Highly organized nanostructures for brain drug delivery--new hope or just a fad?

The blood-brain barrier significantly impedes treatment of central nervous system disorders by preventing drug entry into the brain. Several strategies have been developed to overcome this problem, but progress has been hampered due to a lack of efficacious drug delivery systems (DDS). Now, owing to DDS, therapeutic compounds can be transported to the site of action and accumulate there. This modern approach allows one to decrease the required dose of drug and, therefore, minimize toxicity and side effects. Also, treatment efficiency is increased. Highly organized nanostructures made of biological, polymeric or carbon-based materials are promising carriers in drug delivery to the brain, due to their unique and easily tailorable properties. The drug can be either attached to or entrapped in a carrier. To achieve greater site specificity and selectivity, DDS can be also modified with suitable ligands, providing identification of the molecular site of action. This review illustrates recent advances in using highly-organized structures: dendrimers, fullerenes, liposomes, micelles, nanogels, nanoparticles and nanotubes for this purpose. We also discuss advantages and limitations of each system.

Evidence that FGFR1 loss-of-function mutations may cause variable skeletal malformations in patients with Kallmann syndrome.

PURPOSE: Loss-of-function mutations in FGFR1 have been identified in approximately 10% of the Kallmann syndrome (KS) patients. Previous reports have focused mainly on olfactory, reproductive, and some other features such as cleft lip/palate and dental agenesis. Given the ubiquitous expression of FGFR1 during development, other abnormal phenotypes might, however, have been overlooked in these patients. Here, we demonstrate skeletal phenotypic characterization of patients presented with KS and FGFR1 mutations. MATERIAL AND METHODS: Using the Sanger DNA sequencing technique a cohort of 29 KS patients was screened. RESULTS: Here, we report on 5 KS patients who carry FGFR1 mutations (Gly270Asp, Gly97Ser, Met161Thr, Ser685Phe and Ala167Ser/Ala167Ser). Three patients presented with skeletal abnormalities, i.e. spine (hemivertebra and butterfly vertebra) and limb (oligodactyly of the feet, fusion of the 4th and 5th metacarpal bones) malformations in two patients and one patient, respectively. The hand phenotype found in the patient cannot be thought of as a counter-type of the hand phenotype resulting from FGFR1 gain-of-function mutations. The skeletal anomalies identified in the 3 KS patients are close to those observed in Fgfr1 conditional knockout mice. CONCLUSIONS: This study demonstrates that FGFR1 loss-of-function mutations can be associated with skeletal abnormalities also in humans. Further investigations in KS patients who carry FGFR1 mutations are needed to evaluate the prevalence of skeletal defects in this genetic form of KS. Conversely, the presence of bone malformations in a KS patient should direct the geneticist towards a search for mutations in FGFR1.

Form of dual-specificity tyrosine-(Y)-phosphorylation-regulated kinase 1A nonphosphorylated at tyrosine 145 and 147 is enriched in the nuclei of astroglial cells, adult hippocampal progenitors, and some cholinergic axon terminals.

Compelling lines of evidence indicate that overexpression of dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A) in subjects with trisomy 21 (Down syndrome[DS]) contributes to the abnormal structure and function of the DS brain. In the present study, we used a novel, phospho-dependent antibody recognizing DYRK1A only with nonphosphorylated tyrosine 145 and 147 (DYRK1A Tyr-145/147P(-)), to investigate the expression pattern of this DYRK1A species in trisomic and disomic human and mouse brains. Immunoblotting and dephosphorylation experiments demonstrated higher levels of DYRK1A Tyr-145/147P(-) in postnatal trisomic brains in comparison with controls (by ∼40%) than those of the DYRK1A visualized by three other N- and C-terminally directed antibodies to DYRK1A. By immunofluorescence, the immunoreactivity to DYRK1A Tyr-145/147P(-) was the strongest in the nuclei of astroglial cells, which contrasted with the predominantly neuronal localization of DYRK1A visualized by the three other antibodies to DYRK1A we used. In addition, DYRK1A Tyr-145/147P(-) was enriched in the nuclei of neuronal progenitors and newly born neurons in the adult hippocampal proliferative zone and also occurred in some cholinergic axonal terminals. Our data show a distinctive expression pattern of DYRK1A forms nonphosphorylated at Tyr-145 and Tyr-147 in the brain tissue and suggest that DS subjects may exhibit not only upregulation of total DYRK1A, but also more subtle differences in phosphorylation levels of this kinase in comparison with control individuals.

Estrogen-dependent regulation of PPAR-gamma signaling on collagen biosynthesis in adenocarcinoma endometrial cells.

