Artesunate Affects T Antigen Expression and Survival of Virus-Positive Merkel Cell Carcinoma.

Merkel cell carcinoma (MCC) is a rare and highly aggressive skin cancer with frequent viral etiology. Indeed, in about 80% of cases, there is an association with Merkel cell polyomavirus (MCPyV); the expression of viral T antigens is crucial for growth of virus-positive tumor cells. Since artesunate-a drug used to treat malaria-has been reported to possess additional anti-tumor as well as anti-viral activity, we sought to evaluate pre-clinically the effect of artesunate on MCC. We found that artesunate repressed growth and survival of MCPyV-positive MCC cells in vitro. This effect was accompanied by reduced large T antigen (LT) expression. Notably, however, it was even more efficient than shRNA-mediated downregulation of LT expression. Interestingly, in one MCC cell line (WaGa), T antigen knockdown rendered cells less sensitive to artesunate, while for two other MCC cell lines, we could not substantiate such a relation. Mechanistically, artesunate predominantly induces ferroptosis in MCPyV-positive MCC cells since known ferroptosis-inhibitors like DFO, BAF-A1, Fer-1 and ß-mercaptoethanol reduced artesunate-induced death. Finally, application of artesunate in xenotransplanted mice demonstrated that growth of established MCC tumors can be significantly suppressed in vivo. In conclusion, our results revealed a highly anti-proliferative effect of the approved and generally well-tolerated anti-malaria compound artesunate on MCPyV-positive MCC cells, suggesting its potential usage for MCC therapy.

SERPINB1 expression is predictive for sensitivity and outcome of cisplatin-based chemotherapy in melanoma.

Despite of highly effective new therapeutic strategies, chemotherapy still is an important treatment option in metastatic melanoma. Since predictors of chemotherapy response are rare, drugs and regimens are currently chosen arbitrarily. The present study was aimed at the identification of molecular markers predicting the outcome of chemotherapy in melanoma. Tumor biopsies from metastatic lesions were collected from 203 stage IV melanoma patients prior to chemotherapy onset and used for gene expression profiling (n = 6; marker identification set), quantitative real-time PCR (n = 127; validation set 1), and immunohistochemistry on tissue microarrays (n = 70; validation set 2). The results were correlated to the tumors' in-vitro chemosensitivity and to the patients' in-vivo chemotherapy outcome. SERPINB1 was found to correlate to the in-vitro sensitivity to cisplatin-containing chemotherapy regimens (p = 0.005). High SERPINB1 gene expression was associated with favorable tumor response (p = 0.012) and prolonged survival (p = 0.081) under cisplatin-based chemotherapy. High SERPINB1 protein expression in tumor tissue from cisplatin-treated patients was associated with a favorable survival (p = 0.011), and proved as an independent predictor of survival (p = 0.008) by multivariate analysis. We conclude, that SERPINB1 expression, although not functionally involved, is predictive for the outcome of cisplatin-based chemotherapy in melanoma, and thus may be useful to personalize melanoma chemotherapy.

Galectin-3, Renal Function, and Clinical Outcomes: Results from the LURIC and 4D Studies.

Galectin-3 has been linked to incident renal disease, experimental renal fibrosis, and nephropathy. However, the association among galectin-3, renal function, and adverse outcomes has not been described. We studied this association in two large cohorts of patients over a broad range of renal function. We measured galectin-3 concentrations in baseline samples from the German Diabetes mellitus Dialysis (4D) study (1168 dialysis patients with type 2 diabetes mellitus) and the Ludwigshafen Risk and Cardiovascular Health (LURIC) study (2579 patients with coronary angiograms). Patients were stratified into three groups: eGFR of ≥90 ml/min per 1.73 m(2), 60-89 ml/min per 1.73 m(2), and <60 ml/min per 1.73 m(2). We correlated galectin-3 concentrations with demographic, clinical, and biochemical parameters. The association of galectin-3 with clinical end points was assessed by Cox proportional hazards regression within 10 years (LURIC) or 4 years (4D) of follow-up. Mean±SD galectin-3 concentrations were 12.8±4.0 ng/ml (eGFR≥90 ml/min per 1.73 m(2)), 15.6±5.4 ng/ml (eGFR 60-89 ml/min per 1.73 m(2)), 23.1±9.9 ng/ml (eGFR<60 ml/min per 1.73 m(2)), and 54.1±19.6 ng/ml (dialysis patients of the 4D study). Galectin-3 concentration was significantly associated with clinical end points in participants with impaired kidney function, but not in participants with normal kidney function. Per SD increase in log-transformed galectin-3 concentration, the risks of all-cause mortality, cardiovascular mortality, and fatal infection increased significantly. In dialysis patients, galectin-3 was associated with the combined end point of cardiovascular events. In conclusion, galectin-3 concentrations increased with progressive renal impairment and independently associated with cardiovascular end points, infections, and all-cause death in patients with impaired renal function.

