Comparison of PLA-Based Micelles and Microspheres as Carriers of Epothilone B and Rapamycin. The Effect of Delivery System and Polymer Composition on Drug Release and Cytotoxicity against MDA-MB-231 Breast Cancer Cells.

Co-delivery of epothilone B (EpoB) and rapamycin (Rap) increases cytotoxicity against various kinds of cancers. However, the current challenge is to develop a drug delivery system (DDS) for the simultaneous delivery and release of these two drugs. Additionally, it is important to understand the release mechanism, as well as the factors that affect drug release, in order to tailor this process. The aim of this study was to analyze PLA-PEG micelles along with several types of microspheres obtained from PLA or a mixture of PLA and PLA-PEG as carriers of EpoB and Rap for their drug release properties and cytotoxicity against breast cancer cells. The study showed that the release process of EpoB and Rap from a PLA-based injectable delivery systems depends on the type of DDS, morphology, and polymeric composition (PLA to PLA-PEG ratio). These factors also affect the biological activity of the DDS, because the cytotoxic effect of the drugs against MDA-MB-231 cells depends on the release rate. The release process from all kinds of DDS was well-characterized by the Peppas-Sahlin model and was mainly controlled by Fickian diffusion. The conducted analysis allowed also for the selection of PLA 50/PLA-PEG 50 microspheres and PLA-PEG micelles as a promising co-delivery system of EpoB and Rap.

Lapatinib enhances paclitaxel toxicity in MCF-7, T47D, and MDA-MB-321 breast cancer cells.

Paclitaxel (PTX) is used to treat breast cancer both as a monotherapy and in combination with other anticancer drugs. Chemoresistance is one of the main reasons for the failure of breast cancer treatment. Mechanisms which contribute to multidrug resistance of breast cancer cells to PTX include the active removal of the drug from the cell related to the increased activity of ABC family membrane transporters. Lapatinib (LAP) has been approved by the FDA in combination with other anticancer agents for the treatment of HER2-positive breast cancer. LAP can reverse chemoresistance by interaction with ABC transporters. Therefore the aim of the study was to investigate whether LAP is able to potentiate PTX toxicity in MCF-7, T47D, and MDA-MB-321 breast cancer cells which do not express the HER-2. It was found that LAP inhibited the PTX efflux, increased its intracellular concentration and thus significantly increased the anticancer activity of PTX. The combination of PTX and LAP can be useful in HER2-negative breast cancer treatment.

Cytotoxic effect of targeted biodegradable epothilone B and rapamycin co-loaded nanocarriers on breast cancer cells.

The new therapeutic solutions for breast cancer treatment are needed, for example, combined therapy consisted of several drugs that characterize different mechanisms of action and modern drug delivery systems. Therefore, we used combination of epothilone B (EpoB) and rapamycin (Rap) to analyze the cytotoxic effect against breast cancer cells (MCF-7; MDA-MB-231). Also, the effect of drugs co-delivered in bioresorbable micelles functionalized with biotin (PLA-PEG-BIO; poly(lactide)-co-poly(ethylene glycol)-biotin) was studied. The comparison of effects of the mixture of free drugs and the micelles co-loaded with EpoB and Rap revealed a significant decrease in the cell metabolic activity and survival. Moreover, the dual drug-loaded PLA-PEG-BIO micelles enhanced the cytotoxicity of EpoB and Rap against the tested cells as compared with the free drugs. The blank PLA-PEG-BIO micelles did not affect the tested cells. We expect that mixture of EpoB and Rap may be promising in breast cancer treatment and PLA-PEG-BIO micelles as carrier of these two drugs can be applicable for successful targeted delivery.

Cytotoxic Effect of Paclitaxel and Lapatinib Co-Delivered in Polylactide-co-Poly(ethylene glycol) Micelles on HER-2-Negative Breast Cancer Cells.

