Transporter Proteins as Therapeutic Drug Targets-With a Focus on SGLT2 Inhibitors.

Membrane transporters interact not only with endogenous substrates but are also engaged in the transport of xenobiotics, including drugs. While the coordinated function of uptake (solute carrier family-SLC and SLCO) and efflux (ATP-binding cassette family-ABC, multidrug and toxic compound extrusion family-MATE) transporter system allows vectorial drug transport, efflux carriers alone achieve barrier functions. The modulation of transport functions was proved to be effective in the treatment strategies of various pathological states. Sodium-glucose cotransporter-2 (SGLT2) inhibitors are the drugs most widely applied in clinical practice, especially in the treatment of diabetes mellitus and heart failure. Sodium taurocholate co-transporting polypeptide (NTCP) serves as virus particles (HBV/HDV) carrier, and inhibition of its function is applied in the treatment of hepatitis B and hepatitis D by myrcludex B. Inherited cholestatic diseases, such as Alagille syndrome (ALGS) and progressive familial intrahepatic cholestasis (PFIC) can be treated by odevixibat and maralixibat, which inhibit activity of apical sodium-dependent bile salt transporter (ASBT). Probenecid can be considered to increase uric acid excretion in the urine mainly via the inhibition of urate transporter 1 (URAT1), and due to pharmacokinetic interactions involving organic anion transporters 1 and 3 (OAT1 and OAT3), it modifies renal excretion of penicillins or ciprofloxacin as well as nephrotoxicity of cidofovir. This review discusses clinically approved drugs that affect membrane/drug transporter function.

Solute Carrier Transporters in Synovial Membrane and Hoffa's Pad of Patients with Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a complex autoimmune disease that leads to joint destruction. A number of immune cells that affect joint tissues are involved in the pathogenesis of this disease. This leads to the synthesis of many pro-inflammatory mediators. The transport of drugs, as well as many cytokines involved in the development of inflammation in RA patients, is mediated by membrane transporters. Membrane transporters are proteins that mediate the transfer of substrates across biological membranes. But to date there are no studies examining the expression of solute carrier (SLC) transporters in joint tissues. The aim of the study was to evaluate the expression of individual SLC family transporters in the synovial membranes (SMs) and infrapatellar fat pad (Hoffa's pad) of RA patients. The study included 20 patients with rheumatoid arthritis and 20 with osteoarthritis as the control group who were undergoing joint replacement surgery as a normal part of clinical care. In the SM and Hoffa's pad of RA patients the following 17 membrane transporters were defined at relevant expression levels for SLC transporter superfamily: SLC15A2, SLC16A3, SLC19A1, SLC2A9, SLC22A1, SLC22A3, SLC22A4, SLC22A5, SLC22A18, SLC33A1, SLC47A1, SLC51A, SLC7A5, SLC7A6, SLC01C1, SLC02B1, SLC04A1. The confirmed expression of these transporters in the SMs as well as Hoffa's pad of patients with RA and OA, and the differences in their expression between these groups, suggests the involvement of SLC transporters in both the maintenance of homeostasis under physiological conditions in the tissues of the joints, as well as in the inflammatory process in RA.

Microtubule destabilising activity of selected 7-methoxy-2-phenylbenzo[b]furan derivative against primary and metastatic melanoma cells.

Herein, we report the results of anticancer screening of two 2-phenylbenzo[b]furan derivatives functionalised at the 3-position with 4-hydroxy-3,5-dimethoxybenzoyl (BF2) or 3,4,5-trimethoxybenzoyl (BF3) against 60 different cancer cell lines. The results confirmed the anticancer potential of the tested compounds against different cancer cell types, especially colon cancer, brain cancer and melanoma. BF3 was defined as the most potent (also as a tubulin polymerisation inhibitor). Its anticancer activity against melanoma cell lines that originated from different stages, i.e., primary skin-derived A375 and metastatic WM9/MDA-MB-435S, was evaluated (as the clinical success of melanoma therapy strictly depends on the disease stage). Moreover, to determine the BF3 mode of action and its effect on cell proliferation, intracellular microtubule networks, cell cycle phase distribution and apoptosis were evaluated. Our study revealed that BF3 inhibited cell proliferation in a dose-dependent manner, with IC50 yielding 0.09 ± 0.01 µM, 0.11 ± 0.01 µM and 0.18 ± 0.05 µM for A375, MDA-MB435S and WM9, respectively. The strong antiproliferative activity of compound BF3 correlated well with its inhibitory effect on tubulin polymerisation. Molecular docking proved that BF3 belongs to the colchicine binding site inhibitors (CBSIs), and experimental studies revealed that it disturbs cell cycle progression leading to G2/M arrest and apoptosis.