The link between estrogen and metabolic developmental factors of endometrial carcinoma is well established. PPAR- gamma, (an important modulator of metabolism) and estrogen receptor belong to a family of nuclear hormone receptors that were shown to interact with each other. The interaction may affect transcriptional activity of these transcription factors. The anti-diabetic troglitazone (TGZ) is well known PPAR- gamma ligand. The effect of troglitazone-induced PPAR- gamma activation on estrogen-dependent stimulation of collagen biosynthesis was studied in the Ishikawa endometrial adenocarcinoma cell line. We have found that the presence of estrogen activity in growth medium (1nM) augmented collagen biosynthesis in the cells. An addition of PPAR- gamma agonists, as troglitazone or clofibrat to the growth medium induced inhibition of collagen biosynthesis. The inhibition was effective only when estrogen receptor was stimulated, since removal of estrogen receptor by ICI 182- 780-dependent degradation did not affect collagen biosynthesis. The mechanism of the inhibition was found at the level of NF-kB (known inhibitor of collagen gene expression) and MAPK signaling. PPAR- gamma ligands stimulated expression of NF-kB, while they inhibited expression of p-38 but not ERK1/ERK2. The data document for the first time that inhibitory effect of PPAR- gamma ligands on collagen biosynthesis in endometrial adenocarcinoma cells requires functional estrogen receptor.

Hyaluronic acid counteracts interleukin-1-induced inhibition of collagen biosynthesis in cultured human chondrocytes.

Although, hyaluronic acid (HA) is used in the treatment of osteoarthritis for 30 years, the mechanism of its protective action on collagen metabolism disturbances in tissues during inflammation is not known. Therefore, the present study was undertaken to evaluate the mechanism of IL-1beta action (inductor of experimental inflammation) on deregulation of collagen biosynthesis in cultured human chondrocytes and the effect of HA on the process. It has been found that IL-1beta strongly induced inhibition of collagen biosynthesis, while HA counteracted the process. The mechanism of this phenomenon was found at both transcriptional and post-transcriptional level. IL-1 was found to down regulate the expression of mRNA for type II collagen and to inhibit prolidase activity, an enzyme that plays an important role in collagen biosynthesis at post-translational level. HA was shown to counteract the IL-1beta-dependent inhibition of both processes. During experimental inflammation of chondrocytes cultured in 0.1% FBS there was no differences in the expression of beta(1)-integrin independently of cell number and the presence of HA in growth medium. In chondrocytes cultured in 5% FBS, IL-1beta up-regulated the expression of beta(1)-integrin receptor while HA abolished the effect. The data suggest that HA-dependent up-regulation of collagen biosynthesis in IL-1beta-treated chondrocytes may involve stimulation of prolidase activity in serum "starved" cells and may also originate at the transcriptional level in the cells cultured in standard conditions.

Antimitotic activity of high affinity ligands for peripheral benzodiazepine receptor (PBR) in some normal and neoplastic cell lines.

PURPOSE: Overexpression of PBR has been found in several tumor types including ovarian, colon, breast adenocarcinomas, esophageal cancer. There is evidence suggesting that PBR ligands regulate cell proliferation. However, their action is probably cell-type specific. We decided to evaluate mitotic activity of PBR ligands in some normal and neoplastic cell lines. MATERIAL AND METHODS: The cells were maintained according to standard procedures. Ligand binding assay was performed in cell extract using PK-11195 or Ro-54864 and [N-methyl-3H] Ro-54864 or [N-methyl-3H] PK-11195. Cell proliferation was evaluated using 5-[3H]-thymidine assay. Western Immunoblot assay was conducted using polyclonal anti-PBR antibody. RESULTS: We have found that, macrophages evoked strong binding of both Ro-54864 and PK-11195. This phenomenon was accompanied by drastic decrease in the cell divisions. Similar effect was found only in the case of non-estrogen-dependent breast cancer cells MDA-MB 231. It suggest that PBR-ligand mediated inhibition of mitogenesis may represent a new anticancer strategy in non-estrogen-dependent breast cancer. In respect to macrophages inhibition of the cell division by both PBR ligands may have implication in modulation of inflammatory response. It has been postulated that PBR ligands may have anti-inflammatory activity in rheumatoid arthritis. The presence of peripheral benzodiazepine receptors in chondrocytes, T cells, macrophages and mesenchymal cells suggest that peripheral benzodiazepine receptor ligands may interfere with the cytokine network and thus modulate inflammatory response. CONCLUSIONS: The data suggest that PBR-ligand mediated inhibition of DNA synthesis in non-estrogen dependent breast cancer cells and in macrophages may represent a new therapeutic approach of breast anti-cancer and anti-inflammatory therapy.

Protein kinase C involvement in proliferation and survival of breast cancer cells.

Members of protein kinase C (PKC) family have been widely implicated in the regulation of cell proliferation, differentiation and survival. Increased protein C activity in malignant breast tissue and in most aggressive breast cancer cell lines suggests possible role of PKC in the development and progression of breast cancer. PKC may be therefore a target for breast cancer treatment. In our study we attempted to investigate the effect of: phorbol ester (PMA)-PKC activator, and bisindolylmaleimide II (GF II), a highly selective PKC inhibitor, on the proliferation as well as induction of apoptosis and necrosis in breast cancer cell line MDA-MB-231. Our results provide evidence for multidirectional effects of PKC on the proliferation of this type of breast cancer cells. The effects of both compounds were different after short time of exposition (1-3 h). PMA induced proliferation, while GF II showed an opposite effect. After 24 h, however, both compounds exhibited relatively high inhibitory effect on the proliferation and proved to be effective in induction of necrosis and apoptosis.