Oxime-Linked Peptide-Daunomycin Conjugates as Good Tools for Selection of Suitable Homing Devices in Targeted Tumor Therapy: An Overview.

Chemotherapy is still one of the main therapeutic approaches in cancer therapy. Nevertheless, its poor selectivity causes severe toxic side effects that, together with the development of drug resistance in tumor cells, results in a limitation for its application. Tumor-targeted drug delivery is a possible choice to overcome these drawbacks. As well as monoclonal antibodies, peptides are promising targeting moieties for drug delivery. However, the development of peptide-drug conjugates (PDCs) is still a big challenge. The main reason is that the conjugates have to be stable in circulation, but the drug or its active metabolite should be released efficiently in the tumor cells. For this purpose, suitable linker systems are needed that connect the drug molecule with the homing peptide. The applied linker systems are commonly categorized as cleavable and non-cleavable linkers. Both the groups possess advantages and disadvantages that are summarized briefly in this manuscript. Moreover, in this review paper, we highlight the benefit of oxime-linked anthracycline-peptide conjugates in the development of PDCs. For instance, straightforward synthesis as well as a conjugation reaction proceed in excellent yields, and the autofluorescence of anthracyclines provides a good tool to select the appropriate homing peptides. Furthermore, we demonstrate that these conjugates can be used properly in in vivo studies. The results indicate that the oxime-linked PDCs are potential candidates for targeted tumor therapy.

Neutrophil-to-Lymphocyte Ratio Is an Independent Risk Factor for Coronary Artery Disease in Central Obesity.

Several inflammatory biomarkers were found to be associated with an increased risk of cardiovascular disease. Neutrophil-to-lymphocyte ratio (NLR) is a marker of subclinical inflammation that increases with the stress response. Visceral adiposity index (VAI) calculated as a combination of anthropometric and metabolic parameters reflects both the extent and function of visceral adipose tissue. Given the association of subclinical inflammation with both obesity and cardiovascular diseases, it is plausible that the inflammation-CVD association is modulated by the amount and function of adipose tissue. Thus, our aim was to examine the association between NLR and coronary artery calcium score (CACS), an intermediate marker of coronary artery disease in asymptomatic patients across VAI tertiles. Methods: Data from 280 asymptomatic participants of a cardiovascular screening program were analysed. In addition to the collection of lifestyle and medical history, a non-contrast cardiac CT scan and laboratory tests were performed on all participants. Multivariate logistic regression was conducted with CACS > 100 as the outcome and with conventional cardiovascular risk factors and NLR, VAI, and NLR by VAI tertile as predictors. Results: We found an interaction between VAI tertiles and NLR; NLR values were similar in the lower VAI tertiles, while they were higher in the CACS > 100 in the 3rd VAI tertile (CACS ≤ 100: 1.94 ± 0.58 vs. CACS > 100: 2.48 ± 1.1, p = 0.008). According to multivariable logistic regression, the interaction between NLR and VAI tertiles remained: NLR was associated with CACS > 100 in the 3rd VAI tertile (OR = 1.67, 95% CI 1.06-2.62, p = 0.03) but not in the lower tertiles even after adjustment for age, sex, smoking, history of hypertension, hyperlipidaemia, and diabetes mellitus, as well as high-sensitivity C-reactive protein. Our findings draw attention to the independent association between subclinical, chronic, systemic inflammation and subclinical coronary disease in obesity.

Association between coronary atherosclerosis and visceral adiposity index.

