Cecilia Sahlgren

Targeting of Porous Hybrid Silica Nanoparticles to Cancer Cells

Mesoporous silica nanoparticles functionalized by surface hyperbranching polymerization of poly(ethylene imine), PEI, were further modified by introducing both fluorescent and targeting moieties, with the aim of specifically targeting cancer cells. Owing to the high abundance of folate receptors in many cancer cells as compared to normal cells, folic acid was used as the targeting ligand. The internalization of the particles in cell lines expressing different levels of folate receptors was studied. Flow cytometry was used to quantify the mean number of nanoparticles internalized per cell. Five times more particles were internalized by cancer cells expressing folate receptors as compared to the normal cells expressing low levels of the receptor. Not only the number of nanoparticles internalized per cell, but also the fraction of cells that had internalized nanoparticles was higher. The total number of particles internalized by the cancer cells was, therefore, about an order of magnitude higher than the total number of particles internalized by normal cells, a difference high enough to be of significant biological importance. In addition, the biospecifically tagged hybrid PEI-silica particles were shown to be noncytotoxic and able to specifically target folate receptor-expressing cancer cells also under coculture conditions.

Mesoporous silica nanoparticles in medicine—Recent advances☆

MSNs have attracted increasing interest as drug carriers due to promising in vivo results in small-animal disease models, especially related to cancer therapy. In most cases small hydrophobic drugs have been used, but recent in vitro studies demonstrate that MSNs are highly interesting for gene delivery applications. This review covers recent advances related to the therapeutic use of mesoporous silica nanoparticles (MSNs) administered intravenously, intraperitoneally or locally. We also cover the use of MSNs in alternative modes of therapy such as photodynamic therapy and multidrug therapy. We further discuss the current understanding about the biodistribution and safety of MSNs. Finally, we critically discuss burning questions especially related to experimental design of in vivo studies in order to enable a fast transition to clinical trials of this promising drug delivery platform.

Targeted intracellular delivery of hydrophobic agents using mesoporous hybrid silica nanoparticles as carrier systems.

Targeted nanoparticle-mediated intracellular delivery is demonstrated using two hydrophobic fluorophores as model drug cargo. The presented hybrid carrier system exhibits both cancer cell-targeting ability and capacity to retain a hydrophobic agent with subsequent specific release into the endosomal compartment. Furthermore, the incorporated agent is shown to be able to escape from the endosomes into the cytoplasm, making the particles promising candidates as carriers for targeted drug delivery for cancer treatment.

Nanoparticles in targeted cancer therapy: mesoporous silica nanoparticles entering preclinical development stage

Nanotechnology may help overcome persisting limitations of current cancer treatment and thus contribute to the creation of more effective, safer and more affordable therapies. While some nanotechnology-based drug delivery systems are already being marketed and others are in clinical trial, most still remain in the preclinical development stage. Mesoporous silica nanoparticles have been highlighted as an interesting drug delivery platform, due to their flexibility and high drug load potential. Although numerous reports demonstrate sophisticated drug delivery mechanisms in vitro, the therapeutic benefit of these systems for in vivo applications have been under continuous debate. This has been due to nontranslatable conditions used in the in vitro studies, as well as contradictory conclusions drawn from preclinical (in vivo) studies. However, recent studies have indicated that the encouraging cellular studies could in fact be repeated also in vivo. Here, we report on these recent advances regarding therapeutic efficacy, targeting and safety issues related to the application of mesoporous silica nanoparticles in cancer therapy.

Notch Signaling Regulates Platelet-Derived Growth Factor Receptor-β Expression in Vascular Smooth Muscle Cells

Notch signaling is critically important for proper architecture of the vascular system, and mutations in NOTCH3 are associated with CADASIL, a stroke and dementia syndrome with vascular smooth muscle cell (VSMC) dysfunction. In this report, we link Notch signaling to platelet-derived growth factor (PDGF) signaling, a key determinant of VSMC biology, and show that PDGF receptor (PDGFR)-β is a novel immediate Notch target gene. PDGFR-β expression was upregulated by Notch ligand induction or by activated forms of the Notch receptor. Moreover, upregulation of PDGFR-β expression in response to Notch activation critically required the Notch signal integrator CSL. In primary VSMCs, PDGFR-β expression was robustly upregulated by Notch signaling, leading to an augmented intracellular response to PDGF stimulation. In newborn Notch3-deficient mice, PDGFR-β expression was strongly reduced in the VSMCs that later develop an aberrant morphology. In keeping with this, PDGFR-β upregulation in response to Notch activation was reduced also in Notch3-deficient embryonic stem cells. Finally, in VSMCs from a CADASIL patient carrying a NOTCH3 missense mutation, upregulation of PDGFR-β mRNA and protein in response to ligand-induced Notch activation was significantly reduced. In sum, these data reveal a hierarchy for 2 important signaling systems, Notch and PDGF, in the vasculature and provide insights into how dysregulated Notch signaling perturbs VSMC differentiation and function.