To find better strategies to enhance the cytotoxic effect of paclitaxel (PTX) and lapatinib (LAP) against breast cancer cells, we analyzed the efficacy of a novel delivery system containing polylactide-co-poly(ethylene glycol) (PLA-PEG) filomicelles of over 100 nm in length and spherical micelles of approximately 20 nm in diameter. The ¹H NMR measurements confirmed the incorporation of PTX and LAP into micelles. Analysis of the drug release mechanism revealed the diffusion-controlled release of LAP and anomalous transport of PTX. Drug content analysis in lyophilized micelles and micellar solution showed their good storage stability for at least 6 weeks. Blank micelles, LAP-loaded micelles and free LAP did not affect MCF-7 breast cancer cell proliferation, suggesting that the cytotoxicity of PTX-, PTX/LAP-loaded micelles, and the binary mixture of free PTX and LAP was solely caused by PTX. PTX/LAP-loaded micelles showed greater toxicity compared to the binary mixture of PTX and LAP after 48 h and 72 h. Only free PTX alone induced P-gp activity. This study showed the feasibility of using a LAP and PTX combination to overcome MDR in MCF-7 cells, particularly when co-loaded into micelles. We suggest that PTX/LAP micelles can be applicable not only for the therapy of HER-2-positive, but also HER-2-negative breast cancers.

DHA but not AA Enhances Cisplatin Cytotoxicity in Ovarian Cancer Cells.

Clinical and epidemiological studies show that docosahexaenoic acid (DHA) and arachidonic acid (AA) exert multiple effects on ovarian cancer. While DHA seems to inhibit growth and prevent carcinogenic processes, stimulation of leukotriene B4 receptors BLT1 and BLT2 by several eicosanoids derived from AA plays an important role in mediating cisplatin resistance in ovarian cancer cells. We examined whether DHA and AA exerted antiproliferative effect on epithelial ovarian cancer cells and whether these polyunsaturated fatty acids could alter their susceptibility to cisplatin. Using SKOV3 and OVCAR3 cell lines, we found that DHA but not AA suppressed the cells viability, proliferation, enhanced cell death, and induced activation of caspase-3/7 in the concentration- and time-dependent manner. The OVCAR3 cells were less susceptible to cisplatin than SKOV3 cells. DHA but not AA significantly potentiated cisplatin cytotoxicity in SKOV3 and OVCAR3 cells. We did not observe any significant influence of AA on the above mentioned processes in both cell lines. Similar effect can occur in ovarian cancer patients treated with cisplatin and supplemented with DHA.

Osteogenic differentiation of human mesenchymal stem cells from adipose tissue and Wharton's jelly of the umbilical cord.

Induced osteogenesis of mesenchymal stem cells (MSCs) may provide an important tool for bone injuries treatment. Human umbilical cord and adipose tissue are routinely discarded as clinical waste and may be used as noncontroversial MSCs sources. It still remains to be verified which source of MSCs is the most suitable for bone regeneration. The aim of this research was to investigate the osteogenic potential of human MSCs derived from adipose tissue (AT-MSCs) and Wharton's jelly of the human umbilical cord (WJ-MSCs) differentiated under the same conditions. Osteogenic differentiation of MSCs was detected and quantified by alizarin red S (ARS) staining for calcium deposition and alkaline phosphatase (ALP) activity, osteoprotegerin (OPG), and osteocalcin (OC) secretion measurements. Under osteogenic conditions, after 21 days of differentiation, the measured ALP activity and calcium deposition were significantly higher in the AT-MSCs than in the WJ-MSCs, while the OPG and OC secretion were higher in the WJ-MSCs vs. AT-MSCs. Low concentrations of OPG and high levels of OC in AT-MSCs and WJ-MSCs, prove that these cells reached an advanced stage of the osteogenic differentiation. The levels of OC secreted by AT-MSCs were lower than by WJ-MSCs. Both cell types, AT-MSCs and WJ-MSCs possess a potential to differentiate towards the osteogenic lineage. The observed differences in the levels of osteogenic markers suggest that after 21-days of osteogenic differentiation, the AT-MSCs might have reached a more advanced stage of differentiation than WJ-MSCs.

CYCLOSPORIN A AFFECTS THE PROLIFERATION PROCESS IN NORMAL HUMAN DERMAL FIBROBLASTS.

