Backgroundα3β1 integrin is overexpressed in several types of human cancer and is associated with poor prognosis, metastasis, and resistance to cancer treatment. We previously identified a cyclic peptide ligand LXY1 that specifically binds to the α3β1 integrin on human glioblastoma U-87MG cells. Here, we optimized LXY1 through one-bead one-compound combinatorial library screening and site-specific modifications to improve its in vivo binding property.MethodsThree bead libraries were synthesized and whole-cell binding assays were performed. The binding capacity of individual peptide ligands against different tumor cells was determined by flow cytometry and confirmed by optical imaging. A complex joining biotinylated ligand with streptavidin-Cy5.5 was used for in vivo target imaging in both subcutaneous and orthotopic U-87MG xenograft mouse models.ResultsLXY30, a cyclic peptide with the sequence cdG-Phe(3,5-diF)-G-Hyp-NcR, emerged as the most potent and selective ligand for the α3 subunit of α3β1 integrin with improved in vitro and in vivo tumor-targeting effects compared to LXY1 in U-87MG cells. LXY30 is considerably stable in plasma as demonstrated in an in vitro stability study in 90 % human plasma. LXY30 also binds to several other known α3β1 integrin-expressing glioblastoma, lung, and breast cancer cell lines with various affinities.ConclusionsOur data support further investigating the role of LXY30 as a human tumor-targeting peptide ligand for systemic and intracranial delivery of imaging agents and cancer therapeutics.Electronic supplementary materialThe online version of this article (doi:10.1186/s13550-016-0165-z) contains supplementary material, which is available to authorized users.
Extracellular ATP via the activation of purinergic P2 receptors has an emerging role in cutaneous biology; however, the distribution of these receptors in mouse skin is poorly defined. This study investigated whether murine epidermal cell subpopulations express functional purinergic P2X 7 receptors. P2X 7 expression was examined by immunoblotting and immunofluorescence staining of epidermal cells from C57Bl ⁄ 6 mice. P2X 7 function was evaluated by nucleotide-induced ethidium + uptake measurements in epidermal cells from C57Bl
Natural killer (NK) cells are lymphocytes that provide an important line of defense against many types of microorganisms, viruses and tumors. The development of an efficient gene transfer system for genetically modifying primary murine NK cells will facilitate the studies of NK cell differentiation, acquisition of self-tolerance, and induction of anti-tumor responses. In this study we used an enhanced green fluorescent protein (EGFP)-expressing vector to carry out a systematic evaluation of the efficiency of lentiviral transduction of primary murine NK cells with or without prior interleukin-2 (IL-2) activation. In a single-step transduction protocol, we demonstrated that human immunodeficiency virus type 1-based lentiviral vectors support an average of 40% transduction efficiency on primary NK cells. These genetically modified NK cells are found to maintain stable EGFP transgene expression in vitro, and can be further expanded in IL-2 supplemented culture medium. Lentiviral transduction does not affect NK surface phenotypes or functions (apoptosis, cytokine production and cytotoxicity). We further demonstrated efficient gene transfer into differentiating NK cells derived from the lentiviral-transduced murine hematopoietic progenitor cells in vitro. This study therefore establishes a simple and efficient approach to the genetic engineering of primary murine NK cells, and will prove useful in studying basic NK cell biology and in exploring the therapeutic potential of NK cells in inbred and transgenic mouse models.
The P2X7 receptor is an extracellular ATP-gated cation channel critical in inflammation and immunity, and can be up-regulated by IFN-γ and LPS. This study aimed to examine the effect of TGF-β1 on the up-regulation of P2X7 function and expression in leukemic THP-1 monocytes differentiated with IFN-γ and LPS. Cell-surface molecules including P2X7 were examined by immunofluorescence staining. Total P2X7 protein and mRNA was assessed by immunoblotting and RT-PCR respectively. P2X7 function was evaluated by ATP-induced cation dye uptake measurements. Cell-surface P2X7 was present on THP-1 cells differentiated for 3days with IFN-γ and LPS but not on undifferentiated THP-1 cells. ATP induced ethidium(+) uptake into differentiated but not undifferentiated THP-1 cells, and the P2X7 antagonist, KN-62, impaired ATP-induced ethidium(+) uptake. Co-incubation of cells with TGF-β1 plus IFN-γ and LPS prevented the up-regulation of P2X7 expression and ATP-induced ethidium(+) uptake in a concentration-dependent fashion with a maximum effect at 5ng/ml and with an IC(50) of ~0.4ng/ml. Moreover, ATP-induced YO-PRO-1(2+) uptake and IL-1β release were abrogated in cells co-incubated with TGF-β1. TGF-β1 also abrogated the amount of total P2X7 protein and mRNA induced by IFN-γ and LPS. Finally, TGF-β1 prevented the up-regulation of cell-surface CD86, but not CD14 and MHC class II, by IFN-γ and LPS. These results indicate that TGF-β1 prevents the up-regulation of P2X7 function and expression by IFN-γ and LPS in THP-1 monocytes. This suggests that TGF-β1 may limit P2X7-mediated processes in inflammation and immunity.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.