We present a general approach for the targeting and imaging of cancer cells using dendrimer-entrapped gold nanoparticles (Au DENPs). Au DENPs were found to be able to covalently link with targeting and imaging ligands for subsequent cancer-cell targeting and imaging. The Au DENPs linked with defined numbers of folic acid (FA) and fluorescein isothiocyanate (FI) molecules are water soluble, stable, and biocompatible. In vitro studies show that the FA- and FI-modified Au DENPs can specifically bind to KB cells (a human epithelial carcinoma cell line) that overexpress high-affinity folate receptors and they are internalized dominantly into lysosomes of target cells within 2 h. These findings demonstrate that Au DENPs may serve as a general platform for cancer imaging and therapeutics.

A powerful magnetic nanoprobe with folic acid (FA)‐targeting ligands is fabricated by dendrimer functionalization of Fe3O4 nanoparticles (NPs) precoated with crosslinkable and biocompatible polymer multilayer shells. This magnetic probe allows for magnetic resonance imaging of FA receptor‐overexpressing tumor cells in vitro and of an early‐stage tumor model in vivo (see picture).

Functional interactions between factors bound at multiple sites on DNA often lead to a synergistic or more-than-additive transcriptional response. We previously defined a class of peptide sequences termed synergy control motifs (SC motifs) that function in multiple regulators by selectively inhibiting synergistic activity driven from multiple but not single response elements. By studying the prototypic SC motifs of the glucocorticoid receptor, we show that SC motifs inhibit transcription per se both in cis and in trans, and that a requirement for multiple contacts with DNA renders them selective for compound response elements. Notably, SC motifs are sites for SUMOylation, and the degree of modification correlates strongly with the extent of synergy control. Recruiting SUMO to the promoter either independently or as a fusion to the glucocorticoid receptor is sufficient to recapitulate the in trans and in cis inhibition by SC motifs without apparent changes in subcellular localization. Moreover, we find that the core ubiquitin fold domain of SUMO is sufficient for inhibition and that, independently of their potential for polySUMO chain formation, SUMO-2 and SUMO-3 are more effective inhibitors than SUMO-1. E ukaryotic transcriptional control is highly combinatorial, and the mosaic of response elements in the regulatory elements of a given gene nucleates the assembly of multiprotein complexes where various forms of functional interactions take place (1). One of the most prevalent is the more-than-additive or synergistic response resulting from the recruitment of an activator to multiple copies of a recognition site (compound response element). Despite their importance, the mechanisms that control synergistic effects are poorly understood (2).We have identified a short regulatory motif embedded in a number of sequence-specific regulators that is both necessary and sufficient to limit their transcriptional synergy (3). Disruption of these conserved synergy control (SC) motifs selectively enhances synergistic activation at compound response elements without altering the activity driven from a single site. Structure͞ function analysis of nine SC motifs from the glucocorticoid (GR), mineralocorticoid, and androgen receptors as well as from ETS-1 (3) and C͞EBP␣ (4) revealed a common core sequence (I͞V-K-X-E) and the presence of proline residues within 0-3 aa from either or both ends of the core. Such sequences occur in conserved regions of many factors and, in some cases, these regions have negative regulatory functions (e.g., SP3, SREBP, c-Myb, and C͞EBP). Moreover, a human mutation (P390S) in one of the SC motifs of the androgen receptor is associated with impaired spermatogenesis (5).A clue to the critical role of the Lys residues in SC motifs came from the identification of the consensus motif (⌿-K-x-E͞D) for the posttranslational modification by SUMO. Conjugation of this ubiquitin-like protein follows an analogous pathway to that of ubiquitination and requires dedicated E1 activating (SAE1͞ SAE2) and E2 conjugating ...

Carbon nanotubes hold great promise for their use as a platform in nanomedicine, especially in drug delivery, medical imaging, and cancer targeting and therapeutics. Herein, we present a facile approach to modifying carbon nanotubes with multifunctional poly(amidoamine) (PAMAM) dendrimers for cancer cell targeting and imaging. In this approach, fluorescein isothiocyanate (FI)- and folic acid (FA)-modified amine-terminated generation 5 (G5) PAMAM dendrimers (G5·NH(2)-FI-FA) were covalently linked to acid-treated multiwalled carbon nanotubes (MWCNTs), followed by acetylation of the remaining primary amine groups of the dendrimers. The resulting MWCNT/G5.NHAc-FI-FA composites are water-dispersible, stable, and biocompatible. In vitro flow cytometry and confocal microscopy data show that the formed MWCNT/G5·NHAc-FI-FA composites can specifically target to cancer cells overexpressing high-affinity folic acid receptors. The results of this study suggest that, through modification with multifunctional dendrimers, complex carbon nanotube-based materials can be fabricated, thereby providing many possibilities for various applications in biomedical sensing, diagnosis, and therapeutics.