Cyclosporin A is an immunosuppressant drug that is used not only in solid transplant rejection, but also in moderate and severe forms of psoriasis, pyoderma, lupus or arthritis. Serious side effects of the drug such as skin cancer or gingival hyperplasia probably start with the latent proliferation process. Little is known about the influence of cyclosporin A on molecular signaling in epidermal tissue. Thus, the aim of this study was to estimate the influence of cyclosporin A on the process of proliferation in normal human dermal fibroblasts. Fibroblasts were cultured in a liquid growth medium in standard conditions. Cyclosporin A was added to the culture after the confluence state. Survival and proliferation tests on human dermal fibroblast cells were performed. Total RNA was extracted from fibroblasts, based on which cDNA and cRNA were synthesized. The obtained cRNA was hybridized with the expression microarray HGU-133A_2.0. Statistical analysis of 2734 mRNAs was performed by the use of GeneSpring 13.0 software and only results with p < 0.05 were accepted. Analysis of variance with Tukey post hoc test with Benjamini-Hochberg correction for all three (8, 24, 48 h) culture stages (with and without cyclosporin A) was performed to lower the number of statistically significant results from 679 to 66, and less. Between statistically and biologically significant mRNAs down-regulated were EGRJ, BUBIB, MKI67, CDK1, TTK, E2F8, TPX2, however, the INSIG1, FOSL1, HMOX1 were up-regulated. The experiment data revealed that cyclosporin A up-regulated FOSL1 in the first 24 h, afterwards down-regulating its expression. The HMOX1 gene was up-regulated in the first stage of the experiment (CsA 8 h), however, after the next 16 h of culture time its expression was down-regulated (CsA 24 h), to finally increased in the later time period. The results indicate that cyclosporin A had a significant effect on proliferation in normal human dermal fibroblasts through the changes in the expression of genes related to the cell cycle and transcription regulation process.

In vitro chondrogenesis of Wharton's jelly mesenchymal stem cells in hyaluronic acid-based hydrogels.

BACKGROUND: In this study, we evaluated the usefulness of two commercially available hyaluronic acid-based hydrogels, HyStem and HyStem-C, for the cultivation of Wharton's jelly mesenchymal stem cells (WJ-MSCs) and their differentiation towards chondrocytes. METHODS: The WJ-MSCs were isolated from umbilical cord Wharton's jelly using the explant method and their immunophenotype was evaluated via flow cytometry analysis. According to the criteria established by the International Society for Cellular Therapy, they were true MSCs. We assessed the ability of the WJ-MSCs and chondrocytes to grow in three-dimensional hydrogels and their metabolic activity. Chondrogenesis of WJ-MSCs in the hydrogels was determined using alcian blue and safranin O staining and real-time PCR evaluation of gene expression in the extracellular matrixes: collagen type I, II, III and aggrecan. RESULTS: Chondrocytes and WJ-MSCs cultured in the HyStem and HyStem-C hydrogels adopted spherical shapes, which are characteristic for encapsulated cells. The average viability of the WJ-MSCs and chondrocytes in the HyStem hydrogels was approximately 67 % when compared with the viability in 2D culture. Alcian blue and safranin O staining revealed intensive production of proteoglycans by the cells in the HyStem hydrogels. Increased expression of collagen type II and aggrecan in the WJ-MSCs cultured in the HyStem hydrogel in the presence of chondrogenic medium showed that under these conditions, the cells have a high capacity to differentiate towards chondrocytes. The relatively high viability of WJ-MSCs and chondrocytes in both HyStem hydrogels suggests the possibility of their use for chondrogenesis. CONLUSIONS: The results indicate that WJ-MSCs have some degree of chondrogenic potential in HyStem and HyStem-C hydrogels, showing promise for the engineering of damaged articular cartilage.

Changes in expression of cartilaginous genes during chondrogenesis of Wharton's jelly mesenchymal stem cells on three-dimensional biodegradable poly(L-lactide-co-glycolide) scaffolds.

BACKGROUND: In cartilage tissue regeneration, it is important to develop biodegradable scaffolds that provide a structural and logistic template for three-dimensional cultures of chondrocytes. In this study, we evaluated changes in expression of cartilaginous genes during in vitro chondrogenic differentiation of WJ-MSCs on PLGA scaffolds. METHODS: The biocompatibility of the PLGA material was investigated using WJ-MSCs by direct and indirect contact methods according to the ISO 10993-5 standard. PLGA scaffolds were fabricated by the solvent casting/salt-leaching technique. We analyzed expression of chondrogenic genes of WJ-MSCs after a 21-day culture. RESULTS: The results showed the biocompatibility of PLGA and confirmed the usefulness of PLGA as material for fabrication of 3D scaffolds that can be applied for WJ-MSC culture. The in vitro penetration and colonization of the scaffolds by WJ-MSCs were assessed by confocal microscopy. The increase in cell number demonstrated that scaffolds made of PLGA copolymers enabled WJ-MSC proliferation. The obtained data showed that as a result of chondrogenesis of WJ-MSCs on the PLGA scaffold the expression of the key markers collagen type II and aggrecan was increased. CONCLUSIONS: The observed changes in transcriptional activity of cartilaginous genes suggest that the PLGA scaffolds may be applied for WJ-MSC differentiation. This primary study suggests that chondrogenic capacity of WJ-MSCs cultured on the PLGA scaffolds can be useful for cell therapy of cartilage.

