design: Cross-sectional study of 91 HHV-8 seropositive, HIV seropositive men who have sex with men (57 with KS), and 70 controls at lower risk for KS.

Methods: Patients received clinical evaluations.

Activation of cellular and heterologous promoters by the human herpesvirus 8 replication and transcription activator.

The key regulator of the switch from latent to lytic replication of the human herpesvirus 8 (HHV-8; KSHV) is the replication and transcription activator (Rta). The ability of Rta to regulate cellular gene expression was examined by transient transfection into cells that were not infected with HHV-8. Rta induced some, but not all, NF-kappa B-responsive reporters through mechanisms that did not involve activation of classic forms of NF-kappa B.

Induction of human herpesvirus 8 gene expression in a posttransplantation primary effusion lymphoma cell line.

Human herpesvirus 8 (HHV-8 or Kaposi's sarcoma herpesvirus) is a gamma herpesvirus that is most likely the etiologic agent of both Kaposi's sarcoma and primary effusion lymphoma (PEL), a rare HIV-associated lymphoma. The role of HHV-8 in post-transplant lymphoma is less well characterized. We demonstrate that HHV-8 is constitutively present in LH5-21 cells, an atypical patient derived posttransplant PEL cell line.

Transcriptional activation by the human herpesvirus-8-encoded interferon regulatory factor.

Human herpesvirus-8 (HHV-8), a gammaherpesvirus that is thought to be the viral aetiologic agent of Kaposi's sarcoma and primary effusion lymphoma, encodes a homologue to cellular interferon regulatory factors (IRFs). The HHV-8 IRF homologue (vIRF; ORF K9) has previously been shown to inhibit gene induction by interferons and IRF-1 and to transform NIH3T3 cells or Rat-1 cells. Additionally, expression of antisense to vIRF in BCBL-1 cells results in the repression of certain HHV-8 genes, suggesting that vIRF may also positively regulate gene expression.