Human Breast Tumor-associated Endothelial Cells Favor Interactions with Nanomedicines
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ABSTRACT: Nanomedicines, designed to improve the effectiveness of cancer treatment, have not met anticipated outcomes owing to suboptimal delivery efficiency to tumors. While the enhanced permeability and retention (EPR) effect has been explored, recent research challenges the predominant passive accumulation theory. This study explores the mechanisms governing nanoparticle entry into tumors by comparing human breast tumor-associated endothelial cells (HBTECs) and healthy human mammary macrovascular endothelial cells (HMMECs). Our results demonstrate that HBTECs exhibit higher nanoparticle uptake, involving three major endocytic pathways. Gene expression analysis reveals the upregulation of transport-related genes, especially motor protein genes, in tumor-associated endothelial cells. These insights highlight the need for comprehensive investigations into nanoparticle transport mechanisms and associated factors within the tumor microenvironment, essential for advancing drug delivery and therapeutic outcomes.
Project description:Cancer cells re-program normal lung endothelial cells (EC) into tumor-associated endothelial cells (TEC) that form leaky vessels supporting carcinogenesis. Transcriptional regulators that control reprogramming of EC into TEC are poorly understood. We identified Forkhead box F1 (FOXF1) as a critical regulator of EC-to-TEC transition. FOXF1 was highly expressed in normal lung vasculature but was decreased in TEC within non-small cell lung cancers (NSCLC). Low FOXF1 correlated with poor overall survival of NSCLC patients. In mice, endothelial-specific deletion of FOXF1 decreased pericyte coverage, increased vessel permeability and hypoxia, and promoted lung tumor growth and metastasis. Endothelial-specific over-expression of FOXF1 normalized tumor vessels and inhibited progression of lung cancer. FOXF1-deficiency decreased Wnt/β-catenin signaling in TECs through direct transcriptional activation of Fzd4. Restoring FZD4 expression in FOXF1-deficient TECs through endothelial-specific nanoparticle delivery of Fzd4 cDNA rescued Wnt/β-catenin signaling in TECs, normalized tumor vessels and inhibited progression of lung cancer. Altogether, FOXF1 increases tumor vessel stability, and inhibits lung cancer progression by stimulating FZD4/Wnt/β-catenin signaling in TECs. Nanoparticle delivery of FZD4 cDNA has a promise for future therapies in NSCLC.
Project description:Silencing HoxA1 in vivo by intraductal delivery of nanoparticle-formulated siRNA reduced mammary tumor incidence by 75% , reduced cell proliferation, and prevented loss of ER and PR expression. 8 week wild type FVB mouse whole mammary gland and 8week to 20 week transgenic FVB C3(1)-SV40Tag mouse whole mammary gland
Project description:Silencing HoxA1 in vivo by intraductal delivery of nanoparticle-formulated siRNA reduced mammary tumor incidence by 75% , reduced cell proliferation, and prevented loss of ER and PR expression.
Project description:TNF-alpha has a number of pro-atherogenic effects in macrovascular endothelial cells, including induction of leukocyte adhesion molecules and chemokines. We investigated the role of acyl-CoA synthetase 3 (ACSL3) in the response of cultured human macrovascular endothelial cells to TNF-alpha. TNF-alpha induced ACSL3 both in human umbilical vein endothelial cells (HUVECs) and in human coronary artery endothelial cells (HCAECs). RNA sequencing demonstrated that knockdown of ACSL3 had no marked effects on the TNF-alpha transcriptome in HCAECs. Instead, ACSL3 was required for TNF-alpha-induced lipid droplet formation from fatty acids.
Project description:Endothelial cells (EC) were obtained from commercial source (Lonza), expanded and culture until passage 3. HMVEC-LBI correspond to lung microvascular endothelial cells, HPAEC to lung macrovascular endothelial cells, HAEC to cardiac macrovascular endothelial cells and HMVEC-C to cardia microvascular endothelial cells. These cells presented different capacity to generate endothelial microvesicles. The objective of the study was to identify differentially genes between these cells to understand this difference. Total RNA was extracted using the mirVana miRNA Isolation Kit (Ambion), according to the manufacturer’s recommendations. Cy3-CTP labeled RNA was prepared according to standard Agilent protocol from 400ng total RNA. The hybridization was performed for 17 hrs at 65°C. cRNA labeling and hybridization performance were performed and all parameters checked were found within the manufacturers specifications. Arrays were scanned as described in the manufacturers’ protocol. Signal intensities on 20 bit tiff images were calculated by Feature Extraction software (FE, Version 8.5; Agilent Technologies). Data analyses were conducted with GeneSpring GX software (Vers.12.6; Agilent Technologies)
Project description:Poly(ethylene glycol) (PEG) is widely used in engineering of clinical nanomedicines; however, the emerging PEG immunogenicity is compromising their therapeutic efficacy. Polyoxazolines are gaining popularity as PEG alternatives, since “polyoxazolination” confers stealthing behaviour to nanomedicines in the preclinical mouse model. Here, using nanoparticles coated with either poly-2-methyl-2-oxazoline (PMOXA) or poly-2-ethyl-2-oxazoline (PEOXA), we show species differences in nanoparticle opsonization and phagocytic cell responses. PMOXA- and PEOXA-coated nanoparticles evade murine serum opsonization and phagocytic clearance. Polyoxazolinated nanoparticles, however, undergo differential opsonization in porcine and human sera, resulting in nanoparticle recognition by different phagocytic cells differently. We identify ficolin 2 (FCN2) as a monocyte-specific opsonin in pigs, whereas in human sera, FCN opsonization is isoform-dependent with inter-individual variability. Polymer recognition is through the S2 domain of FCN2 which sense hydrogen bond forming chemical moieties and hydrophobic patterns of polyoxazolines. Contrary to the porcine model, both FCN2 and complement promote nanoparticle uptake by human monocytes. On the other hand, nanoparticle uptake by human and porcine macrophage is complement-dependent. These findings highlight the importance of species differences in innate immune recognition of nanomaterials’ molecular patterns, and are relevant to the selection and chemical design of polymers for engineering of the next generation of stealth nanoparticles.
Project description:Endothelial cells represent the key cellular components of the blood-brain barrier, whose selective permeability presents a challenge for drug delivery. A better understanding of molecular proteins specific for brain endothelial cells compared to endothelial barries in different tissues can highlight novel blood-brain barrier transport targets.
Project description:We sequenced the transcroptome of isolated nanoparticle positive and nanoparticle negative tumour endothelial cells from mouse 4T1 mammary tumours