Anticancer properties of bacterial cellulose membrane containing ethanolic extract of Epilobium angustifolium L.

Epilobium angustifolium L. is a medicinal plant well known for its anti-inflammatory, antibacterial, antioxidant, and anticancer properties related to its high polyphenols content. In the present study, we evaluated the antiproliferative properties of ethanolic extract of E. angustifolium (EAE) against normal human fibroblasts (HDF) and selected cancer cell lines, including melanoma (A375), breast (MCF7), colon (HT-29), lung (A549) and liver (HepG2). Next, bacterial cellulose (BC) membranes were applied as a matrix for the controlled delivery of the plant extract (BC-EAE) and characterized by thermogravimetry (TG), infrared spectroscopy (FTIR), and scanning electron microscopy (SEM) images. In addition, EAE loading and kinetic release were defined. Finally, the anticancer activity of BC-EAE was evaluated against the HT-29 cell line, which presented the highest sensitivity to the tested plant extract (IC50 = 61.73 ± 6.42 µM). Our study confirmed the biocompatibility of empty BC and the dose and time-dependent cytotoxicity of the released EAE. The plant extract released from BC-2.5%EAE significantly reduced cell viability to 18.16% and 6.15% of the control values and increased number apoptotic/dead cells up to 37.53% and 66.90% after 48 and 72 h of treatment, respectively. In conclusion, our study has shown that BC membranes could be used as a carrier for the delivery of higher doses of anticancer compounds released in a sustained manner in the target tissue.

Kidney Drug Transporters in Pharmacotherapy.

The kidney functions not only as a metabolite elimination organ but also plays an important role in pharmacotherapy. The kidney tubule epithelia cells express membrane carriers and transporters, which play an important role in drug elimination, and can determine drug nephrotoxicity and drug-drug interactions, as well as constituting direct drug targets. The above aspects of kidney transport proteins are discussed in the review.

Detection of key mRNAs in liver tissue of hepatocellular carcinoma patients based on machine learning and bioinformatics analysis.

One methodology extensively used to develop biomarkers is the precise detection of highly responsive genes that can distinguish cancer samples from healthy samples. The purpose of this study was to screen for potential hepatocellular carcinoma (HCC) biomarkers based on non-fusion integrative multi-platform meta-analysis method. The gene expression profiles of liver tissue samples from two microarray platforms were initially analyzed using a meta-analysis based on an empirical Bayesian method to robust discover differentially expressed genes in HCC and non-tumor tissues. Then, using the bioinformatics technique of weighted correlation network analysis, the highly associated prioritized Differentially Expressed Genes (DEGs) were clustered. Co-expression network and topological analysis were utilized to identify sub-clusters and confirm candidate genes. Next, a diagnostic model was developed and validated using a machine learning algorithm. To construct a prognostic model, the Cox proportional hazard regression analysis was applied and validated. We identified three genes as specific biomarkers for the diagnosis of HCC based on accuracy and feasibility. The diagnostic model's area under the curve was 0.931 with confidence interval of 0.923-0.952.•Non-fusion integrative multi-platform meta-analysis method.•Classification methods and biomarkers recognition via machine learning method.•Biomarker validation models.

Response of Skin-Derived and Metastatic Human Malignant Melanoma Cell Lines to Thymoquinone and Thymoquinone-Loaded Liposomes.