BACKGROUND AND AIMS: Visceral obesity is a marker of dysfunctional adipose tissue and ectopic fat infiltration. Many studies have shown that visceral fat dysfunction has a close relationship with cardiovascular disease. For a better identification of visceral adiposity dysfunction, the visceral adiposity index (VAI) is used. Coronary artery calcium score (CACS) is known to have a strong correlation with the total plaque burden therefore provides information about the severity of the coronary atherosclerosis. CACS is a strong predictor of cardiac events and it refines cardiovascular risk assessment beyond conventional risk factors. Our aim was to evaluate the association between VAI and CACS in an asymptomatic Caucasian population. METHODS AND RESULTS: Computed tomography scans of 460 participants were analyzed in a cross-sectional, voluntary screening program. A health questionnaire, physical examination and laboratory tests were also performed. Participants with a history of cardiovascular disease were excluded from the analysis. Mean VAI was 1.41 ± 0.07 in men and 2.00 ± 0.15 in women. VAI showed a positive correlation with total coronary calcium score (r = 0.242) in males but not in females. VAI was stratified into tertiles by gender. In males, third VAI tertile was independently associated with CACS>100 (OR: 3.21, p = 0.02) but not with CACS>0 after the effects of conventional risk factors were eliminated. CONCLUSION: VAI tertiles were associated with calcium scores and the highest VAI tertile was an independent predictor for the presence of CACS>100 in males but not in females.

Serum Uric Acid Is Independently Associated with Coronary Calcification in an Asymptomatic Population.

Detecting early-stage atherosclerosis is an important step towards cardiovascular disease prevention. Coronary artery calcium (CAC) score is a sensitive and non-invasive tool for detecting coronary atherosclerosis. Higher serum uric acid (SUA) levels are known to be associated with cardiovascular diseases. However, there is inconsistency regarding the independence of the association. The aim of our study was to assess the association of CAC and SUA in an asymptomatic population. CAC scans of 281 participants were analyzed in a voluntary screening program. A health questionnaire, physical examination, and laboratory tests were also performed. Participants with a history of cardiovascular disease were excluded from the analysis. 36.3% (n = 102) of the participants had no detectable CAC and 13.9% (n = 39) had a CAC score of > 300. SUA showed positive correlation with CAC score (0.175, p < 0.01). SUA was independently associated with Ca score > 300 (OR 5.17, p = 0.01) after the effects of conventional risk factors were eliminated.

Drug targeting to decrease cardiotoxicity - determination of the cytotoxic effect of GnRH-based conjugates containing doxorubicin, daunorubicin and methotrexate on human cardiomyocytes and endothelial cells.

Background: Cardiomyopathy induced by the chemotherapeutic agents doxorubicin and daunorubicin is a major limiting factor for their application in cancer therapy. Chemotactic drug targeting potentially increases the tumor selectivity of drugs and decreases their cardiotoxicity. Increased expression of gonadotropin-releasing hormone (GnRH) receptors on the surface of tumor cells has been reported. Thus, the attachment of the aforementioned chemotherapeutic drugs to GnRH-based peptides may result in compounds with increased therapeutic efficacy. The objective of the present study was to examine the cytotoxic effect of anticancer drug-GnRH-conjugates against two essential cardiovascular cell types, such as cardiomyocytes and endothelial cells. Sixteen different previously developed GnRH-conjugates containing doxorubicin, daunorubicin and methotrexate were investigated in this study. Their cytotoxicity was determined on primary human cardiac myocytes (HCM) and human umbilical vein endothelial cells (HUVEC) using the xCELLigence SP system, which measures impedance changes caused by adhering cells on golden electrode arrays placed at the bottom of the wells. Slopes of impedance-time curves were calculated and for the quantitative determination of cytotoxicity, the difference to the control was analysed. Results: Doxorubicin and daunorubicin exhibited a cytotoxic effect on both cell types, at the highest concentrations tested. Doxorubicin-based conjugates (AN-152, GnRH-III(Dox-O-glut), GnRH-III(Dox-glut-GFLG) and GnRH-III(Dox=Aoa-GFLG) showed the same cytotoxic effect on cardiomyocytes. Among the daunorubicin-based conjugates, [4Lys(Ac)]-GnRH-III(Dau=Aoa), GnRH-III(Dau=Aoa-YRRL), {GnRH-III(Dau=Aoa-YRRL-C)}2 and {[4N-MeSer]-GnRH-III(Dau-C)}2 had a significant but decreased cytotoxic effect, while the other conjugates - GnRH-III(Dau=Aoa), GnRH-III(Dau=Aoa-K(Dau=Aoa)), [4Lys(Dau=Aoa)]-GnRH-III(Dau=Aoa), GnRH-III(Dau=Aoa-GFLG), {GnRH-III(Dau-C)}2 and [4N-MeSer]-GnRH-III(Dau=Aoa) - exerted no cytotoxic effect on cardiomyocytes. Mixed conjugates containing methotrexate and daunorubicin - GnRH-III(Mtx-K(Dau=Aoa)) and [4Lys(Mtx)]-GnRH-III(Dau=Aoa) - showed no cytotoxic effect on cardiomyocytes, as well. Conclusion: Based on these results, anticancer drug-GnRH-based conjugates with no cytotoxic effect on cardiomyocytes were identified. In the future, these compounds could provide a more targeted antitumor therapy with no cardiotoxic adverse effects. Moreover, impedimetric cytotoxicity analysis could be a valuable technique to determine the effect of drugs on cardiomyocytes.