We develop a facile approach to fabricating multifunctional dendrimer-stabilized gold nanoparticles (Au DSNPs) for cancer cell targeting and imaging. In this work, amine-terminated generation 5 (G5) poly(amidoamine) (PAMAM) dendrimers pre-functionalized with folic acid (FA) and fluorescein isothiocyanate (FI) are complexed with Au(III) ions, followed by acetylation of the amine groups on the dendrimer surfaces. This one-step process leads to the spontaneous formation of 6 nm-sized Au nanoparticles stabilized by multifunctional dendrimers bearing both targeting and imaging functionalities. The multifunctional Au DSNPs are characterized by UV-Vis spectrometry, 1 H NMR, and transmission electron microscopy (TEM). The formed Au DSNPs are water-soluble, stable, and biocompatible. Combined flow cytometry, confocal microscopy, silver staining, and inductively coupled plasma-mass spectrometry (ICP-MS) analyses show that the FAand FI-functionalized Au DSNPs can specifically target to cancer cells expressing high-affinity FA receptors in vitro. This approach to functionalizing Au DSNPs may be extended to other targeting molecules, providing a unique nanoplatform for targeting and imaging of a variety of biological systems.

The proteins encoded by cellular and viral src genes are believed to be involved in the transmission of mitogenic signals, the nuclear recipients of which are largely unknown. In this work, we report that four different v-src-transformed cell lines from three different species possess elevated levels ofjunB transcripts.Transient expression ofjunB promoter-chloramphenicol acetyltransferase constructs in NIH 3T3 cells was used to demonstrate that the increase injunB transcripts was specifically associated with v-src expression and could not be recapitulated with a c-src, v-H-ras, or v-rafexpression vector. Deletion mutants were used to localize the v-src-responsive region in thejunB promoter to a 121-nucleotide region encompassing the CCAAT and TATAA elements. This region is distinct from one in the 5' untranslated region of thejunB gene which is required to maintain its high-level basal expression. Point mutagenesis of thejunB TATAA box completely abolished v-src responsiveness, suggesting that proteins which bind to this element are modified by src transformation. Several v-src and c-src mutants were used to demonstrate that elevated tyrosine kinase activity of src proteins is required for the observed effects on junB expression. Finally, homology between the TATAA box regions of junB and the unrelated but src-responsive gene 9E3/CEF-4 suggests that modulation of gene activity through proteins which bind to this region may be a recurrent, although not exclusive, theme in src transforming action. Our results suggest that src proteins may modulate some nuclear effectors through pathways not involving cellular ras or raf gene products.Numerous studies indicate that the normal pp60csrc protein and its oncogenic viral counterpart, pp6Ovsrc, are intimately involved in cell growth control and differentiation (8,28,37). Some progress has been made in identifying and assigning a putative order to intermediates along the src pathway. For example, the activation of the platelet-derived growth factor receptor in quiescent cells by ligand binding is accompanied by the physical association of pp60csrc with the receptor and by increased pp60csrc phosphorylation and intrinsic tyrosine kinase activity (1,20,32,34). In addition, pp6Oc-src and pp60Osrc associate with and phosphorylate ras

The proteins encoded by cellular and viral src genes are believed to be involved in the transmission of mitogenic signals, the nuclear recipients of which are largely unknown. In this work, we report that four different v-src-transformed cell lines from three different species possess elevated levels of junB transcripts. Transient expression of junB promoter-chloramphenicol acetyltransferase constructs in NIH 3T3 cells was used to demonstrate that the increase in junB transcripts was specifically associated with v-src expression and could not be recapitulated with a c-src, v-H-ras, or v-raf expression vector. Deletion mutants were used to localize the v-src-responsive region in the junB promoter to a 121-nucleotide region encompassing the CCAAT and TATAA elements. This region is distinct from one in the 5' untranslated region of the junB gene which is required to maintain its high-level basal expression. Point mutagenesis of the junB TATAA box completely abolished v-src responsiveness, suggesting that proteins which bind to this element are modified by src transformation. Several v-src and c-src mutants were used to demonstrate that elevated tyrosine kinase activity of src proteins is required for the observed effects on junB expression. Finally, homology between the TATAA box regions of junB and the unrelated but src-responsive gene 9E3/CEF-4 suggests that modulation of gene activity through proteins which bind to this region may be a recurrent, although not exclusive, theme in src transforming action. Our results suggest that src proteins may modulate some nuclear effectors through pathways not involving cellular ras or raf gene products.