Thymoquinone has been proved to be effective against neoplasms, including skin cancer. Its high lipophilicity, however, may limit its potential use as a drug. Melanoma remains the deadliest of all skin cancers worldwide, due to its high heterogeneity, depending on the stage of the disease. Our goal was to compare the anti-cancer activity of free thymoquinone and thymoquinone-loaded liposomes on two melanoma cell lines that originated from different stages of this cancer: skin-derived A375 and metastatic WM9. We evaluated the proapoptotic effects of free thymoquinone by flow cytometry and Western blot, and its mitotoxicity by means of JC-1 assay. Additionally, we compared the cytotoxicity of free thymoquinone and thymoquinone in liposomes by WST-1 assay. Our results revealed a higher antiproliferative effect of TQ in WM9 cells, whereas its higher proapoptotic activity was observed in the A375 cell line. Moreover, the thymoquinone-loaded liposome was proved to exert stronger cytotoxic effect on both cell lines studied than free thymoquinone. Differences in the response of melanoma cells derived from different stages of the disease to thymoquinone, as well as their different responses to free and carrier-delivered thymoquinone, are essential for the development of new anti-melanoma therapies. However, further research is required to fully understand them.

Protein Abundance of Drug Transporters in Human Hepatitis C Livers.

Transmembrane drug transport in hepatocytes is one of the major determinants of drug pharmacokinetics. In the present study, ABC transporters (P-gp, MRP1, MRP2, MRP3, MRP4, BCRP, and BSEP) and SLC transporters (MCT1, NTCP, OAT2, OATP1B1, OATP1B3, OATP2B1, OCT1, and OCT3) were quantified for protein abundance (LC-MS/MS) and mRNA levels (qRT-PCR) in hepatitis C virus (HCV)-infected liver samples from the Child-Pugh class A (n = 30), B (n = 21), and C (n = 7) patients. Protein levels of BSEP, MRP3, MCT1, OAT2, OATP1B3, and OCT3 were not significantly affected by HCV infection. P-gp, MRP1, BCRP, and OATP1B3 protein abundances were upregulated, whereas those of MRP2, MRP4, NTCP, OATP2B1, and OCT1 were downregulated in all HCV samples. The observed changes started to be seen in the Child-Pugh class A livers, i.e., upregulation of P-gp and MRP1 and downregulation of MRP2, MRP4, BCRP, and OATP1B3. In the case of NTCP, OATP2B1, and OCT1, a decrease in the protein levels was observed in the class B livers. In the class C livers, no other changes were noted than those in the class A and B patients. The results of the study demonstrate that drug transporter protein abundances are affected by the functional state of the liver in hepatitis C patients.

Gene Expression and Protein Abundance of Nuclear Receptors in Human Intestine and Liver: A New Application for Mass Spectrometry-Based Targeted Proteomics.

BACKGROUND: Unwanted drug-drug interactions (DDIs), as caused by the upregulation of clinically relevant drug metabolizing enzymes and transporter proteins in intestine and liver, have the potential to threaten the therapeutic efficacy and safety of drugs. The molecular mechanism of this undesired but frequently occurring scenario of polypharmacy is based on the activation of nuclear receptors such as the pregnane X receptor (PXR) or the constitutive androstane receptor (CAR) by perpetrator agents such as rifampin, phenytoin or St. John's wort. However, the expression pattern of nuclear receptors in human intestine and liver remains uncertain, which makes it difficult to predict the extent of potential DDIs. Thus, it was the aim of this study to characterize the gene expression and protein abundance of clinically relevant nuclear receptors, i.e., the aryl hydrocarbon receptor (AhR), CAR, farnesoid X receptor (FXR), glucocorticoid receptor (GR), hepatocyte nuclear factor 4 alpha (HNF4α), PXR and small heterodimer partner (SHP), in the aforementioned organs. METHODS: Gene expression analysis was performed by quantitative real-time PCR of jejunal, ileal, colonic and liver samples from eight human subjects. In parallel, a targeted proteomic method was developed and validated in order to determine the respective protein amounts of nuclear receptors in human intestinal and liver samples. The LC-MS/MS method was validated according to the current bioanalytical guidelines and met the criteria regarding linearity (0.1-50 nmol/L), within-day and between-day accuracy and precision, as well as the stability criteria. RESULTS: The developed method was successfully validated and applied to determine the abundance of nuclear receptors in human intestinal and liver samples. Gene expression and protein abundance data demonstrated marked differences in human intestine and liver. On the protein level, only AhR and HNF4α could be detected in gut and liver, which corresponds to their highest gene expression. In transfected cell lines, PXR and CAR could be quantified. CONCLUSIONS: The substantially different expression pattern of nuclear receptors in human intestinal and liver tissue may explain the different extent of unwanted DDIs in the dependence on the administration route of drugs.