Downregulation of MHC-I expression is prevalent but reversible in Merkel cell carcinoma.

Merkel cell carcinoma (MCC) is an aggressive, polyomavirus-associated skin cancer. Robust cellular immune responses are associated with excellent outcomes in patients with MCC, but these responses are typically absent. We determined the prevalence and reversibility of major histocompatibility complex class I (MHC-I) downregulation in MCC, a potentially reversible immune-evasion mechanism. Cell-surface MHC-I expression was assessed on five MCC cell lines using flow cytometry as well as immunohistochemistry on tissue microarrays representing 114 patients. Three additional patients were included who had received intralesional IFN treatment and had evaluable specimens before and after treatment. mRNA expression analysis of antigen presentation pathway genes from 35 MCC tumors was used to examine the mechanisms of downregulation. Of note, 84% of MCCs (total n = 114) showed reduced MHC-I expression as compared with surrounding tissues, and 51% had poor or undetectable MHC-I expression. Expression of MHC-I was lower in polyomavirus-positive MCCs than in polyomavirus-negative MCCs (P < 0.01). The MHC-I downregulation mechanism was multifactorial and did not depend solely on HLA gene expression. Treatment of MCC cell lines with ionizing radiation, etoposide, or IFN resulted in MHC-I upregulation, with IFNs strongly upregulating MHC-I expression in vitro, and in 3 of 3 patients treated with intralesional IFNs. MCC tumors may be amenable to immunotherapy, but downregulation of MHC-I is frequently present in these tumors, particularly those that are positive for polyomavirus. This downregulation is reversible with any of several clinically available treatments that may thus promote the effectiveness of immune-stimulating therapies for MCC.

Type I and II IFNs inhibit Merkel cell carcinoma via modulation of the Merkel cell polyomavirus T antigens.

Merkel cell carcinoma (MCC) is a rare and highly aggressive skin cancer associated with the Merkel cell polyomavirus (MCV). As MCC cell lines show oncogene addiction to the MCV T antigens, pharmacologic interference of the large T antigen (LTA) may represent an effective therapeutic approach for this deadly cancer. In this study, we investigated the effects of IFNs on MCC cell lines, especially on MCV-positive (MCV(+)) lines. Type I IFNs (i.e., Multiferon, a mix of different IFN-α subtypes, and IFN-ß) strongly inhibited the cellular viability. Cell-cycle analysis showed increased sub-G fractions for these cells upon IFN treatment indicating apoptotic cell death; these effects were less pronounced for IFN-γ. Notably, this inhibitory effect of type I IFNs on MCV(+) MCC cell lines was associated with a reduced expression of the MCV LTA as well as an increased expression of promyelocytic leukemia (PML) protein, which is known to interfere with the function of the LTA. In addition, the intratumoral application of Multiferon resulted in a regression of MCV(+) but not MCV(-) MCCs in vivo. Together, our findings show that type I IFNs have a strong antitumor effect, which is at least in part explained by modulation of the virally encoded LTA.

Effect of pitcher age on trapping efficiency and natural prey capture in carnivorous Nepenthes rafflesiana plants.

BACKGROUND AND AIMS: Nepenthes pitchers are sophisticated traps that employ a variety of mechanisms to attract, capture and retain prey. The underlying morphological structures and physiological processes are subject to change over the lifetime of a pitcher. Here an investigation was carried out on how pitcher properties and capture efficiency change over the first 2 weeks after pitcher opening. METHODS: Prey capture, trapping efficiency, extrafloral nectar secretion, pitcher odour, as well as pH and viscoelasticity of the digestive fluid in N. rafflesiana pitchers were monitored in the natural habitat from pitcher opening up to an age of 2 weeks. KEY RESULTS: Pitchers not only increased their attractiveness over this period by becoming more fragrant and secreting more nectar, but also gained mechanical trapping efficiency via an enhanced wettability of the upper pitcher rim (peristome). Consistently, natural prey capture was initially low and increased 3-6 d after opening. It was, however, highly variable within and among pitchers. At the same time, the pH and viscoelasticity of the digestive fluid decreased, suggesting that the latter is not essential for effective prey capture. CONCLUSIONS: Prey capture and attraction by Nepenthes are dynamic processes strongly influenced by the changing properties of the pitcher. The results confirm insect aquaplaning on the peristome as the main capture mechanism in N. rafflesiana.