Cdk5 regulates the organization of nestin and its association with p35

ABSTRACT The intermediate filament protein nestin is characterized by its specific expression during the development of neuronal and myogenic tissues. We identify nestin as a novel in vivo target for cdk5 and p35 kinase, a critical signaling determinant in development. Two cdk5-specific phosphorylation sites on nestin, Thr-1495 and Thr-316, were established, the latter of which was used as a marker for cdk5-specific phosphorylation in vivo. Ectopic expression of cdk5 and p35 in central nervous system progenitor cells and in myogenic precursor cells induced elevated phosphorylation and reorganization of nestin. The kinetics of nestin expression corresponded to elevated expression and activation of cdk5 during differentiation of myoblast cell cultures and during regeneration of skeletal muscle. In the myoblasts, a disassembly-linked phosphorylation of Thr-316 indicated active phosphorylation of nestin by cdk5. Moreover, cdk5 occurred in physical association with nestin. Inhibition of cdk5 activity—either by transfection with dominant-negative cdk5 or by using a specific cdk5 inhibitor—blocked myoblast differentiation and phosphorylation of nestin at Thr-316, and this inhibition markedly disturbed the organization of nestin. Interestingly, the interaction between p35, the cdk5 activator, and nestin appeared to be regulated by cdk5. In differentiating myoblasts, p35 was not complexed with nestin phosphorylated at Thr-316, and inhibition of cdk5 activity during differentiation induced a marked association of p35 with nestin. These results demonstrate that there is a continuous turnover of cdk5 and p35 activity on a scaffold formed by nestin. This association is likely to affect the organization and operation of both cdk5 and nestin during development.

A nestin scaffold links Cdk5/p35 signaling to oxidant‐induced cell death

The intermediate filament protein, nestin, has been implicated as an organizer of survival‐determining signaling molecules. When nestin expression was related to the sensitivity of neural progenitor cells to oxidant‐induced apoptosis, nestin displayed a distinct cytoprotective effect. Oxidative stress in neuronal precursor cells led to downregulation of nestin with subsequent activation of cyclin‐dependent kinase 5 (Cdk5), a crucial kinase in the nervous system. Nestin downregulation was a prerequisite for the Cdk5‐dependent apoptosis, as overexpression of nestin efficiently inhibited induction of apoptosis, whereas depletion of nestin by RNA interference had a sensitizing effect. When the underlying link between nestin and Cdk5 was analyzed, we observed that nestin serves as a scaffold for Cdk5, with binding restricted to a specific region following the alpha‐helical domain of nestin, and that the presence and organization of nestin regulated the sequestration and activity of Cdk5, as well as the ubiquitylation and turnover of its regulator, p35. Our data imply that nestin is a survival determinant whose action is based upon a novel mode of Cdk5 regulation, affecting the targeting, activity, and turnover of the Cdk5/p35 signaling complex.

Cancer‐Cell‐Specific Induction of Apoptosis Using Mesoporous Silica Nanoparticles as Drug‐Delivery Vectors

Targeted delivery of the chemotherapeutic agent methotrexate (MTX) to cancer cells using poly(ethyleneimine)-functionalized mesoporous silica particles as drug-delivery vectors is reported. Due to its high affinity for folate receptors, the expression of which is elevated in cancer cells, MTX serves as both a targeting ligand and a cytotoxic agent. Enhanced cancer-cell apoptosis (programmed cell death) relative to free MTX is thus observed at particle concentrations where nonspecific MTX-induced apoptosis is not observed in the nontargeted healthy cell line, while corresponding amounts of free drug affect both cell lines equally. The particles remain compartmentalized in endo-/lysosomes during the time of observation (up to 72 h), while the drug is released from the particle only upon cell entry, thereby inducing selective apoptosis in the target cells. As MTX is mainly attached to the particle surface, an additional advantage is that the presented carrier design allows for adsorption (loading) of additional drugs into the pore network for therapies based on a combination of drugs.

Multifunctional Mesoporous Silica Nanoparticles for Combined Therapeutic, Diagnostic and Targeted Action in Cancer Treatment

The main objective in the development of nanomedicine is to obtain delivery platforms for targeted delivery of drugs or imaging agents for improved therapeutic efficacy, reduced side effects and increased diagnostic sensitivity. A (nano)material class that has been recognized for its controllable properties on many levels is ordered mesoporous inorganic materials, typically in the form of amorphous silica (SiO2). Characteristics for this class of materials include mesoscopic order, tunable pore dimensions in the (macro)molecular size range, a high pore volume and surface area, the possibility for selective surface functionality as well as morphology control. The robust but biodegradable ceramic matrix moreover provides shelter for incorporated agents (drugs, proteins, imaging agents, photosensitizers) leaving the outer particle surface free for further modification. The unique features make these materials particularly amenable to modular design, whereby functional moieties and features may be interchanged or combined to produce multifunctional nanodelivery systems combining targeting, diagnostic, and therapeutic actions. This review covers the latest developments related to the use of mesoporous silica nanoparticles (MSNs) as nanocarriers in biomedical applications, with special focus on cancer therapy and diagnostics.

Specific and Innervation-Regulated Expression of the Intermediate Filament Protein Nestin at Neuromuscular and Myotendinous Junctions in Skeletal Muscle

The intermediate filament proteins nestin, vimentin, and desmin show a specific temporal expression pattern during the development of myofibers from myogenic precursor cells. Nestin and vimentin are actively expressed during early developmental stages to be later down-regulated, vimentin completely and nestin to minimal levels, whereas desmin expression begins later and is maintained in mature myofibers, in which desmin participates in maintaining structural integrity. In this study we have analyzed the expression levels and distribution pattern of nestin in intact and denervated muscle in rat and in human. Nestin immunoreactivity was specifically and focally localized in the sarcoplasm underneath neuromuscular junctions (NMJs) and in the vicinity of the myotendinous junctions (MTJs), ie, in regions associated with acetylcholine receptors (AChRs). This association prompted us to analyze nestin in neurogenically and myogenically denervated muscle. Immunoblot analysis disclosed a marked overall increase of accumulated nestin protein. Similar to the extrajunctional redistribution of AChRs in denervated myofibers, nestin immunoreactivity extended widely beyond the NMJ region. Re-innervation caused complete reversion of these changes. Our study demonstrates that the expression levels and distribution pattern of nestin are regulated by innervation, ie, signal transduction into myofibers.