Membrane Transporters and Carriers in Human Seminal Vesicles.

Seminal vesicles play an important role in the male reproductive system, producing seminal fluid and thus adequate environment for sperm. However, mechanisms underlying secretory functions of the seminal vesicles' epithelium have not been defined yet. The aim of the present study was to characterize expression and immunolocalization of selected membrane transporters and carriers in the seminal vesicles. The study included biopsy specimens collected from non-affected parts of seminal vesicles from 53 patients of Caucasian origin subjected for prostatectomy. RT-PCR was used to define expression of 15 genes coding for ABC-family and 37 genes encoding 37 SLC-family transporters/carriers. Immunohistochemistry was used to define localization of 6 transporters. In the seminal vesicles, the following membrane transporters and carriers were defined: ABCA1, ABCB1, ABCB5, ABCB6, ABCC1, ABCC2, ABCC3, ABCC4, ABCC5, ABCC6, ABCG2, SLC01C1, SLC02B1, SLC04A1, SLC04C1, SLC10A1, SLC15A1, SLC15A2, SLC16A1, SLC16A3, SLC19A1, SLC22A1, SLC22A3, SLC22A11, SLC22A18, SLC22A4, SLC22A5, SLC28A1, SLC2A9, SLC33A1, SLC47A1, SLC47A2, SLC51A, SLC51B, SLC7A5, SLC7A6. Age-dependent expression was evidenced for ABCB1, ABCG2, SLC04C1, SLC15A1, SLC16A1, SLC22A11, SLC22A18, SLC47A1 and SLC47A2. ABCG2, P-gp, MRP1, MRP3, MCT1 and LAT1 were localized in the apical membrane and P-gp in the basolateral membrane of the seminal vesicle epithelium. The expression of the membrane transporters and carriers in the seminal vesicle epithelium confirms its secretory and barrier functions.

The reference liver-CYP450 and UGT enzymes in healthy donor and metastatic livers: the impact of genotype.

BACKGROUND: Hepatic enzymes involved in drug metabolism vary markedly in expression, abundance and activity, which affects individual susceptibility to drugs and toxicants. The present study aimed to compare mRNA expression and protein abundance of the most pharmacologically relevant drug-metabolizing enzymes in two main sources of the control liver samples that are used as the reference, i.e. organ donor livers and non-tumorous tissue from metastatic livers. An association analysis of the most common genetic variants with mRNA and protein levels was also performed. METHODS: The CYP450 and UGT enzymes (CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, CYP3A5, UGT1A1, UGT1A3, UGT2B7 and UGT2B15) were analyzed for mRNA (qPCR) and protein abundance (LC-MS/MS) in healthy donors (n = 11) and metastatic (n = 13) livers. Genotyping was performed by means of TaqMan assays and pyrosequencing. RESULTS: Significantly higher protein abundance in the metastatic livers was observed in case of CYP2C9, CYP2D6, and UGT2B7, and for UGT1A3 the difference was only significant at mRNA level. For all the enzymes except CYP2E1 some significant correlation between mRNA and protein content was observed, and for UGT1A1 an inverse correlation with age was noted. CYP2C19, CYP3A5 and CYP2D6 were significantly affected by genotype. CONCLUSION: The selection of a control group for the study on drug-metabolizing enzymes (e.g. in pathological states) may possibly affect its conclusions on differences in mRNA and protein content. Genotyping for common functional variants of CYP450 enzymes should be performed in all studies on drug-metabolizing enzymes.

Mass spectrometry-based targeted proteomics method for the quantification of clinically relevant drug metabolizing enzymes in human specimens.

