ABSTRACT
The Notch ligand, JAG1 is associated with breast cancer recurrence. Herein, we report on a genomics approach to elucidate mechanisms downstream of JAG1 that promote breast cancer growth. In a survey of 46 breast cancer cell lines, we found that triple negative (TN; basal and mesenchymal ER-, PR-, and Her2-negative) lines express JAG1 at significantly higher levels than do HER2(+) or luminal (ER(+)) Her2(-) cell lines. In contrast to the luminal lines tested (T47D and MCF7), TN breast cancer cell lines (HCC1143 and MDA MB231) display high-level JAG1 expression and growth inhibition with RNA interference-induced JAG1 down-regulation. We used microarray profiling of TN tumor cells transfected with JAG1 siRNA to identify JAG1-regulated genes (P
Subject(s)
Breast Neoplasms/metabolism , Calcium-Binding Proteins/metabolism , Cyclin D1/metabolism , Gene Expression Regulation, Neoplastic , Intercellular Signaling Peptides and Proteins/metabolism , Membrane Proteins/metabolism , Receptors, Notch/metabolism , Signal Transduction/genetics , Blotting, Western , Breast Neoplasms/genetics , Calcium-Binding Proteins/genetics , Cell Line, Tumor , Chromatin Immunoprecipitation , Cyclin D1/genetics , Female , Gene Expression , Humans , Intercellular Signaling Peptides and Proteins/genetics , Jagged-1 Protein , Membrane Proteins/genetics , Oligonucleotide Array Sequence Analysis , Promoter Regions, Genetic/genetics , RNA, Small Interfering , Receptor, ErbB-2/biosynthesis , Receptor, ErbB-2/genetics , Receptors, Estrogen/biosynthesis , Receptors, Estrogen/genetics , Receptors, Notch/genetics , Receptors, Progesterone/biosynthesis , Receptors, Progesterone/genetics , Reverse Transcriptase Polymerase Chain Reaction , Serrate-Jagged Proteins , TransfectionABSTRACT
A novel conjugate of human hemoglobin (Hb) and the nucleoside analogue ribavirin (RBV) was synthesized to demonstrate the utility of Hb as a biocompatible drug carrier for improved drug delivery in the treatment of liver disease. RBV is used in combination with interferon for the treatment of hepatitis C, but its side effects can result in dose limitation or discontinuation of treatment. Targeted delivery of RBV may help to prevent or minimize its toxicity. The hemoglobin-ribavirin conjugate (Hb-RBV) was designed to release bioactive drug upon endocytosis by cells and tissues involved in extracellular Hb catabolism and clearance. Ribavirin-5'-monophosphate (RBV-P) was prepared from RBV and activated as the 5'-monophosphorimidazolide (RBV-P-Im) for reaction with carbonmonoxyhemoglobin to yield Hb-RBV consisting of multiple RBV drugs covalently attached as physiologically labile phosphoramidates via their 5'-hydroxyl groups. A molar drug ratio of six to eight RBV molecules per Hb tetramer was obtained with near complete haptoglobin (Hp) binding of the drug modified Hb maintained. The conjugate complex (Hp-Hb-RBV) was selectively taken up in vitro by cells that express the hemoglobin-haptoglobin receptor, CD163. Recovered ribavirin enzymatically cleaved from Hb-RBV showed equipotent antiproliferative activity compared to control unconjugated RBV against human HepG2 and mouse AML12 liver cell lines. Based upon the reported high level of Hb uptake in the liver, Hb-RBV may be useful in the treatment of certain liver diseases, as well as inflammatory disorders associated with CD163-positive macrophages.