Toxic effects of n-3 polyunsaturated fatty acids in human lung A549 cells.

N-3 polyunsaturated fatty acids (PUFAs), particularly eicosapentaenoic acid (EPA, 20:5) and docosahexaenoic acid (DHA, 22:6) are crucial for the prevention of lung cancer. PUFAs may act through alteration of membrane fluidity and cell surface receptor functions; modulation of cyclooxygenase activity; and increased cellular oxidative stress, which may induce apoptosis and autophagy. Therefore the aim of the study was to investigate whether EPA and DHA (25-100 µM) are able to reduce human lung cancer cell growth through oxidative stress influence on autophagy and apoptosis. It was found that both EPA and DHA in the concentration-dependent manner suppressed the cell viability, enhanced cell death, induced activation of caspase-3/7 and potentiated intracellular oxidative DNA and protein damage. In response to PUFAs intracellular autophagic vacuolization occurred and the observed effect was reverted when the autophagy inhibitor 3-methyladenine (3-MA) was applied. The inhibition of the autophagic process enhanced the cell viability, suppressed cell death, and decreased activation of caspase-3/7 indicating that EPA and DHA-induced autophagy amplified A549 apoptotic cell death.

Influence of cyclosporin A on expression pattern of genes associated with dna repair in human dermal fibroblasts.

Cyclosporin A (CsA) is a cyclic nonribosomal peptide with immunosuppressive activity. Chronic immunosuppressive medication is associated with time distant side effects and is the cause of the different secondary diseases, including cancers (especially skin cancers). Anomalies in the functioning of DNA repair mechanisms are closely related to the processes of neoplastic transformation. The object of this study was to assess the impact of CsA exposure (8 h, early cell response) on expression of genes associated with DNA repair in normal human dermal fibroblasts (NHDF). NHDF from CC-2511 cell line were routinely maintained in FBM medium. Transcriptional activity of genes associated with DNA repair in NHDF after 8 h of cells exposition to CsA (C = 100 ng/mL) in relation to control cells was compared using Affymetrix HG-U133A 2.0 oligonucleotide microarray technique. GeneSpring GX fluorescence signals analysis of 1514 probes, which represented the expression of 875 genes selected from the NetAffx Analysis Center database for "DNA repair" query, demonstrated the inhibited expression of 32 probes (p-value < 0.05; Fold Change > 2.0), including: BRCA1, RAD51, TOP2A, EXO1, RRM2, CDK1 and POLE2. The obtained results suggest that CsA can have a silencing effect on DNA repair genes. Therefore, the risk of skin cancer development during CsA therapy can result not only from immunosuppressive effects of the drug, but is also likely to arise from inhibition of DNA repair pathways.

Polyunsaturated fatty acids potentiate cytotoxicity of cisplatin in A549 cells.

In normal and tumor cells, polyunsaturated fatty acids (PUFAs) act as intracellular second messengers, which play a role in signaling, proliferation and cell death. PUFAs have selective tumoricidal action and may alter sensitivity of tumor cells to cisplatin (CDDP), a commonly used anticancer agent. The aim of this study was to evaluate the influence of arachidonic acid (AA, 20:4 n-6), eicosapentaenoic acid (EPA, 20:5 n-3), docosahexaenoic acid (DHA, 22:6 n-3) and CDDP on autophagy and apoptosis in A549 human lung adenocarcinoma cells. Viability of A549 cells treated with CDDP and PUFAs was measured using the XTT tetrazolium salt based assay. Caspase-3/7 activity was estimated using ApoTox-Glo kit (Promega). Autophagic vac- uoles were detected by Cyto-ID Autophagy Detection Kit (Enzo). The results were compared to control cultures maintained in the absence of CDDP and PUFAs. PUFAs, in particular EPA and DHA, added to the cultivation medium, increased the antitumor activity of CDDP in A549 cells in a concentration dependent manner. In case of AA this effect was observed at the highest of the concentrations tested only (100 µM). Both, EPA and DHA, but not AA, significantly increased the amount of autophagic vacuoles and induced caspase-3/7 activity. The obtained results suggest that the antiproliferative effect of CDDP in A549 cells can be enhanced by AA and in particular by EPA and DHA through their influence on autophagic and apoptotic cell death. It is likely that BPA and DHA incorporated to the tumor cells may improve outcomes in lung cancer patients.