Biotransformation by phase I and II metabolizing enzymes represents the major determinant for the oral bioavailability of many drugs. To estimate the pharmacokinetics, data on protein abundance of hepatic and extrahepatic tissues, such as the small intestine, are required. Targeted proteomics assays are nowadays state-of-the-art for absolute protein quantification and several methods for quantification of drug metabolizing enzymes have been published. However, some enzymes remain still uncovered by the analytical spectra of those methods. Therefore, we developed and validated a quantification assay for two carboxylesterases (CES-1, CES-2), 17 cytochrome P450 enzymes (CYP) (CYP1A1, CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C18, CYP2C19, CYP2D6, CYP2E1, CYP2J2, CYP3A4, CYP3A5, CYP3A7, CYP4F2, CYP4F12, CYP4A11) and five UDP-glucuronosyltransferases (UGTs) (UGT1A1, UGT1A3, UGT2B7, UGT2B15, UGT2B17). Protein quantification was performed by analyzing proteospecific surrogate peptides after tryptic digestion with stable isotope-labelled standards. Chromatographic separation was performed on a Kinetex® 2.6 µm C18 100 Å core-shell column (100 × 2.1 mm) with a gradient elution using 0.1% formic acid and acetonitrile containing 0.1% formic acid with a flow rate of 200 µl/min. Three mass transitions were simultaneously monitored with a scheduled multiple reaction monitoring (sMRM) method for each analyte and standard. The method was partly validated according to current bioanalytical guidelines and met the criteria regarding linearity (0.1-25 nmol/L), within-day and between-day accuracy and precision as well as multiple stability criteria. Finally, the developed method was successfully applied to determine the abundance of the aforementioned enzymes in human intestinal und liver microsomes. Our work offers a new fit for purpose method for the absolute quantification of CES, CYPs and UGTs in various human tissues and can be used for the acquisition of data for physiologically based pharmacokinetic modelling.

Hepatic drug-metabolizing enzymes and drug transporters in Wilson's disease patients with liver failure.

BACKGROUND: Wilson's disease is a genetic disorder inherited in a recessive manner, caused by mutations in the copper-transporter ATP7B. Although it is a well-known disease, currently available treatments are far from satisfactory and their efficacy varies in individual patients. Due to the lack of information about drug-metabolizing enzymes and drug transporters profile in Wilson's disease livers, we aimed to evaluate the mRNA expression and protein abundance of selected enzymes and drug transporters in this liver disorder. METHODS: We analyzed gene expression (qPCR) and protein abundance (LC-MS/MS) of 14 drug-metabolizing enzymes and 16 drug transporters in hepatic tissue from Wilson's disease patients with liver failure (n = 7, Child-Pugh class B and C) and metastatic control livers (n = 20). RESULTS: In presented work, we demonstrated a downregulation of majority of CYP450 and UGT enzymes. Gene expression of analyzed enzymes ranged between 18 and 65% compared to control group and significantly lower protein content of CYP1A1, CYP1A2, CYP2C8, CYP2C9, CYP3A4 and CYP3A5 enzymes was observed in Wilson's disease. Moreover, a general decrease in hepatocellular uptake carriers from SLC superfamily (significant at protein level for NTCP and OATP2B1) was observed. As for ABC transporters, the protein abundance of BSEP and MRP2 was significantly lower, while levels of P-gp and MRP4 transporters were significantly higher in Wilson's disease. CONCLUSIONS: Altered hepatic expression of drug-metabolizing enzymes and drug transporters in Wilson's disease patients with liver failure may result in changes of drug pharmacokinetics in that group of patients.

Arterial Blood Pressure Variability and Other Vascular Factors Contribution to the Cognitive Decline in Parkinson's Disease.

Dementia is one of the most disabling non-motor symptoms in Parkinson's disease (PD). Unlike in Alzheimer's disease, the vascular pathology in PD is less documented. Due to the uncertain role of commonly investigated metabolic or vascular factors, e.g., hypertension or diabetes, other factors corresponding to PD dementia have been proposed. Associated dysautonomia and dopaminergic treatment seem to have an impact on diurnal blood pressure (BP) variability, which may presumably contribute to white matter hyperintensities (WMH) development and cognitive decline. We aim to review possible vascular and metabolic factors: Renin-angiotensin-aldosterone system, vascular endothelial growth factor (VEGF), hyperhomocysteinemia (HHcy), as well as the dopaminergic treatment, in the etiopathogenesis of PD dementia. Additionally, we focus on the role of polymorphisms within the genes for catechol-O-methyltransferase (COMT), apolipoprotein E (APOE), vascular endothelial growth factor (VEGF), and for renin-angiotensin-aldosterone system components, and their contribution to cognitive decline in PD. Determining vascular risk factors and their contribution to the cognitive impairment in PD may result in screening, as well as preventive measures.

