Imaging and therapy of tumors induced to express somatostatin receptor by gene transfer using radiolabeled peptides and single chain antibody constructs

2004 ◽  
Vol 34 (1) ◽  
pp. 32-46 ◽  
Author(s):  
Donald J Buchsbaum
Keyword(s):  
Gene Transfer ◽  
Somatostatin Receptor ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Radiolabeled Peptides

Targeting Gene Transfer Selectively to Vascular Endothelial Cells Using Peptides Isolated by Phage Display: Implications for Development of Gene Therapy in Hypertension

Hypertension ◽  
2000 ◽  
Vol 36 (suppl_1) ◽  
pp. 691-691
Author(s):  
Stuart A Nicklin ◽  
Steve J White ◽  
Andrew H Baker
Keyword(s):  
Endothelial Cells ◽  
Gene Transfer ◽  
Phage Display ◽  
Vascular Endothelial Cells ◽  
Ldl Receptor ◽  
Vascular Endothelial ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Adenoviral Gene Transfer ◽  
Low Efficiency

75 Current gene transfer vectors are extremely limited for selective vascular cell delivery due to their promiscuous tropism and low efficiency of gene delivery to the vasculature. We have sought to improve the efficiency of gene transfer to vascular endothelial cells using phage display. Using bio-panning on whole cells, we have isolated a panel of 60 7-mer peptides that have the ability to bind endothelial cells but not to non-endothelial cells including vascular smooth muscle cells and hepatocytes. One candidate peptide was cloned upstream of a single chain antibody (scFv) generated against the knob domain of adenovirus type 5, expressed in bacteria and purified. While the scFv alone inhibited adenoviral fiber-dependent infection of all cell types tested (to >95% in hepatocytes), the scFv-peptide fusion mediated selective infection into endothelial cells without infection into non-endothelial cells types. Furthermore, the level of infection achieved in endothelial cells was 15 fold higher than that achieved with fiber-mediated gene transfer alone. We have additionally isolated 15 individual peptides that have the ability to target the endothelial leptin-like oxidised LDL receptor (LOX-1), a receptor highly expressed in endothelial cells in hypertensive models and in atherosclerotic lesions, by phage bio-panning on cells ectopically expressing the LOX-1 receptor. Candidate peptides mediated significantly higher binding to LOX-1 expressing cells compared to LOX-1 negative cells. Their ability to re-target adenoviral gene transfer is being tested. Our results demonstrate that small, novel peptides isolated by phage display have the ability to retarget gene transfer selectively and efficiently to vascular endothelial cells. This has important implications for targeting gene transfer to endothelial cells for molecular and therapeutic protocols in hypertension.

Efficient gene transfer into human primary blood lymphocytes by surface-engineered lentiviral vectors that display a T cell–activating polypeptide

Blood ◽  
2002 ◽  
Vol 99 (7) ◽  
pp. 2342-2350 ◽  
Author(s):  
Marielle Maurice ◽  
Els Verhoeyen ◽  
Patrick Salmon ◽  
Didier Trono ◽  
Stephen J. Russell ◽  
...  
Keyword(s):  
T Cell ◽  
Gene Transfer ◽  
Gene Delivery ◽  
Genetic Material ◽  
Lentiviral Vectors ◽  
Target Cells ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Blood Lymphocytes ◽  
Hiv 1

In contrast to oncoretroviruses, lentiviruses such as human immunodeficiency virus 1 (HIV-1) are able to integrate their genetic material into the genome of nonproliferating cells that are metabolically active. Likewise, vectors derived from HIV-1 can transduce many types of nonproliferating cells, with the exception of some particular quiescent cell types such as resting T cells. Completion of reverse transcription, nuclear import, and subsequent integration of the lentivirus genome do not occur in these cells unless they are activated via the T-cell receptor (TCR) or by cytokines or both. However, to preserve the functional properties of these important gene therapy target cells, only minimal activation with cytokines or TCR-specific antibodies should be performed during gene transfer. Here we report the characterization of HIV-1–derived lentiviral vectors whose virion surface was genetically engineered to display a T cell-activating single-chain antibody polypeptide derived from the anti-CD3 OKT3 monoclonal antibody. Interaction of OKT3 IgGs with the TCR can activate resting peripheral blood lymphocytes (PBLs) by promoting the transition from G0 to G1 phases of the cell cycle. Compared to unmodified HIV-1–based vectors, OKT3-displaying lentiviral vectors strongly increased gene delivery in freshly isolated PBLs by up to 100-fold. Up to 48% transduction could be obtained without addition of PBL activation stimuli during infection. Taken together, these results show that surface-engineered lentiviral vectors significantly improve transduction of primary lymphocytes by activating the target cells. Moreover these results provide a proof of concept for an approach that may have utility in various gene transfer applications, including in vivo gene delivery.

scholarly journals Structure Determination of Inactive-State GPCRs with a Universal Nanobody

2021 ◽  
Author(s):  
Michael J. Robertson ◽  
Feng He ◽  
Justin G. Meyerowitz ◽  
Alpay B. Seven ◽  
Ouliana Panova ◽  
...  
Keyword(s):  
Somatostatin Receptor ◽  
Protein Structures ◽  
Single Chain ◽  
Intracellular Loop ◽  
Single Chain Antibody ◽  
Inactive State ◽  
X Ray Crystallography ◽  
High Resolution Map ◽  
Μ Opioid Receptor

