Biomedical subjects

Zhenping Zhu

Publications and source records attributed to Zhenping Zhu.

60 records · Page 4Linked to original sources

PubMed2002

Fab-scFv fusion protein: an efficient approach to production of bispecific antibody fragments.

The clinical development of bispecific antibodies (BsAb) as therapeutics has been hampered by the difficulty in preparing the materials in sufficient quantity and quality by traditional methods. Here, we describe an efficient approach for the production of a novel bispecific antibody fragment by genetically fusing a single-chain Fv (scFv) to the C-terminus of either the light chain or the heavy chain of a Fab fragment of different antigen-binding specificity. The bispecific Fab-scFv fragments were expressed in a single Escherichia coli host and purified to homogeneity by a one-step affinity chromatography. Two different versions of the bispecific Fab-scFv fragments were constructed using two antibodies directed against the two tyrosine kinase receptors of vascular endothelial growth factor. These bispecific antibody fragments not only retained the antigen-binding capacity of each of the parent antibodies, but also are capable of binding to both targets simultaneously as demonstrated by a cross-linking ELISA. Further, the bispecific antibodies were comparable to their parent antibodies in their potency in blocking ligand binding to the receptors and in inhibiting ligand-induced biological activities. This design for BsAb fragments should be applicable to any pair of antigen specificities.

Animals↗

PubMed2002

Placental growth factor reconstitutes hematopoiesis by recruiting VEGFR1(+) stem cells from bone-marrow microenvironment.

The mechanism by which angiogenic factors recruit bone marrow (BM)-derived quiescent endothelial and hematopoietic stem cells (HSCs) is not known. Here, we report that functional vascular endothelial growth factor receptor-1 (VEGFR1) is expressed on human CD34(+) and mouse Lin(-)Sca-1(+)c-Kit(+) BM-repopulating stem cells, conveying signals for recruitment of HSCs and reconstitution of hematopoiesis. Inhibition of VEGFR1, but not VEGFR2, blocked HSC cell cycling, differentiation and hematopoietic recovery after BM suppression, resulting in the demise of the treated mice. Placental growth factor (PlGF), which signals through VEGFR1, restored early and late phases of hematopoiesis following BM suppression. PlGF enhanced early phases of BM recovery directly through rapid chemotaxis of VEGFR1(+) BM-repopulating and progenitor cells. The late phase of hematopoietic recovery was driven by PlGF-induced upregulation of matrix metalloproteinase-9, mediating the release of soluble Kit ligand. Thus, PlGF promotes recruitment of VEGFR1(+) HSCs from a quiescent to a proliferative BM microenvironment, favoring differentiation, mobilization and reconstitution of hematopoiesis.

Animals↗

PubMed2002

Efficient inhibition of human B-cell lymphoma xenografts with an anti-CD20 x anti-CD3 bispecific diabody.

Bispecific antibodies have been exploited both as cancer immunodiagnostics and as cancer therapeutics, and have shown promise in several clinical trials in cancer imaging and therapy. A number of bispecific antibodies against B-cell markers have been shown to be effective in vitro in mediating tumor cell lysis and in vivo in inhibiting tumor growth in animal models. We have constructed a bispecific diabody from the variable genes encoding two hybridoma-derived monoclonal antibodies directed against human CD20 on B cells and CD3 on T cells. The anti-CD20 x anti-CD3 diabody was expressed in a single Escherichia coli host and purified by a one-step affinity chromatography. The bispecific diabody bound as efficiently to both CD20- and CD3-positive cells as the respective parental antibodies, and was capable of cross-linking CD20-positive tumor cells and human T lymphocytes as shown by cellular rosetting. The diabody effectively lysed human B-lymphoma cells in the presence of T-enriched human peripheral blood lymphocytes (PBL). Further, when combined with human PBL and interleukin-2, the diabody significantly prolonged the survival of nude mice inoculated with human B-lymphoma cells. Taken together, our results suggest that an anti-CD20 x anti-CD3 diabody may have significant clinical application in the treatment of human CD20-positive B-cell malignancies.

Animals↗

PubMed2002

Selection of high affinity human neutralizing antibodies to VEGFR2 from a large antibody phage display library for antiangiogenesis therapy.