Functional cyclophilin D moderates platelet adhesion, but enhances the lytic resistance of fibrin.

In the course of thrombosis, platelets are exposed to a variety of activating stimuli classified as 'strong' (e.g. thrombin and collagen) or 'mild' (e.g. ADP). In response, activated platelets adhere to injured vasculature, aggregate, and stabilise the three-dimensional fibrin scaffold of the expanding thrombus. Since 'strong' stimuli also induce opening of the mitochondrial permeability transition pore (MPTP) in platelets, the MPTP-enhancer Cyclophilin D (CypD) has been suggested as a critical pharmacological target to influence thrombosis. However, it is poorly understood what role CypD plays in the platelet response to 'mild' stimuli which act independently of MPTP. Furthermore, it is unknown how CypD influences platelet-driven clot stabilisation against enzymatic breakdown (fibrinolysis). Here we show that treatment of human platelets with Cyclosporine A (a cyclophilin-inhibitor) boosts ADP-induced adhesion and aggregation, while genetic ablation of CypD in murine platelets enhances adhesion but not aggregation. We also report that platelets lacking CypD preserve their integrity in a fibrin environment, and lose their ability to render clots resistant against fibrinolysis. Our results indicate that CypD has opposing haemostatic roles depending on the stimulus and stage of platelet activation, warranting a careful design of any antithrombotic strategy targeting CypD.

Signs of subclinical atherosclerosis in asymptomatic patients at increased risk of type 2 diabetes mellitus.

AIMS: We aimed to study carotid intima media thickness (CIMT) in asymptomatic patients with an increased risk of type 2 diabetes mellitus (T2DM) and in a pre-diabetic state. METHODS: Diabetes risk assessment was performed in 2420 participants in a voluntary screening program between 2011 and 2013. The risk of T2DM was estimated by the Findrisc scoring system (FR). A FR≥12 was considered as increased risk. HbA1c% between 5.7 and 6.4% signified a pre-diabetic state. Carotid duplex scan was performed and CIMT above 0.9 mm was regarded as pathological. Patients with T2DM or a history of cardiovascular disease were excluded. RESULTS: Overall 1475 subjects were included. Four groups were compared: "control" (normal HbA1c, FR<12), "HbA1c only" (HbA1c: 5.7-6.4%, FR<12), "Findrisc only" (normal HbA1c, FR≥12) and "combined" (HbA1c: 5.7-6.4%, FR≥12). Frequency of pathological maximal CIMT was 9.4%, 19.7%, 27.4% and 36.4% in the groups, respectively (p<0.001). Logistic regression analysis revealed that compared to control subjects, sex and risk factor-adjusted Odds Ratios for the presence of pathological maximal CIMT were 2.2 (p<0.001), 3.4 (p<0.001) and 5.1 (p<0.001) for the groups, respectively. CONCLUSIONS: Evaluation of Findrisc score and HbA1c at population level may facilitate early recognition of subclinical vascular complications even in the pre-diabetic state.

Impedimetric Analysis of the Effect of Decellularized Porcine Heart Scaffold on Human Fibrosarcoma, Endothelial, and Cardiomyocyte Cell Lines.