Synthesis and Anticancer Activity of Mitotic-Specific 3,4-Dihydropyridine-2(1H)-thiones.

Most anticancer drugs target mitosis as the most crucial and fragile period of rapidly dividing cancer cells. However the limitations of classical chemotherapeutics drive the search for new more effective and selective compounds. For this purpose structural modifications of the previously characterized pyridine aalog (S1) were incorporated aiming to obtain an antimitotic inhibitor of satisfactory and specific anticancer activity. Structure-activity relationship analysis of the compounds against a panel of cancer cell lines allowed to select a compound with a thiophene ring at C5 of a 3,4-dihydropyridine-2(1H)-thione (S22) with promising antiproliferative activity (IC50 equal 1.71 ± 0.58 µM) and selectivity (SI = 21.09) against melanoma A375 cells. Moreover, all three of the most active compounds from the antiproliferative study, namely S1, S19 and S22 showed better selectivity against A375 cells than reference drug, suggesting their possible lower toxicity and wider therapeutic index. As further study revealed, selected compounds inhibited tubulin polymerization via colchicine binding site in dose dependent manner, leading to aberrant mitotic spindle formation, cell cycle arrest and apoptosis. Summarizing, the current study showed that among obtained mitotic-specific inhibitors analogue with thiophene ring showed the highest antiproliferative activity and selectivity against cancer cells.

Identifying Differentially Expressed MicroRNAs, Target Genes, and Key Pathways Deregulated in Patients with Liver Diseases.

Liver diseases are important causes of morbidity and mortality worldwide. The aim of this study was to identify differentially expressed microRNAs (miRNAs), target genes, and key pathways as innovative diagnostic biomarkers in liver patients with different pathology and functional state. We determined, using RT-qPCR, the expression of 472 miRNAs in 125 explanted livers from subjects with six different liver pathologies and from control livers. ANOVA was employed to obtain differentially expressed miRNAs (DEMs), and miRDB (MicroRNA target prediction database) was used to predict target genes. A miRNA-gene differential regulatory (MGDR) network was constructed for each condition. Key miRNAs were detected using topological analysis. Enrichment analysis for DEMs was performed using the Database for Annotation, Visualization, and Integrated Discovery (DAVID). We identified important DEMs common and specific to the different patient groups and disease progression stages. hsa-miR-1275 was universally downregulated regardless the disease etiology and stage, while hsa-let-7a*, hsa-miR-195, hsa-miR-374, and hsa-miR-378 were deregulated. The most significantly enriched pathways of target genes controlled by these miRNAs comprise p53 tumor suppressor protein (TP53)-regulated metabolic genes, and those involved in regulation of methyl-CpG-binding protein 2 (MECP2) expression, phosphatase and tensin homolog (PTEN) messenger RNA (mRNA) translation and copper homeostasis. Our findings show a novel panel of deregulated miRNAs in the liver tissue from patients with different liver pathologies. These miRNAs hold potential as biomarkers for diagnosis and staging of liver diseases.

Effects of the IL6 -174G>C promoter polymorphism and IL-6 serum levels on the progression of cutaneous malignant melanoma.

