טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentZhitomirsky Binyamin
SubjectThe Role of Lysosomes in Cancer Multidrug Resistance
DepartmentDepartment of Biology
Supervisor Professor Yehuda Assaraf
Full Thesis textFull thesis text - English Version


Abstract

Cancer multidrug resistance (MDR), a phenomenon in which cancer cells present with an intrinsic chemoresistant phenotype or become irresponsive to cytotoxic drug treatment following chemotherapy, remains one of the leading causes of treatment failure and consequent dismal prognosis of cancer patients. Deciphering the underlying mechanisms of MDR has been a major goal of cancer research over the last Jubilee. Multiple molecular mechanisms by which cancer cells acquire drug resistant phenotypes have been described. These include reduced drug uptake, active drug efflux from the cytoplasm or the cell membrane, qualitative and/or quantitative alterations in drug targets, drug compartmentalization, drug inactivation via metabolism and anti-apoptotic mechanisms.

Herein we explored the role of lysosomes as mediators of cancer MDR. Due to their acidic lumen, lysosomes have been known to accumulate drugs with hydrophobic weak base properties via a mechanism known as ion trapping. These lysosomotropic drugs travel via passive diffusion across the cell membrane and the lysosomal membrane. However, once in lysosomes, they become readily protonated and thus sequestered in the acidic lumen of lysosomes. Lysosomal sequestration of cytotoxic anticancer drugs has been suggested to contribute to cancer drug resistance by acting as a sink that markedly reduces the effective concentration of the drug at its target site and thus preventing it from exerting its cytotoxic activity. In this respect, we herein demonstrate that cancer cells with an intrinsically higher number of lysosomes per cell, are able to sequester lysosomotropic anticancer drugs more efficiently in their lysosomes, thereby efficiently reducing the cytotoxic drug concentration at their intracellular target sites. We demonstrate the key importance of the pH gradient between the cytosol and the lysosome for efficient lysosomal drug sequestration. We further show that lysosomal accumulation of lysosomotropic anticancer drugs triggers the activation and nuclear translocation of the master transcription factor regulator TFEB, triggering the transcriptional activation of the CLEAR pathway and activation of lysosomal biogenesis, which results in an elevation in lysosome number per cell. Based on these findings we propose a novel model for drug-induced lysosome-mediated cancer MDR, in which lysosomal accumulation of lysosomotropic anticancer drugs activates TFEB-mediated lysosomal biogenesis, resulting in an elevated lysosome number per cell, thus facilitating a more efficient lysosomal sequestration of anticancer drugs away from their cellular target sites.

We next determined the fate of anticancer drugs sequestered in lysosomes. We demonstrate that

lysosomotropic drugs induce lysosomal exocytosis via active translocation of lysosomes towards the plasma membrane on microtubule tracks, followed by fusion of the lysosomal membrane with the plasma membrane and the consequent release of the lysosomal content into the extracellular milieu. We propose that this drug-induced activation of lysosomal exocytosis can serve as an important line of resistance to chemotherapeutic drug treatment, in which the drug is not only sequestered in the lysosome but also extruded from the cell via lysosomal exocytosis. We discuss the implications of chemotherapy-induced lysosomal exocytosis on the secretion of several lysosomal enzymes that have been shown to partake in key malignant processes such as tumor invasion and metastasis.