Bridgette Connell

University of Twente | UT · Strategic Business Development

The HIV-1 envelope gp120, which features both the virus receptor (CD4) and coreceptor (CCR5/CXCR4) binding sites, offers multiple sites for therapeutic intervention. However, the latter becomes exposed, thus vulnerable to inhibition, only transiently when the virus has already bound cellular CD4. To pierce this defense mechanism, we engineered a se...

By targeting cells that provide protection against infection, HIV-1 causes acquired immunodeficiency syndrome. Infection starts when gp120, the viral envelope glycoprotein, binds to CD4 and to a chemokine receptor usually CCR5 or CXCR4. As many microorganisms, HIV-1 also interacts with heparan sulfate (HS), a complex group of cell surface associate...

Chemokines stimulate signals in cells by binding to G protein (heterotrimeric guanine nucleotide–binding protein)–coupled receptors. These chemoattractant cytokines alsointeract with heparan sulfate (HS), which provides positionalinformation within tissues in the form of haptotactic gradients along which cells can migrate directionally. To inves-ti...

HIV-1 cell entry is mediated by binding to the CD4-receptor and chemokine co-receptors CCR5 (R5) or CXCR4 (X4). R5-tropic viruses are predominantly detected during early infection. A switch to X4-tropism often occurs during the course of infection. X4-tropism switching is strongly associated with accelerated disease progression and jeopardizes CCR5...

On-chip sample preparation in self-contained microfluidic devices is a key element to realize simple, low-cost, yet reliable in vitro diagnostics that can be carried out at the point-of-care (POC) with minimal training requirements by unskilled users. To address this largely unmet POC medical need, we have developed an optimized polysaccharide matr...

CCR5 plays immune functions and is the coreceptor for R5 HIV-1 strains. It exists in diverse conformations and oligomerization states. We interrogated the significance of the CCR5 structural diversity on HIV-1 infection. We show that envelope glycoproteins (gp120s) from different HIV-1 strains exhibit divergent binding levels to CCR5 on cell lines...

Different HIV-1 gp120s differentially recognize antigenically distinct populations of CCR5 in a cell-type dependent manner. A The anti-CCR5 mAbs CTC5, 2D7 and 45531 used in the displacement experiments of 35S-gp120 binding map distinct epitopes of CCR5. B Theoretical picture of gp120 binding competition by mAbs. In these experiments, assuming that...

Effects of WT- or L196K-CCR5 on entry of BlaM-vpr containing virus #25 or #34 into HEK-CD4 cells. CD4-expressing HEK 293 T cells (described in ref 42) in 6-well plates (5 x 105 cells / well) were incubated for 48 h with 25 or 400 ng Gag p24 of FLAG/SNAP-tagged WT-CCR5- or L196K-CCR5-expressing lentiviral particles, respectively. (A) Expression leve...

Binding parameters of the different HIV-1 gp120s used in the study. The KD values represent the equilibrium dissociation constants of the 35S-gp120-sCD4/CCR5 complexes deduced in the saturation binding experiments to membranes from HEK 293T cells expressing CCR5 (HEK-R5 cells). The Bmax values represent the maximum numbers of receptors binding the...

Related to the competition experiments of 35S-gp120 #34 binding by unlabeled gp120s presented in Fig 2. (DOCX)

Binding of 35S-gp120s to intact HEK-CD4 cells. Experiments were carried out as in Fig 1F using 1 x 105 cells in the assay buffer. A representative experiment out of two independent determinations is shown. (PPTX)

The levels of gp120 binding to CCR5 vary differentially between different cell-types. A Specific binding of 10 nM of the indicated 35S-gp120s (+ 200 nM sCD4) to membranes from HEK-R5 cells or the CD4 negative, human primary glioblastoma cell line U87 in which we ectopically expressed CCR5 (U87-R5 cells). U87-R5 cells showing comparable labeling wit...

Distinct HIV-1 gp120s differentially interact with antigenically distinct populations of CCR5. This text is related to S3A, S3B, S3C and S3D Fig. (DOCX)

Saturation and competition binding experiments of gp120s from the Bx08 and 1f HIV-1 strains to membranes from HEK-R5 cells. A The saturation experiments showed that 35S-gp120 1f has a three-fold lower Bmax value compared to 35S-gp120 Bx08. Despite this, however, unlabeled gp120 1f produced full displacement of 35S-gp120 Bx08 in competition experime...

