To examine reciprocal or unilateral implications between two cell destruction processes, autophagy and apoptosis, in 5-Fluorouracil (5-FU)-treated tumor cells, a combination of chemical inhibitors, RNAi and genetic approaches were used. In contrast to cancer cells harboring obstructed apoptosis, either at the DISC or the mitochondrial level, p53-deficiency generated signs of autophagy deregulation upon chemotherapy. On the other, hand disruption of lysosomal function by chloroquine, caused a profound decrease in apoptotic markers appearing in response to 5-FU. DR5, which is essential for 5-FU-induced apoptosis, accumulated in lysosomes and autophagosomes upon chloroquine treatment. Since neither 3-MA, RNAi of critical autophagy regulators or inhibition of cathepsins reversed apoptosis in a similar manner, it is likely that not autophagy per se but rather correct receptor transport is an important factor for 5-FU cytotoxicity. We found that apoptosis generated by TRAIL, the cognate ligand for DR5, remained unchanged upon chloroquine lysosomal interference, indicating that 5-FU activates the receptor by a discrete mechanism. In support, depletion of membrane cholesterol or hampering cholesterol transport drastically reduced 5-FU cytotoxicity. We conclude that targeting of lysosomes by chloroquine deregulates DR5 trafficking and abrogates 5-FU- but not TRAIL-stimulated cell elimination, hence suggesting a novel mechanism for receptor activation.

• Klíčová slova
• DR5, apoptosis, autophagy, chloroquine, lysosomes,
• MeSH
• apoptóza MeSH
• autofagie * MeSH
• buněčná membrána metabolismus   MeSH
• chlorochin chemie   MeSH
• cholesterol chemie   MeSH
• fagozomy MeSH
• fluoruracil chemie   MeSH
• HCT116 buňky MeSH
• lidé MeSH
• ligandy MeSH
• lyzozomy metabolismus   MeSH
• makrolidy chemie   MeSH
• mitochondrie metabolismus   MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• protein TRAIL farmakologie   MeSH
• protinádorové antimetabolity farmakologie   MeSH
• RNA interference MeSH
• signální transdukce účinky léků   MeSH
• TRAIL receptory metabolismus   MeSH
• transport proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bafilomycin A MeSH Prohlížeč
• chlorochin MeSH
• cholesterol MeSH
• fluoruracil MeSH
• ligandy MeSH
• makrolidy MeSH
• nádorový supresorový protein p53 MeSH
• protein TRAIL MeSH
• protinádorové antimetabolity MeSH
• TNFRSF10B protein, human MeSH Prohlížeč
• TP53 protein, human MeSH Prohlížeč
• TRAIL receptory MeSH

PURPOSE: DNA repair defects due to detrimental BRCA2-mutations confer increased susceptibility towards DNA interstrand-crosslinking (ICL) agents and define patient subpopulations for individualized genotype-based cancer therapy. However, due to the side effects of these drugs, there is a need to identify additional agents, which could be used alone or in combination with ICL-agents. Therefore, we investigated whether BRCA2-mutations might also increase the sensitivity towards TRAIL-receptors (TRAIL-R)-targeting compounds. EXPERIMENTAL DESIGN: Two independent model systems were applied: a BRCA2 gene knockout and a BRCA2 gene complementation model. The effects of TRAIL-R-targeting compounds and ICL-agents on cell viability, apoptosis and cell cycle distribution were compared in BRCA2-proficient versus-deficient cancer cells in vitro. In addition, the effects of the TRAIL-R2-targeting antibody LBY135 were assessed in vivo using a murine tumor xenograft model. RESULTS: BRCA2-deficient cancer cells displayed an increased sensitivity towards TRAIL-R-targeting agents. These effects exceeded and were mechanistically distinguishable from the well-established effects of ICL-agents. In vitro, ICL-agents expectedly induced an early cell cycle arrest followed by delayed apoptosis, whereas TRAIL-R-targeting compounds caused early apoptosis without prior cell cycle arrest. In vivo, treatment with LBY135 significantly reduced the tumor growth of BRCA2-deficient cancer cells in a xenograft model. CONCLUSIONS: BRCA2 mutations strongly increase the in vitro- and in vivo-sensitivity of cancer cells towards TRAIL-R-mediated apoptosis. This effect is mechanistically distinguishable from the well-established ICL-hypersensitivity of BRCA2-deficient cells. Our study thus defines a new genetic subpopulation of cancers susceptible towards TRAIL-R-targeting compounds, which could facilitate novel therapeutic approaches for patients with BRCA2-deficient tumors.

• Klíčová slova
• BRCA2, TRAIL, apoptosis, gene targeting, targeted therapy,
• MeSH
• apoptóza účinky léků   MeSH
• kontrolní body buněčného cyklu účinky léků   MeSH
• lidé MeSH
• malá interferující RNA genetika   MeSH
• myší monoklonální protilátky farmakologie   MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• oprava DNA genetika   MeSH
• proliferace buněk MeSH
• protein BRCA2 genetika   MeSH
• protinádorové látky farmakologie   MeSH
• reagencia zkříženě vázaná farmakologie   MeSH
• RNA interference MeSH
• TRAIL receptory antagonisté a inhibitory   MeSH
• viabilita buněk účinky léků   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• BRCA2 protein, human MeSH Prohlížeč
• LBY135 monoclonal antibody MeSH Prohlížeč
• malá interferující RNA MeSH
• myší monoklonální protilátky MeSH
• protein BRCA2 MeSH
• protinádorové látky MeSH
• reagencia zkříženě vázaná MeSH
• TNFRSF10B protein, human MeSH Prohlížeč
• TRAIL receptory MeSH

Glycosylation abnormalities have been observed in autoimmune diseases and cancer. Here, we compare mechanisms of aberrant O-glycosylation, i.e., formation of Tn and sialyl-Tn structures, on MUC1 in breast cancer, and on IgA1 in an autoimmune disease, IgA nephropathy. The pathways of aberrant O-glycosylation, although different for MUC1 and IgA1, include dysregulation in glycosyltransferase expression, stability, and/or intracellular localization. Moreover, these aberrant glycoproteins are recognized by antibodies, although with different consequences. In breast cancer, elevated levels of antibodies recognizing aberrant MUC1 are associated with better outcome, whereas in IgA nephropathy, the antibodies recognizing aberrant IgA1 are part of the pathogenetic process.

• MeSH
• adenokarcinom imunologie   MeSH
• glykosylace MeSH
• IgA nefropatie imunologie   MeSH
• imunoglobulin A chemie   imunologie   MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• mucin 1 chemie   imunologie   MeSH
• nádory prsu imunologie   MeSH
• polysacharidy chemie   imunologie   MeSH
• protilátky chemie   imunologie   MeSH
• prsy imunologie   MeSH
• sacharidové sekvence MeSH
• sekvence aminokyselin MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• imunoglobulin A MeSH
• mucin 1 MeSH
• polysacharidy MeSH
• protilátky MeSH