Cutaneous malignant melanoma (CMM) is one of the most immunogenic types of cancer, with a 6-fold higher rate of spontaneous regression than any other malignancy. In addition to responsiveness to different immunotherapies, the immunogenicity of CMM highlights the important role of the host immune system in the response to CMM. The present study aimed to explore the role of two functional promoter polymorphisms [IL6 -174G>C (rs1800785) and TNFA -308G>A (rs1800629)] in the regulation of the genes encoding the pro-inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor-α, specifically in patients with CMM. A total of 76 patients with CMM and 200 control subjects were genotyped using PCR-restriction fragment length polymorphism. The genotype frequencies for both single nucleotide polymorphisms (SNPs) did not differ significantly between the patients and controls (P=0.358 and P=0.810 for IL6 and TNFA, respectively). However, compared with carriers of C-allele genotypes (CG+CC), patients with the IL6 -174GG genotype exhibited more advanced melanoma (Clark scale ≥3; P=0.037) and shorter survival times, particularly those who worked outdoors (in conditions with increased sunlight exposure; P=0.016). Furthermore, the serum IL-6 levels of patients with CMM were significantly higher than those of the control subjects, which were associated with unfavorable blood and serum characteristics and tumor progression (development of new distant metastases; P=0.004), and with a shorter overall survival time (P=0.042). Using a Cox proportional hazard model, the IL6 -174GG genotype was found to be an independent prognostic factor for reduced survival time (P=0.030), together with sex (being male; P=0.004) and occupations with higher exposure to sunlight (P=0.047). In conclusion, the results of the present study indicated that the promoter polymorphisms IL6 -174G>C and TNFA -308G>A are not predisposing factors for CMM. However, the IL6 -174G>C SNP and IL-6 serum concentrations are likely to influence the progression of the disease, and the GG genotype and higher IL-6 serum levels may indicate shorter survival.

Double-Emulsion Copolyester Microcapsules for Sustained Intraperitoneal Release of Carboplatin.

Despite on-going medical advances, ovarian cancer survival rates have stagnated. In order to improve IP delivery of platinum-based antineoplastics, we aimed to develop a sustained drug delivery system for carboplatin (CPt). Toward this aim, we pursued a double emulsion process for obtaining CPt-loaded microcapsules composed of poly(ethylene terephthalate-ethylene dilinoleate) (PET-DLA) copolymer. We were able to obtain PET-DLA microspheres in the targeted size range of 10-25 µm (median: 18.5 µm), to reduce intraperitoneal clearance by phagocytosis and lymphoid transit. Empty microspheres showed the lack of toxicity in vitro. The double emulsion process yielded 2.5% w/w CPt loading and obtained microcapsules exhibited sustained (>20 day) zero-order release. The encapsulated CPt was confirmed to be bioavailable, as the microcapsules demonstrated efficacy against human ovarian adenocarcinoma (SK-OV-3) cells in vitro. Following intraperitoneal injection in mice, we did not observe adhesions, only mild, clinically-insignificant, local inflammatory response. Tissue platinum levels, monitored over 14 days using atomic absorption spectroscopy, revealed low burst and reduced systemic uptake (plasma, kidney), as compared to neat carboplatin injection. Overall, the results demonstrate the potential of the developed microencapsulation system for long-term intraperitoneal sustained release of carboplatin for the treatment of ovarian cancer.

The reference liver - ABC and SLC drug transporters in healthy donor and metastatic livers.

BACKGROUND: Analysis of results and conclusions in studies dedicated to pathology of the liver are usually based on comparison of pathological liver specimens and control/reference (considered as healthy) tissues. There are two main sources of the control liver samples used as the reference livers, i.e. deceased organ donor livers and non-tumorous tissue from metastatic livers, which are also applied for drug transporter investigations. However, no information has yet been published on drug transporters in these two major types of reference livers. METHODS: We explored ABC (P-gp, MRP1, MRP2, MRP3, MRP4, BCRP, BSEP) and SLC (NTCP, MCT1, OCT1, OCT3, OAT2, OATP1B1, OATP1B3, OATP2B1) family transporters expression (qPCR) and protein abundance (LC-MS/MS) in healthy donors (n = 9) and metastatic (n = 13) livers. RESULTS: The analysis of mRNA content revealed significant differences in ABCB11, ABCC1, ABCG2, SLC10A1, SLC16A1, SLCO1B1 and SLCO2B1 gene expression between livers from organ donors and patients who underwent surgical resection of metastatic tumors. The protein abundance of NTCP was significantly higher, whereas of P-gp significantly lower in non-tumorous tissues from metastatic livers. Greater inter-individual variability in protein abundance of all studied transporters in subjects with metastatic colon cancer was also observed. CONCLUSIONS: The results suggest that final conclusions in liver pathology studies may depend on the reference liver tissue used, especially in gene expression studies.