Compelling evidence suggests that vascular endothelial growth factor (VEGF) and its receptors play an important role in angiogenesis associated with tumor growth and metastasis. VEGF exerts its biologic activities through 2 transmembrane tyrosine kinase receptors: the fms-like tyrosine kinase receptor (Flt-1, or VEGFR1) and kinase insert domain-containing receptor (KDR or VEGFR2). We have previously produced a panel of antibodies directed against KDR from mice immunized with the recombinant form receptor. These antibodies efficiently neutralized VEGF-induced KDR activation and mitogenesis of human umbilical vascular endothelial cells (HUVEC). Murine antibodies, however, may not be suitable candidates for human therapy because of their propensity to elicit human anti-mouse antibody response. Here we isolated several high-affinity human Fab antibody fragments directed against KDR from an antibody phage display library constructed from the pooled B lymphocytes of nonimmunized healthy human donors. These human Fab fragments bind specifically to KDR with nanomolar affinity and block KDR/VEGF interaction with IC(50) of approximately 2-20 nM. Further, they effectively inhibit VEGF-stimulated mitogenesis of HUVEC and migration of human leukemia cells. Epitope mapping studies demonstrated that all neutralizing human antibodies bound the epitope(s) located within the first 3 N-terminal immunoglobulin-like domains of KDR, the same region that encompasses the binding site of VEGF. Our results suggest that these human anti-KDR antibodies may have potential application in the treatment of cancer and other diseases in which pathologic angiogenesis occurs.

Amino Acid Sequence↗

PubMed2002

Clinical development of angiogenesis inhibitors to vascular endothelial growth factor and its receptors as cancer therapeutics.

Angiogenesis, the formation of new blood vessels, is essential for both tumor growth and metastasis. Recent advances in our understanding of the molecular mechanisms underlying the angiogenesis process and its regulation have led to the discovery of a variety of pharmaceutical agents with anti-angiogenic activity. The potential application of these angiogenesis inhibitors is currently under intense clinical investigation. Compelling evidence suggests that vascular endothelial growth factor (VEGF) and its receptors play critical roles in tumor-associated angiogenesis, and that they represent potential targets for therapeutic intervention. This has been demonstrated in a variety of animal tumor models in which disabling the function of VEGF and its receptors was shown to inhibit both tumor growth and metastasis. A number of agents designed specifically for targeting VEGF and/or its receptors are being evaluated in various clinical trials in cancer patients. This review will discuss the biology of the VEGF and its receptors, the mechanisms of action as well as the current status in clinical development of antagonistic agents to VEGF and its receptors. Included in this review are antagonistic antibodies, ribozymes, immunotoxins, and synthetic small molecular inhibitors.

Angiogenesis Inhibitors↗

PubMed2002

Localization of VEGF receptor-2 (KDR/Flk-1) and effects of blocking it in oxygen-induced retinopathy.

PURPOSE: Vascular endothelial cell growth factor (VEGF) has been implicated in vascular development and in proliferative retinopathies. The goal of this study was to examine the immunohistochemical localization and relative levels of VEGF receptor-2 (KDR) in canine retina during postnatal vasculogenesis and during angiogenesis in oxygen-induced retinopathy (OIR) and to investigate the effects of neutralizing KDR on these processes. METHODS: Eyes from normal dogs ranging from 1 to 22 days of age and age-matched oxygen-treated animals were snap frozen for immunohistochemical analysis with antibodies against human KDR. To examine the effects of blocking KDR, 6-day-old air-reared control and oxygen-treated animals were surgically implanted with slow release polymer pellets containing control IgG or anti-KDR. Material eluted from pellets was assessed using a binding assay (measures binding to soluble KDR) to determine the kinetics of anti-KDR release and endothelial cell proliferation to measure bioactivity. Animals were killed at 22 days of age and tissues examined with adenosine diphosphatase (ADPase) histochemical staining of blood vessels. RESULTS: KDR immunoreactivity was only weakly associated with developing retinal vessels and was not observed in angioblasts throughout normal postnatal development. Immunoreactivity was very strong in reforming retinal vessels and intravitreal neovascularization in oxygen-treated animals. Anti-KDR had no effect on vessel morphology or growth in air-reared control animals. In oxygen-treated animals, anti-KDR significantly inhibited revascularization of the retina (P = 0.005) and formation of intravitreal neovascularization compared with control IgG pellet eyes (P < 0.04). CONCLUSIONS: KDR/Flk-1 was only weakly associated with normal developing primary retinal vessels but was strongly expressed by proliferating endothelial cells in reforming retinal vessels and intravitreal neovascularization after hyperoxic insult. Anti-KDR antibody delivered by slow-release pellets had no effect on normal vasculogenesis, but it inhibited the formation of intravitreal neovascularization and retinal vessel development in OIR. The study suggests that blocking KDR may be beneficial for treating pathologic angiogenesis in adult tissue.

Animals↗