Phytic acid inhibits lipid peroxidation in vitro.

Phytic acid (PA) has been recognized as a potent antioxidant and inhibitor of iron-catalyzed hydroxyl radical formation under in vitro and in vivo conditions. Therefore, the aim of the present study was to investigate, with the use of HPLC/MS/MS, whether PA is capable of inhibiting linoleic acid autoxidation and Fe(II)/ascorbate-induced peroxidation, as well as Fe(II)/ascorbate-induced lipid peroxidation in human colonic epithelial cells. PA at 100 µM and 500 µM effectively inhibited the decay of linoleic acid, both in the absence and presence of Fe(II)/ascorbate. The observed inhibitory effect of PA on Fe(II)/ascorbate-induced lipid peroxidation was lower (10-20%) compared to that of autoxidation. PA did not change linoleic acid hydroperoxides concentration levels after 24 hours of Fe(II)/ascorbate-induced peroxidation. In the absence of Fe(II)/ascorbate, PA at 100 µM and 500 µM significantly suppressed decomposition of linoleic acid hydroperoxides. Moreover, PA at the tested nontoxic concentrations (100 µM and 500 µM) significantly decreased 4-hydroxyalkenal levels in Caco-2 cells which structurally and functionally resemble the small intestinal epithelium. It is concluded that PA inhibits linoleic acid oxidation and reduces the formation of 4-hydroxyalkenals. Acting as an antioxidant it may help to prevent intestinal diseases induced by oxygen radicals and lipid peroxidation products.

Polyunsaturated fatty acids inhibit melanoma cell growth in vitro.

Human malignant melanoma is a highly aggressive and incurable cancer due to intrinsic resistance to apoptosis and reprogramming proliferation and survival pathways during progression. Numerous studies, including our own, linked arachidonic acid (AA, 20:4 n-6), eicosapentaenoic acid (EPA, 20:5 n-3), and docosahexaenoic acid (DHA, 22:6 n-3) supplementation to induction of apoptosis and decreased proliferation of various cancer cells. The cytotoxic effects result from lipid peroxidation and formation of reactive oxygen species (ROS), which modify proteins and nucleic acids. DNA damage by ROS causes mutations and genomic instability, leading to uncontrolled proliferation or cell death. In the present work, four human melanoma cell lines differing in origin, doubling time, metastatic potential, and melanin content (A375, A2058, G361, and C32) were exposed to AA, EPA or DHA added into culture media in the concentrations ranging from 0 (control) to 100 mM. After 24 h incubation cytotoxicity of the analyzed acids was determined with TOX-2 (In Vitro Toxicology Assay Kit XTT Based, TOX-2, Sigma) test. The oxidative protein modifications were measured using Aldehyde Site (DNA and Protein) Detection Kit (Cayman). All the acids tested showed marked inhibition of cell proliferation. The observed effects were statistically significant and depended on the concentration. Decrease of proliferation, associated by oxidative protein and DNA damage (measured as aldehyde sites in cells), was observed for EPA and DHA (50 mM and 100 mM) in A375, A2058, and G361 cells. In case of C32 cell line, which is amelanotic melanoma, EPA and DHA inhibited cell proliferation at 100 mM only. The effect of DHA was more pronounced. AA did not show its antiproliferative action in this cell line. The obtained results suggest that antiproliferative effects of the fatty acids in cultured human melanoma cells depend on the type of acid, its concentration and may be diverse when different melanoma cell lines are used.

Effects of 300 mT static magnetic field on IL-6 secretion in normal human colon myofibroblasts.