Fusion kinetics of virus #25 or #34 with WT-CCR5- or L196K-CCR5-expressing A3.01 T-cells. Each of the fusion kinetics shown in Fig 5 was expressed as percent of fusion relative to the maximal extent of fusion measured at t = 360 min. Results are presented for all cells (A) or the cells expressing high (B), intermediate (C) or low (D) amounts of rec...

Amino acid sequence alignment of the gp120s used in this study. The HIV-1 gp120 consists of constant regions separated by five variable domains (V1 to V5). The V3 loop (red box) and regions forming the bridging sheet (green-colored) that play a crucial role in binding to CCR5 are shown. Numbering of amino acids is performed relative to the HxB2 ref...

We demonstrate the fabrication of fully printed microfluidic CD4 counting chips with complete on-chip sample preparation and their applicability as a CD4 counting assay using samples from healthy donors and HIV-infected patients. CD4 counting in low-income and resource-limited point-of-care settings is only practical and affordable, if disposable t...

Background: Switch from CCR5 (R5) to CXCR4 (X4) co-receptor tropism of HIV-1 subtype B occurs in ±60% of untreated patients and is associated with rapid progression to AIDS. The etiology of this switch is largely unknown, however CD4+ T-cell activation (CD38-HLADR) has been shown to correlate with X4 tropism in HIV-1 subtype B. We investigated the...

Amongst the many strategies aiming at inhibiting HIV-1 infection, blocking viral entry has been recently recognized as a very promising approach. Using diverse in vitro models and a broad range of HIV-1 primary patient isolates, we report here that IND02, a type A procyanidin polyphenol extracted from cinnamon, that features trimeric and pentameric...

—Fig A. HPLC analysis of IND02. The IND02, isolated from cinnamon extract, was analysed by HPLC, and followed by UV detection at 280 nm, demonstrating the presence of procyanidin trimer (~ 4% of the eluted material) and pentamer polyphenols (~ 92% of the eluted material). Black triangles indicate the beginnings and ends of each peak (see Table A)....

Available data on the use of maraviroc (MVC) in clinical settings are limited. In this cohort study, the clinical outcomes of HIV-1-infected patients treated with MVC were analysed and the predictive values of different tropism assays were compared. Baseline viral tropism was assessed and compared by phenotypic (Trofile and MT-2) and genotypic assa...

Cell-associated Heparan Sulphate (HS) binds the V3 loop of gp120 of HIV-1 thus aiding in viral infectivity. However, a soluble polyanion (HS12) has anti-viral properties once conjugated to CD4 (mCD4-HS12), and showed nM activity against HIV-1 in vitro. Due to the structural complexity of HS, screening differently sulphated-oligosaccharides to impro...

CCR5 has preferentially been used by all circulating HIV-1 subtype C viruses for cell entry. Recently, we reported the highest proportion of CXCR4-utilizing primary isolates among a cohort of 20 South African AIDS patients. This study describes and compares the Env genotypic characteristics from these 20 HIV-1 subtype C (and unique CD recombinant)...

This study investigated the genotype and phenotype of HIV-1 isolates of 20 South African AIDS patients. We found the highest percentage of CXCR4 usage among primary isolates, in which 30% efficiently utilized CXCR4 and exhibited the syncytium-inducing phenotype. Phylogenetic analysis of env confirmed that 19 of the 20 were subtype C, and syncytium-...

This study investigated the genotype and phenotype of HIV-1 isolates of 20 South African AIDS patients. We found the highest percentage of CXCR4 usage among primary isolates, in which 30% efficiently utilized CXCR4 and exhibited the syncytium-inducing phenotype. Phylogenetic analysis of env confirmed that 19 of the 20 were subtype C, and syncytium-...

HIV-1 Vif, Vpr, and Vpu proteins have a profound effect on efficient viral replication and pathogenesis. This study describes the genotypic characterisation of vif , vpr and vpu from 20 South African HIV-1 subtype C primary isolates, and extensive analysis and comparison of known motifs. All HIV-1 subtype C Vif, Vpr and Vpu proteins revealed the pr...