BACKGROUND Experiments on porcine heart scaffold represent significant assays in development of immunoneutral materials for cardiac surgery. Characterization of cell-cell and cell-scaffold interactions is essential to understand the homing process of cardiac cells into the scaffolds. MATERIAL AND METHODS In the present study, the highly sensitive and real-time impedimetric technique of xCELLigence SP was used to monitor cell adhesion, which is the key process of recellularization in heart scaffolds. Our objectives were: (i) to characterize the effect of decellularized porcine heart scaffold on cell adhesion of human cardiovascular cells potentially used in the recellularization process; and (ii) to investigate cell-extracellular matrix element interactions for building artificial multi-layer systems, applied as cellular models of recellularization experiments. Human fibrosarcoma, endothelial, and cardiomyocyte cells were investigated and the effect of decellularized porcine heart scaffold (HS) and fibronectin on cell adhesion was examined. Adhesion was quantified as slope of curves. RESULTS Heart scaffold had neutral effect on cardiomyocytes as well as on endothelial cells. Adhesion of cardiomyocytes was increased by fibronectin (1.480±0.021) compared to control (0.745±0.029). The combination of fibronectin and HS induced stronger adhesion of cardiomyocytes (2.407±0.634) than fibronectin alone. Endothelial and fibrosarcoma cells showed similarly strong adhesion profiles with marked enhancer effect by fibronectin. CONCLUSIONS Decellularized porcine HS does not inhibit adhesion of human cardiovascular cells at the cell biological level, while fibronectin has strong cell adhesion-inducer effect, as well as an enhancer effect on activity of HS. Consequently, decellularized porcine hearts could be used as scaffolds for recellularization with cardiomyocytes and endothelial cells with fibronectin acting as a regulator, leading to construction of working bioartificial hearts.

[Cardiovascular screening programme in the Central Hungarian region. The Budakalász Study]. / Cardiovascularis szuroprogram a közép-magyarországi régióban. Budakalász Vizsgálat.

INTRODUCTION: The reduction in mortality due to prevention programmes observed in some European countries is not currently reached in Hungary. Effective prevention is based on the screening of risk factors and health state of the population. AIM: The goal of this study was to develop a longitudinal, population-based screening programme in the Central Hungarian region in order to collect information on the health state and cardiovascular risk profile of the citizens and discover new potential cardiovascular risk factors. METHOD: The Budakalász Study is a self-voluntary programme involving the adult population (>20 yrs, approx. 8000 persons), and it consists of questionnaires, non-invasive tests (anthropometry, cardiac echo, carotid duplex scan, blood pressure measurement, ankle-brachial index), venous blood sample collection and laboratory tests. RESULTS: Until January, 2014, 2420 persons (30% of the population, male: 41.2%, average age 54.8 years) participated in the programme. Cardiovascular morbidity was higher in contrast to a former national survey. The number of risk factors and, therefore, 10-year cardiovascular risk were also elevated in this population. CONCLUSIONS: These findings underline the importance of screening programmes and effective therapies.

Basic cell physiological activities (cell adhesion, chemotaxis and proliferation) induced by selegiline and its derivatives in Mono Mac 6 human monocytes.

Selegiline (R-deprenyl), a monoamine oxidase-B (MAO-B) inhibitor, has complex pharmacological effect that contributes to treatment of neurodegenerative diseases such as Parkinson's and presumably Alzheimer's disease and might work as an inhibitor of tumor growth. In respect of tumorigenesis and metastasis formation, the controlled modifications of adhesion and migration have high therapeutic significance. In the present study, our purpose was to investigate cell physiological responses (adhesion, chemotaxis and proliferation) induced by selegiline, its metabolites and synthetic derivatives and to find some correlations between the molecular structure and the reported antitumor behavior of the derivatives. Our results demonstrated that both R- and S-deprenyls have the potency to elicit increased adhesion and a chemorepellent activity in monocyte model (Mono Mac 6 cell line derived from monoblastic leukemia); however, only the R-enantiomer proved to be cytotoxic. Among the metabolites R-amphetamine has retained the adhesion inducer and the chemorepellent effect of the parent drug on the most significant level. In contrast, a reversed chemotactic effect and an improved cytotoxic character were detected in the presence of fluoro group (p-fluoro-S-deprenyl). In summary, the adhesion inducer activity, chemorepellent and advantageous cytotoxic effects of selegiline and some derivatives indicate that these drug molecules might have inhibitory effects in metastasis formation in primary tumors.