The development of multidrug resistance (MDR) is a major problem during cancer treatment. Drug efflux via ATP-binding cassette (ABC) transporters is the main mechanism responsible for resistance to chemotherapeutics. We have recently observed that statins enhance susceptibility to doxorubicin-induced apoptosis in human rhabdomyosarcoma cells, which is now also observed in human SH-SY5Y neuroblastoma cells. We have therefore investigated the ABC transporter activity to confirm a possible inhibition by statins in SH-SY5Y cells. Indeed, simvastatin directly inhibited dye transport at equimolar concentrations of the ABC transporter inhibitor, verapamil. Making use of the fluorescence behavior of doxorubicin the accumulation of anthracycline was monitored in real-time confocal microscopy. The intracellular doxorubicin accumulation was immediately enhanced by statins in SH-SY5Y cells and also in a MYCN-amplified neuroblastoma cell line STA-NB-10. The heavily glycosylated P-glycoprotein (ABCB1, P-gp) transporter appeared as a 180-and 140-kDa species. Atorvastatin and simvastatin reduced the 180-kDa form of P-gp, but not verapamil. Thereby the fully glycosylated species is shifted to the core glycosylated species (140 kDa), which was only seen at statin exposure times longer than 24 hr. The functional importance of glycosylation of the transporter was highlighted by exogenous application of N-glycosidase F, which was sufficient to enhance doxorubicin accumulation. Hence, these novel findings of statins' dual impact on P-gp have clinical implications. The enhanced intracellular accumulation of chemotherapeutics or other ABC transporter substrates in the presence of statins may represent a novel concept to overcome MDR in cancer therapy and improve drug safety.Neuroblastoma is the most common solid tumor in early childhood. About 55% of patients older than 1 year already have metastases at the time of diagnosis and therefore have a poor survival rate despite intensive therapy. 1 In $20% of the neuroblastoma a MYCN oncogene amplification occurs, which is the most unfavorable prognostic factor. 2,3 The MYCN amplification is the best established clinical and biological marker, which allows determination of the prognosis of the tumor, with numerous reports confirming the association with rapid tumor progression, advanced clinical stage and poor outcome. 3,4 Studies have shown that the expression of the multidrug resistance-associated protein 1 (MRP1, ABCC1) in neuroblastoma also correlates with poor clinical outcome and prognosis. 5,6 Recently, a prospective study has provided further evidence that high levels of MRP1 expression are strongly associated with MYCN oncogene amplification and predictive of poor outcome. 7 ATP-binding cassette (ABC) transporters, also termed multidrug resistance (MDR) proteins, play an important role in the efflux of many drugs. 8,9 Moreover, these ABC transporters are overexpressed in a wide range of tumors, for example, colon, kidney, adrenocortical or hepatocellular carcinomas, and thereby conf...

Tumours, which are initially sensitive to cytotoxic agents, often develop resistance to a broad spectrum of structurally unrelated drugs. The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have been shown to inhibit ATP-binding cassette (ABC) transporters but have also impact on glycosylation of such proteins. Doxorubicin is a substrate for ABC transporters like P-glycoprotein (ABCB1) which is present in human RD rhabdomyosarcoma cells. It was therefore the aim of this study to identify the compartmentalisation and action of doxorubicin in simvastatin-treated RD cells. Due to autofluorescence of doxorubicin, intracellular distribution was monitored by confocal microscopy. The biological effects were traced on the level of colony formation, caspase activation and DNA injury. Here we show that simvastatin treatment leads to ABCB1 inhibition and down-regulation of the transporter. Consequently, these cells accumulate significant amounts of doxorubicin, predominantly in the nucleus and lysosomes. While clearance of the anthracycline into lysosomes is not altered by simvastatin treatment, it significantly enhanced nuclear accumulation in a HMG-CoA reductase-independent manner. Thus, in such treated cells, topoisomerase II activity is significantly inhibited, which is further corroborated by augmented double-strand DNA breaks. Moreover, colony formation was synergistically inhibited by the combination of simvastatin and doxorubicin. Given the fact that ABCB1 expression correlates with an adverse prognosis in many tumours, adjuvant chemotherapy including statins might represent a novel therapeutic concept to overcome ABCB1-mediated multidrug resistance by direct inhibition and down-regulation.

Autosomal dominant familial Alzheimer's disease (AD) is associated with mutations in the AβPP, PSEN1, and PSEN2 genes. The clinical phenotype associated with AβPP mutations is mainly characterized by dementia or by strokes related to cerebral amyloid angiopathy (CAA). We present a comprehensive clinical, neuropathological, genetic, and biochemical study on a patient affected by familial AD associated with the I716F mutation in the AβPP gene. The clinical phenotype was characterized by early age of onset of 47 years, and rapidly progressive cerebellar ataxia, myoclonic jerks, rigidity, and dementia reminiscent of Creutzfeldt-Jakob disease (CJD), followed by a prolonged persistent vegetative state. Neuropathological evaluation of the proband revealed AD-related pathology but also α-synucleinopathy compatible with dementia with Lewy bodies neocortical stage or Parkinson's disease corresponding to Braak stage 6. Tau-pathology in the form of neurofibrillary degeneration corresponded to stage VI according to the Braak classification. The severe Aβ pathology included CAA, numerous plaques, and deposition of N-truncated pyroglutamate-modified Aβ peptides. Remarkably, pyroglutamate Aβ oligomers were also present intracellularly in Purkinje cells corresponding to the ataxic phenotype. The detection of a CJD-like phenotype expands the spectrum of clinical presentations associated with familial AD. Our study supports the concept that the neuropathology of familial AD expands beyond the classical AD-related pathology as defined by plaques and tangles. Finally, we provide evidence for the first time that oligomeric pyroglutamate Aβ is present in a specific pattern correlating with the clinical symptoms of a patient with AβPP I716F mutation.

The interleukin (IL)-6 inhibits the growth of early-stage melanoma cells, but not metastatic cells. Metastatic melanoma cells are susceptible to statin-induced apoptosis, but this is not clear for early-stage melanoma cells. This study aimed to investigate the IL-6 susceptibility of melanoma cells from different stages in the presence of simvastatin to overcome loss of growth arrest. ELISA was used to detect secreted IL-6 in human melanoma cells. The effects of IL-6 were measured by western blots for STAT3 and Bcl-2 family proteins. Apoptosis and proliferation were measured by caspase 3 activity, Annexin V staining, cell cycle analysis, and a wound-healing assay. Human metastatic melanoma cells A375 and 518A2 secrete high amounts of IL-6, in contrast to early-stage WM35 cells. Canonical IL-6 signaling is intact in these cells, documented by transient phosphorylation of STAT3. Although WM35 cells are highly resistant to simvastatin-induced apoptosis, coadministration with IL-6 enhanced the susceptibility to undergo apoptosis. This proapoptotic effect of IL-6 might be explained by a downregulation of Bcl-XL, observed only in WM35 cells. Furthermore, the IL-6 receptor blocking antibody tocilizumab was coadministered and unmasked an IL-6-sensitive proportion in the simvastatin-induced caspase 3 activity of metastatic melanoma cells. These results confirm that simvastatin facilitates apoptosis in combination with IL-6. Although endogenous IL-6 secretion is sufficient in metastatic melanoma cells, exogenously added IL-6 is needed for WM35 cells. This effect may explain the failure of simvastatin to reduce melanoma incidence in clinical trials and meta-analyses.

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