Cryogenic electron microscopy (cryo-EM) has widened the field of structure-based drug discovery by allowing for routine determination of membrane protein structures previously intractable. However, despite representing one of the largest classes of therapeutic targets, most inactive-state G protein-coupled receptors (GPCRs) have remained inaccessible for cryo-EM because their small size and membrane-embedded nature impedes projection alignment for high-resolution map reconstructions. Here we demonstrate that a camelid single-chain antibody (nanobody) recognizing a grafted intracellular loop can be used to obtain cryo-EM structures of different inactive-state GPCRs at resolutions comparable or better than those obtained by X-ray crystallography. Using this approach, we obtained the structure of human neurotensin 1 receptor (NTSR1) bound to antagonist SR48692, of μ-opioid receptor (MOR) bound to the clinical antagonist alvimopan, as well as the structure of the previously uncharacterized somatostatin receptor 2 (SSTR2) in the apo state; each of these structures yields novel insights into ligand binding and specificity. We expect this rapid, straightforward approach to facilitate the broad structural exploration of GPCR inactive states without the need for extensive engineering and crystallization.

Single-chain Antibody Against Reg4 Suppresses Gastric Cancer Cell Growth and Enhances 5-FU-induced Cell Death in vitro

2019 ◽  
Vol 19 (5) ◽  
pp. 610-619 ◽  
Author(s):  
Xue-Qing Zhang ◽  
Lu-Ting Yu ◽  
Pei Du ◽  
Tian-Qi Yin ◽  
Zhi-Yuan Zhang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Cell Death ◽  
Cancer Cell ◽  
Gastric Cancer Cell ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Ags Cells ◽  
Thiazolyl Blue Tetrazolium Bromide

Background:Regenerating islet-derived gene family member 4 (Reg4), a well-investigated growth factor in the regenerative pancreas, has recently been reported to be highly associated with a majority of gastrointestinal cancers. Pathological hyper-expression or artificial over-expression of Reg4 causes acceleration of tumor growth, migration, and resistance to chemotherapeutic 5-Fluorouracil (5-FU). Until now, no method has been successfully established for eliminating the effects of Reg4 protein.Methods:This study reports the production of an engineered immunoglobin, a single-chain variable fragment (scFv-Reg4), to specifically bind Reg4 and block the bioactivity. The complementary-determining regions (CDRs) against Reg4 were assigned using MOE and ZDOCK servers. The binding affinity (KD) was determined by bio-layer interferometry (BLI). MKN45 and AGS cell proliferation was determined by Thiazolyl blue tetrazolium bromide (MTT) method and the cell apoptosis was detected by flow cytometry assay.Results:The KD of scFv-Reg4 to Reg4 was determined to be 1.91×10-8. In MKN45 and AGS cell lines, scFv- Reg4 depressed Reg4-stimulated cell proliferation and the inhibitory rates were 27.7±1.5% and 17.3±2.6%, respectively. Furthermore, scFv significantly enhanced 5-FU-induced cell death, from 23.0±1.0% to 28.4±1.2% in MKN45 and 28.2±0.7% to 36.6±0.6% in AGS cells. Treatment with scFv alone could lyse cancer cells to a certain extent, but no significance has been observed.Conclusion:The single-chain antibody (scFv-Reg4) significantly inhibited gastric cancer cell proliferation and synergistically enhanced the lethal effect of 5-FU. Thus, traditional chemo-/radio- therapeutics supplemented with scFv-Reg4 may provide advances in the strategy for gastrointestinal cancer treatment.

Single-chain antibody-decorated Au nanocages@liposomal layer nanoprobes for targeted SERS imaging and remote-controlled photothermal therapy of melanoma cancer cells

2021 ◽  
Vol 124 ◽  
pp. 112086
Author(s):  
Ghazal Farahavar ◽  
Samira Sadat Abolmaali ◽  
Foroogh Nejatollahi ◽  
Amin Safaie ◽  
Sanaz Javanmardi ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Photothermal Therapy ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Melanoma Cancer ◽  
Sers Imaging

scholarly journals Computational and Rational Design of Single-Chain Antibody against Tick-Borne Encephalitis Virus for Modifying Its Specificity

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1494
Author(s):  
Ivan K. Baykov ◽  
Pavel Y. Desyukevich ◽  
Ekaterina E. Mikhaylova ◽  
Olga M. Kurchenko ◽  
Nina V. Tikunova
Keyword(s):  
Rational Design ◽  
Fold Increase ◽  
Encephalitis Virus ◽  
Chimeric Antibody ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Glycoprotein E ◽  
Tick Borne Encephalitis ◽  
Tick Borne Encephalitis Virus ◽  
Far Eastern

Tick-borne encephalitis virus (TBEV) causes 5−7 thousand cases of human meningitis and encephalitis annually. The neutralizing and protective antibody ch14D5 is a potential therapeutic agent. This antibody exhibits a high affinity for binding with the D3 domain of the glycoprotein E of the Far Eastern subtype of the virus, but a lower affinity for the D3 domains of the Siberian and European subtypes. In this study, a 2.2-fold increase in the affinity of single-chain antibody sc14D5 to D3 proteins of the Siberian and European subtypes of the virus was achieved using rational design and computational modeling. This improvement can be further enhanced in the case of the bivalent binding of the full-length chimeric antibody containing the identified mutation.

Sign in / Sign up

Export Citation Format

Share Document