Differential response of the endo-lysosomal system to chloroquine in mammary tumor cells

PEREIRA LAURA; PERALTA S; VARGAS-ROIG L; SOSA MA; CARVELLI, L.

Palmas de Mallorca

Workshop; 2nd International Workshop on Translational Cancer Research; 2021

Institut d´Investigació Illes Balears

Treatments on ER-positive breast cancer cells are more effective than on the triple-negative breast cancer subtype, where patients have a higher likelihood of recurrence and a poorest survival prognosis. Some tumoral cells have shown an increased lysosomal biogenesis, together with an altered lysosomal integrity and/or functionality, and increased levels of lysosomal proteases such as cathepsin-D (catD). In most cell types, lysosomal proteins are selectively transported from the trans-Golgi-network (TGN) to lysosomes by the mannose-6-phosphate receptors (CD-MPR and CI-MPR). Alterations in the lysosomal membrane permeability induce a release of catD into the cytoplasm, triggering apoptotic processes. Thus, lysosomes are considered as potential therapeutic targets for antitumor drugs. It is thought that acidotropic amines could accumulate in lysosomes, increasing the lysosomal membrane permeability, and leading to leakage of enzymes into the cytoplasm. The acidotropic amine chloroquine is known to affect lysosomal acidification and it is used as an adjuvant for chemotherapeutic treatments. The aim of this study was to evaluate the effect of chloroquine on the cellular distribution of catD and CD-MPR in cells derived from tumors with different grade of malignancy. MCF-7 and MDA-MB-231 cell lines were incubated either in the presence or the absence of chloroquine during 6, 12 or 24 h, and processed for catD, CD-MPR and Golgin97 (TGN marker) detection by indirect immunofluorescence and confocal microscopy. In MCF-7 cells (ER-positive), the signal for catD was decreased at 6 h of incubation with chloroquine, and a redistribution to a perinuclear area was observed, whereas the CD-MPR was mostly redistributed in the cytoplasm. This could indicate an impaired recycling of CD-MPR to TGN and an increased secretion of catD by default. In turn, the TGN appears disorganized and co-localizes with the CD-MPR. Surprisingly, after 12 h of incubation with chloroquine, catD, CD-MPR and Golgin97 returned to their initial condition denoting a loss of chloroquine effect over time. By contrast, chloroquine did not induce changes on the distribution of the proteins in MDA-MB-231 cells (triple-negative), indicating a differential response of breast tumor cells (with distinctive malignancy features) to chloroquine. This should be taken into account when using this drug as an adjuvant to antitumor drugs.