Chemotherapy drug response in ovarian cancer cells strictly depends on a cathepsin D-Bax activation loop.

The ovarian cancer cell lines A2780 (wild-type p53) and NIHOVCAR3 (mutated p53) showed, respectively, sensitivity and resistance towards several chemotherapy drugs. We hypothesized that the two cell lines differ in their ability to activate the intrinsic death pathway and have, therefore, dissected the lysosome-mitochondrion signalling pathway by pharmacological inhibition or genetic manipulation of key regulators and executioners. Biochemical and morphological confocal fluorescence studies showed that: (1) In A2780 cells bcl-2 is expressed at an undetectable level, whereas Bax is expressed at a rather high level; by contrast, bcl-2 is highly expressed and Bax is expressed at extremely low levels in NIHOVCAR3 cells; (2) Chemotherapy treatment reduced the expression of bcl-2 in NIHOVCAR3 cells, yet these cells resisted to drug toxicity; (3) Cathepsin D (CD), not cathepsin B or L, mediates the activation of the mitochondrial intrinsic death pathway in A2780 cells; (4) Lysosome leakage and cytosolic relocation of CD occurs in the chemosensitive A2780 cells, not in the chemoresistant NIHOVCAR3 cells; (5) Bax is essential for the permeabilization of both lysosomes and mitochondria in A2780 cells exposed to chemotherapy drugs; (6) CD activity is mandatory for the oligomerization of Bax on both mitochondrial and lysosomal membranes; (7) Bax activation did not occur in the resistant NIHOVCAR3 cells despite their high content in CD.

Prolactin promotes the secretion of active cathepsin D at the basal side of rat mammary acini.

Cathepsin D (CD), a lysosomal aspartic protease present in mammary tissue and milk in various molecular forms, is also found in the incubation medium of mammary acini in molecular forms that are proteolytically active on prolactin at a physiological pH. Because prolactin controls the vesicular traffic in mammary cells, we studied, in vivo and in vitro, its effects on the polarized transport and secretion of various forms of CD in the rat mammary gland. CD accumulated in vesicles not involved in endocytosis in the basal region of cells.

High yield synthesis and characterization of phosphorylated recombinant human procathepsin D expressed in mammalian cells.

We used a vaccinia virus expression system for the production of recombinant human cathepsin D (CD), a lysosomal protease implicated in various patho-physiological processes including cancer, neurodegeneration, and development. The recombinant protein was successfully expressed in various human and non-human cells. It was correctly synthesized as a glycosylated 53 kDa precursor (proCDrec) that reacted with a polyclonal antibody against residues 7-21 of the propeptide sequence.

Preconditioning-induced cytoprotection in hepatocytes requires Ca(2+)-dependent exocytosis of lysosomes.

A short period of hypoxia reduces the cytotoxicity produced by a subsequent prolonged hypoxia in isolated hepatocytes. This phenomenon, termed hypoxic preconditioning, is mediated by the activation of adenosine A2A-receptor and is associated with the attenuation of cellular acidosis and Na+ overload normally occurring during hypoxia. Bafilomycin, an inhibitor of the vacuolar H+/ATPase, reverts the latter effects and abrogates the preconditioning-induced cytoprotection.

Destination 'lysosome': a target organelle for tumour cell killing?

Lysosomes and lysosome-related organelles constitute a system of acid compartments that interconnect the inside of the cell with the extracellular environment via endocytosis, phagocytosis and exocytosis. In recent decades it has been recognized that lysosomes are not just wastebaskets for disposal of unused cellular constituents, but that they are involved in several cellular processes such as post-translational maturation of proteins, degradation of receptors and extracellular release of active enzymes. By complementing the autophagic process, lysosomes actively contribute to the maintenance of cellular homeostasis.

Lysosomal proteases as potential targets for the induction of apoptotic cell death in human neuroblastomas.

Neuroblastoma is the most common type of cancer in infants. In children this tumor is particularly aggressive; despite various new therapeutic approaches, it is associated with poor prognosis. Given the importance of endosomal-lysosomal proteolysis in cellular metabolism, we hypothesized that inhibition of lysosomal protease would impact negatively on neuroblastoma cell survival.