Intestinal subepithelial myofibroblasts play crucial role in the growth and development of the intestine. Colitis, small bowel injury, gastric ulcer disease and inflammatory bowel disease (IBD) accompany the increase of number of activated myofibroblasts. In the last few years, the increasing production of electromagnetic (EMF) and static magnetic fields (SMF), due to the expanding use of electronic devices in everyday life, has led to a number of studies on the effects of these fields on living organisms. EMF therapy, because of its anti-inflammatory properties, may be used in medicine in IBD treatment. This mechanism has not been elucidated yet. In the present work normal human colon myofibroblasts CCD-18Co were exposed to SMF with a flux density of 300 mT. After 24 h incubation TNF-alpha-dependent IL-6 secretion was determined with ELISA kit (RandD Systems).The influence of magnetic field and its effect on cell proliferation were determined with TOX-2 (In Vitro Toxicology Assay Kit XTT Based, TOX-2, Sigma) and CyQUANT NF cell proliferation assay kit (Molecular Probes). It was shown that SMF inhibited TNF-alpha-dependent IL-6 secretion. The observed effects were statistically significant and depended on the time of incubation. Moreover, SMF triggered cell proliferation whereas it did not alter cell viability. IL-6 belongs to pro-inflammatory cytokines family and plays a crucial role in IBD. Inhibition of IL-6 secretion by SMF and lack of its cytotoxic effect seem to be advantageous whilst SMF is implicated in the treatment of inflammatory diseases associated by increase in number of activated myofibroblasts.

Synthesis, structure and antitumor activity of [RhCl3(N-N)(DMSO)] polypyridyl complexes.

The [RhCl(3)(N-N)(DMSO)] complexes, the N-N being 2,2'-bipyridine (1), 1,10-phenanthroline (2), 4,7-diphenyl-1,10-phenanthroline (3), 4,4'-dimethyl-2,2'-bipyridine (4) and 1,10-phenanthroline-5,6-dione (5), have been synthesized and characterized with spectroscopic methods. The compounds 2-5 adopt mer- and complex 1 fac-structure. The molecular and electronic structure studies of mer- and fac-complexes with bpy and phen ligands at the DFT B3LYP level with 3-21G( * *) basis set showed that mer-isomers are more stable. The cytostatic activity of the [RhCl(3)(N-N)(DMSO)] complexes against Caco-2 and A549 tumor cells have been studied. Their antibacterial activity have also been investigated. It has been found that the very promising biological activity show complexes 2, 3 and 4.

Preliminary study of the expression of genes connected with the orexigenic and anorexigenic system using microarray technique in anorexia nervosa.

The pathogenesis of anorexia nervosa (AN) is still poorly understood. The Diagnostic and Statistical Manual of Mental Disorders (4th edition) classification differentiates 2 AN types: the restricting type (AN-R) and the binge eating/purging type (AN-BP). We investigated 4 young women suffering from AN (2 with AN-R and 2 with AN-BP). Four women, age matched, with other psychiatric disorders (paranoid schizophrenia, adjustment disorder, mental retardation) served as the reference group. The oligonucleotide microarray method (HG-U133A, Affymetrix) was used to determine the expression profile of 13 genes connected with the orexigenic and anorexigenic system: leptin, leptin receptor-coding gene, hypocretin (orexin) receptor-coding gene, hypocretin (orexin) neuropeptide precursor-coding gene and growth hormone secretagogue receptor. A hierarchical analysis of the results showed that AN-BP and AN-R patients were grouped into different clusters. Also, expression levels of leptin receptor-coding gene showed significant differences between AN-BP and AN-R patients and between AN-R and control subjects. This preliminary study suggests that the microarray technique may contribute to elucidating molecular genetics of the pathogenesis of both types of AN.

Aldehydic lipid peroxidation products in human brain astrocytomas.

Among mediators of oxidative stress, highly reactive secondary aldehydic lipid peroxidation products can initiate the processes of spontaneous mutagenesis and carcinogenesis and can also act as a growth-regulating factors and signaling molecules. We explored whether these aldehydes and histone H3 mRNA levels could serve as biomarkers of malignancy and predictive factor in human brain astrocytomas. Histone H3 mRNA, a biomarker of cellular proliferation, was analyzed by QRT-PCR (TaqMan). Aldehydic lipid peroxidation products were determined as their dinitrophenylhydrazone derivatives in specimens obtained from 26 adult patients with brain astrocytomas. RP-HPLC with diode array detector and MSMS spectrometer were used for the analysis. H3 mRNA, 2-hydroxyhexanal, and 4-hydroxynonenal levels were higher in high-grade astrocytomas compared to low-grade astrocytomas and showed negative correlation with survival. Higher levels of 2-hydroxyhexanal and 4-hydroxynonenal, and lower levels of n-hexanal were associated with poorer patient prognosis. Our data suggest that tissue concentrations of aldehydic lipid peroxidation products can assist grading and predicting the clinical outcome in patients with astrocytic brain tumors. Possibly, this parameter will enhance optimal selection of patients for individualized treatment protocols, tailored to unique biochemical and molecular profile of the tumor.